List of hits from the EZbiocloud 16S database.
\\n\\n
Released this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\\n\\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
\\n"}]',published:!0,mainMedia:{caption:"Highly Cited",originalUrl:"/media/original/117"}},components:[{type:"htmlEditorComponent",content:'IntechOpen is proud to announce that 191 of our authors have made the Clarivate™ Highly Cited Researchers List for 2020, ranking them among the top 1% most-cited.
\n\nThroughout the years, the list has named a total of 261 IntechOpen authors as Highly Cited. Of those researchers, 69 have been featured on the list multiple times.
\n\n\n\nReleased this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\n\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
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Owing to the dramatic prevalence of obesity, there is a drastic increase in obesity-associated diseases, including hypertension, type 2 diabetes, dyslipidemia, kidney disease, heart disease, cancer, obstructive sleep apnea, and osteoarthritis. The adipose tissue, also called fat, includes white adipose tissue (WAT) and brown adipose tissue. Obesity is mainly caused by an excessive accumulation of fat mass in WAT to the extent that health may be adversely affected. WAT functions as both a metabolic and an endocrine organ that lies at the heart of a complex network participating in the regulation of various biological responses, including inflammation, immunity, endocrine, and metabolism [2].
Adipose tissue is a dynamic tissue, and the mass of adipose is generally determined by a balance of adipocyte proliferation, differentiation, enlargement, and lipolysis. Increased adipose mass may result from the enhancement of adipocyte formation (adipocyte differentiation, also called adipogenesis), enlargement of adipocyte (lipogenesis), and/or reduction of lipolysis that contribute to fat accumulation in WAT. Adipocyte differentiation is defined as the formation of new adipocytes from multipotent stem cells or preadipocyte precursors [3]. In brief, adipogenesis is divided into four steps, including initial growth arrest, mitotic clonal expansion (MCE), early differentiation, and terminal differentiation-development of mature adipocyte phenotype [4]. White and brown adipocyte differentiation share common pathways and possess specific characteristics. Unless specified, “adipocyte differentiation or adipogenesis” indicates white adipocyte differentiation here.
It is viewed that obesity is a state of chronic oxidative stress. In obese patients, increased oxidative metabolic products of DNA, protein and lipid, and/or decreased antioxidant activity are closely associated with excessive fat accumulation. In high fat (HF) diet-fed animals, oxidative stress precedes the onset of insulin resistance and obesity. In obese animals, levels of reactive oxygen species (ROS) in mature adipocytes are higher than that in other tissues, including liver, skeletal muscle, and aorta. In the course of adipogenesis, intracellular redox potentials tend to a significant oxidizing state. It is suggested that increased ROS level is associated with the development of obesity and may be required for the formation of new adipocytes.
ROS plays a controversial role in adipogenesis. On the one hand, ROS is a required signal molecule in the activation of key transcription factors responsible for adipogenesis. On the other hand, ROS may also act as a negative regulator of the formation of adipocytes.
It is originally identified that ROS is produced physiologically during adipogenesis. In the last decades, accumulating data prove that endogenously generated ROS acts as an essential mediator for adipogenic differentiation. Compared with that of preadipocytes, adipocytes have a relatively higher rate of spontaneous intra- and extracellular ROS production. Adipogenic hormonal cocktails could increase ROS production in preaipocytes and enhance the process of differentiation. C/EBPβ and PPARγ are the main factors that mediate the proadipogenic effect of ROS.
It is shown that oxidative stress inhibits fat cell formation by reducing the C/EBP DNA-binding activity [5]. Mitochondrial ROS could inhibit preadipocyte proliferation, indicating that it can influence adipocyte differentiation through inhibiting MCE. In 3T3-F442A preadipocytes treated with various pharmacological inhibitors, Carriere et al. [6] demonstrated that mitochondrial ROS was negatively correlated with adipocyte differentiation through increasing adipogenic repressor CHOP-10/GADD153.
In a cell, ROS are mainly generated by mitochondrial respiration system, nicotinamide adenine dinucleotide phosphate (NADPH) oxidases, endoplasmic reticulum (ER), and certain metabolic and detoxifying enzymes. Effect of ROS-producing systems on adipocyte differentiation is shown in Figure 3.
Approximate 1–3% of O2 consumed in a cell is incompletely metabolized and diverted into superoxide in complex I and complex III in the respiratory chain. ROS production in mitochondria is obligatory during mitochondrial oxidative metabolism. In addition, ROS can also be produced by several other enzymes that locate in mitochondria. It is demonstrated that most of mitochondrial alterations is associated with redox oscillation.
Mitochondria are highly dynamic organelles that can be rapidly restructured to meet the metabolic demands timely. Notable cytoplasmic remodeling occurs during the process of adipocyte differentiation, and reorganization of mitochondrial network is one of the main events. In preadipocytes, mitochondria are filamentous and organized as continuous reticulum. In contrast, in differentiated adipocytes, they are fragmented, punctual, and redistributed around lipid droplets. Mitochondrial fusion and fission have direct influence on lipid accumulation in adipocytes. Induction of mitochondrial fusion by silencing of mitochondrial fission proteins, including dynamic-related protein (Drp1) and fission 1 homologue protein (Fis1), decreases cellular TG content. In contrast, silencing mitochondrial fusion proteins, including mitofusin 2 (Mfn2) and optic atrophy-1 (OPA1), increased cellular TG content followed by fragmentation of mitochondria.
The subunits of mitochondrial respiratory complexes I–IV increase as the differentiation progressed. There is a significant “burst” of expression level of proteins involved in the tricarboxylic acid (TCA) cycle, especially pyruvate carboxylase, at the very beginning of differentiation, which was followed by a more gradual and less intensive increase in the expression of genes associated with the electron transport chain, fatty acid metabolism, and mitochondrial transport. Above results indicate that during the transition from preadipocytes to adipocytes, cells enter a state of metabolic-overdrive characterized by simultaneous increase of flux through the TCA cycle and fatty acid oxidation.
Besides the depictive studies, several genetic and pharmacological treatments have been introduced to evaluate the role of mitochondria in adipogenesis. Depletion of mtDNA using ethidium bromide inhibits lipid accumulation in 3T3-L1 adipocytes. Carbonyl-cyanide-trifluoromethoxy-phenyl-hydrazone, an uncoupler of oxidative phosphorylation (OXPHOS), suppresses adipogenesis. Inhibition of electron transmission chain by several agents, such as rotenone, suppresses adipogenesis by modulating the expression of the key transcription factors C/EBPα, PPARγ, and sterol regulatory element binding protein (SREBP)-1c. Several factors critical for mitochondrial function have been reported to be required for adipocyte differentiation, such as CR6-interacting factor 1 and prohibitin.
Although specific changes of mitochondria during adipocyte differentiation are variant in different laboratories, we still could depict the common pattern: mitochondrial fission, replication, and metabolism are transiently enhanced by adipogenic stimuli, and then the drastic transformation gradually “calms down” when the adipocytes are terminally differentiated. As an essential signal or by-products, enhancement of mitochondrial activity and boost of OXPHOS may result in more ROS production. In addition, direct evidence support that ROS-generating enzymes in mitochondria were found to be activated during adipocyte differentiation, such as monoamine oxidases and dihydrolipoamide dehydrogenase. Moreover, p66Shc, a main isoform of SHC-transforming protein 1 (SHC1) that locates in mitochondria and functions as a potent redox regulator through production of ROS, could be activated by insulin, a key component of DMI cocktail. After the MCE phase, p66Shc phosphorylation is reduced in differentiated adipocytes.p66Shcand consequent mitochondrial ROS generation is also implicated in the adipogenic conversion of muscle satellite cells (SCs), forming intermuscular adipose tissue. Despite the complexity of the involvement of mitochondrial alterations in adipocyte differentiation, there is a common view that mitochondria may function to influence the process of differentiation, at least, through the following ways. First, increased TCA and fatty acid oxidation meet the energy need. Second, mitochondria can provide key substrates necessary to support the massive lipogenesis during adipogenesis.
Endoplasmic reticulum (ER) plays an important role in regulating redox homeostasis. An exclusive oxidizing-folding environment exists in ER, which facilitates disulfide bond formation and generally produces approximately 25% of ROS in a cell. ROS could be generated in ER by uncoupling protein reaction (UPR), NADPH oxidase (NOX) 4, and microsomal monooxygenase (MMO) system.
