Most Common Erythroid Metabolic Inherited Diseases. BM transplantation and gene therapy approaches
\r\n\tAnimal food additives are products used in animal nutrition for purposes of improving the quality of feed or to improve the animal’s performance and health. Other additives can be used to enhance digestibility or even flavour of feed materials. In addition, feed additives are known which improve the quality of compound feed production; consequently e.g. they improve the quality of the granulated mixed diet.
\r\n\r\n\tGenerally feed additives could be divided into five groups:
\r\n\t1.Technological additives which influence the technological aspects of the diet to improve its handling or hygiene characteristics.
\r\n\t2. Sensory additives which improve the palatability of a diet by stimulating appetite, usually through the effect these products have on the flavour or colour.
\r\n\t3. Nutritional additives, such additives are specific nutrient(s) required by the animal for optimal production.
\r\n\t4.Zootechnical additives which improve the nutrient status of the animal, not by providing specific nutrients, but by enabling more efficient use of the nutrients present in the diet, in other words, it increases the efficiency of production.
\r\n\t5. In poultry nutrition: Coccidiostats and Histomonostats which widely used to control intestinal health of poultry through direct effects on the parasitic organism concerned.
\r\n\tThe aim of the book is to present the impact of the most important feed additives on the animal production, to demonstrate their mode of action, to show their effect on intermediate metabolism and heath status of livestock and to suggest how to use the different feed additives in animal nutrition to produce high quality and safety animal origin foodstuffs for human consumer.
",isbn:"978-1-83969-404-2",printIsbn:"978-1-83969-403-5",pdfIsbn:"978-1-83969-405-9",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,hash:"8ffe43a82ac48b309abc3632bbf3efd0",bookSignature:"Prof. László Babinszky",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/10496.jpg",keywords:"Technological Feed Additives, Feed Industry, Quality of Compound Feed, Non-Antibiotic Growth Promoter, Product Quality, Additive Enzymes, Digestibility of Nutrients, NSP Enzymes, Farm Animals, Livestock, Immunity, Microbiome",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:null,numberOfDimensionsCitations:null,numberOfTotalCitations:null,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"November 24th 2020",dateEndSecondStepPublish:"December 22nd 2020",dateEndThirdStepPublish:"February 20th 2021",dateEndFourthStepPublish:"May 11th 2021",dateEndFifthStepPublish:"July 10th 2021",remainingDaysToSecondStep:"2 months",secondStepPassed:!0,currentStepOfPublishingProcess:4,editedByType:null,kuFlag:!1,biosketch:"Professor Emeritus from the University of Debrecen, Hungary who authored 297 publications (papers, book chapters) and edited 3 books. Member of various committees and chairman of the World Conference of Innovative Animal Nutrition and Feeding (WIANF).",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"53998",title:"Prof.",name:"László",middleName:null,surname:"Babinszky",slug:"laszlo-babinszky",fullName:"László Babinszky",profilePictureURL:"https://mts.intechopen.com/storage/users/53998/images/system/53998.jpg",biography:"László Babinszky is Professor Emeritus of animal nutrition at the University of Debrecen, Hungary. From 1984 to 1985 he worked at the Agricultural University in Wageningen and in the Institute for Livestock Feeding and Nutrition in Lelystad (the Netherlands). He also worked at the Agricultural University of Vienna in the Institute for Animal Breeding and Nutrition (Austria) and in the Oscar Kellner Research Institute in Rostock (Germany). From 1988 to 1992, he worked in the Department of Animal Nutrition (Agricultural University in Wageningen). In 1992 he obtained a PhD degree in animal nutrition from the University of Wageningen.He has authored 297 publications (papers, book chapters). He edited 3 books and 14 international conference proceedings. His total number of citation is 407. \r\nHe is member of various committees e.g.: American Society of Animal Science (ASAS, USA); the editorial board of the Acta Agriculturae Scandinavica, Section A- Animal Science (Norway); KRMIVA, Journal of Animal Nutrition (Croatia), Austin Food Sciences (NJ, USA), E-Cronicon Nutrition (UK), SciTz Nutrition and Food Science (DE, USA), Journal of Medical Chemistry and Toxicology (NJ, USA), Current Research in Food Technology and Nutritional Sciences (USA). 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Clinical trials performed during last two decades have demonstrated its usefulness in the treatment of several genetic diseases [1] but also the need to improve vector delivery, expression and safety [2]. New vectors should reduce genotoxicity (genomic alteration due to vector integration), immunogenicity (immune response to gene delivery vectors and/or trangenes) and cytotoxicity (induced by ectopic expression and/or overexpression of the transgene).
In mature erythrocytes, most metabolic needs are covered by glycolysis, oxidative pentose phosphate pathway and glutathione cycle. Hereditary enzyme deficiencies of all these pathways have been identified, being most of them associated with chronic non-spherocytic hemolitic anemia (CNSHA). Hereditary hemolytic anemia exhibits a high molecular heterogeneity with a wide number of different mutations involved in the structural genes of nearly all affected enzymes. Deficiency in metabolic enzymes impairs energy balance in the erythrocytes, with or without changes in oxygen affinity of hemoglobin and delivery to the tissues. Despite of having a better understanding of their molecular basis, definitive curative therapy for Red Blood Cells (RBC) enzyme defects still remains undeveloped.
Conventional bone marrow transplantation allows the generation of donor-derived functional hematopoietic cells of all lineages in the host, and represents the standard of care or at least a valid therapeutic option for many inherited diseases [3]. However, complications associated to allogeneic transplantation can be as severe as the enzymatic deficiency. The recessive inheriting trait of most of these metabolic diseases and the confined enzymatic defect to the hematopoietic/erythropoietic system, make them suitable diseases to be treated by gene therapy. Correction by gene therapy requires the stable transfer of a functional gene into the autologous self-renewing Hematopoietic stem cells (HSCs) and their mature progeny. Autologous BM transplantation of genetically corrected cells shows several advantages over the allogeneic procedure. First, it overcomes the limitation of human leukocyte antigen (HLA)-compatible donor availability, so it can be applied to every patient. Second, the reduction of morbidity and mortality associated with the transplant procedure, as there is no risk of graft versus host disease (GvHD) and consequently no need for post-transplant immunosuppression.
To date, gene therapy approaches for the treatment of inherited metabolic deficiencies are still limited, mainly because of the frequent lack of selective advantage of genetically corrected cells. This implies that high levels of transgene expression are required, as well as an efficient transduction of HSCs. This requirement have already been described in different RBC diseases as in the erytropoietic protoporphyria (EPP) [4] caused by the deficiency of the last enzyme of the heme biosintesis pathway or in the piruvate kinase deficiency (PKD) [3], where there is an impairment in the final yield of ATP in RBC. Additionally, some RBC pathologies require switching on expression of the transgene at only the proper stage of differentiation, which represents another challenge in the development of new gene therapy protocols.
Although more than 14 metabolic deficiencies have been identified causing CNSHA, approaches of gene therapy have been done only in a few of them (Table 1). Below, we are including a short description of the different diseases and the attempts addressed.
Among glycolytic defects causing CNSHA, Glucose 6-phosphate dehydrogenase (G6PD) deficiency is the most common genetic disease. More than 400 million people are affected world wide, showing a vast variability of clinical features. G6PD catalyzes the first reaction of the pentose phosphate pathway, in which Glucose 6-phosphate (G6P) is oxidized and Nicotinamide adenine dinucleotide phosphate is reduced (NADPH) resulting in decarboxylation of CO2 and pentose phosphate. G6PD plays a central role in the cellular physiology as it is the major source of NADPH, required by many essential cellular systems including the antioxidant pathways, nitric oxide synthase, NADPH oxidase, cytochrome p450 system and others. Indeed, G6PD is essential for cell survival. G6PD is a 20 kb X-linked gene that maps to the Xq28 region, consisting of 13 exons and 12 introns, which encode a 514 amino acids protein with ubiquitous expression. More than 100 missense mutations in the G6PD gene have been identified [14], being most of them single-point mutations causing an amino acid substitution. Frequently, these mutations cause mild symptoms or no disease, except when patients are challenged by increased oxidative stress or fava beans. However, some mutations provoke severe instability of the G6PD and, as a result, lifelong CNSHA with a variable severity [15,16]. Through genetic studies it has been observed that severe clinical manifestations appear preferentially in exons 7, 10 and 11. As G6PD is X-linked, the defect is fully expressed in affected males (hemizygotes who inherit the mutation only from the mother), whereas in homozygous females the mutations are transmitted from both parents. Thereby, female heterozygotes represent a red blood cell mosaic population, causing a wide range clinical picture.
\n\t\t\t\tDisease\n\t\t\t | \n\t\t\t\n\t\t\t\tGene\n\t\t\t | \n\t\t\t\n\t\t\t\tChrom.\n\t\t\t | \n\t\t\t\n\t\t\t\tInheritance\n\t\t\t | \n\t\t\t\n\t\t\t\tOther sympthoms\n\t\t\t | \n\t\t\t\n\t\t\t\tBone Marrow Transplantation\n\t\t\t | \n\t\t\t\n\t\t\t\tGene Therapy\n\t\t\t | \n\t\t
Glucose-6 Phosphate Dehydrogenase Deficiency (G6PD) | \n\t\t\t\n\t\t\t\tG6PD\n\t\t\t | \n\t\t\tXq28 | \n\t\t\tX-linked | \n\t\t\tjaundice, spleno- and hepatomegaly, hemoglobinuria, leukocyte disfunction, and susceptibility to infections | \n\t\t\t\n\t\t\t | D: C57BL/6 mice P: Transduction of 5-FU treated BM cells with MMLV-hG6PD or MPSV-hG6PD vectors and subsequent transplantation. R: lethally irradiated C57BL/6 mice [5] | \n\t\t
Pyruvate Kinase Deficiency (PKD) | \n\t\t\t\n\t\t\t\tPKLR\n\t\t\t | \n\t\t\t1q21 | \n\t\t\tA.R | \n\t\t\tReticulocytosis, splenomegaly, hidrops foetalis, and death in neonatal period | \n\t\t\tD: normal CBA/N+/+ mice + 5FU R: CBA Pk-1slc/ PK-1slc mice C: minimal (100 or 400 cGy) [6] | \n\t\t\tD: WT mice P: Transduction of 5-FU treated BM cells with pMNSM-hLPK retroviral vector and subsequent transplantation R: lethally irradiated mice [7] | \n\t\t
D: normal CBA/N+/+ mice R: CBA Pk-1slc/ PK-1slc mice C: no conditioning [8] | \n\t\t\tD: CBA PK-1slc/PK-1slc mice P: Transgenic rescue using the μLCR-PKLR-hRPK construct [9] | \n\t\t|||||
D: normal Basenji dogs R: PKD Basenji dogs C: sublethal dose (200 cGy) + mycophenolate memofetil + cyclosporine [10] | \n\t\t\tD: WT mice P: Transduction of Lin-Sca1+ BM cells with a MSFV-hRPK retroviral vector and subsequent transplantation R: lethally irradiated WT mice [11] | \n\t\t|||||
D: HLA-identical sister R: PKD severe patient C: busulfan + cyclophosphamide [12] | \n\t\t\tD: AcB55 mice P: Transduction of Lin-Sca1+ BM cells with a MSFV-hRPK retroviral vector and subsequent transplantation R: lethally irradiated AcB55 mice [13] | \n\t\t|||||
Glucose Phosphate Isomerase Deficiency (GPI) | \n\t\t\t\n\t\t\t\tGPI\n\t\t\t | \n\t\t\t19q13.1 | \n\t\t\tA.R | \n\t\t\tneuromuscular disturbances | \n\t\t\t\n\t\t\t | \n\t\t |
Triose Phosphate Isomerase Deficiency (TPI) | \n\t\t\t\n\t\t\t\tTPI1\n\t\t\t | \n\t\t\t12p13 | \n\t\t\tA.R | \n\t\t\tneuromuscular disorders, mental retardation, frecuent infections and death in utero\n\t\t\t | \n\t\t\t\n\t\t\t | \n\t\t |
Hexokinase Deficiency (HK) | \n\t\t\t\n\t\t\t\tHK1\n\t\t\t | \n\t\t\t10q22 | \n\t\t\tA.R | \n\t\t\tdefects in platelets | \n\t\t\t\n\t\t\t | \n\t\t |
Phosphofructokinase Deficiency (PFK) | \n\t\t\t\n\t\t\t\tPFKL\n\t\t\t | \n\t\t\t21q22.3 | \n\t\t\tA.R | \n\t\t\tmyopathy, storage disease type VII | \n\t\t\t\n\t\t\t | \n\t\t |
Bisphosphoglycerate Mutase Deficiency (BPGM) | \n\t\t\t\n\t\t\t\tBPGM\n\t\t\t | \n\t\t\t7q31-q34 | \n\t\t\tA.R | \n\t\t\terythrocytosis | \n\t\t\t\n\t\t\t | \n\t\t |
Glutathion Synthetase Deficiency (GSD) | \n\t\t\t\n\t\t\t\tGSS\n\t\t\t | \n\t\t\t20q11.2 | \n\t\t\tA.R | \n\t\t\t5-oxoprolinuria, metabolic acidosis, central nervous system impairment | \n\t\t\t\n\t\t\t | \n\t\t |
Most Common Erythroid Metabolic Inherited Diseases. BM transplantation and gene therapy approaches
A.R, autosomic recessive; D, donor; R, receptor; C, conditioning; P, protocol
Patients with CNSHA suffer anemia and jaundice, but often tolerate their condition well. However, G6PD variants with low activity are related with alterations in the erythrocyte membrane facilitating its breakdown and causing intravasal hemolysis. These symptoms are often accompanied by spleno- and hepatomegaly and hemoglobinuria. Besides, leukocyte dysfunctions caused by lower concentration of NADPH appear when G6PD activity is below 5% of the normal activity, leading to an immune depression [17]. Vives et al. and other groups have also observed an increased susceptibility to infections [18,19].
Preclinical work from Rovira et al demonstrates that hG6PD gene transfer into HSCs may be a viable strategy for the treatment of severe G6PD deficiency [5]. Through the transplantation of pluripotent hematopetic stem cells transduced with γ-retroviral vectors carrying the wild type human G6PD cDNA, they achieved a stable and lifelong expression of hG6PD in all the hematopoietic tissues of primary and secondary receptor mice. In this study, transgene expression was driven by the 3’ LTR from either the Moloney murine leukemia virus (MMLV) or the myeloproliferative sarcoma virus (MPSV), obtaining an efficient transduction in murine hematopoietic progenitors. The corrected cells were then injected into lethally irradiated syngeneic mice, increasing 2-fold the enzyme activity in peripheral blood cells in comparison with non-transplanted control mice. Long-term hG6PD expression derived from the vector was also observed, which was similar to that of the endogenous enzyme activity. Similar expression was detected in RBC and in White Blood Cells (WBC) in different hematopoietic organs, as expected due to the use of a viral ubiquitous promoter. These results support gene therapy as a suitable strategy for the treatment of severe CNSHA due to G6PD deficiency. Additionally, they also demonstrated the efficacy of this gene therapy vector in human embryonic stem cells (hESC) in which the G6PD gene had been inactivated by targeted homologous recombination, which implies the potential application of gene therapy to G6PD hESCs. Moreover, although a selective advantage in favor of G6PD corrected cells has not been reported because the mice used showed normal G6PD activity, Rovira et al observed a strong selection after transduction of G6PD-deficient ES cells with their vectors. In this regard, the development of G6PD deficient mouse models would be a valuable tool to test new protocols. Furthermore, the mouse strain recently developed by Hay Ko et al may be useful, although it does not reproduce all the features of the human G6PD-deficiency [20].
Pyruvate kinase deficiency (PKD), the second most frequent abnormality of glycolysis causing CNSHA, has also been proposed as a potential disease to be treated by gene therapy. Pyruvate kinase (PK) catalyzes the second ATP generation reaction of the glycolysis pathway by converting phosphoenolpyruvate (PEP) into pyruvate, yielding nearly 50% of the total ATP production in red blood cells. PK plays a crucial role in erythrocyte metabolism, since mature RBC are absolutely dependent on the ATP generated by glycolysis, giving the loss of mitochondria, nucleus and endoplasmic reticulum in their mature state. RPK is therefore necessary for maintaining cell integrity and function. Reduced levels of erythrocyte Pyruvate kinase (RPK) lead to an accumulation of glycolytic intermediates that ultimately shortens the life span of mature RBC by metabolic block [21]. Four tissue-specific isoenzymes of PK (M1, M2, R and L) encoded by two different genes (PK-M and PK-LR) have been identified in humans [22]. The PK-LR gene, located on chromosome 1 (1q21) [23] encodes for both LPK (expressed in liver, renal cortex and small intestine) and RPK (restricted to erythrocytes) through the use of alternative promoters [24]. PK-M1 is expressed in adult nomal tissue, like brain or muscle. The PK-M2 isoform is typically expressed in proliferating tissues like fetal, tumoral and several other adult tissues [25] and during the maturation of the erythroblasts, gradually decreases, giving rise to the RPK isoform.
The codifying region of PK-LR gene is split into twelve exons, ten of which are shared by the two isoforms, while exons 1 and 2 are specific for the erythrocyte and the hepatic isoenzyme respectively [26]. However, clinical symptoms caused by PK-LR mutations are confined to RBC because the hepatic deficiency is usually compensated by the persistent enzyme synthesis in hepatocytes [27]. To date, more than 150 different mutations in the PK-LR have been associated with CNSHA, being most of them missense mutations, splicing and codon stop. Only two variants, -72 G and -83 C, have been identified in the promoter regions so far [26,27]. Molecular studies indicate that severe syndrome is commonly associated with disruptive mutations and missense mutations involving the active site or protein stability [28].
PK deficiency is transmitted as an autosomal recessive trait and although its global incidence is still unknown, it has been estimated in 1:20000 in the general caucasian population [29]. Clinical symptoms appear in homozygous and compound heterozygous patients, which lead to a very variable clinical picture, ranging from mild or fully compensated forms to life-treating neonatal anemia necessitating exchange transfusions and subsequent continuous support [28]. Pathological manifestations are usually observed when enzyme activity falls below 25% of normal PK activity [30], and severe disease has been associated with a high degree of reticulocytosis [31]. Hydrops foetalis and death in the neonatal period have also been reported in rare cases [32,33]. PK deficiency treatment is based on supportive measures since no specific therapy for severe cases is available to date. Periodic cell transfusions may be required in severe anemic cases, often impairing their quality of life. Splenectomy can be clinically useful in some patients increasing the hemoglobin levels, as well as iron chelation to decrease the common iron overload observed in PKD patients [34]. However, in some severe cases, allogeneic bone marrow transplantation is required and it has been successfully performed in one severe affected child [12].
The feasibility of gene therapy in PKD was first reported by the group of Asano, who introduced the human LPK cDNA into C57BL/6 mouse bone marrow cells using a retroviral vector [7]. They demonstrated the expression of the LPK transgene mRNA in both peripheral blood and hematopoietic organs after bone marrow transplantation. However, viral-derived expression in peripheral blood was detectable no longer than 30 days post-transplantation, indicating an insufficient transduction efficacy of the retroviral vector used or transduction of non-pluripotent BM cells. In a hemolytic anemia dog model, bone marrow transplantation of minimal conditioned receptors failed to correct the hematological symptoms [10]. Other approaches to rescue RPK phenotype through a gene addition strategy have been also addressed using a PKD transgenic mouse model (CBA/N PK-1SLC/PK-1SLC) [9]. In this assay, the hemolytic anemia and reticulocytosis was fully corrected when the human gene was highly expressed by means of pronuclear injection, although splenomegaly was still present. Interestingly, the authors observed a negative correlation between RBC PK activity and the number of apoptotic erythroid progenitors in the spleen, providing evidence that the metabolic alteration in PK deficiency affects not only the survival of RBC, but also the maturation of erythroid progenitors, resulting in ineffective erythropoiesis [35]. Further studies from this group indicate that RPK plays an important role as an antioxidant during erythrocyte differentiation, since glycolytic inhibition by mutations in Pklr gene increased the oxidative stress in SLC3 cells (established from Pk-1slc mouse) and led to the activation of hypoxia-inducible factor-1 (HIF1), as well as the expression of downstream proapoptotic genes [36].
In addition, our work carried out in mouse models supported the therapeutic potential of viral vectors for the gene therapy of PK deficiency. Throughout the transduction of bone marrow cells using γ-retroviral vectors that carry the human RPK cDNA and subsequent transplantation, we reported a long-term expression of the human protein in RBC obtained from primary and secondary receptor mice, without detectable adverse effects [11]. Recently, we have also reported a successful gene therapy approach using the same retroviral vectors in the congenital mouse strain AcB55, identified by Min-Oo in studies of alleles involved in malaria susceptibility [37]. These mice carry a loss-of-function mutation (269T-> A) resulting in the amino acid substitution I90N in the Pklr gene, which yields a similar RBC phenotype to that observed in PKD patients, including splenomegaly and constitutive reticulocytosis. Retroviral-derived expression was capable of fully resolving the pathological phenotype in terms of hematological parameters, anemia, reticulocytosis and splenomegaly, together with normalization of bone marrow and spleen erythroid progenitors, erythropoietin (EPO), PK activity and ATP levels. Interestingly, despite a strong viral promoter was used to drive the expression of the transgene, metabolic energy balance was no modified in white blood cells. Moreover, we observed that values above 25% of genetically corrected cells were needed to fully rescue the deficiency [3], suggesting that RPK transfer protocols will always require a significant extent of gene-complemented HSC. Nevertheless, other experiments performed in the CBA/N PK-1SLC/PK-1SLC mouse model of PKD have reveled that 10% of normal BM renders RBC expressing nearly normal RPK protein levels [5]. Differences in the genetic defect of the mouse models used could account for these discrepancies, reinforcing the need for high transduction efficiencies to address the disease in the heterogeneous human population. Additionally, we have proposed the in utero transplantation of gene corrected cells as an alternative option for the treatment of PKD. The transplantation of RPK deficient lineage negative fetal liver cells transduced with lentiviruses (LVs) expressing the human wild type version of the RPK in 14.5 day-old fetuses partially restored the anemic phenotype, mainly due to a low engraftment of corrected cells [13]. Improved in utero cell transfer would allow therapeutic levels, thus offering an alternative therapeutic option for prenatally diagnosed severe PKD. Following our results in the AcB55 mouse model of PKD, phenotype correction could be reached if the percentage of engraftment of corrected cells is significant. We are currently developing improved lentiviral vectors that could be applied in future clinical settings.
Glucose phosphate isomerase (GPI) deficiency is the third most common hereditary cause of CNSHA, due to mutations in GPI gene located on the long arm of chromosome 19. The prevalence of this disease is still unknown, with no more than 50 cases reported so far, and with a higher incidence in the black population. The enzyme catalyzes the reversible isomerization from glucose 6-phosphate to fructose 6-phosphate, an equilibrium reaction of the glycolysis pathway. Glucose turnover is affected only in deficiencies below a very low critical residual GPI activity, but with a drastic decline of lactate formation. As no isoenzyme does exist, patients suffer not only from CNSHA and tissue hypoxia, but also from neuromuscular disturbances. In some cases, GPI deficiency has been found in PKD patients, increasing the severity of the clinical scenario and reflecting the degree of the perturbation of glycolysis. The lack of ATP leads to a destabilization of the erythrocyte membrane causing earlier lysis of the RBC and hemolytic anemia of variable degrees [38]. Animal models of GPI deficiency have been described, showing similar symptoms to the human disease [39]. Until now, no gene therapy attempt has been applied to this deficiency.
Other enzyme deficiencies causing CNSHA are Triose phosphate isomerase (TPI) deficiency, associated with neuromuscular disorders, mental retardation and frequent infections, Hexokinase deficiency (HK), affecting also platelet metabolism, phosphofructokinase (PFK) deficiency, 2,3-bisphosphoglycerate mutase (BPGM) deficiency and Glutathione synthetase (GS) deficiency (reviewed in [17,40,41]). Although the incidence of these diseases can be high (ie. TPI is considered as a frequent enzymopathy affecting 0.1% for caucasian populations and even 4.6% for black populations), they are considered rare or very rare diseases, because only few cases (~25 patients in the case of TPI) are diagnosed due to the severity of the clinical manifestations. No gene therapy approaches have been addressed up to now to treat these enzymopaties. However, due to their common characteristics, strategies developed in the other enzyme deficiencies could be applied directly to the treatment of all of them.
The introduction of a cDNA, encoding for the correct version of the target mutated gene into patient cells using retroviral vectors has been successful for several inherited diseases. The initial integrative vectors for gene therapy design and used in clinical trials were based on Gamma(γ)-retroviral vectors in which the transgene expression was driven by the viral LTR promoter. γ-retroviruses preferably integrate in regions adjacent to the transcription initiation site [42]. The expression of the transgene is promoted by the viral LTR, which drives a high expression that can affect gene regulation of the surrounding genes. Although a high efficiency of transduction and therapeutic effects have been described with these vectors in various monogenic disorders such as immunodeficiencies, adverse effects associated with insertional mutagenesis have also been observed. This has led to the development of the next generation of integrative vectors using self-inactivating-LTR lentiviral backbones. SIN-Lentiviral vectors tend to integrate in intergenic transcribing areas, which represent a safer integrative pattern than γ-retroviral vectors. Aditionally, the expression of the transgene is driven by internal promoters, offering a more physiological expression and a less genotoxic profile when using weak promoters [43]. Current efforts to reduce the mutagenic potential of gene therapy vectors are focussed on not only the use of new viral backbones [44] but also on tissue-specific promoters to restrict the transgene expression to target cells [45] and insulators to confer position-independent expression [46]. Additional regulatory DNA elements such as locus control regions (LCR), enhancers, or silencers have also been used to increase lineage specificity.
Gene therapy for RBC disorders requires, ideally, high erythroid-specific transgene expression in order to avoid side effects in progenitors or hematopoietic lineages other than the erythroid one. In inherited enzymophaties, the overexpression of metabolic enzymes in non-erythroid cells could provide these cells with a potential energetic advantage, with the consequent risk of disturbing the physiological generation of ATP in WBC. Also, the restriction of transcriptional activity to target cells with the use of either tissue-specific or physiologically regulated vectors decreasees the effect of the integrative vectors in the host genome. This goal is particularly important for erythrocyte metabolic deficiencies, as all the affected enzymes are highly regulated and connected with central metabolic pathways. Indeed, an expression limited to the erythroid progeny would reduce the genotoxic risk, as RBC become transcriptionally inactive during differentiation, and finally extrude their nucleus. To study tissue-specific gene therapy strategies for RBC diseases, hemoglobinophaties have been the most widely used.
Erythroid regulatory elements have been extensively used to manage targeted expression to RBC using reporter genes (Table 2). The Locus Control Regions (LCR), defined by their ability to enhance the expression of linked genes to physiological levels in a tissue-specific and copy number-dependent manner at ectopic chromatin sites are commonly used. The components of the LCR normally colocalize to sites of DNase I hypersensitivity (HS) in the chromatin of expressing cells. Individual HS are composed of arrays of multiple ubiquitous and lineage-specific transcription factor-binding sites. In early experiments performed with retroviral backbones, the group of Ferrari developed an erythroid-specific vector by the replacement of the constitutive retroviral enhancer in the U3 region of the 3’ LTR with the HS2 autoregulatory enhancer of the erythroid GATA-1 transcription factor gene. The expression of this vector was restricted to the erythroblastic progeny of both human progenitors and mouse-repopulating stem cells [47,48]. Later, they showed that the addition of the HS1 enhancer to HS2, both from the GATA-1 gene, within the LTR of the retroviral vector significantly improved the expression of the reporter gene. Another enhancer element that has been used to achieve erythroid-specific expression is HS40, located upstream of the ζ-globin gene, since it is able to enhance the activity of heterologous promoters in a tissue-specific manner [49]. It has been shown to be genetically stable in MMLV vectors and enhances expression comparable to that of a single -globin gene [50], although HS40 lacks some of the properties of the LCR, like position independence [51] or copy number dependence [52].