The changes of the ER environment for protein-folding can cause the accumulation of misfolded or unfolded proteins and activate the uncoupling protein reaction (UPR), a condition also called ER stress. ER stress is a pathophysiological characteristic of obesity and exhibits notable impact on adipogenesis. On the one hand, activation of ER stress could repress adipocyte differentiation. On the other hand, UPR pathways are found to be required for adipogenic differentiation. Eukaryotic initiation factor 2 α (eIF2α)-CHOP pathway may explain the inhibitory effect of ER stress on adipogenesis. In contrast, the inositol-requiring enzyme 1 alpha/X-box binding protein 1 (IRE1alpha/XBP1) pathway of the UPR is required for adipogenesis. In addition, glucose-regulated protein 78 (GRP78), an important molecular chaperone in ER, is reported to play an essential role in adipogenesis. Protein kinase RNA-like ER kinase (PERK) utilizes its intrinsic lipid kinase activity to generate phosphatidic acid, mediates Akt activation, and promotes adipocyte differentiation. PERK can promote adipocyte differentiation through activation of Akt. Activating transcription factor 6 (ATF6) α pathway is also involved in adipogenesis. Thus the three arms of UPR-mediated pathways, PERK, XBP1, and ATF6, are involved in adipogenesis, and blocking of ER stress prevents adipocyte differentiation and weight gain in mice. Regarding the inevitable ROS production in ER stress, the dual roles of ER stress in adipogenic differentiation could be attributed to different signal pathways, such as eIF2α-CHOP versus PERK, XBP1, and ATF6, which may be differentially regulated by ROS. Further studies are needed to test this hypothesis.
The NADPH oxidases (NOXs) are important enzymatic sources of ROS in a cell. NOXs family members include catalytic subunits, known as NOX1 to NOX5, Duox1, and Duox2, and interacting partners, such as p22phox, p47phox, and p67phox. Among those NOXs, only NOX4 and p47phox subunits have been observed to be associated with adipocyte differentiation. Role of NOX4 in adipogenesis is controversial. Compared with preadipocytes, both white and brown adipocytes possess lower expression of NOX4. In adipose tissue in vivo, NOX4 is mainly expressed in cells localized within SVF rather than mature adipocytes accumulated area. In apolipoprotein E knockout (apoE−/−) mice, AT2 receptor deficiency-attenuated adipocyte differentiation is accompanied by an increase in NOX activity. In this trend, NOX4-deficient mice display latent adipose tissue accumulation and are susceptible to diet-induced obesity and early onset of insulin resistance, which could be attributed to accelerated adipocyte differentiation and hypertrophy. Above evidence suggests that NOX4 acts as a negative regulator of adipocyte differentiation.
However, opposite data have been reported. In hypertrophied adipose tissue, ROS level is increased accompanied by an increase of NOX. In 3T3-L1 adipocytes, NOX4 is required for proper insulin signaling transduction through ROS generation. Thus, pharmacological inhibition of NOX4 or KD of NOX4 by RNA interference inhibits ROS production and adipocyte differentiation. As a component of the NOX, p47phox is shown to be positively related with adipocyte differentiation. Mice lacking p47phox are protected against HF diet-induced increase of adiposity, adipocyte size, and hepatic and alcohol-induced hepatic steatosis. Whether other NOX isoforms are involved in adipogenesis is currently unclear.
In some cases, activation of xanthine oxidoreductase (XOR) contributes to ROS generation through the enzymatic form of xanthine dehydrogenase (XDH) or xanthine oxidase (XO). It is reported that XOR colocalizes with the lipid-binding protein adipophilinon the milk-fat globule, functioning as a major component of the milk-fat globule membrane, and is required for fat-droplet secretion in mice. Cheung et al. first show that XOR is highly expressed in adipose tissue compared to other tissues and is transiently induced during 3T3-L1 adipocyte differentiation in vitro. Under obese state, XOR expression is increased in the adipose tissue. In vivo, adipose mass is reduced by 50% in XOR−/− mice versus wild-type littermates. KD of XOR inhibits PPARγ activity and adipogenesis in vitro. The deficiency of adipogenesis in cells with XOR down-regulation is fully restored by an addition of rosiglitazone, implicating that XOR is at the upstream of PPARγ. However, constitutive overexpression of XOR increases PPARγ activity but inhibits adipogenesis. The authors explain that XOR may be transiently required for adipogenic differentiation, and thus prolonged expression of XOR may exert unexpected results. In addition, C/EBPβ is shown to transcriptionally regulate XOR level in several cell types. These results support the notion that XOR lies downstream of C/EBPβ and upstream of PPARγ in the cascade of factors that control adipogenesis and is required for adipocyte differentiation. Moreover, although lacking direct evidence, the finding that inhibition of XOR’s dehydrogenase activity suppresses PPARγ activation and inhibits adipogenesis, indicates that NADH-oxidizing activity of XOR and the concomitant generation of ROS might be involved in adipogenic regulatory role of XOR.
NO, a ubiquitous signaling molecule belonging to reactive nitrogen species (RNS), is synthesized by nitric oxide synthases (NOS). There are three isoforms of NOS, including neuronal NOS (nNOS), inducible NOS (iNOS), and endothelial NOS (eNOS). NO is found to promote differentiation of rat brown and white preadipocytes. The mechanism of NO-promoted adipogenesis involves the activation of guanylyl cyclase and increase of cGMP, which in turn up-regulates cAMP and leads to enhancement of adipogenesis. In addition, NO is shown to induce mitochondrial biogenesis in brown adipocytes, which is dependent on induction of PGC-1α expression.
Both eNOS and iNOS exist in adipose tissue. iNOS expression is increased in adipogenic differentiation. Clinical studies have suggested that increase of NOS and NO level strongly correlate with body fat in obese humans, and eNOS expression is increased in omental versus subcutaneous adipose tissue in obese human subjects. In contrast, it is also found that eNOS levels are reduced in the adipose tissue of obese humans, db/db mice, and HF diet-fed mice. Moreover, overexpression of eNOS prevents HF diet-induced obesity and attenuates hypertrophy of WAT. It is more likely that iNOS and eNOS play differential roles in adipocyte differentiation through regulating NO generation under different conditions. Inducible NO generation by iNOS may be a required event for adipogenic differentiation.
The cytochrome P450 enzymes (CYPs) is a major source of ROS in various tissues and cells. AA is metabolized by several CYP isoforms to produce 20-HETE and epoxyeicosatrienoic acids (EETs). It has been shown that 20-HETE promotes ROS production, whereas EETs is associated with inhibition of ROS production. Under obese condition, CYPs family could be altered in a complicated and isozyme-specific way. CYP3A4 is decreased and CYP2E1 is increased in obesity. The changing patterns of CYP1A2, 2C9, 2C19, and 2D6 isozymes under obese condition are inconclusive. In addition, CYP2E1 expression and activity are increased in female ob/ob mice, while that of CYP1A2 are decreased in male ob/ob mice regardless of gender. Inhibition of CYP27A1 activity, or KD and/or deletion of the Cyp27a1 gene induce adipocyte differentiation through regulating oxysterol 27-hydroxycholesterol. During the differentiation of MSCs to adipocytes, CYP4A11 and CYP4F2 expression are decreased to nearly undetectable level. CYP2J5 is decreased in visceral adipose tissue isolated from HO-2 null mice, in which adipocyte differentiation is enhanced.
The enzymatic antioxidants are capable of counteracting ROS/RNS and oxidant insults through catalyzing a variety of redox reactions.
Regarding the dual roles of superoxide dismutase (SOD) in eliminating superoxide and generating hydrogen peroxide, SOD may play opposite roles in adipocyte differentiation. It is reported that SOD1 expression is decreased in adipose tissue of obese mice. However, in Sod2−/− BMSCs, adipogenesis is found to be increased. Amifostine, an antioxidant, was shown to decrease the numbers of adipocytes in Sod2−/− BMSCs in both basal and adipogenic media and reduce the expression of PPARγ and LPL. In contrast, in Sod2+/+ BMSCs, adipogenesis was decreased in adipogenic medium. During 3T3-L1 adipocyte differentiation, SOD expression and activity were also found to be increased. In mice fed with HF diet, MnTBAP, a pharmacological analogue of SOD, attenuates weight gain and adiposity through a reduction in adipocyte hypertrophy, adipogenesis, and fatty acid uptake in epididymal (eWAT) but not in inguinal (iWAT) adipose tissue.
Catalase (CAT) catalyzes the decomposition of hydrogen peroxide to water and oxygen. mRNA expression of CAT is decreased in the adipose tissue in obese mice. The decreased CAT activity resulting from its promoter mutation is positively associated with childhood obesity and obesity biomarkers such as weight, BMI, and aP2, along with a tendency toward significance with insulin resistance biomarkers. However, in inguinal WAT in obese Zucker rats, CAT activities were found to be increased significantly. Similarly, CAT expression and activity are greatly elevated along with the progression of 3T3-L1 adipocyte differentiation. Furthermore, a remote enhancer region containing two functional PPARγ binding sites in mouse CAT gene was found, suggesting that increased CAT expression during adipocyte differentiation may be a subsequent consequence of PPARγ activation. Thus, whether there is a causal role of CAT alteration in adipocyte differentiation is still unelucidated.
Thiol-dependent antioxidant system mainly consists of glutathione (GSH), thioredoxins (Trxs), and peroxiredoxins (Prxs). The interactions between these antioxidant proteins form a potent antioxidant defense system against potential oxidant threat.