An additional improvement to provide safer vectors for RBC gene therapy was provided by the use of insulators elements, which have been shown to reduce position effects in transgenic animals [60]. Insulators are genomic elements that can shelter genes from their surrounding chromosomal environment, by either blocking the action of a distal enhancer on a promoter [60,61], or by acting as barriers that protect the gene from the silencing effect of heterochromatin [61]. The most well studied element is the chicken hypersensitive site 4 (cHS4), an insulator sequence of the chicken -like globin cluster. Studies performed by Chung et al with the γ-globin promoter and the neo reporter gene on selected cells lines, demonstrated the ability of cHS4 to insulate the expression cassette from the effects of a strong -globin LCR element [63] and therefore reducing its genotoxicity. Experiments from Arumugam et al showed a two-fold reduction in transforming activity with insulated LCR-containing lentiviral vectors comparing with vectors lacking the cHS4 element [68].
\n\t\t\t\tErythroid tissue-specific vectors\n\t\t\t | \n\t\t|||
\n\t\t\t\tPromoter / enhancer\n\t\t\t | \n\t\t\t\n\t\t\t\ttransgene\n\t\t\t | \n\t\t\t\n\t\t\t\tVector type\n\t\t\t | \n\t\t\t\n\t\t\t\tReference\n\t\t\t | \n\t\t
HS2 GATA-1 enhancer within the LTR | \n\t\t\tΔLNGFR and NeoR / EGFP | \n\t\t\tSFCM retroviral vector | \n\t\t\t[47] \n\t\t\t | \n\t\t
HS1 to HS2 GATA-1 enhancer within the LTR | \n\t\t\tEGFP and hΔLNGFR | \n\t\t\tSFCM retroviral vector | \n\t\t\t[48] | \n\t\t
Ankyrin-1 and α-spectrin promoters combined or not with HS40, GATA-1, ARE and intron 8 enhancers | \n\t\t\tEGFP | \n\t\t\tHIV-1 based vectors | \n\t\t\t[53] | \n\t\t
α-globin HS40 enhancer and Ankyrin-1 promoter | \n\t\t\tGFP / FECHcDNA | \n\t\t\tHIV-1 based vectors | \n\t\t\t[4] | \n\t\t
IHK, IHβp and HS3βp chimeric enhancers/promoters | \n\t\t\thβ-globin cDNA | \n\t\t\tSleeping beauty transposon | \n\t\t\t[54] | \n\t\t
\n\t\t\t\tPhysiologically regulated vectors\n\t\t\t | \n\t\t|||
\n\t\t\t\tPromoter / enhancer\n\t\t\t | \n\t\t\t\n\t\t\t\ttransgene\n\t\t\t | \n\t\t\t\n\t\t\t\tVector type\n\t\t\t | \n\t\t\t\n\t\t\t\tReference\n\t\t\t | \n\t\t
HSFE and β-globin promoter | \n\t\t\thβ-globin cDNA | \n\t\t\tMSCV retroviral vector | \n\t\t\t[55] | \n\t\t
LCR and β-globin promoter | \n\t\t\thβ-globin cDNA or EGFP | \n\t\t\tHIV-1 based vectors | \n\t\t\t[56,57] | \n\t\t
β-globin and θ-globin promoters combined or not with HS40, GATA-1, ARE and intron 8 enhancers | \n\t\t\tEGFP | \n\t\t\tHIV-1 based vectors | \n\t\t\t[53] | \n\t\t
LCR HS4, HS3, HS2, β-globin promoter and truncated β-globin intron 2 | \n\t\t\tEGFP | \n\t\t\tHIV-1 based vectors | \n\t\t\t[58] | \n\t\t
LCR, cHS4 and β-globin promoter | \n\t\t\thβ-globin cDNA | \n\t\t\tHIV-1 based vectors | \n\t\t\t[46] | \n\t\t
β-globin promoter, LCR HS2, HS3, HS4 | \n\t\t\thβ-globin cDNA | \n\t\t\tAAV2 | \n\t\t\t[59] | \n\t\t
Specific vectors for gene therapy of erythroid inherited diseases.
LTR, long terminal repeats; HS: hypersensitive site; IHK, human ALAS2 intron 8 enhancer, HS40 from αLCR and ankyrin-1 promoter; Ihβp, human ALAS2 intron 8 enhancer, HS40 from αLCR and β-globin promoter; HS3βp, HS3 core element form human βLCR and β-globin promoter; LCR, locus control region. Modified from Toscano et al., 2011
Tissue-specific expression using alternative human promoters can be convenient or more efficient for some diseases, but driving the expression of the therapeutic genes using own promoters is still the most physiological approach to reduce the genotoxic risk of integrating gene vectors [62]. The use of physiologically regulated vectors has been limited mainly because the promoter and the enhancer elements have to be obtained from the affected genes and they are often too large to be included in a lentiviral backbone, and also because the gene expression pattern depends partially on chromatin positioning [63]. -globin LCR has been widely used when attempting to solve this limitation. The -globin LCR consists of 5 HS regions located upstream of the entire cluster of human -like globin genes, each containing a high density of erythroid-specific and ubiquitous transcription binding elements [64]. Much of the transcriptional activity of the -globin LCR resides in HS2 and HS3 sites, but site 4 is important in adult globin expression [65]. Previous studies in vitro and in vivo have shown that -globin LCR can enhance erythroid-specific expression from heterologous non-erythroid promoters [66,67]. First approaches using -globin LCR and 3’ enhancers were based on murine γretroviral vectors [74,75], but the limited packaging capacity of these vectors (up to 8 kb) did not allow the presence of such as large regulatory sequences. Several vector designs including different combinations of regulatory sequences and a deletion of a cryptic polyadenylation site within intron 2 of -globin gene [68], flanked by an extended promoter sequence and the -globin 3’ proximal enhancer were developed. The combination of the LCR elements (3’2 kb) spanning HS2, 3 and 4, were the best amongst several possibilities [69] to achieve a high titer retroviral vector capable of expressing high levels of the transgene.
Other approaches to achieve consistent long-term expression of a transgene have been based on the use of HSFE element, an erythroid-specific chromatin remodelling element derived from the human β-globin LCR which contains binding sites for the erythroid-specific factors NF-E2, GATA-1, EKLF and the ubiquitous factor Sp-1, all of which are necessary to establish a hypersensitive chromatin domain. Work by Nemeth et al., demonstrated that the HSFE can mediate functional tissue-specific “opening” of a minimal human β-globin promoter and increases expression of a human β-globin gene in both MEL cell clones and in transgenic mice. Their results indicated that the most effective vector included tandem copies of the HSFE and produced a 5-fold increase in expression compared to the promoter alone [55] in the context of an integrated retroviral vector.
Gene therapy for RBC metabolic diseases can also benefit from the new technologies based on the modification in mRNA stability or translation efficacy of the transgenes. The use of the post-transcriptional regulatory element (Wpre) from the woodchuck hepatitis virus (WHV) has significantly increased transgene expression in target cells [64,65], even in HSC [70] by stabilization of mRNA at post-transcriptional level. However, it may raise safety concerns, since it contains a truncated form of the WHV X gene, which has been implicated in animal liver cancer [71]. Therefore, Wpre has subsequently been improved by a mutation of the open reading frame of the X gene [72]. Combination of erythroid promoters like ankyrin-1 or -spectrin with Wpre sequence increased 2-fold the expression in unilineage erythroid cultures [53], and when combined also with erythroid enhancers inserted in tandem: HS40 and GATA-1 or HS40 and I8 enhancers [53]. RNA targeting strategies have mainly been used to down regulate expression of cellular genes using vectors expressing interference RNAs (iRNAs). They can be also used to control the expression of integrating vectors knocking down the transgene by the endogenous microRNA cellular machinery. Following this strategy, engineered microRNA target sequences in the vector (miRTs-vector) are recognized by a cell specific microRNA (miRNA), avoiding the expression of the therapeutic gene in undesired cell populations [63]. Several miRNAs are differentially expressed during hematopoiesis and their specific expression regulates key functional proteins involved in hematopoietic lineage differentiation. Particularly, miR-223 has been proposed as a myeloid-specific regulator that negatively regulates progenitor proliferation and granulocyte differentiation and activation [73]. Moreover, Felli et al observed that hematopoietic progenitor cells transduced with miR-223 showed a significant reduction of their erythroid clonogenic capacity, suggesting that down-modulation of this miRNA is required for erythroid progenitor recruitment and commitment [79]. Further studies may determine if the use of miRNA-223 target in lentiviral vectors could be useful to achieve a desirable erythroid-specific expression for gene therapy of red blood cell diseases.
In addition, the erythroid-specificity of short segments of the -globin LCR element has been documented in adeno-associated virus 2 (AAV2) system. Their efficacy to mediate an erythroid-restricted expression has been proved by Tan et al., who reported a successful AAV2-mediated high and stable transduction of the human -globin gene in HSCs from -thalassemia mouse model, which were then transplanted into recipient and rescued them of the disease [59]. These vectors have gained attention as potential useful vectors for human gene therapy, mainly because of their non-pathogenic nature in humans and their relativly easy production. Besides, AAV2 vectors are easily purified to high titers and are able to transduce dividing and non-dividing cells. However, most of proviral AAV2 genomes remain episomal and the insert size is restricted to just over 4kb. Further studies are still needed to know whether they would be a better option than current lentiviral vectors. Also, long-term genotoxic risk of recombinant AAV2 therapy in human is not known up to the date.
In addition, the efficacy of some of these erythroid-specific elements and promoters has also been tested in non-viral vectors, such as transposons. Zhu et al, for instance, studied several hybrid promoters driving the expression of the human -globin gene using the sleeping beauty transposon (SB-Tn). They combined several erythroid elements to develop different chimeric promoters. Their results indicated that the ankyrin-1 minimum promoter was stronger than -globin’s, and the hALAS I8 enhancer (IH) was significantly more powerful that HS3 core element from -LCR and -globin promoter [54]. SB-Tn system is a promising non-viral vector for efficient genomic insertion, even with erythroid-specificity. However, its efficiency for delivering transgenes into HSCs is still much lower than other engineering viral vectors.
Since Yamanaka et al first reported the generation of mouse induced Pluripotent Stem Cells (iPSC) in 2006 by the ectopic expression of four transcription factors (Oct4, Sox2, Klf4 and cMyc) [74] and one year later in human cells together with Thompson’s group [75,76], many laboratories around the world have been able to reprogram a large range of somatic cells into pluripotent stem cells, from neural stem cells [77] to terminally differentiated B-lymphocytes [78]. The reproducibility and potentiallity (unlimited self-renewal and ability to differentiate into any cell type) of this technology has made the iPSC field to advance very rapidly. The human iPSC (hiPSC) technology brings together all the potential of hESC in terms of pluripotency without any ethical issue and the immunotolerance of the autologous cell treatment. Therefore, hiPSC technology is one of the most promising fields for future therapies for many human diseases. Safer reprogramming approaches have been designed and many patient specific hiPSC have been generated both to model human diseases and to correct by gene therapy approaches. Depending on the cell type to be reprogrammed, the number of factors used could be reduced and, what is more important, oncogenes or tumor related proteins included in the reprogramming cocktail, like c-MYC or KLF4 [79] could be removed from the original reprogramming cocktail [80-82]. Several groups developed excisable polycistronic lentiviral vectors [83,84] or transposon-based reprogramming systems [85,86], which could be removed after getting the hiPSC clones. Similar results have been obtained using recombinant proteins [82], synthetic mRNAs [87], and non integrating RNA Sendai Virus vectors [88]. Except for Sendai viruses, non integrating methods show a reduced reprogramming efficiency and the range of cells reprogrammed is not as large as with lentiviral or retroviral vectors.
iPSC technology makes feasible the availability of patient specific cells to study the biology of the disease and develop advanced tools to cure the phenotype and could potentially be used as a therapeutic option (Figure 1). Focussing on metabolic diseases, the first reported metabolic disease patient specific hiPSC line was obtained one year after the first generation of hiPSCs. It was from a 42-year old female that suffered from Type I Diabetes mellitus [89] and it showed no differences compared to a wild type hiPSC line in terms of pluripotency. Next report in which liver metabolic disease patient samples were reprogrammed was carried out by the group of Ludovic Vallier [90], and showed the potential of this kind of approaches for disease modelling and new drug discovery. They reprogrammed fibroblast obtained from α-1 Antitrypsin deficiency (A1ATD), Familiar Hypercholesterolemia (FH), Glucose-6-Phosphate deficiency (G6PD), Crigler-Najjar Syndrome and hereditary Tyrosinemia Type 1 patients, and generated hepatocytes that showed characteristics of mature hepatic cells, like albumin secretion or cytocrome p450 metabolism. Three of the five cell lines (A1ATD, FH, and GSD1a hiPSCs) were capable of recapitulating the disease phenotype in vitro. Disease modelling in erythroid diseased induced pluripotent cell lines has been performed for -Thalassemia [91,92] and sickle cell anemia [93,94]. In these reports the phenotype was corrected by LVs integrated in areas of the genome that were considered safe for viral integration [83] or by gene editing using homologous recombination of the affected locus [91,93,94].