GSH, also named γ-L-glutamyl-L-cysteinyl-glycine, is the most important hydrophilic antioxidant. Glutamate-cysteine ligase (GCL) is the rate-limiting enzyme for de novo synthesis of GSH from the precursor amino acids cysteine, glutamate, and glycine in cooperation with glutathione synthase (GS). GCL consists of a modulatory or light subunit (GCLm) and a catalytic or heavy subunit (GCLc). GSH could eliminate oxidant radicals through nonenzymatic reaction. GSH-mediated reduction of hydroperoxides needs the enzymatic catalysis by GSH peroxidase (GPx). Glutathione-S-transferases (GSTs) detoxify xenobiotic compounds through catalyzing the conjugation of GSH to nonpolar substrates. After the reduction of oxidants or the detoxification of toxicants, GSH form GSSG (oxidized form of GSH) or GSSR (glutathionylated-cysteine derivative), which can be reduced back to GSH catalyzed by NADPH-dependent GSH reductase (GR) and Trxsystem.
Compared with WT littermates, GSH level in adipose tissue is higher in ob/ob mice. Sulfhydryl depletor DEM significantly inhibits adipogenesis, suggesting GSH is involved in the adipogenic process. Moreover, in vivo evidence shows that pharmacological depletion of the GSH in mice prevents diet-induced obesity and increases energy expenditure.
However, controversial data have been reported that up-regulation of GSH was involved in carnosic acid (CA) and carnosol (CS)-inhibited adipocyte differentiation. As a precursor of cysteine, intracellular NAC can be converted into GSH. NAC can be used for the treatment of acetaminophen-induced hepatic injury and for the prevention of radio contrast-induced nephropathy due to its potent antioxidant property. NAC dose-dependently inhibits ROS level and attenuates DMI-induced adipogenesis in 3T3-L1 cells, accompanied by inhibited expression of C/EBPβ and PPARγ. NAC also blocks adipocyte differentiation and the transcriptional activation of CREB in MSC. During ST2 cell adipogenesis, the intracellular GSH redox potential becomes more oxidized. Furthermore, intraperitoneal administration of NAC to rats and mice resulted in reduction of bodyweight. These data suggest that dysregulation of GSH may disturb the physiological role of GSH redox in adipogenic differentiation.
GCL expression in adipose tissue is higher in ob/ob mice in comparison to WT littermates. BSO, a specific and irreversible inhibitor of GCL, deletes GSH and inhibits the differentiation of 3T3-L1preadipocytes and preadipocytes derived from the SVF of inguinal as well as epididymal fat pads. BSO inhibits adipocyte differentiation through inhibition of MCE via down-regulation of E2F-dependent transactivation of the MCM7 target promoter. However, up-regulation of GCLc was proposed to be involved in CA and CS-inhibited adipocyte differentiation.
GPx1, 3, 4, and 7 are identified to be abundantly expressed in mature adipocytes and WAT. In parallel with lipid accumulation during adipocyte differentiation, GPx activity is increased. However, controversial data are also reported.
Compared with WT counterparts, mice over-expressing GPx1 develop hyperglycemia, hyperinsulinemia, and obesity. In patients with nonalcoholic fatty liver disease (NAFLD), the increase in GPx1 expression level is an independent variable associated with NAFLD progression. Chung et al. suggest that GPx3 is required for the regulation of PPARγ-mediated antioxidant effects. Although GPx3 is abundantly expressed in adipose tissues, its expression is reduced selectively in the adipose tissue of several obese animal models. Antioxidant NAC and the antidiabetic drug rosiglitazone increase adipose GPx3 expression in obese and diabetic db/db mice.
GPx4 is stimulated in the course of 3T3-L1 adipocyte differentiation. GPx7 is highly enriched in WAT and is mainly expressed in preadipocytes but not mature adipocytes of WAT. In response to induction medium, GPx7 level in 3T3-L1 preadipocytes declines rapidly within the first 24 h. GPx7 deficiency in 3T3-L1 preadipocytes increases the expression of key transcription factors involved in adipogenesis, including C/EBPβ, C/EBPα, and PPARγ. GPx7 deficiency increases C/EBPβ expression through PKA signaling pathway in a ROS-dependent manner. In addition, deficiency of GPx7 promoters C/EBPβ dimerization increases its binding to the promoter of C/EBPα, leading to an increase of cell proliferation and expression of downstream targets C/EBPα and PPARγ. Furthermore, human genetic variant association results suggest that the subjects carrying risky GPx7 alleles may have reduced GPx7 expression in adipose tissue and higher BMI. However, GPx7 level was increased in the late period of adipocyte differentiation in human liposarcoma cell line SW872.
GSTA3 is over-expressed at the late stage of adipogenesis in human liposarcoma cell line SW872. Jowsey et al. identified GSTA3 as a novel adipocyte differentiation-associated protein whose expression was enhanced manyfolds during the conversion of mouse preadipocytes into adipocytes. During the terminal phase of adipogenesis in 3T3-L1 preadipocytes, GSTζ is found to be increased in C/EBPα- and PPARγ-dependent manner. However, in diet-induced obese rats, GSTYc2 subunit, GST8, and GSTP subunits were found to be down-regulated. Up-regulation of GSTA2 may participate in CA and CS-inhibited adipocyte differentiation.
The Trxs system is composed of NADPH, Trx, thioredoxin reductase (TrxR), thioredoxin interacting protein (Txnip), and novel nucleoredoxin (Nrx). TrxR reduces oxidized Trx. TXNIP is a negative regulator of Trx, resulting in oxidative stress. There are significant alterations of Trx system during adipogenesis. Rajalin et al. showed that the protein level of TrxR1, TrxR2, and Trx2 was elevated in the course of 3T3-L1 adipocyte differentiation. Chutkow et al. found that Txnip protein dramatically disappeared within minutes after DMI stimulation, indicating that Txnip degradation is required for the onset of adipocyte differentiation. Over-expression of Txnip in preadipocytes prevents adipocyte differentiation, whereas its down-regulation enhances adipogenesis. Compared with WT littermates, Txnip KO mice gain significantly more adipose mass due to calorie intake and adipogenesis. In Txnip-silenced preadipocytes and Txnip−/− mouse embryonic fibroblasts (MEFs), adipogenesis is markedly increased, whereas Txnip over-expression impaired adipocyte differentiation. Txnip deletion could also augment PPARγ-stimulated adipogenesis. Txnip negatively regulates the expression and activation of PPARγ, which in turn, suppresses Txnip expression, reflecting reciprocal feedback inhibition between them. In 3T3-L1 preadipocytes, Trx over-expression stabilized Txnip protein levels and thus promoted inhibition of adipogenesis. As an important component of adipogenic stimulants, insulin promoted the dissociation of Txnip-Trx into a more labile state.
Nrx level is higher in WAT of ob/ob mice and is increased in the early adipogenic stage of 3T3-L1 preadipocyte differentiation. KD of Nrx decreases and its over-expression increases adipogenic differentiation of 3T3-L1 cells. In transgenic mice with adipose tissue-specific Nrx over-expression, adipocyte size as well as number is increased compared with WT mice. Moreover, inhibition of Wnt/β-catenin pathway is involved in Nrx-exerted regulation of adipogenic differentiation.
As one of the most abundant antioxidant proteins, Prxs are able to reduce hydrogen peroxide and organic hydroperoxides to water and alcohol. Genetic variations of Prx3 are associated with the risk of obesity. In WAT of db/db mice and in perirenal WAT of human subjects with BMI > 25, Prx3 expression is significantly decreased. Huh et al. first studied the role of Prxs in adipocyte differentiation in detail and showed a gradual increase inPrx1, Prx3, and Prx5 expression in the course of 3T3-L1 adipocyte differentiation. Compared with Prx1 and Prx5, Prx3 was increased by a larger extent. At the age of 20 months, Prx3 KO mice display larger fat pads in epididymal WAT with hypertrophied adipocytes. In differentiated ADSCs, expression of aP2, C/EBPα, and PPARγ, adipophilin, and FAS are increased in Prx3 KO mice. In 3T3-L1 preadipocytes, siRNA-mediated KD of Prx3 increased aP2, C/EBPα, and PPARγ expression, whereas Prx3 over-expression inhibited their expression. Moreover, ROS level in mitochondria was increased in WAT of Prx3 KO mice. In MSCs, over-expression of Prx2 also shows inhibitory effect on the lipid accumulation during adipocyte differentiation.