Potential utilities of hiPSC and iPS technology
The future therapeutic application of hiPSC will not only require non-integrative reprogramming system, but also a more precise gene correction. During last years, the cooperation between hiPSC technology and gene editing is being explored. Human iPSC technology has led to the opportunity to control the integration of viral vectors at a clonal level. As we have mentioned before, the analysis of lentiviral integration sites in β-thalassemia hiPSC allowed the identification of corrected hiPSC clones expressing β-globin transgene from a safe genomic site (also called Safe harbour), a site in which integration does not disturb the expression of any neighbouring genes during their erythroid differentiation [83]. The therapeutic use of patient-specific hiPSC emerges then from the combination of gene and cell therapy. From this new research field,future gene therapy protocols will emerge.
Gene editing is a process in which a DNA sequence is introduced into a specific locus or a chromosomal sequence is replaced. This site-specific precise introduction requires an accurate recognition mechanism of the target site on the genome. Under normal conditions, the maintenance of the integrity of the genome requires that the cells repair DNA damage with high fidelity. One of the most harmful DNA damage is the generation of double-strand breaks (DSB). DSB are often resolved by non-homologous end joining (NHEJ), which joins the two ends of the DSB. However this DNA repair mechanism could introduce mutations. On the contrary, homologous recombination (HR) is a truly accurate DNA repair mechanism because it is basically a “copy and paste” mechanism. This process uses an undamaged homologous segment of DNA that can be exogenously provided as a template to copy the information across the DSB. The fidelity of HR gives us the specificity and accuracy that gene editing requires.
The natural HR process has been adapted by researchers to get the desirable addition of an exogenous cassette into the targeted locus. This techniques have been widely use for the generation of knock-out and knock-in transgenic animals [95]. To correct or insert and express a transgene by HR we can consider three different strategies: i) Gene correction, a base or some bases can be substituted from the original strand using an homologous sequence where this base or bases are modified; it is the way to introduce/repair point or small mutations; ii) Safe harbour integration, a complete expression cassette (promoter, transgene and regulatory signals) is inserted in a safe place of the genome, without altering the expression of the surrounding genes and without being silenced by epigenetic mechanisms; this is the case for AASV1 and CCR5 loci. Additionally to these well known safe harbours, there is a wide research focused on finding potential new safe harbour places. iii) Knock-in insertion, the cDNA of a gene is introduced in the same site of the endogenous gene, linked by splicing mechanisms to the endogenous gene assuring the expression of the inserted sequence by the endogenous regulatory elements of the locus where it is integrated.
Gene editing process can be separated in two different steps, generation of DSB and HR. The efficacy of gene editing in human cells depends on the generation of DSB at the specific target site and on the DNA repair mechanism that the cell uses to resolve the DSB. Unfortunately, NHEJ is the dominant pathway to solve these DNA lesions in human cells. Additionally, HR varies in different cell types and requires transit through S-G2 phase of the cell cycle [96]. These limitations make gene editing in human cells difficult to achieve. However, different approaches are being used to improve gene editing by HR, like increasing the length of the DNA sequences homologous to target site (homology arms) [97], the use of adeno-associated vectors [98], the improvement of selection methods for edited cells or the stimulation of HR by inducing DSB using DNA nucleases.
Recently, engineered DNA nucleases have been developed to specifically induce DSB at a unique and defined sequence in the cell genome. These proteins are formed by a nuclease domain and a DNA binding domain whose sequence specificity can be engineered. The most widely used DNA nucleases are Zinc finger nucleases (ZFN), homing meganucleases (MN) and transcription activator-like effector nucleases (TALEN). They identify a potentially unique sequence in the genome and generate DSBs in the desired genomic site, aiming to promote the repair of the DSB by the cell machinery and, ideally by HR. The DNA binding domain of a ZFNs is derived from zinc-finger proteins and is linked to the nuclease domain of the restriction enzyme Fok-I. DNA-biding domain is a tandem repeat of Cys2His2 zinc fingers, each of which recognizes three nucleotides. ZFNs work as pairs of two monomers of ZFN, one in reverse orientation. This ZFN dimer can be designed to bind to genomic sequences of 18-36 nucleotides long. TALENs have a similar structure to ZFNs, but the DNA-binding domain comes from transcription activator-like effector proteins. The DNA-binding domain in TALENs is a tandem array of amino acid repeats. Each of these units is able to bind to one of the four possible nucleotides and this makes that the DNA binding domain can be designed to recognize any desired genomic sequence. TALENs also cleave as dimers. Contrary to these synthetic DNA-nucleases, MNs are a subset of homing endonucleases which recognize a DNA sequence from 14 to 40 nucleotides. Current MNs have been engineered from natural homing endonucleases to increase the number of target DNA sequences.
ZNFs have been widely used for gene editing in hESC and hiPSC. In 2007, Dr. Naldini’s laboratory showed the insertion of GFP into the CCR5 safe harbour in human stem cells (HSC and hESC) after inducing HR by ZFN expression. The CCR5-ZFN and donor DNAs were delivered into hESC by intergrative deficient lentiviruses. More interestingly, targeted hESC were able to differentiate into neurons keeping GFP expression [99]. Soon, the proof of principle for the clinical application of ZFN-mediated gene editing was tested in hiPSC from patients affected by different genetic diseases. The first pre-clinical use of ZFN for gene therapy of a metabolic disease was performed by Yusa et al. In this report, gene correction was performed at the α1-antitrypsin (A1AT) locus to revert A1AT deficiency in hiPSC derived from a patient with a point mutation. This group included a Puromycin resistence cassette flanked by piggyBac sites, so that the Puromycin selection facilitated the isolation of corrected A1ATD-iPSC clones. Afterwards, the selection cassette was removed by piggyBac transposon, obtaining corrected hiPS clones without any additional sequence. These corrected hiPS clones were then differentiated into hepatocyte-like cells to confirm the complete correction of the A1ATD [101]. Other hiPSC gene editing approaches and functional correction of erithroid diseases include gene correction of Sickle Cell Anemia [94] and -Thalassemia [91].
One of the major limitations of ZFN is the generation of “off-target” DSB, due to unspecific sequence recognition. Different studies have highlighted this as a possible limitation in the clinical use of ZFN-mediated HR [100,101]. Recent works have explored the potential of other types of DNA-nucleases in order to prevent the “off-target” cleave limitations of the ZFN, being TALEN and MN the most promising ones. The feasibility of TALEN to mediate HR in hESC and hiPSC was assess by Jaenisch’s group when they designed TALEN targeting the PPP1R12C (at AAVS1 locus), POU5F1 and PITX3 genes at precisely the same positions as the one targeted by ZFN in their previous work [102]. The authors described a gene editing efficiency similar to the one achieved by ZFN with a low level of “off-targets” [103]. A strategy to minimize the potential number of “off-targets” is to design TALEN to work as obligatory heterodimers, which has beeing already done in the engineered MNs. The application of the TALEN and MN as tools to improve HR is still on going. We are exploring the pre-clinical use of TALEN and MN to correct erythroid metabolic genetic diseases, such as PKD.
Another challenge for the clinical application of gene therapy relates to vector targeting. To achieve successful gene therapy, the appropriate gene must be delivered to target cells and specifically expressed in them, without harming non-targeted cells. The most common and easiest way to target specific cells is by ex vivo infection of the desired cell population. Therefore, cells can be directly exposed to the viral vectors facilitating viral-cell interaction. These interactions are driven by the envelope protein which can be adapted from other viruses redirecting the tropism of the vector. The most widely used vectors are lentiviral vectors pseudotyped with the attachment glycoprotein of the vesicular stomatitis virus (VSV-G), which allows the production of high-titre vectors and confers a broad host range [104]. In comparison with them, engineered LVs capable of delivering genes of interest to predetermined cells, can reduce the targeting of undesirable cell types and improve the safety profile, which will further enhance the use of this vector system for gene therapy applications [105,106]. As we have mentioned above, the use of promoters and regulatory sequences that are only active in target cells adds lineage specific expression, although integration of the viruses in non desired cells is still possible. Ex vivo-targeted gene delivery, as commonly used in HSCs transduction, is associated with a risk of inducing cell differentiation and loss of the engraftment potential of these cells [107]. On the contrary, in vivo gene transfer could target HSCs in their stem cell niche, a microenvironment that regulates HSC survival and maintenance [105]. To accomplish this, the vector must display a suitable system to selectively infect the desired population, for example the introduction of a specific ligand to bind a target-cell receptor [106].
Many attempts have been made to develop targeted transduction systems using retroviral and lentiviral vectors by altering the envelope glycoprotein (Env), which is responsible for the binding of the virus to the cell surface receptors and for mediating viral entry into the cell. The plasticity of the surface domain of Env allows insertion of ligands, peptides or single-chain antibodies that can direct the vectors to specific cell types [108]. However, this type of manipulation negatively affects the fusion domain of Env, resulting in low viral titers. To overcome this downside, a method to engineer lentiviral vectors has been developed. These vectors transduce specific cell types by breaking up the binding and fusion functions of the envelope protein into two distinct proteins [108]. Instead of pseudotyping lentiviral vectors with a modified viral envelope protein, these lentiviral vectors co-display a targeting antibody and a fusogenic molecule on the same viral vector surface. Based on molecular recognition, the targeting antibody should direct lentiviral vectors to the specific cell type. The binding between the antibody and the corresponding cellular antigen should induce endocytosis resulting in the transport of lentiviral vectors into the endosomal compartment. Once inside the endosome, the fusogenic molecule should undergo a conformational change in response to the decrease in pH, thereby releasing the viral core into the cytosol [109]. The use of fusion domain of the binding defective Sindbis virus glycoprotein together with an anti-CD20 antibody has been shown to mediate the targeted transduction of lentiviral vectors to CD20-expressing B cells [110].
However, two major challenges for in vivo gene delivery are LVthe exposure to the host immune/complement system and off-target cell transfer after systemic administration. For these reasons, second generation of early-acting-cytokine-displaying LVs has been developed, that circumvents these obstacles by specifically targeting hCD34+ cells [111,112]. For example, RDTR/SCFHA-LV, consisting of RD114 glycoprotein and stem cell factor (SCF) fused to the Influenza hameglutinin env protein, is resistant to degradation by human complement and efficiently transduces very immature hCD34+ HSCs [113]. This new generation of HSC-targeted LVs should improve current gene therapy protocols through the transduction of primitive HSCs directly in the bone marrow of patients with genetic diseases.
Periodical blood transfusion is the previous to the last therapeutic option for severe cases of CNSHA patients. However, this clinical practice involves also adverse effects related to the immuneresponse against minor erythrocyte antigens which makes the patients refractory to additional blood transfusions in the long run. The availability of genetically corrected patient-specific iPSC would allow the possibility of generating disease free erythrocytes ready for transfusion, avoiding the adverse immune effects.
There have been numerous attempts to produce RBC in vitro from different sources of stem cells. To date, the most successful protocol has been developed by the group of Luc Douay [113,114]. Using peripheral blood CD34+ cells, these authors were able to expand and generate RBC with in vitro and in vivo features of native RBC, and were also capable of transfusing a patient with in vitro generated erythrocytes. Notably, the same authors reported a protocol to generate RBC from hiPSC as an alternative source of HSC [114]. Other groups have described similar protocols to generate erythrocytes from hESC or hiPSC [115-118], although in all these studies the RBC generated from embryonic cells expressed embryonic and foetal hemoglobins but low levels of adult hemoglobin. Additional efforts should be done to make this possibility a therapeutic option.
Erythroid metabolic diseases are well defined and well known diseases which main symptom is CNSHA. As they are monogenic diseases that can be cured by allogeneic bone marrow transplantation, they are very good candidates to be treated by gene therapy. However, the low number of patients with poor prognosis requiring BM transplantation and the absence of an apparent selective advantage of the corrected cells over the diseased ones have made their approach for gene therapy less attractive than other erythropaties. Up to now, no gene therapy clinical trial for erythroid metabolic diseseases has been accomplished. Gene therapy attempts in animal models have been applied to G6PD and PKD with successful results, emphasizing the usefulness of a gene therapy approach for these diseases. Although adverse effects due to ectopic expression of the metabolic enzyme have not been observed, an erythroid specific expression is preferred. Many developments have been made for the specific expression of globin genes that could be adapted to vectors developed for the discussed erythroid metabolic diseases. Similarly, any attempt directed to the improvement of HSC transduction, including the possibility of in vivo targeted gene therapy could be applied. On the other hand, the combination of cell reprogramming and gene editing opens a new world of possibilities that could be easily applied to these diseases. hESC and hiPSC are helping in the development of the next generation of gene therapy, which implies a precise gene targeting. Gene editing by HR is the best and safest gene therapy procedure because avoids any perturbation in the targeted genome. Besides the combination of hiPSC and gene editing could be the future therapy for many genetic-based diseases. The hiPSC technology is the springboard for the development of more efficient HR protocols applicable to other types of stem cells such as hematopoietic stem cells. The combination of methods for obtaining big amounts of RBC from HSC or embryonic cells, along with the improvement of the different gene therapy approaches described in this chapter, opens up the possibility of the therapeutic application involving the infusion of RBC differentiated in vitro from genetically corrected patient specific stem cells.
5-FU 5-fluorouracil
A1ATD-1 antitripsin deficiency
AAV Adeno-associated virus
BM Bone marrow
BPGM 2,3-bisphosphoglycerate mutase
CNSHA Chronic non spherocytic hemaolotyc anemia
DSB Double strand breaks
Env Viral envelope
FH familiar hypercholesterolemia
G6P Glucose-6-phosphate
G6PD Glucose-6-phopahate dehydrogenase
GPI Glucose phosphate isomerase
GS Glutathione synthetase
hESC human embryonic stem cell
hIF1 hypoxia-inducible factor-1
hiPSC Human induced pluripotent stem cell
HK Hexokinase
HR Homologous recombination
HS DNase I hypersensitive sites
HSC Hematopoietic stem cell
iPSC Induced pluripotent stem cell
kb kilobases
LCR Locus control region
LTR Long terminal repeats
LV Lentivirus
MN homing meganuclease
NHEJ non-homologous end joining
PFK phosphofructokinase
RBC Erythrocytes
SIN-LV Self-inactivated lentiviral vector
TALEN transcription activator-like effector nuclease
TPI Triose phosphate isomerase
WT wild-type
ZFN zinc finger nuclease
The authors thank L. Cerrato, M.A. Martín and I. Orman for their technical assistance. We would also like to thank Dr. J. Bueren for careful reading and suggestion of the manuscript. M.G.G. was partially supported by a short-term fellowship from the European Molecular Biology Organization (EMBO ASTF 188.00-2010). Z.G. is a fellowship of the PhD program of the Departamento de Educación, Universidades e Investigación del Gobierno Vasco. This work was funded by grants from the Ministerio de Economía y Competitividad (SAF2011-30526-C02-01), Fondo de Investigaciones Sanitarias (RD06/0010/0015) and the PERSIST European project. The authors also thank the Fundación Botín for promoting translational research at the Hematopoiesis and Gene Therapy Division-CIEMAT/CIBERER.