NADPH:quinone oxidoreductase 1 (NQO1) may generate quinones, quinoneimines, and andazodyes accompanied by inhibition of ROS generation. NQO1 protein is present in human adipocytes, and NQO1 mRNA level is higher in adipocytes than in adipose-derived stromal vascular cells. NQO1 expression is positively correlated with BMI and weight loss results in a decrease in NQO1 mRNA content in adipose tissue. In comparison to WT mice, NQO1−/− mice exhibit lower abdominal adipose tissue. Using 3T3-L1 preadipocytes, Vomhof-DeKrey et al. investigated NQO1 expression in the course of adipocyte differentiation. They found an increase of NQO1 protein at limited MCE and postmitotic growth arrest (Days 1–3) stages and a decrease in terminally differentiated (Day 8) adipocytes that lasted for several days afterward. In contrast to the mapping of protein, NQO1 mRNA expression was increased in differentiated adipocytes (Days 11–14), indicating a discrepancy between steady-state of mRNA and resulting protein levels. Sulforaphane (SFN) enhanced NQO1 protein level and blunted TG accumulation in a NQO1-dependent manner.
Heme oxygenase or haem oxygenase (HO) is an enzyme that catalyzes the degradation of heme and produces free iron, biliverdin, and carbon monoxide. Three isozymes of HO have been defined, including inducible heme oxygenase-1 (HO-1), constitutive heme oxygenase-2(HO-2) and not fully defined HO-3. HO-1 exhibits antioxidant capacity due to its products bilirubin/biliverdin and carbon monoxide. The HO system acts as a key cellular antioxidant defense system in obesity and diabetes. Decreased HO-1 expression levels were observed in type 2 diabetic patients. In addition, products of HO-1 catalysis, CO, and bilirubin are decreased in humans and animal models of type 2 diabetes. Zucker fat (ZF) rats display a decrease in both HO activity and HO-1 and HO-2 protein expression.
A number of studies have demonstrated that genetic inhibition of HO expression and activity increase adipogenesis. In vitro induction of HO-1 decreases adipogenesis. siRNA-mediated inhibition of either HO-2 or HO-1 leads to an increase in adipogenesis. Up-regulation of HO-1 induces a decrease of adipocyte differentiation in MSCs. BMSCs from HO-2−/− mice displayed an increase in adipogenesis and accumulation of lipid droplets, resulting in adipocyte hypertrophy. Using a lentivirus construct of the human HO-1 under the control of theaP2 promoter, Cao et al. revealed that adipocyte-specific over-expression of HO-1 attenuated adiposity and adipogenesis.
In addition, pharmacological studies support the notion that HO-1 negatively regulates adipocyte differentiation. Cobalt protoporphyrin (CoPP) or L-4F, a systemic apomimetic peptide inducer of HO-1, promotes chronic body weight loss in various species, decreases visceral and subcutaneous fat content, and reduces the number of enlarged adipocytes, and increases adiponectin levels and small adipocytes, without any effect on food intake, or other metabolic changes such as energy expenditure and O2 consumption. In addition, other HO-1 inducers, such as D-4F, an apolipoprotein A1 mimetic peptide, replicate the body weight-lowering effect of HO-1 induction. In ZF animals and human bone marrow-derived adipocytes, HO-1 expression and activity was found to be increased by CoPP treatment. In vivo, HO-1 up-regulation was associated with decrease in superoxide levels, visceral and subcutaneous fat content and weight gain, and increase in plasma adiponectin. In vitro, up-regulation of HO-1 decreased superoxide level, adipose remodeling, smaller adipocytes, and increased adiponectin secretion in the culture medium. Burgess et al. found that induction of HO-1 by CoPP slowed the rate of weight gain in male obese mice and produced a significant decrease of IL-6, TNFα, and IL-1β. In B6v-Lep obese/J mice, HO-1 induction increases the number, whereas decreases the size of adipocytes. Moreover, the chronic treatment of carbon monoxide, the product of HO metabolism, prevents the development of HF diet-induced obesity via stimulating metabolism and remodeling adipocytes. In addition, over-expression of HO-1 increases pAMPK and eNOS levels and promotes human osteoblastic differentiation, another direction of cell differentiation. It was found that induction of HO-1 was associated with reduction of C/EBPα, PPARγ, Peg-1/Mest, aP2, CD36, Wnt5b expression and the increase of β-catenin, pGSK3β, Wnt10b, Pref-1, shh, and adiponectin, indicating that those key factors were responsible for HO-exerted effect on adipogenesis.
However, Huang et al. reported that over-expression of HO-1 did not protect against HF diet-induced body weight gain and insulin resistance in mice. Vanella et al. reported that L-4F increased early adipocyte differentiation markers and decreased Peg-1/Mest through activation of HO-1. In human MSCs, HO-2 depletion results in increase of adipogenesis and inflammatory cytokines, with lower expression of HO-1. Sofalcone, a chalcone derivative, inhibits the differentiation of 3T3-F442A preadipocytes into adipocytes, which is restored by SnPP treatment, indicating the involvement of HO-1.
Glucose-6-phosphate dehydrogenase (G6PD) generates NADPH, providing the substrate for NOX and reducing equivalent for GSH and Trx system. Thus, G6PD could affect ROS level in a bidirectional manner. NADPH is also required for the biosynthesis of fatty acids and cholesterol. The expression and enzymatic activity of G6PD are significantly elevated in WAT of obese models, including db/db, ob/ob, and diet-induced obese mice. In 3T3-L1 cells, G6PD over-expression stimulates the expression of most adipocyte markers and elevates the levels of cellular FFA, triglyceride, and FFA release, indicating an important role of G6PD in adipogenesis.
Selenoprotein P (SeP), a circulating selenium carrier, functions as an antioxidant enzyme. Although it is expressed most abundantly in liver, SeP also plays a role in the regulation of lipid metabolism in adipocyte through redox modulation. Zhang et al. reported that SeP1 gene expression was significantly reduced in adipose tissue of ob/ob and HF diet-induced obese mice as well as in primary adipocytes isolated from ZF rats. SeP expression is induced in the course of 3T3-L1 adipocyte differentiation. In adipose tissue of obese mice, rosiglitazone treatment increased SeP protein expression. In contrast, exposure to either TNFα or high level of H2O2 reduces SeP1 expression in a time- and dose-dependent manner in differentiated 3T3-L1 adipocytes. Moreover, KD of SeP1 reduces GPx activity, whereas increases the expression inflammatory cytokines, such as MCP-1 and IL-6 in preadipocytes. These characteristics contribute to the inductive role of SeP in 3T3-L1 adipocyte differentiation.
Metallothioneins (MTs) are a family of proteins with abundant cysteine residues. MTs are capable of regulating oxygen respiration, possessing potential antioxidant activities. MTs function as regulators of various cellular processes including gene expression, apoptosis, proliferation, and differentiation. Evidences suggest that MTs are altered during adipocyte differentiation and play important roles in regulation of adipocyte differentiation. In human, metallothionein-2A (MT-2A) mRNA level in fat tissues is significantly elevated in obese subjects. MT-2 expression is higher in omental than in subcutaneous adipose tissue. Moreover, MT could be secreted by fat cells in WAT. However, Sato et al. discovered that MT−/− mice fed with a HF diet exhibited more fat mass and a larger adipocyte volume, indicating that MT may have a preventive role against HF diet-induced obesity.
In addition to those enzymatic antioxidants, endogenous and exogenous nonenzymatic antioxidants also play important roles in regulating lipid homeostasis. They are usually classified into water-soluble antioxidants, including GSH, vitamin C (VC), lipoic acid (LA), and uric acid (UA), or lipid-soluble antioxidants, such as β-carotene, vitamin E (VE), and coenzyme Q (CoQ). The adipose tissue of ob/ob mice exhibits higher content of hydrophilic molecules (GSH, VC) in a lower redox state, which is associated with lower content of lipophilic molecule (VE, CoQ) and lipid peroxidation. In particular, CoQ is deficient in WAT of ob/ob mice. In rodents as well as in humans, CoQ content is strongly and negatively correlated with subcutaneous adipose tissue and obesity indexes. In 3T3-F442A cell line, pharmacological inhibition of CoQ synthesis by chlorobenzoic acid strongly triggers adipose differentiation. In contrast, over-expression of 4-hydroxybenzoate acid polyprenyltransferase increases CoQ level and inhibits adipogenesis of 3T3-F442A cell. These data suggest that CoQ is an anti-adipogenic factor. NADPH is an essential reducing equivalent for numerous enzymes. Cytosolic NADP+-dependent isocitrate dehydrogenase (IDPc) is a key enzyme for providing cytosolic NADPH. In 3T3-L1 preadipocytes and mice, KD of IDPc reduces adipogenesis and lipid accumulation.
In the last decades, dietary supplementation of natural antioxidants is prevalent worldwide. The dramatic discrepancies about the biological outcomes of antioxidant consumption have been reported.