Man is not an indifferent hanger for carrying the disease. Man is the larger part of the disease. Temperament affects even skull fractures. It affects the coloration of acute diseases and especially certain chronic and maybe non-exogenous disease types, which cannot be properly understood or judged unless we follow the internal and typically more significant threads of their etiology into the unique tangle of individual characteristics (László Németh [1]).
Psychosomatic medicine, as a philosophical frame and practical approach of the diagnostic and therapeutical agency, had been undergone several renewals and reframing in the past. If we try to explore the archeology of psychosomatics, we can trace its orientation back to Galenus, Hippocrates, or the Chinese The Yellow Emperor’s Classic of Medicine.
Galenus wrote about the connection between melancholy and mammary carcinoma following theories by Hippocrates, attributing the cause of breast cancer to an “excess of black bile,” implying more than a disbalance of humor, and pointing to the habitus, the emotional and behavioral character. Emotional disorders and mental illnesses also had been considered to constitute a significant part of diseases in Chinese medicine, where such illnesses were classified as Qing Zhi disorders. The so-called emotion-wills implied the Qi Qing:-seven emotion, namely happiness, anger, anxiety, pensiveness, sorrow, fear, and fright and the Wu Zhi five wills: happiness, anger, thinking, sorrow, and fear. According to the ancient Chinese approach, they play a primary role in the onset, progress, and prognosis of most of the diseases.
As emotions are deep human ecological representations of the environment depending on perceptions, evaluation, and interpretation of the outer and internal environment, we can realize that psychosomatics is also a human ecological approach immersed into external and internal networks of social, psychological, neuroendocrine-immune and molecular layers.
The emotional response to environmental challenges depends on personality (A, C, D type) as a result of personal history (early mother–child attachment, adverse childhood experiences); personal development; Pavlov’s, Skinner’s, and Bandura’s learning processes and system-like social influences (family relationships in frame of Milano School, worksite mental health issues); competition and frustration; domination and submission; social rank; and self-evaluation (shame, feeling guilty).
In an overview of the history of psychosomatic concepts regarding human suffering, we find changing frames for the connection between mind and body in a multilayered human ecological setting. The psychoanalytic and psychodynamic frameworks, the hypnotic phenomena, stress medicine based on Cannon’s fight-or-flight reaction, Selye’s stress, Lipowski’s consultation-liaison medicine, the Engelian biopsychosocial concept, and the paradigm of behavioral medicine have network features in common.
In the background, the clinical psychophysiological research emerges with the clinical fields of psychoneuroimmunology, psychocardiology, and biobehavioral oncology. This way, the so-called mind-body medicine and stress medicine frameworks reflect a converging pluralism. The frames are diverse, and the foci are common. Information flow through the social, cognitive-emotional, psychological, neural, endocrine, and immune interfaces and the molecular transcriptomic interfaces and backward. These paths and regulative networks have shared evolutionary origins. These are the structural-functional, patterned heritage of ours, organizing biopsychosocial adaptation and the structural wisdom of the human body. Their adaptive/maladaptive potential depends on the changing environmental context.
Drawing a Venn diagram of different historical or competing schools of psychosomatic medical philosophy, we find many overlapping themes, which might also be considered as hubs of multilayered network organization of psychosomatic phenomena, working as a network of networks (Figure 1). The letters sign some of the evolutionary steps of psychosomatics without a claim for the whole picture (Table 1).
Venn diagram of different psychosomatic discourses.
A | 1818 | Heinroth | The name “psychosomatics” | Mind-body network |
B | 1843 | Le Cabanis | Relationship between body and spirit | Mind-body network |
C | 1880 | Fabre | A nervous system disorder can cause organ damage, and the abnormal functioning of an organ always affects the nervous system | Neurovisceral network |
D | 1886 | Bernheim | Physical symptoms of hypnotic suggestions are results of ideosensory and motoric reflexes | Neurovisceral network |
E | 1889 | Janet | The block, the dissociation, and reversible amnesia between the conscious and unconscious results in several psychopathological phenomena that can be treated by hypnosis | Neurovisceral network |
F | 1892 | Male | Pathology of emotions, the organ symptoms that result from emotions, is similar to those caused by physical factors | Neurovisceral network |
G | 1896 | Freud | Psychoanalysis based on the theory of dynamic psychiatry, libido theory, conversion neurosis, hysteria, symbolic organ speech | Mind-body network |
Gy | 1905 | Ferenczi | Short dynamic psychotherapy | Mind-body network |
H | 1905 | Pavlov | Paradigm of conditioned reflexes providing a framework for neurobiological learning theory | Neurovisceral network |
I | 1909 | Eppinger and Hess | Description of sympathicotony and vagotony | Neurovisceral network |
J | 1928 | Heileg and Hoff | Relationship between environmental pressure and infection incidence | Neuroimmune network |
K | 1926–1935 | Metalnikov and Chorin 1926 Speransky 1935 | The conditioned neutral stimulus might provoke inflammation “immune conditioning” | Neuroimmune network |
L | 1932 | Cannon | The alarm reaction Fight-or-flight versus tend or mend | Neurovisceral network |
M | 1932 | Erickson M.H. | Traumatic amnesia and psychosomatic symptoms are psychoneuro-physiological dissociations that can be resolved by “internal resynthesis” using hypnotherapy | Neurovisceral network |
N | 1935 | Breur | Relationships between tuberculosis relapses and life events | Neuroimmune network |
O | 1936 | Selye | Designates the information pathways of HPA axis as mechanism of stress-related psychosomatic problems | Neuroimmune network Neuroendocrine network |
P | 1936 | Bergmann | Pathology of functional internal medicine | Neurovisceral network |
Q | 1937 | Hetényi | Autonomous nervous system-based diseases of internal medicine | Neurovisceral network |
R | 1937 | Papez | Mental experience is transformed into the psychophysiological pattern of emotions by the limbic-hypothalamic system | Neurovisceral network |
S | 1940 | Scharrer | The central nervous system controls the hormone production of the endocrine system through the hypothalamus | Neuroendocrine network |
T | 1942 | Bykow | Corticovisceral pathology | Neurovisceral network |
TY | 1943 | Dunbar | Relationship between personality and psychosomatic disease | Mind-body network |
X | 1950 | Alexander | Psychosomatic medicine | Neuroimmune network |
Y | 1955 | Charva | The system model of neurohumoral integration | Neurovisceral network Neuroendocrine network |
U | 1955 | LeShan | Specific pattern of cancer risk based on a biographical history and personality survey of cancer patients | Neuroimmune network |
Ü | 1957 | Bálint | Doctor-patient relationship and communication as a factor in healing. Bálint groups | Mind-body network |
V | 1972 | Weiner | Psychosomatic problems as disorders of information transmission between the limbic-hypothalamic–pituitary system | Neuroendocrine network |
W | 1974 | Ader | Psychoneuroimmunology | Neuroimmune network |
Sz | 1977 1978 | Matarazzo Schwartz/Weiss | Behavioral medicine | Mind-body network |
Z | 1984 | Caccioppo | Development of social neuroscience | Mind-body network |
ZS | 1995 | Meaney | Social epigenomics | Mind-body network |
Contemporary psychosomatic medicine broke away from the psychoanalytic foundations, and its research directions reflected a turn toward networking with other disciplines, as an interdisciplinary approach named behavioral medicine. The behavioral medicine and the concept of consultation-liaison psychosomatics bound to Lipowski [2] brought psychosomatics closer to mainstream biomedicine, enhancing their collaboration. The Engelian turn of the biopsychosocial paradigm explicitly expressed the importance of information flow through the network of networks that built up dynamically connected social, psychological, somatic, and molecular-genetic layers.
In 1977, the Yale Conference on Behavioral Medicine had a strong impact to the history of psychosomatic medicine. The participants, like Joseph Matarazzo, Redford Williams, David Shapiro, and Gary Schwartz, defined a new framework for the former psychosomatic medicine, as the study and treatment of diseases, disorders, or abnormal states in which psychological processes and reactions are believed to play a prominent role. There were several opinions regarding the identification of psychosomatics with behavioral medicine. Some considered it identical; others expressed the opinion that behavioral medicine was only a fraction of psychosomatics, while others viewed behavioral medicine implying psychosomatic medicine and additional areas of medical and psychological concern. The wider definition of behavioral medicine extended the former borders of psychosomatics, proposing behavioral medicine as “the field concerned with the development of behavioral-science knowledge and techniques relevant to the understanding of physical health and illness and the application of this knowledge and these techniques to prevention, diagnosis, treatment, and rehabilitation. Psychosis, neurosis, and substance use are included only insofar as they contribute to physical disorders as an endpoint” [3]. Further extension of former psychosomatics included social and institutional spheres and deep biological system and network insights as well.
The disciplines contributing to the study of behavioral phenomena include psychology, sociology, anthropology, education, epidemiology, biostatistics, and psychiatry. These disciplines must be coupled with the biological and medical sciences relevant to understanding the disease processes under study. The networking position of behavioral medicine is apparent from epistemological perspective, too. The following matrix clarifies the deep connection with network medicine (Figure 2).
Matrix of problems with which behavioral medicine is concerned [3].
In this matrix, a disease is indexed at the sociological, epidemiological, anthropological, psychological, biomedical, and physiological levels of networks, and this is a multidimensional analysis with reference to different times (risks, pathophysiology, prognosis, treatment, and rehabilitation) and agency.
While behavioral medicine extended the core psychosomatic view to the broadest hermeneutical frame, the consultation-liaison psychosomatic medicine was connected to the mainstream psychiatry, with the following scope of interest:
The role of psychosocial variables in the development of illness (etiology)
The examination of the causative connections between the changes of psychosocial variables and physiological parameters (psycho-endocrinology, psychoimmunology, psychocardiology)
The fundamental psychological changes accompanying illness (illness behavior)
The psychological and mental concomitants of specific somatic processes
The effect of therapeutic methods affecting behavior on somatic conditions and somatic variables
Research into neuroimmunomodulation in immune laboratories had an explosion in the 1970s, in addition to research into the physiology of stress, creating the basis for a new “network” field of psychosomatics, the psychoneuroimmunology.
The rise of psychoneuroimmunology is a typic example of behavioral and medical interpretation of human disease. Its core explanatory model is based on insights of neuroimmune modulation, the bidirectional communication between neuroendocrine and immune system enabled by shared receptors and cross talk of messengers, and their integrated neuroendocrine-immune information pathways consisting of neurotransmitters, interleukins, neuropeptides, and hormone, including even myokines and adipokines [6]. These evolutionary patterned communication networks create a network of networks throughout the whole body, including the brain and all the organs. In the social-psychological, cognitive-evaluative, emotional, neurovisceral associations, connections, and circles, regulative positive and negative feedback loops create unity of rational, emotive, visceral, molecular, receptoral, and transcriptomic-genetic levels. The prehistory of psychoimmunology is mostly shared with psychosomatics, and its hermeneutic and heuristic features are close to what network medicine offers [6].
As early as at the end of the nineteenth century, we see data about the effect of damaging the nervous system on the loss of protection against anthrax. At the beginning of the twentieth century, Salomondsen and Mandsen already connected vagotomy and the atopic and anaphylactic reactions, and Hatiegan first described the effect of adrenaline on increasing the amount of lymphocytes in 1925, which was confirmed by Frey and Tonietty in 1927.
In Metalnikov and Chorine’s 1926 work, they already discussed the conditionability of immune phenomena. The general immunological influence of emotions was described by Erich Wittkower, when he detected an increase in the number of white blood cells in the states of anxiety, anger, grief, and heightened mood. He coined the term “Affektleukocytose” to describe this phenomenon, which he explained with the stimulation of the sympathetic nervous system. A group of physiologists from Cluj-Napoca, Csaba Hadnagy and the Romanian Baciu, also joined this trend when they examined the effects of emotions and the autonomous nervous system on the number of white blood cells at the beginning of the 1940s.
Locke had already prepared a bibliography of more than 1500 articles in 1983 under the title Behavioral Immunology. If we take into account the names given to the scientific field discussed in these articles, the first “christening” took place in 1974 and is connected to Robert Ader, who used the term psychoimmunology and in 1981 extended it as psychoneuroimmunology. The term of neuroimmunomodulation is connected to Herbert Spector, while Berczi and Szentiványi used the term neuroimmune biology. They all include the overlap of different networks thought to be autonomous.
Even in the comprehensive work of Franz Alexander, psychosomatic medicine includes also internal diseases which, some decades later, turned to be understood in psychoneuroimmune contexts, like IBD, bronchial asthma, rheumatoid arthritis, peptic ulcer, Graves disease, neurodermatitis, and, as recent data show, hypertension which is not an exception at all. Although Alexander did not realize the neuroimmune information pathways and the networked features behind these diseases, his “psychosomatic” internal medicine was strongly attached to psychosocial relationships and conflicts including clinical phenomena generated by neuro-immunobiological networks.