A large number of experimental and clinical investigations have shown that many antioxidants possess potent anti-obesity effects. α-lipoic acid, a famous ROS scavenger for hydroxyl radicals and singlet oxygen, exerts reproducible anti-obesity effects in several independent clinical trials. The inhibition of adipocyte differentiation by α-lipoic acid involves the regulation of pro-adipogenic transcription factors via MAPK pathways. Dietary orlistat supplementation induces a reduction of body weight in rats fed with HF diet. Bardoxolone methyl, a novel antioxidant capable of activating the Keap1-Nrf2 pathway, shows unexpected weight-reducing effects in a recent clinical trial. Brazilein, a natural biologically active compound derived from
Phenolic compounds, which are secondary plant products, are consumed regularly as part of the human diet and are helpful for disease prevention. Oleuropein is the principal phenolic compound in olive tree products, which is the main source of healthy Mediterranean diet ingredients. Hydroxytyrosol is a bioactive substance after the hydrolysis of oleuropein. It is reported that oleuropein and hydroxytyrosol exhibit antioxidant activities and inhibit adipogenesis in stem cells derived from human bone marrow and in 3T3-L1 preadipocytes. In 3T3-L1 preadipocytes, oleuropein and hydroxytyrosol dose-dependently suppressed fat accumulation and differentiation-related markers, including PPARγ, C/EBPα, SREBP-1c, GLUT4, CD36, and FASN through suppression of MCE. Cigarette smoking aggravates Graves’ orbitopathy through oxidative stress-mediated adipogenesis. Quercetin, as an antioxidant, inhibits adipogenesis by reducing ROS in cultured orbital fibroblasts from Graves’ orbitopathy patients treated by cigarette smoke extract.
However, there is also solid evidence supporting the conception that consumption of antioxidants promotes adipocyte differentiation and obesity. Abe et al. prepared a 100% methanol fraction of methanolic extract from unripe kiwi fruit (
Beyond the direct regulation of ROS by ROS-generating enzymes and enzymatic and nonenzymatic antioxidants, redox balance is controlled by a wide class of transcription factors, including nuclear factor erythroid 2-related factor 2 (Nrf2), peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α), p53, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and forkhead box O (FoxO). These transcription factors could respond to subtle redox changes, determining the reaction toward oxidation or reduction through transcriptionally modulating ROS-generating enzymes and antioxidant enzymes. The large amount of transcription factors influencing adipocyte differentiation play particularly important roles in a cooperative and interactive way.
Nrf2 is the most thoroughly studied redox-sensitive transcription factor. Under normal conditions, Nrf2 is inactivated by kelch-like ECH-associated protein 1 (Keap1), which targets Nrf2 for proteasomal degradation. Once activated, Nrf2 binds to antioxidant response elements (AREs) or electrophile response elements (EpREs), inducing the expression of more than 100 genes involved in the response to cellular stress. Functioning as a “supreme director” in redox regulation, Nrf2 can regulate both ROS/RNS-generating enzymes (such as SOD and iNOS ) and antioxidant enzymes. However, Nrf2 more likely acts as an antioxidant regulator in most cases. Redox proteins regulated by Nrf2 are listed as follows: NQO1, GCLc and GCLm, GPxs, GR, GSTs, HO-1, SOD, CAT, Trxs, TrxRs, Prxs, UCPs, NOS, and G6PD [7, 8].
A large number of papers have revealed contradictory roles of Nrf2 in the regulation of metabolism. On the one hand, Nrf2 is critical for antioxidant defense system, innate immunity, protection against inflammation, insulin resistance and diabetes, and cancer. On the other hand, Nrf2 deficiency is beneficial for the attenuation of atherosclerosis, HF diet-induced obesity and insulin resistance, and cancer.
The Nrf2/Keap1 pathway is linked to the development of obesity and hepatic lipid accumulation in animal models. Various natural and synthetic substances with the ability to stimulate Nrf2 have been verified to exert anti-obesity activity, including 1-[2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oyl]imidazole (CDDO-Im), oleanolic acid, oltipraz, ellagic acid, quercetin, curcumin, resveratrol, and chromium histidinate. Keap1 KD mice with a constitutive increase in Nrf2 activity have lower body weight and epididymal fat masse than WT mice. Enhanced expression of Nrf2 in mice attenuates the fatty liver induced by special diet. However, using Nrf2 KO mice, Nrf2 deficiency has been found to protect against HF diet-induced obesity. However, no significant relationship between Nrf2 and obesity has also been reported. Tanaka et al. found that in both WT and Nrf2-null mice fed with a HF diet for up to 4 weeks, weight gain shows no difference. Consistent with its role in obesity, Nrf2 functions as a double-edged sword in the pathogenesis of insulin resistance. Studies using genetic manipulation have shown that Nrf2 deficiency is beneficial for glucose homeostasis and insulin sensitivity.
Nrf2 is highly expressed in the adipose tissue and animals fed with HF diet exhibit higher Nrf2 expression. As an important constituent of adipogenic stimuli, insulin could significantly activate Nrf2 pathway [9]. These findings suggest a potential role of Nrf2 in the modulation of adipogenesis. Indeed, genetic manipulation and pharmacological activator treatment have been introduced to investigate the role of Nrf2 in adipogenesis and lipid metabolism, which is contradictory. Role of the Nrf2/ARE pathway in energy metabolism and storage has been reviewed by Vomhof-DeKrey et al. In this review, we focus on the involvement of Nrf2/Keap1 pathway in the regulation of adipocyte differentiation.
Pi and his colleagues have made great contributions to the identification of the proadipogenic effect of Nrf2. In their studies, Nrf2 was found to be activated in the early stage (<12 h) of adipogenesis [10]. Consistent with the notion that Nrf2 is a key regulator for GSH synthesis, intracellular GSH level follows a similar pattern as Nrf2-ARE. However, in another study, Nrf2 and Keap1 mRNA levels were found to be increased in differentiated adipocytes (Days 11–14), indicating a programmed control of Nrf2 during adipogenic differentiation. Pi et al. showed that KD of Nrf2 could totally block the enhancement of adipogenesis in Keap1-KD cells, confirming the contribution of Nrf2 activation to above process. To directly determine the regulatory role of Nrf2 in adipogenesis, preadipocytes derived from WAT of Nrf2-KO (Nrf2−/−) and WT (Nrf2+/+) mice were used. Hormonal cocktail-induced adipogenesis in Nrf2−/−preadipocytes shows substantially reduced lipid accumulation. Moreover, Nrf2−/− preadipocytes exhibit significantly decreased expression of adipogenic genes, including PPARγ1, PPAR2, adipsin, and aP2. In 3T3-L1 preadipocytes and human subcutaneous preadipocytes, shRNA-mediated KD of Nrf2 expression inhibits adipogenesis. Pi et al. also found that mice Nrf2 deficiency displayed reduced fat mass in association with small adipocytes and increased adipocyte numbers, and were resistant to diet-induced obesity.
In contrast to the discovery by Hou et al. [10], Shin and colleagues [11] demonstrated an inhibitory role of Nrf2 in adipogenesis using Nrf2−/−-immortalized MEFs. In Keap1−/−-primary MEFs, adipogenesis is inhibited by enhanced Nrf2 signaling, compared to WT MEFs. In Nrf2−/−-immortalized MEFs, ectopic expression of dominant-positive Nrf2 delays differentiation. HF diet feeding decreased the expression of Nrf2 and its target genes, such as NQO1 and GSTm6, and Nrf2 inhibited lipid accumulation in mouse liver after feeding a HF diet. Chartoumpekis et al. found a decrease in nuclear abundance and DNA binding activity of Nrf2 during adipogenesis in ST2 cells. Vomhof-Dekrey et al. speculate that these controversial data may be mainly due to the differential use of primary versus immortalized MEFs and the different experimental periods. It is highly recommended to construct adipose tissue-specific Nrf2 knockout mice to investigate its role in adipogenesis.
From an evolutionary standpoint, obesity protects humans from potential threat of food shortage through energy storage. Adipocyte differentiation is a protective mechanism essential for survival, in which energy is stored in the form of TG in the adipose tissue. In this tendency, adipocyte differentiation is a protective mechanism essential for survival. However, this protective mechanism would lead to obesity without control, causing a series of unhealthy problems. In obese individuals, WAT release proinflammatory factors, resulting in metabolic dysfunctions [461]. Moreover, ectopic accumulation of fat in other tissues is detrimental for normal physiological functions, leading to several serious diseases.
In aerobic organisms, the capability of oxygen consumption precedes the onset of advanced life and promotes the biological evolution. Three decades ago, ROS was considered to be generated as by-products of oxygen consumption. Current concepts support that ROS may be evolutionally utilized as essential factors for the regulation of various biological processes that reflect the essence of “concise” in life activity. In the last decades, more and more evidences have shown the active role of ROS in the regulation of life processes rather than the passive role in inducing oxidative injury.
Based on the current results and the complexity of redox system, we propose that there is a complex and interactive “Redox Chain,” consisting of “ROS-generating Enzyme Chain,” “Combined Antioxidant Chain,” and “Transcription Factor Chain” (Figure 1). On the oxidant side, “ROS-generating Enzyme Chain” consisted of enzymes that are responsible for intracellular ROS generation, including ROS-generating enzymes in mitochondria and ER, NOXs, XOR, NOS, LOX, COX, CYPs, and other undefined enzymes that may be involved in ROS production. In response to physiological and/or pathophysiological stimuli, these enzymes could be activated in a temporal and spatial sequence and ROS could be generated in different time periods and at different positions from various sources in a cell. ROS is probably produced by the activation of certain enzymes that could quickly respond to intra- or extracellular stimulation, such as NOX. Under continued pathological conditions, it is likely that ROS would mainly come from metabolic burden, such as OXPHOS and ER stress. On the antioxidant side, “Non-enzymatic Antioxidant Chain” comprises SOD, CAT, GCL, GPx, GSTs, Trxs, Prxs, NQO1, HO-1, SeP, MTs, and other undefined enzymes that may exhibit antioxidant activities, in combination with nonenzymatic antioxidants, constituting the first in vivo defense line against potential ROS insult. This defense chain could be reinforced by the consumption of naturally extracted antioxidants that are commercially available. On a higher level, the antioxidant enzymes constitute the “Antioxidant Enzyme Chain,” a more efficient defense line against potential ROS damage. These enzymatic and nonenzymatic antioxidants comprise the “Combined Antioxidant Chain,” which sequentially and cooperatively regulate redox state. As the super directors, the redox-regulating transcription factors, such as Nrf2, PGC-1α, p53, NF-κB, and FoxO, form the “Transcription Factor Chain,” which direct the battle between oxidants and antioxidants through transcriptionally regulating both ROS-generating enzymes and antioxidant enzymes. Moreover, there are additional mechanisms of redox regulation, including epigenetic and posttranslational regulation. The regulation of “Redox Chain” could be implemented in a temporal and spatial way. In all, more work is needed to clarify the role of each member in redox family and its regulatory mechanisms under normal and pathophysiological conditions.
A hypothesis of “Redox Chain.” The innermost line on the left indicates “Nonenzymatic Antioxidant Chain”. The middle line on the left indicates “Antioxidant Enzyme Chain”. The innermost line on the right indicates “Oxidant Chain”. The middle line on the right indicates “ROS-generating Enzyme Chain”. The outermost line indicates “Transcription Factor Chain”.
Disturbance of “Redox Chain” results in either oxidative stress or blockage of ROS signaling transduction, leading to various disorders. In the context of obesity or enhanced adipocyte differentiation, redox state is altered as reflected by disorganized “Redox Chain” at different levels. However, we only observed changes or roles of single or several members of “Redox Chain” in adipocyte differentiation. Clarification of temporal and spatial changing pattern of “Redox Chain” in an integrated way will undoubtedly contribute to the understanding of the mechanisms underlying adipocyte differentiation and obesity.
Based on the large amount of paradoxical results, it is difficult to draw a “clear” conclusion of the role of redox regulation in adipocyte differentiation. What we know is that adipocyte differentiation is definitely determined either directly by redox system or indirectly by redox-sensitive transcriptional, posttranslational, or epigenetic regulation. Compared with preadipocytes, mature adipocytes are under relatively more oxidizing state. In the physiological process of adipocyte differentiation, ROS is a definitely required and essential signal for the initiation and maintenance of adipogenic events. However, ROS-generating signals may be limited to a certain concentration and sources in different stages of adipogenic differentiation. We propose that in addition to the differential experimental backgrounds, including differences in experimental subjects and interventions, the paradoxical results from different laboratories may be attributed to the “narrow safety window” of concentration, time and compartment for the redox-regulated adipogenic differentiation. The difference of genetic or dietary background of the subjects, doses or periods of interventions or “time window” selected for detection may severely affect the outcomes. Concentration is a key determinant of the effect of ROS on adipocyte differentiation. Turker et al. reported that 1 and 10 μM H2O2 resulted in a marked decrease in adipocyte differentiation, while higher doses of H2O2 markedly increased differentiation. Both deficiency and over-expression of XOR have been shown to inhibit adipocyte differentiation. In the course of adipocyte differentiation, cells undertake dramatic transformation of cell fate, involving drastic changes of organelles and compartments. In differential stages of adipogenesis, redox state in different organelles and compartments may be differential. These data demonstrated that redox state required for adipocyte differentiation must be controlled in a fine-tuned way, including appropriate concentration, time, and space. The paradoxical results indicate the importance of delicate regulation of redox system in adipocyte differentiation. Due to the quick redox reaction and extremely short half-life period, to date, the absolute amount of ROS could not be determined accurately in biological samples, and thus the ROS level we evaluated is a relative index. In different experiments, ROS level could not be compared. Thereof, over-expression or inhibition of a certain molecule (transcription factor, antioxidant enzyme, or ROS-generating enzyme) could not reflect the accurate level of ROS or redox state. Moreover, the adipogenic differentiation requires physiologically and endogenously generated ROS to create an oxidizing environment and to transduce molecular signals. The differentiation of adipocytes needs the accurate generation of ROS at the appropriate “time window” and “place.” However, under pathological and stress conditions, ectopic and excessive generation of ROS may influence the redox environment and disturb adipogenesis. It is proposed that ROS acts as a double-edged sword in the course of adipocyte differentiation and thus in the pathogenesis of obesity and related metabolic disorders (Figure 2) [12].
Role of redox system in the regulation of adipocyte differentiation.
During adipocyte differentiation, cells undertake dramatic and extensive morphological, physiological, and biochemical changes. These great changes render redox alteration the most probable “candidate” that could mediate those extensive reactions. The large amount and extensive distribution of members in redox family provide a general and microenvironment for thousands of reactions.
Adipocyte differentiation is a time-dependent process, which consists of different stages, including confluence, MCE, early differentiation, and late differentiation. In different stages of adipocyte differentiation, redox status is dramatically altered. It appears that redox system works as an “on-off switch” and orchestrates the transition from preadipocyte to adipocyte. During this process, redox regulation on the expression of key proteins for adipocyte differentiation may occur in diverse levels, including transcriptional, epigenetic, and posttranslational modulations (Figure 3). Indeed, the “tridimensional” redox regulatory mechanism is not limited in the process of adipocyte differentiation, but has general implications in a variety of biological processes.
The “tridimensional” redox regulatory mechanism in the process of adipocyte differentiation.
A specific molecular mechanism would definitely play a role in a given background. However, one molecule or even one signaling pathway could not account for all the redox-sensitive events. Thereof, a more integrated view of redox biology should be highlighted. Considering the burst of various omics concept and technology, we suggest that redoxomics should be applied extensively and widely to help us understand the biological effects of redox alterations in a more integrated way. We need a stable and standardized “redox index” to evaluate the general redox status under a certain condition. Just like BMI, “redox index” represents a value to assess the overall situation of redox state in an organ or in a whole body. In our future work, we need to find the representative and easily obtained indicators and establish a simple equation to calculate “redox index.” Moreover, adipose tissue-targeted redox interventions may be more attractive because disturbance of insulin signaling in other main organs, such as liver, could also result in insulin resistance and metabolic syndromes. Differential redox state in different organs may influence the overall metabolic end-points by using antioxidants. Our aim is to establish and maintain general and fine-tuned redox balance rather than to conduct simple prooxidant and antioxidant interventions. Individualized intervention according to their respective redox state should be emphasized to treat obesity and other redox-related metabolic disorders [13].
This work was supported by National Natural Science Foundation of China (No. 31400724) and Natural Science Foundation of Shaanxi Province (2014JQ4135). We reused some content we published in Free Radical Biology and Medicine and got the permission.
Megadiverse countries constitute exceptional areas on Earth where most of the planetary biodiversity is present. The complexity of these areas is huge, but in most of the cases, two major points are key: (1) the geographical location, and (2) the abiotic and biotic elements present. Mexico is one of the top five megadiverse countries in the world and its macrodiversity and endemism are well represented by amphibians, mammals, plants, and reptiles [1]. However, the knowledge of microscopic organisms such as archaea, bacteria, protozoa, microscopic algae, and microscopic fungi, that inhabit aquatic, atmospheric, marine, and soil ecosystems is neither poorly known, studied nor understood.
The vision of this chapter is to contribute to the knowledge, research, and study of microscopic life in different Mexican ecosystems, as they are often ignored or poorly mentioned in federal texts or even in biotic inventories. Our examples are some members of the class actinobacteria [2], and we aim to demonstrate why it is so important to study these bacteria in such detail to fully explore and untap the unknown actinobacterial diversity with potential in biology. Using a dynamic isolation strategy on air, soil, and marine sediments and sponges collected from yet unexplored sites of the Mexican territory, we have been able to cultivate novel actinobacteria. Our findings showed that expertise, patience, and talent of the techniques applied are keys in the hunt for new potential microbes.
The isolation of microorganisms, including actinobacteria, is not new but a dynamic strategy that is continuously changing, and the developed to date is a powerful tool. For more than two centuries, researchers from Japan, the UK, and USA have shown that beneficial microorganisms isolated from the soil are important to Biology. In recent years the isolation of the first genus of actinobacteria from the marine origin [3] and novel marine species [4] have shown the importance of exploring the marine environment. Extreme or unexplored sites have also shown the isolation of actinobacteria including putative novel actinobacteria [5].