Reviewing contributions to the prehistory of psychoimmunology, we can find the researchers’ sensitivity toward the neuro-immunobiological network response to environmental challenges, as a shared feature in oeuvre of Hungarian scientists, like Selye, Berczi and Nagy, Bertók, Bohus, or Jancsó Jr. Selye presented the first neuroendocrine-immunological insight to human adaptation in 1936 when he proved the somatic triad of general adaptation syndrome, including peptic ulcer, adrenal hypertrophy (endocrine), and thymic and lymphoid atrophy. Selye confirmed the effect of the adrenocortical extract on inducing thymic atrophy in rats in 1943, and he called attention to the role of corticosteroids in regulating the inflammatory response in 1949. This research resulted in the development of the medication that is so important for autoimmune or allergic patients. However, Selye’s Hungarian students also achieved important results in the field of endocrine immunology. István Berczi became a colleague of Selye in the 1960s, and he was exploring the immunological effects of hypophysis hormones together with Éva Nagy. They were among the first to confirm that not only cortisol but other stress hormones, such as the growth hormone or prolactin, also participate in the regulation of the hemo-lymphopoietic system and the immune functions. Lóránd Bertók, a guest researcher of Selye in the 1960s, can also be considered one of the pioneers of natural immunity research. He examined the protective role of bile acids against bacterial lipopolysaccharides. The toxic effect of the lipopolysaccharide endotoxins released by bacteria is an essential promoter of inflammation since their membrane-disrupting, capillary-penetrating, shock-inducing, and fever-inducing effects provide dramatic components of the illness. During endotoxic shock, the levels of ACTH, corticoids, and beta-endorphin increase; however, the levels of prolactin, TSH, T3, and T4 are reduced. These effects are mediated by immune mediators such as IL-1, IL-6, and TNF, which are secreted by the macrophages activated by endotoxins and monocytes. Lóránd Bertók’s research also confirmed that the radiotherapeutic treatment of the endotoxins reduces their toxic effect and this harmless product, the so-called Tolerin, can increase the natural immune reaction and mobilize stem cells.
Hungarian researchers played a pioneering role in the discovery of another system of connections, the “neuroimmune network.” Besides István Berczi, Andor Szentiványi also played a role in discovering the nature of the connections between neuroendocrine-immune networks when he prevented the anaphylactic response by lesions created in the tubular area of the hypothalamus. The work of Miklós Jancsó Jr. was also important. He investigated the effect of histamine on the endothelial vascular cells and the reticuloendothelial system as early as in the 1940s and identified histamine as the physiological activator of the reticuloendothelial system. However, he also identified another neuroimmune network, which played a large role in the understanding of the neuroimmune processes taking place on the internal and external surface of the body. Through research sensory neurons in the 1950s, Miklós Jancsó Jr. concluded for the first time that a neuroimmune network must exist, in which the sensory fibers play an important role. The antidromic electric excitation of the sensory nerves triggered an inflammatory response, which the researcher was able to prevent with capsaicin treatment and the selective destruction of C fibers. On the internal surfaces of the gut or joints, sensory fibers and the increase of substance P play an essential role in inflammatory processes. The discovery of Miklós Jancsó Jr. still provides a paradigmatic network interpretation framework for psycho-immune research today. This network might play a role in trigger point and referred pain theory of Janet Travell and other reflextherapy theories, too.
Béla Bohus and his colleagues also brought a new slice of reality into the range of interpretation of PNI, examining the correlations between social interactions, dominance, subordination, behavioral traits, and physiological indicators. Social hierarchy is a network structure at ethological/social levels, which is transferred to neuroendocrine-immune networks through cognitive behavioral networks.
We should mention further Hungarian think tanks as well, such as the works of Elemér Endrőczy Csaba Nyakas and Lajos Korányi, or the research group led by Szilveszter Vizy, among which Ilia Elenkov’s or Judit Szelényi can be mentioned as outstanding representatives of the field. In the field of applied psychoimmunology, we have to mention the pioneering role of György Németh and András Guseo.
The turning points and parallel evolutionary pathways of psychosomatics and psychoneuroimmunology, listed in Table 1, share covert network logic.
Networks are stand-alone factors in themselves, displayed by graphs depicting symmetrical or asymmetrical relations between cells, molecules, organs, and social relationships and life events. The network-type depiction is also warranted by the need to present regulatory cycles in block schematics, system theory modeling, and the communication and information paths and logical relationships. The neuroimmune networks are graphs, and the task is to identify the nodes (sometimes hubs) connecting them and the system of relations between them. However, under the socio-psychoimmunological approach, the limits of the graph’s validity exceed the levels of the systems of molecules, organs, and organ systems and bypass the individual and personal as well. Under this approach, partnerships, social support, control, power, the territorial principle, dominance and submission, and social ranking are all presented in a set of relationships that can be outlined by graphs, edges, and hubs. However, connections over time are also aligned to the psychoimmunological interpretation of diseases as a graph and network, in the narrative framework of psychosomatics. Therefore, the particular “metagraph network” of socio-psychoimmunology lies across several layers of graphs.
The anamnesis and history disclosed by the patient, the writing or conversation therapy for exploring and disclosing traumas, the research for early traumas, and the investigation of infection chains, learning about the dramatic dynamics underlying socio-somatic relations, are all possible using the toolset of this expanded, narrative network analysis.
This is the anatomy of experience embodied in text, the crystallography of the petrified personal suffering. Identifying the persons and events included in the fate-text and exploring the system of their relations pose the same kind of challenge for network theory as the exploring the “small-world” networks of relevant mediators, and comparison of the neural, endocrine, and immune networks, and locating the hubs that connect them and drawing the graph lines of the relationships in them.
The identification of key players, dominant communities, groups, and the system of relationships between them, based on the personal narrative, and the understanding of tensions of social rank are an inevitable part of “decoding” the socio-psycho-immune network. This is how actual dramatic hubs connect patterns of vulnerable personality reactions, traumatic life events, social rank, and dominance relations and neuroimmune stress networks. As it is the person who explores and reveals it in the therapeutic process, psychoimmunotherapy includes a rearrangement of the representations of the external set of relations and the set of relations hidden by time generated in mind. This means a network analysis of social behavioral cognitive and clinical psychophysiological networks of relevance. This might offer revelatory rearrangements between the related set of life events, personality, and psychological network pattern and the narrative representation network, which may reach even the neuroimmune networks in the deep. This is why the social networks and their narrative reflection in life history should be analyzed together with biological networks of the socio-psychoimmunological network model.
Situation assessment, psychophysiological, and neuroendocrine networks make up such extended networks, and so do the neuroendocrine and neuropeptide patterns, cytokine networks, extracellular messengers, and intracellular molecular paths, genetic programs, and transcription processes. Overrepresented hubs and edges that determine the dynamics and types of physiological and pathological events, as well as possible therapies, are also outlined here.
George Freeman Solomon was one of those pioneers who established the scientific paradigm of psychoimmunology in the 1960s and 1970s, pointing to the connections between brain, behavior, and immunity. He gathered the psychoimmunological revelations in a corpus of postulates [7]. We can test these postulates from the network perspective.
Graph of neuroendocrine-immune networks might be seen as real small-world networks in which most nodes are not neighbors of one another, but the neighbors of any given node are likely to be neighbors of each other, and most nodes can be reached from every other node by a small number of hops or steps.
IL-1, paraventricular NA secretion- CRH-ACTH-adrenocortical cortisol/(sickness behavior, neuroendocrine-immune feedback, inflammation theory of depression based on depletion of dopamine, or diminishing serotonin secretion)
Immunological abnormalities may be accompanied by psychological or mental disorders.
Activation of the immune system can lead to changes in the activity of the central nervous system.
Immune signaling can also affect the central nervous system.
Cytokines of the immune system, as part of the neuroimmune endocrine axis, play a role in endocrine regulation, including stress-induced endocrine processes.
Cytokines influence psychological processes and cause psychiatric symptoms. Immunity influences behavior, and behavior can aid in immune regulatory functions.
Psychological states/traits-neural networks-stress hormones-immune system (C-type personality, right frontal hemispherical dominance, chronic stress, depression, shame, submissive status)
Adaptive coping styles and enduring characteristics can improve the prognosis of immune diseases and protect susceptible patients from the disease.
Stress coping and traits, including personality traits that influence stress management, may influence the immune response to exogenous antigenic stimuli.
Emotional changes and distress (state characteristics) can influence the onset, severity, and course of disorders controlled by immune processes or resulting from disturbed immune processes (allergies, autoimmune, diseases, AIDS).
Severe emotional and mental disorders can cause immunological disorders.
Immune functions may also be affected by altered states of consciousness.
Experimental behavioral effects may lead to immunological changes.
Damage to and stimulation of some regions of the central nervous system may lead to immunological changes.
Substances produced and regulated by the central nervous system (neurotransmitters, neuropeptides, other neuroendocrine factors) must influence immune processes.
In extensive prospective studies, specific patterns of psychological risk should be associated with a higher incidence of immune disease.
Genetic, gender, and behavioral factors influence the immunological effects of stress.
In addition to the influence on the adult psyche, early injury and the traumatic mother-child relationship can affect the adult immune system.
Therapeutic influencing behavior (psychotherapy, relaxation, biofeedback, and hypnosis) may also affect immune function.
Positive emotions stimulate immune function.
Immunocompetent cells have receptors for receiving neuropeptides, neurotransmitters, and endocrine signals.
Central nervous and hormonal factors may play a role in the regulation of feedback processes in the immune system.
Lymphocyte receptors are also affected by changes in transducer sensitivities that are characteristic of mental disorders and cells of the central nervous system.
Thymic hormones that regulate immune function may be under central nervous system influence.
CRH plays a role in the processes and symptoms of depression and immunosuppression associated with depression.
Certain cell groups of the nervous and immune systems occur together.
The prenatal hormonal environment has an effect on CNS and the development of the immune system, which can have lasting effects on both behavioral and immune functions.
Sleep affects both CNS and immune processes.
Immunological processes and specific personality characteristics (coping style, “hardiness”) may play a role in longevity.
Enzymes for the synthesis of brain neurotransmitters are found in immunocompetent cells and neurons.
Melatonin, a neuronal hormone involved in the regulation of circadian rhythmicity and affected by stress, affects immune function.
Mitogens, potent, non-specific immunostimulants, also act on the nervous system.
Immune cells influence the development and function of the surrounding nervous system.
Lymphokines may affect pituitary hormones directly and via the central nervous system.
Some cells of the central nervous system are capable of lymphokine production.
Cytokines of the immune system, as part of the neuroimmune endocrine axis, play a role in endocrine regulation, including stress-induced endocrine processes.
These psychoimmunological facts support the profound relevance of social and psychological network changes exerting deep visceral influence through psychophysiological networks. It also supports the connection between psychosomatics and broad areas of internal medicine.
The network-based interpretation of crucial issues of psychosomatics mapped in Table 1 follows the above principles. Social networks and neuro-immunobiological networks are linked with psycho developmental hubs. Common, hub-like narrative foci are the early mother–child relationships and the adverse childhood experiences, just like the syndrome of post-traumatic stress. Distorted early mother-child attachment organizations have an impact on the so-called internal working model and other personality features creating enhanced risks for some somatic diseases. A-type anxious avoidant secondary attachment organization might diminish empathy and hypothetically create a tendency toward A-type personality development, strive for dominance, competition, and hostility and tendencies for cardiovascular vulnerability. In contrast, C-type secondary anxious/ambivalent attachment organization creates lower self-esteem, behavioral inhibition of aggression, and expression of emotions, high anxiety, and psychophysiological arousal [8, 9].
Epigenetic consequences of distorted mother-child attachment, like downregulation of hippocampal GR receptors via histone methylation, distorted HPA feedback, and distorted estrogen regulation with consequences on adult maternal behavior also prove the hub-like role of the mother-child relationship between socio-psychological and developmental personality networks, stress physiological networks, and neuroimmune network.
Relations between transactional events, traumas, feelings of submission, and loss of control, just as chronic psychosocial stressors, that carry psychological meaning are explored in the networks of the socio-psychological layer. Alexithymia or social inhibition and the psychological network patterns of the C-type personality convert all this into increased HPA activity, high arousal, and increased LC/NAerg activity, so that all this is eventually embodied in the disruptions of the immune cell network controlled by cytokines. Then, IL-1 and IL-6 as result of modified protein synthesis in the cell reach the central nervous system, and via modified dopaminerg and serotoninerg molecular network changes is transcribed into psychological network patterns and depression. The disturbances of the rank position experienced in social networks (reduced motivation, lack of adequacy in the workplace, family conflicts, loss of socioeconomic status) are also embodied this way (via neural networks and proinflammatory cytokines).
Human social relationships might be occasionally the source of severe conflicts. In the light of social exchange theory, it is apparent that the individual is often exposed to severe distress in the high-cost medium of the temporal, monetary, and emotional strain of social interactions. Plenty of evidence is available for presenting the social-psychoimmune consequences of distorted human relationships.
The negative or ambivalent social relationships and the resulting conflicts and associated negative emotions can influence immune processes. Hostility, which we primarily know as a cardiovascular risk factor of psychosomatics, promotes inflammatory processes as well, which is indicated by elevation of CRP and IL-6, according to the work of Suarez [10], and the increased level of the pro-inflammatory cytokines typical of depression as well.
Depression highlights a distinct area of research within social-psychoimmunology, taking the correlations between depression and social integration into account, as well as the relationship conflicts and its negative effect on social perception. Depression is proven to be a mediating factor between socioeconomic patterns (personal income) and physical consequences (number of sick days) in behavioral and epidemiological research called Hungarostudy, verified by route analysis [11]. Depression is proven to be also an independent risk factor of myocardial infarction [12].