Our research studying actinobacteria started in 1999 [6, 7], but until 2009 we properly started the exploration of the Mexican (marine) ecosystems [8] as an independent group. We followed bioprospecting, diversity, and systematic approach but designing a selective isolation strategy was the first step for a complete full project or protocol [9].
Actinobacteria is a complex group of bacteria, they present forms such as rods or bacilli, many differentiate in vegetative mycelium, aerial hyphae, and chain of spores, and in a few genera fragmentation of the hyphae is present. In general, the Gram reaction is positive and the content of guanine plus cytosine is above 69%mol. The morphological characteristics within the class showed how complex this group is. Actinobacteria are considered saprophytes or beneficial microbes, but a small number of species have been shown to be either pathogenic [10] or opportunistic [11]. This microbial group has been isolated or cultivated using classical methods from almost every sample taken on Earth and they are always detected when using molecular methods to study this group in a given environmental sample.
Actinobacteria also have the innate ability to produce secondary metabolites with biological activity, to date, this class encompasses 80% of the microbes that produce the antibacterial compounds used in medicine. Complete Genome Sequencing of some genera of actinobacteria such as
The more we study and discover actinobacteria the more important they become in pass, present, and future assignments. Microorganisms and microbial biomass, including actinobacteria, represent the major resource for biotechnology and biological areas. We should continue exploring their role in nature in order to understand their biology, ecology, and bioprospecting potential [13, 14, 15, 16].
The Earth’s atmosphere is divided into six specific layers with completely different characteristics: (1) Troposphere, (2) Stratosphere, (3) Mesosphere, (4) Thermosphere, (5) Ionosphere, and (6) Exosphere. It has been established that the atmosphere plays an important role to transport microorganisms, place to place, continent to continent. The latter has been established using scientific tools in the last 200 years and in the last 15 years, NASA has monitored mineral dust particles from the Sahara desert with a robust precision using spaceborne satellites. These Saharan dust plumes contain microorganisms and enter mainland Mexico by the Yucatan Peninsula [17].
The atmosphere is a hostile environment for microorganisms though there are a significant number of them in the troposphere, with air as their main dispersion pathway. The abundance, diversity, survival, and transport of microorganisms, as passive drivers, and how they get stressed severely by the conditions presented in the atmosphere have fully been reported [18]. Most of the microorganisms in the atmosphere are present as spores, while others have adapted to resist desiccation or high/low temperatures [19]. Recent reports have also shown that some microorganisms (i.e., by using specific proteins) can act as ice nucleating particles [20] and that they may play an important role in cloud formation [21]. In general, bacteria (including actinobacteria) present in the atmosphere are attached to suspended particles [17], and their concentration change notably during the dry or wet seasons of each year [22].
As part of the African Dust and Biomass Burning Over Yucatan (ADABBOY) Project [23] in the city of Merida (N 21°02´75.4´´ W 89°65´44.8´´) a selective isolation strategy was carried out in order to cultivate/recovered putative actinobacteria in May 2017. Air samples were impacted using a Quick Take 30 Sample Pump® and a BioStage® SKC (Figure 1) in Petri dishes prepared with a slightly modified Glucose Yeast Malt extract agar (GYM medium; Appendix A; Medium 65: DSMZ; www.dsmz.de) supplemented with Rifampicin (5 μg/mL; Sigma-Aldrich, USA) and Nystatin (50 μg/mL; MICOSTATIN® Bristol Myers Squibb, Mexico).
The device used for the air particles.
Plates were incubated in two different laboratories and conditions. The first laboratory was in the city of Merida at the Universidad Autónoma de Yucatán, using an aerobic incubator set at 25°C and the plates were incubated for 24 hours. For the second procedure, the plates were transported to a laboratory in Mexico City where the incubation time continued aerobically at 30°C (IncuMax IC-320, Amerex USA) for 8 weeks with eye observation each week. One microorganism with the production of aerial hyphae, a gray mass of spores, and a very deep purple diffusible pigment (Figure 2) was selected from the isolation plates for further studies.
Morphology and purple diffusible pigment of an airborne streptomycete.
The selected isolate was coded C6-CCA-May-1. After a purification process using GYM medium and two different techniques (cross streak and serial dilutions), bacterial biomass and spores of the strain were ultra-preserved in 20% glycerol. Morphological characterization was carried out using a GYM medium (Figure 3) and a Gram staining procedure (Figure 4) was carried out following well-known universal protocols.
Aerial hyphae and spore mass of isolate C6-CCA-May-1.
Gram staining of the airborne streptomycete.
Molecular identification of strain C6-CCA-May-1 was carried out following protocols previously published [8, 24]. First, the DNA of strain C6-CCA-May-1 was extracted and used as a template for PCR amplification using the 16S rRNA gene (Appendix B). The sequence of the 16S rRNA gene PCR product confirmed that strain C6-CCA-May-1 belongs to the genus
Hit taxon name | Hit strain name | Accesion | Similarity | Hit taxonomy | Completeness (%) |
---|---|---|---|---|---|
NBRC 133559T | AB18350 | 99.51 | Bacteria;Actinobacteria;Actinobacteria_c;Streptomycetales;Streptomycetaceae;Streptomyces | 99.6 | |
NBRC 13404T | AB18381 | 98.88 | 99.9 | ||
KNN 35.1bT | LT621750 | 98.77 | 95.5 | ||
NBRC 13000T | AB184249 | 98.66 | 99.9 | ||
NBRC 15423T | AB184670 | 98.66 | 99.0 |
List of hits from the EZbiocloud 16S database.
A Bayesian phylogenetic tree was constructed in order to establish the taxonomic position of
Phylogenetic tree of the 16S rRNA gene of the airborne streptomycete.
Streptomycetes are an ecologically important group capable of producing diverse bioactive compounds. However, their taxonomy and diversity in air samples remain unknown. For almost two centuries the genus
Seventy percent of our planet is covered by the ocean but from one marine research project, there are 10 of terrestrial origin. Little is still known about marine biodiversity (including microorganisms) though their potential is extraordinary and needs to be fully studied and exploited. Mexico is surrounded by the Pacific Ocean, the Sea of Cortez (
In the present project, a total collection of 34 marine sediments or sponges were collected at RANP during two expeditions (December 2017 and January 2018). A selective isolation strategy using 11 of the marine sediments and two different media was developed following a previously reported study [24]. In order to isolate marine obligate and nonobligate actinobacteria, 1 g of wet sediments was transferred to tubes containing 9 mL of saline solution (0.9%; NaCl; Sigma-Aldrich, Mexico), four dilutions were prepared (10−1 to 10−4) and 100 μL (Gilson, France) of each dilution were spread onto marine GYM medium and 1:10 marine GYM medium (Appendix A); both media supplemented with Rifampicin [15, 25 and 50 μg/mL] and Nystatin (100 μg/mL). Plates were then aerobically incubated at 30°C (IncuMax IC-320, Amerex USA) for up to 16 weeks. Starting at week eight, the isolation plates were checked by eye looking for actinobacterial colonies. Once putative colonies were noticed each was then streaked in new GYM plates without antibiotics or antifungal compounds until an axenic culture was obtained. The conditions of incubation were as mentioned above. Because of the pressure set in the isolation strategy, not many microbes were able to grow but actinobacteria were successfully cultivated.
A preliminary test to quickly select marine obligate actinobacteria was carried out using marine GYM medium (Figure 6A and C) and GYM medium (Figure 6B and D). A positive result was considered when nonmicrobial biomass was observed growing on the surface of GYM medium after 4 weeks of incubation (Figure 6B). The ones that presented growth only in the marine GYM medium were considered those marine obligate actinobacteria (Figure 6A). It should be pointed out, however, that we were also able to isolate nonobligate actinobacteria that showed the typical characteristics of members of the family Micromonosporaceae [32] (Figure 6C and D). Up to date there is only one genus that is considered halophile within the Phylum Actinobacteria, and this is
Screening of obligate and nonobligate marine actinobacteria.
The molecular identification using the 16S rRNA gene of obligate and nonobligate marine actinobacteria confirmed that they belong to the genera
Code of microorganism | Identity (%) | Hit taxonomy | Accesion |
---|---|---|---|
C114 col. 1 | 99 | GD145235.1 | |
C60 b bca col. 1 | 100 | MH299440.1 | |
C60a col. 3 | 100 | KX394599.1 | |
C60 col. 4 | 99 | KX394598.1 | |
C72 col. 2 | 100 | AG506245.1 | |
C134 col. 4 | 99 | CF133005.1 |
Taxonomic identification of some of the obligate and nonobligate marine actinobacteria from RANP.
To isolate obligate and nonobligate marine actinobacteria from the sponge samples, five different species of
Strategy to isolate actinobacteria from six marine sponges.
Starting at week eight, the isolation plates were checked by eye looking for actinobacterial colonies (Figure 8). Once putative colonies were selected they were streaked in new GYM plates until axenic culture were obtained. The conditions of incubation were the same as mentioned before.