The attitudes and emotions increasing the stress of social interaction and interpersonal emotional relationships, such as anger or the hardships caused by depression, are also reflected in the differences in the immune response. Social conflict influences the course of rheumatoid arthritis, in which case catecholamine plays an important role among the neurohormonal factors mediating psychosocial distress. In the social network of ambivalent individuals (those who exert positive and negative influence as well), the ambivalent persons cause increased adrenergic reactions based on the work of Uchino et al. [13], and the contact and conflict with ambivalent persons can provoke an increase in systolic blood pressure. Long-term tight ambivalent human relationships, rich in conflicts, are common in bad marriages, where worse health indicators are also often observed according to the findings of Kiecolt-Glaser and Newton [14]. The dissatisfaction indicator of marriage is accompanied by worse immune indicators, as seen in the case of the ratio of anti-EBV antibodies, CD4+, or CD8+ cells. The hostile behavior typical of bad marriage, impulsivity resulting in cutting each other off when speaking, as well as critical and judgmental impatience, can be indicators of physiological differences and increased blood pressure and endocrine values, based on the work of Malarkey et al. [15]. Among newlywed couples, those who are more prone to adverse, hostile reactions, and this is recalled during a short, 30-min discussion, suppressed immune function was shown in samples taken 24 h later. Kiecolt-Glaser et al. [16] stated that the discrepancy indicating dysfunction of the endocrine-immune regulation was true to older couples as well during discussions where they had to recall their conflicts. The amount of negativistic behavior was in direct correlation with the weakening of the immune response. Mayne et al. [17] confirmed that as much as 45 minutes of exploratory discussion of conflict was enough to reduce lymphocyte proliferation in the examined women. During prospective research, Levenstein et al. [18] found a connection between ulcerative inflammation of the oral cavity and marital stress, while Kiecolt-Glaser et al. observed significantly approximately 60% longer wound healing in the case of couples exhibiting hostile behavior.
Trait-like hostility, characterized by aggression, anger, and cynicism, causes an even more evident immune regulatory disorder in the event of family conflicts, according to Mayne et al. [17]. Miller et al. [19] found a distinct correlation between hostile and cynical attitudes and behavior during conflict management and the cardiovascular response, cortisol, and immune discrepancies. Social stressors induce a rise of pro-inflammatory mediators as well and cause systematic inflammation in the body, based on the work of Steptoe et al. [20]. Partnership conflicts, rejection, and exclusion have significant pro-inflammatory effects even compared to depression and various life events, according to the findings of Denson et al. [21].
The neuroendocrine effects triggered in the brain by threatening environmental stimuli can create a preparatory pathogen-host defense effect on the native immune system, as a result of which the redistribution of the cells of the native immune system and their migration toward the exposed area are detected. All of this ensures the increased rate of healing after an injury. This response can be mobilized by both the presence of predators and the emergence of a significant conflict situation. In the opinion of Slavich and Cole [22], the mobilization of innate immunity is not only an evolutionary remnant but something that can be triggered by symbolic threats, social conflict, rejection, isolation, and exclusion as well. If we consider the genetic basis of the neuroendocrine and immune systems of mammals when investigating their immune system, we can identify a typical pro-inflammatory/anti-inflammatory response pattern by examining the transcriptome of the leukocytes circulated in the body that is the set of RNA typical of the cell.
Under normal circumstances, the activity of the sympathetic nervous system increases the conserved transcriptional response to adversity (CTRA) with the help of the adrenergic receptors, and the activity of the HPA axis reduces the CTRA-dependent inflammatory response as a result of the released cortisol.
However, in the case of chronic social isolation, the threat of grief, and post-traumatic stress, reduced activity of the anti-inflammatory glucocorticoid receptor (GR) can be detected. Therefore, the so-called conserved transcriptional response to adversity is triggered by threatening, stressful, or permanently uncertain adversity as well, as indicated in Antoni’s report [23].
As mentioned above, the threat of grief, traumatic stress, social isolation, low socioeconomic status, or a cancer diagnosis all result in pro-inflammatory transcription disorders. In experimental animal models, social instability, low social rank, and repeated defeat also resulted in a CTRA. Such challenges increase the activity of the genes responsible for the inflammatory immune response to extracellular pathogens and bacterial infections and inhibit the genes responsible for the antiviral immune response to intracellular pathogens. The selective evolutionary advantage of all this is indicated by the fact that it increases the rate of CTRA, wound healing, and response to infection in the event of an actual physical threat. However, it is apparent from the observations that the CTRA is activated by several symbolic, social, anticipated, or imagined emergencies experienced in everyday life. In the event of prolonged perceived or real danger, social or physical threat, a glucocorticoid resistance might develop, which may lead to more severe inflammation or depression.
These phenomena had evolved as a result of the coevolution of hierarchic layers of social, cognitive, neural, immune, and transcriptomic, genetic layers of this hierarchical construction of different networks linked to each other. The highly conserved biological response to adversity, described above, is crucial to overcome the physical threats or injury. Modern-day social, symbolic, or perceived, even imagined, threats might also lead to a pro-inflammatory phenotype of (mal)adaptive answer. The elevation of pro-inflammatory cytokines, such as IL-1 and IL-6, may contribute to elicit depressive symptoms. The overlap of depression with several physical conditions, including asthma, rheumatoid arthritis, chronic pain, metabolic syndrome, cardiovascular disease, obesity, and neurodegeneration, shows the psychosomatic significance and network character of this civilizational paradox. It is a central issue of psychosomatics and roots in the nonadaptive linking of different, environmental, social, cognitive-emotional, neuroimmune, and genetic networks. Psychosomatics deals with this network of networks, where the informational pathways are the edges between nodes, hubs, and the more extensive network “patches.” The so-called social signal transduction theory of depression is a proper example to track how social-environmental information activate biological processes that lead to depression.
The hypothesis that experiences of social threat and adversity upregulate components of the immune system involved in inflammation is central to the social signal transduction theory of depression. The key mediators or messengers, called pro-inflammatory cytokines, play a hub-like role in the network, which might induce profound changes in behavior like psychomotor retardation and social behavioral withdrawal, and influencing immune networks, and neural regulations of mood, anhedonia, and fatigue as symptoms of depression. Self-perceived/perceived lower social status is associated with higher pro-inflammatory cytokines (IL-6) in the dorsomedial prefrontal cortex (DMPFC) activity. The DMPFC plays a crucial role in the so-called mentalizing network, which is active in brain processes that model the thoughts and feelings of others, as well as in evaluating the social status associated with this process. The ventromedial prefrontal cortex (VMPFC) plays an essential role in detecting and assessing signs of dominance. VMPFC damage leads to insensitivity to the social hierarchy and a lack of respect for age and gender. The amygdala plays an integrative role in the perception of dominance, learning processes are related to the social hierarchy, the perceived value of the individual within the group are linked to the amygdala, and its relationships with the hippocampus and striatum are productive. The lateral prefrontal cortex (LPFC) integrates social hierarchy information from the intraparietal sulcus and hippocampus, while VMPFC is responsible for organizing adaptive behavior. The network approach gives these centers a real social-psychoneuroimmune “hub” position.
On the other hand, diverse anatomical connections connect it to the amygdala, hypothalamus, and periaqueductal gray matter, thus reaching the stress pathways affecting the immune processes. Beyond its role in empathy and mentalization, it is also part of the so-called “aversive amplification” subnetwork, which activates the appropriate limbic areas in the event of threatening stress. In this regard, it plays a role in the processing of social impulses, perceiving others’ higher positions in social rank as a source of critical, negative, exclusionary, and punitive social impulses, as well as in their qualification of danger. The inferior social status presumption is associated with increased activity in this brain area.
The hypothesis of social signal transduction theory of depression regarding experiences of psychoimmunological effects of social threat and adversity is central in our network theory of psychosomatics.
In the network medicine, identification of networks, hubs, and edges represents a true “big data” challenge, as the protein synthesis is determined by nearly 25,000 genes and the network hubs of “interactomes” are created by numerous proteins and functional RNS molecules as cell builders, on a scale of thousands. The number of network interactions with functional relevance is even higher. Learning about these interactions and identifying biological networks are the tasks for network medicine. However, the logic of network pathology is followed by research on the connections between the brain, hormone organs, and the immune system, on physiological networks, exploration of which is also a mapping task for interactomes of different levels. Such a map, comprising nearly 7000 interactions, is drawn by the protein-protein interaction network map of Rual et al. [24], the metabolic network summary by Duarte et al. [25], as well as the cytokine maps. However, the concept of meaningful narratives, life events, personality types susceptible to disease, or Berne’s transaction analysis also strives to draw up such maps. Exploring the connection between anamnesis and disease progression is also a similar effort at representation.
The task is to identify interactomes as networks, within which the network patterns and relevant connection paths associated with the disease should be identified. Probably, the analysis of the socio-psychoimmunology paths is necessary as well, given that the “interactomes” of this mappable system of network relations can be identified. It is possible to explore the relationships between these factors, causal relations, and multidirectional pathways of influence, the network characteristics of the personality and the body, and the dynamics of the evolution and progression of diseases. Low socioeconomic status, discrimination, and subordination are accompanied by an increased level of pro-inflammatory cytokines, with the mediation of neurohumoral pathways, as demonstrated by Dickerson et al. [26], for instance. Anxiety, depression, and post-traumatic stress disease, along with the accompanying social and behavioral phenomena, are connected to neurohumoral and immune network anomalies, such as increased pro-inflammatory cytokines (e.g., IL-6) or the activation of the NF-kappa B path, which has central significance in the activity of inflammatory networks, according to Haroon et al. [27].
Identifying the degree of distribution and identifying the hubs characterized by several connections are needed to find the characteristics of these networks. At a molecular level, these can be TLR4, NF-kappa B, caspase, or, at the cellular level, macrophages or the cellular elements of the HPA axis representing nodes of the network. More abstract network modeling makes neurological structures participating in the assessment of controllability, the psychological processes of social perception and assessment, and neurophysiological structures that organize the personality also such as hubs. This way, the early mother–child relationship, which is vital for the development of personality, in the evolution of neurobiological structures, and carries permanent immunobiological consequences, becomes a network hub, as described above. Similarly, several neural networks as centers might create a greater network system responsible for translating social events.
The network itself is held together by a few hubs that have many connections. This is why socio-psychoimmunology explores lifetime hubs pointing in so many directions (mother-child relationship, separation, loss of object, loss of control), positive or negative traits (pessimism, C-type personality, active or passive coping) that are sensitive in psychoimmunological terms, pathologic network patterns (blunted HPA activity, deregulation of glucocorticoid receptors, TH1/TH2 shift), and allergic or autoimmune disease patterns in patient narratives at the social level. (Figure 3). These various key “hubs” may connect several types of networks of correlations. These small worlds are features of complex networks. The interconnected molecular networks are surrounded by relatively short path connections where a large portion of the component proteins are responsible for a low number of interactions, while they may be along main routes affecting the entire body, influencing the entire network.
Psychosomatic network of networks.
Therefore, the hubs responsible for specific local cellular processes may be deemed to be “party” hubs, while they may also be “date” hubs interconnecting processes and associating relationships that organize the interactome. Further characteristics of the network are the “subgraphs” having motif power and in charge of biological functions such as negative or positive feedback or the oscillator function. These subgraphs are the totality of the interconnected hubs that make up a subnetwork within the network. Most networks may be described by a substantial creation of beams and are accompanied by the generation of topology modules characterized by the emergence of a high local region with mutual connections. Hubs are characterized by a high betweenness centrality that describes the number of the shortest paths running through the hub, otherwise referred to as “bottleneck.” This is the nature of regulatory networks with vector edges.
An essential part of network analysis is link analysis, which looks primarily at the relationships between factors, hubs, and objects. Psychoimmunology itself offers an excellent example for the analysis of key relationships and links between the various objects, as it identifies and maps relations between networks of different characters (wired neural, endocrine propagated by blood flow, immune cells and mediators moving through tissues). Socio-psychoimmunology allocates the anamnestic narrative network relations, life events, and the social and symbolic cultural hub networks, through personality patterns, social-neuroscientific insight to responsible neural circuits deep to the cellular transcriptomic level of neuroendocrine-immune networks, exposing their mutual interactions. This network approach provides a new framework of cognitive mapping for anamnesis, diagnosis, and therapy. The result is a transversal metanetwork appearing through a series of information transcripts and translation mechanisms, which weaves a psychosomatic disease pattern through the network layers with its own heteronomous hubs.
The integrated internal medicine/psychosomatic/lifestyle medicine method is beneficial in improving the treatment of the disease, including the psychosocial factors to be taken into account [28]. Such are loneliness, chronic stress, the role of life events, the loss of object, and the personal characteristics of coping. Psychosomatic diagnosis is supported by the Diagnostic Criteria for Psychosomatic Research (DCPR), which incorporates relevant psychological variables into the diagnostic system along the lines of the most important psychosomatic syndromes, like anxiety, functional physical and conversion symptoms, somatic symptom formation of psychiatric origin, somatic and hypochondriac perceptions and fears (disease phobia and fear of death), and suppression of the disease that refers to psychosomatically colored disease behavior. In contrast, alexithymia; risk patterns of A-, C-, and D-type personality; and behavior patterns include trait and state features of personality characteristics that affect the patient’s condition, including psychophysiological risks [29, 30]. Patients may require appropriate anxiety-reducing therapeutic support or cognitive behavioral therapy for psychophysiological involvement of chronic diseases. Clinical psychoimmunology offers new explanatory model and therapeutic framework for bronchial asthma, inflammatory bowel diseases, rheumatoid arthritis, specific autoimmune endocrine pathologies, and psychosomatic skin diseases. It is crucial if 25% of cardiovascular patients suffer from untreated depression, and the chronic inflammational process fed by depression, or similar psychoimmunological processes might contribute to the atherosclerotic vascular processes. Oncological patients have similar problems with untreated depression and the immunosuppressive effects of depression (in the case of NK cells) on the disease process.
Significant evidence-based research has indicated the increasing importance of a psychosocial approach in the field of internal medicine diseases, such as the SPIRR-CAD study among depressed patients suffering from coronary arterial disease [31]; the PISO study, somatoform disorders [32]; or DAD study, diabetes [33]. Although the SPIRR-CAD study did not demonstrate the overall benefit of cascading interventional psychotherapy among depressed coronary artery patients, it showed the success of therapy in a “bond-damaged” group of patients and among adult bypass-linked ISB patients [34]. Katon et al. [35] integrated “behavioral medicine” and psychosomatic approach to primary care. Lower HbA1c, blood pressure, and serum cholesterol was demonstrated among diabetic patients in the TEAM-care program than the control group receiving average care. Psychosomatics is not an alternative but an extension of the perspective of internal medicine. Lipowski [2] emphasizes that “psychosomatics” is an expression of the inseparability and interdependence of psychosocial and biological (physiological, somatic) aspects of human existence. This extension includes the extension of networks, too.
Independent risk factors that increase the risk of internal medical diseases are also objects of a separate discipline, lifestyle medicine, addressing the relationship between avoidable risk factors and lifestyle. However, lifestyle medicine links biomedicine and psychosomatics, also. Obesity; distorted coping that escalates into addictions (smoking, alcohol, drugs, sedatives, chemical comforters); sedentary lifestyle, or, on the contrary, overtraining due to a distorted body image; eating disorders; and high carbohydrate and fat intake may affect the risk of developing cardiovascular and metabolic disorders.
The Framingham study was one of the early follow-up studies that demonstrated the role of hypertension, smoking, and high blood fats among independent risk factors for coronary sclerosis. Today, depression and anxiety must also be considered an independent risk factor for coronary artery disease [12].
Life events can also affect the development and course of the disease. Bereavement, divorce with high values in the Holmes-Rahe scale, and common everyday stress experience called daily hassles all might play a role. For example, in respiratory diseases, a correlation is observed between stressful life events, perceived stress, and upper respiratory symptoms. The risk of provoking asthma is known for severe adverse life events. Rheumatoid arthritis often flares up after bereavement, divorce, and job loss. Serious conflict, divorce, bereavement, or love disappointment might induce or worsen course of ulcerative colitis.
Lifestyle medicine offers network-like interventions along with behavioral modification. One of these is regular exercise. In a plague of sedentarism, physical exercise might be a panacea for many lifestyle problems. The active muscles are part of the neuroendocrine-immune network of the human organism and exert significant influence on the metabolic system, the immune system, the brain, and the abdominal fat, which is also part of the complex informational network. Exercise induces endorphin secretion. Myokines exert their influence by the presence of their receptors on muscle, fat, liver, pancreatic, bone, heart, immune, and brain cells. Myokines like myostatin, IL-6, IL-8, IL-15, FGF21, follistatin-like 1, brain-derived neurotrophic factor (BDNF), hepatocyte growth factor, fibroblast growth factor, and insulin-like growth factor play role in metabolism and tissue regeneration. IL-15 reduces abdominal adipose tissue, while in heavy physical exercise, the secreted IL-6 as myokine rises to 100-fold of resting level and increases IL-1 and IL-10 as an anti-inflammatory mediator. Brain-derived neurotrophic factor might be secreted as a myokine, and muscle-derived BDNF enhances fat oxidation.
On the other side, psychosomatic medicine as a unique professional medical specialization is not universal at all. In Europe, one can find such psychosomatic professional specialization only in Germany, while psychotherapy applied by somatic experts is practiced in many other countries. The new niches for psychosomatic orientation offered by integrative frameworks of stress medicine, mind-body medicine, or lifestyle medicine and network medicine are based on the above insights of linked biopsychosocial networks. Is this a trans/interdisciplinary challenge or a constraint for networking of different disciplines? If we compare the definition of behavioral medicine by Schwartz and Weiss in 1978 [36] and the 2019 proposal for its renewal by the ISBM consensus boards, we find meaningful shifts. The “interdisciplinary field” was exchanged to “field characterized by the collaboration among multiple disciplines” with the meaning of networking of disciplines instead of filling the intermediary disciplinal gaps.
This way, network medicine means double challenge, to see the patient as socio-psycho-biological “network of networks” and organize his/her healing in networks of disciplines, discourses, and institutions.
Psychosomatic medicine has its permanent revival fed by new findings in social neuroscience, clinical psychophysiology, or the new public health; nevertheless, its institutional network shows a narrow picture. Mental disorders (depression, chronic stress) proved to be independent risk factors in the development of autoimmune, allergic and neoplastic diseases, and myocardial infarction. It has been confirmed by evidence-based basic research (social neuroscience, psychoneuroimmunology, psychocardiology) and epidemiological analyses.
However, the institutionalization of psychosomatic clinical discourse showed a rather marginal status in the shadow zone of the high-tech, evidence-based practical development of biomedicine in the frontline. The discourse dynamics reflects the power inequities of health economic, academic, educational, and clinical health service networks.
In some countries (e.g., Germany, Japan), psychosomatic medicine can be practiced as a specialist field, with specialized psychosomatic clinical departments, separate institutes, and somatic and psychotherapeutic care in a joint framework, in teamwork. In Germany, there are over 5000 specialist physicians with psychosomatic and psychotherapist certifications. Outpatient care employs 3058 psychosomatic professionals, while 10,269 physicians hold the title of the psychotherapist, and a total of 21,312 physicians with somatic background have the title of psychotherapist. There are 120 psychosomatic institutions in Germany with a total of approx. 20,000 beds (Statistik-Portal, 2014). Institutional care is also highly developed, and psychosomatic wards providing regional care in regional central hospitals provide patient care. Although the number of hospital beds is limited (9 to 36 beds), the units also provide consultation-liaison psychosomatic care for other clinical departments. The university and teaching hospitals (20–70 beds) have a higher supply capacity, where in addition to healing, there is research and education.
In March 2016, the Japanese Psychosomatic Society had 3300 members, 71.6% of whom were physicians (general practitioners, psychiatrists, pediatricians, obstetricians-gynecologists, dentists, and dermatologists). Psychosomatic internal therapists also formed a separate association with 1200 members (Japanese Society of Psychosomatic Internal Medicine) [37].
Although there are widespread organized discourse communities, academic associations of psychosomatic experts from gynecology and obstetrics, internal medicine, cardiology, and gastroenterology, just as numerous clinical departments all around the medical world, one can find significant disproportion between psychosomatic medical specialization and mainstream organized health care in most of the contemporary medical systems.
Psychosomatics might be absorbed by psychiatry, as C-L psychiatry might be seen as a branch of mainstream psychiatry. It shows the significant disciplinary distance from internal medicine and other disciplines, while issues of psychoimmunology and psychocardiology are deeply embedded in the health-care system of internal medicine. This way, the emerging network centered renewal of behavioral medicine remains only an ideology than everyday clinical practice.
If clinical practice incorporates psychosomatics as part of mainstream medical discourse, guaranteeing the possibility of specialist examinations and specialized care and creating such specialist care units and scenes, the institutional and economic “emancipation” of the field is assured. In 2004, the so-called DAK/AHG study weighed the cost/benefit of long-term institutional psychosomatic treatment burdened with costly hotel services in 338 insured persons treated in psychosomatic hospital wards between January 1999 and February 2000. The results supported its “raison d’etre” and profitable values for health economic point of view [38].
One might see the reason of these contradictions even in nature of psychosomatic disease, as a patient complaining of somatic symptoms used to be reluctant to classify his or her complaints as psychiatric. One can overcome this situation by an invited consultation-liaison psychiatrist, as the patient is not seeking psychological treatment but a somatic care provider for his/her psychosomatic disorders. All of this requires collaboration, a psychosomatically informed professional organizational culture, and a genuinely competent psychosomatic therapeutic delivery environment for the other treatments offered. Psychosomatic patients travel through routes of somatic care with their symptoms because of their interpretation. Once treated in a somatic ward, they are strongly attached to the physical origin of their complaints based on their explanatory model. Psychological assessment of symptoms is often considered offensive. Therefore, psychosomatic care is highly dependent on patient choice. If the primary and specialist care systems do not offer this type of care, the patient will not make such a decision either. Few people turn to psychiatric care providers for physical complaints of psychological origin, and the fear of stigmatization is a barrier, too. It follows that the internal structural features of institutionalized discourse impede the proper care of a large group of patients. Whereas in general medical practice, about one-third of patients suffer from psychiatric symptoms, and 23% of patients in primary care experience depression, 22% with anxiety, and 20% with somatization, it may be relevant for primary care physicians to have additional psychosomatic licensure training. One-third of cardiological patients have mild depression without treatment; oncological patients have a similar situation, frequently. Beyond these institutional difficulties, there is a great need for integrating psychosomatic to biomedicine, as argued above.
Katon et al. [35] also demonstrated that the so-called TEAM-care program, integrating behavioral and psychosomatic approaches with the somatic practice of primary care, lowered HbA1c, blood pressure, and serum cholesterol levels. Psychosomatic patients also increase the costs of somatic care because of hotel costs and ineffective, sometimes unnecessary diagnostics efforts. This costly, unnecessary “evidence-driven” defensive medical practice consumes energy, time, and space in patients requiring care. Specified psychosomatic care is mostly related/reducted to clinical “elite institutions” and does not form part of general public hospital and outpatient practice. A few psychosomatic centers are connected to the university education (e.g., like the Psychosomatic Outpatient Department at the Institute of Behavioral Sciences, Semmelweis University) or occasionally as a department of the psychiatry clinic or elements of hospital psychiatric wards.
The concept of networked medicine in medical systems can also create new theoretical “niches” for psychosomatic clinical thinking. All of this may be important to connect biomedicine with social neuroscience, clinical psychophysiology (e.g., psychoimmunology), stress medicine, or mind-body medicine. All these conceptual spaces, theoretical niches, also designate real institutional niches. There are vacant clinical spaces that can be filled with training, a new competent workforce, and purely organizational innovation. As the affected patient population is unaware of the psychological roots or modifiers of their complaints, and even this non-knowledge often forms the mechanics of symptom formation (suppression, complexation, alexithymia, traumatic learning,), therefore their care is closely linked to extension of somatic specialists’ competence toward the psychosomatic horizon (specialist exam, license exam). On the other hand, the involvement of highly trained psychologists with clinical psychology specialization might also have an essential part of this organizational change. Such psychosomatic development can also affect oncology, dermatology, rheumatology, cardiology, and gastroenterology networks.
The occupational health services offer a wide surface for preventive network medicine, too. Occupational health might have an important priority area for psychosomatic preventive work and early disease detection. Recognizing the increasing work-related stress in the industrial space of globalization and the consequent economic loss of nearly EUR 40 billion to European Member States’ budgets has prompted European Union decision-makers to do the management of work-related stress management and mental health support, as a Member State’s duty from 2007. Preventive stress management can be part of health promotion and may be of interest to both the employee and the employer in health psychological and psychosomatic practice, linked to screening and other public health preventive practices. The use of de-medicalized cognitive behavioral elements of mind-body preventive agency might be applied as worksite stress management training (like in case of Williams Life Skills training), new screening ways of psychophysiological risks, and available psychometric methods might help to implement worksite and community-based prevention and intervention.
Psychologists with such skills, and occupational health practitioners sensitized in this regard, would achieve economically demonstrable results. Occupational health is the apparent scene for preventive and early psychosomatic intervention, as such screening of employees is easy to do and suits to the personal and corporate interest.
Psychosomatic diagnostic and counseling work or psychosomatic “lifestyle medicine” might have their niches in spa health, wellness network. They are, like the occupational health or specialist network, empty niches to fill with psychosomatics. The map of diverse, nevertheless, coherent discourses of psychosomatics can be reframed by the network medicine concept, a common denominator. If clinical practice incorporates psychosomatics as part of mainstream medical discourse, guaranteeing the possibility of specialist examinations and specialized care and creating such specialist care units and scenes, the institutional and economic “emancipation” of the field is assured. The hermeneutic bridge, which had been already established in the biopsychosocial framework, did not lead to closer hybridization. The neuroimmune biological network frame might help the social and psychological aspects join to the evidence-based biomedical disciplines including the molecular and genomic transcriptomic level.
Steps in the history of psychosomatics share common heuristics in connecting different levels of environmental, psychological, neural, and visceral phenomena. This “multilayer” approach reflects the scientific will to follow the information flow from the social through the psychoneural and the visceral down to the molecular and genetic sphere and back. The psychophysiological core of psychosomatics has a human ecological context and deals with regulative network patterns of evolutionary roots. Concept of behavioral medicine shifted psychosomatics from a comprehensive psychodynamic explanatory model toward an integrative, multidisciplinary framework including levels of social, psychological, and somatic networks. Specific subfields of behavioral medicine, like psychoneuroimmunology, offer insights to the multilayered network-based interpretation of diseases. Dysregulation of evolutionary-based adaptive network activities like the conserved transcriptional response to adversity or the social signal transduction theory of depression reflects the clinical significance of network approach.
Depression itself is proven to be a mediating element between SES and sick days, between social and somatic, just as between the immunological and the psychological networks. Network theory offers an inclusive metanarrative for the description of the different social, narrative, and psychosomatic network layers and their interconnections as well anamnestic, diagnostic, and therapeutic significance. Behavioral medicine has shifted from an “interdisciplinary field” to the promoter of the collaboration among multiple disciplines, so this collaboration might be reframed by the extended and comprehensive network approach. Network medicine [39] as shared conceptual explanatory frame might bring closer behavioral epidemiology, the preventive lifestyle medicine, behavioral medicine, and occupational health and biomedicine. The exploring and implementing efforts based on the above defined “networks of networks” includes medical sociology, medical ecology, behavioral epidemiology, new public health, health promotion on the social side, and clinical psychophysiological depth of psychosomatic therapies including several cognitive behavioral approaches, hypnosis, psychodynamic approaches, and narrative medicine on the psychological side. Internal medicine, behavioral medicine, and psychosomatics with related disciplines overlap in the different social and psychophysiological network layers; network medicine might be the common denominator and the widest inclusive conceptual framework for collaboration.
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