Morphology of the marine obligate actinobacteria.
The preliminary test to select marine obligate actinobacteria was carried out as mentioned previously. Obligate marine actinobacteria (Figure 9A) were isolated from
Morphology of obligate and nonobligate marine actinobacteria.
We isolated marine obligate actinobacteria that were preliminarily assigned to the genus
The microbial communities of marine obligate and nonobligate actinobacteria associated with marine sediments remain poorly characterized [34] and we must continue searching for these gifted microorganisms [35]. Culture-dependent methods captured approximately 3% of the total count of the microbes and in some reports around 39 genera have been only detected in culture. The latter shows the importance to carry out/improve, innovative and original selective isolation techniques since these may be more effective than previously recognized.
Soil is one of the most complex ecosystems on Earth and its amount of organic matter, mineral composition, and diversity of microorganisms will determine its ecology. There are different kinds of soils but in general, those with less anthropogenic impact will be richer in microorganisms. Mexico encompasses 26 types of soil out of the 32 recognized in the world [36, 37] and this is due to several causes, namely: (1) the complexity of the topography originated from the volcanic activity in the Cenozoic Era, (2) the wide altitudinal gradient (from 0 to 5, 600 m.a.s.l.), (3) by the five main climates present according to the Köppen classification [38], (4) the enormous diversity of landscapes present and, finally (5) the different kind of rocks that the Mexican territory enclose. It is well recognized that actinobacteria are abundant in soils and that they play an important role in the degradation and recycling of organic matter. Soil microorganisms have a remarkable ability to produce compounds with biological activity such as antibiotics, and historically this has been exemplified by streptomycin which is produced by a streptomycete named
Mexico encompasses nearly 4, 000 insular regions of outstanding natural beauty, their biodiversity is remarked by high number of endemism (plants and animals) and most of these regions are federal protected. Revillagigedo Archipelago National Park (RANP) [39] encompasses four tropical volcanic Islands: (1) Socorro, (2) Clarion, (3) San Benedicto, and (4) Roca Partida. The RANP is considered as one of the best-preserved areas in the world.
In the present project, soil samples were collected at five different sites of Socorro Island (SI) (Figure 10) in 2016 and 2017, respectively (Table 3). A selective isolation strategy using five soil samples and three different media was developed in order to isolate actinobacteria. One gram of each soil was transferred to tubes containing 9 mL of saline solution (0.9%; NaCl; Sigma-Aldrich, Mexico). Modified Pikovskaya agar (Appendix A), GYM without antibiotics or antifungal compounds, and marine GYM supplemented with Rifampicin [5 μg/mL] and Nystatin (100 μg/mL) were used as isolation media. The dilutions used for modified Pikovskaya and marine GYM were 10−1 to 10−4 and for GYM 10−6 to 10−8. One hundred microliters (Gilson, France) of each dilution were spread in the media and then aerobically incubated at 30°C (IncuMax IC-320, Amerex USA) for up to 4 weeks.
Sites of sampling in Socorro Island.
Code of sampling site | Name of sampling site | Meter of above sea level (masl) | Date of sampling | ||
---|---|---|---|---|---|
S1 | Camping place | South of Socorro Island | 450 | 25 | December 2016 |
S2 | Cave | 550 | 28 | ||
S3 | North camping place | North of Socorro Island | 941 | 30 | |
S4 | Everman volcano | South East of Everman volcano | 850 | 3 | January 2017 |
S5 | Parrot camping place | 600 |
General information of the sampling sites.
The isolation plates were checked by eye looking for actinobacteria and selected colonies were streaked in new GYM media plates until axenic cultures were obtained (Figure 11). The conditions of incubation were the same as mentioned above. The cultivable actinobacteria diversity was remarkable but only 215 isolates were selected from the isolation plates. Eighty-six isolates presented morphological differentiation based on aerial hyphae and spore mass so that they were considered as streptomycetes (Figure 12).
Isolation plate (a), selected actinobacteria (b), and axenic culture (c).
Morphological diversity of actinobacteria from Socorro Island soils.
Molecular identification using the 16S rRNA gene of 10 selected strains morphologically resembling streptomycetes confirmed that they indeed belong to this extensive and important genus (Table 4). The phylogenetic tree constructed using five of sequences of the selected strains showed that they are different amongst them and from those more related
Code of microorganism | Hit taxonomy | Identity (%) |
---|---|---|
C1-S1-1 | 99 | |
C10-S2-14 | ||
C67-S2-1 | ||
C43-S3-1 | ||
C43-S3-2 | ||
C58-S5-2 | ||
C59-S5-1 | ||
C59-S5-2 | ||
C27-S4-3 | ||
C57-S5-1 |
Preliminary identification of selected streptomycetes from the different sites.
Phylogenetic tree of the 16S rRNA gene selected streptomycetes isolates.
Soil actinobacteria, particularly
More than two centuries of work to isolate actinobacteria from natural resources have ended in major discoveries, academic contributions, and important recognitions. Novel actinobacteria represent the entree of new natural compounds of significant importance. For more than 15 years our research group has developed and applied selective isolation strategies exploring distinct natural unexplored sites or less studied ecological niches in Mexico. To avoid the isolation of the “same bugs,” it is needed to use different selective isolation strategies, pre-treatments of the environmental sample, supplementation of the media with a specific concentration of antibiotics and antifungal compounds while carefully selecting the target organisms.
The results presented in this chapter support the proposal that the isolation of microorganisms is not “
Actinobacteria is one most of the diverse and complex groups of bacteria and produces more than 80% of the antibiotics used in medicine today. Their ability to produce novel natural compounds, such as antibiotics and novel cancer compounds is widely recognized. According to the World Health Organization (WHO) there is an urgent need to discover novel antibiotics for priority pathogenic bacteria and emerging pathogenic organisms [43] and the first step to respond and to contribute with this global initiative is to isolate novel actinobacteria. To our knowledge, our reports here are one of the few research projects in our country that are dedicated to study the ecological role and the genetic potential of novel actinobacteria from unexplored sites or less studied ecological niches in Mexico.
Our research was supported by Instituto Politécnico Nacional (IPN)—Secretaría de Investigación y Posgrado (SIP), Grants SIP 20170432, 20170434, 20170410, 20181167, 20181528, 20181803, 20196605, 20196630, 20196649, 20201026, 20201893, 20202083, 20210987, 20211209, and 20211740. L.C.-C. was supported by a Ph.D. Scholarship from Consejo Nacional de Ciencia y Tecnología (CONACyT, Mexico) no. 270230 and Beca de Estímulo Institucional de Formación de Investigadores Program (BEIFI-IPN). ETQ, CJHG and JCCD acknowledge Comisión de Operación y Fomento de Actividades Académicas del Instituto Politécnico Nacional (COFAA), Estímulo al Desempeño de los Investigadores (EDI) and Sistema Nacional de Investigadores (SNI-CONACYT) fellowships. A.A-V ackcnowledges a Mexican Postdoctoral Scholarship Program 3 and 4, Program 4, 2020–2021 and Program 1 and 2, Program 2, 2021–2022 (CONACyT, Mexico).
The authors declare no conflict of interest.
Glucose Yeast Malt Extract Agar -GYM medium- (DSM medium 65) | |
---|---|
Dextrose (Bacto™, BD) | 4 g |
Yeast extract (Bacto™, BD) | 4 g |
Malt extract (Bacto™, BD) | 10 g |
Calcium carbonate (SIGMA-ALDRICH) | 2 g |
Agar (Bacto™, BD) | 12 g |
Distilled water | 1000 mL |
pH 7.2 |
Marine media was prepared by replacing distilled water with artificial seawater (Ocean™). 1:10 marine GYM medium was prepared using an aliquot of marine GYM. All the media was sterilized at 121°C, 1.5 Lb. for 15 min.
Reagents | Volume |
---|---|
10× DNA polymerase buffer [50 mM stock solution] (Bioline, USA) | 5 μL |
MgCl2 [50 mM] (Bioline, USA) | 1.5 μL |
dNTPs [10 mM stock mixture] (Bioline, USA) | 1.25 μL |
Primer 27f [20 μM stock solution] (Invitrogen) | 0.5 μL |
Primer 1525r [20 μM stock solution] (Invitrogen) | 0.5 μL |
DNA [100 ng/μL] | 1 μL |
Taq polymerase [5 U] (Bioline, USA) | 1 Unit |
Ultra-pure Milli-Q water | Up to 50 μL |
Amplification was achieved using a Techno 512 gradient PCR machine.
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In this review, we outline the evidence of gender differences related to PTSD, and the factors of resilience and susceptibility differ between men and women.",book:{id:"5472",slug:"gender-differences-in-different-contexts",title:"Gender Differences in Different Contexts",fullTitle:"Gender Differences in Different Contexts"},signatures:"Jingchu Hu, Biao Feng, Yonghui Zhu, Wenqing Wang, Jiawei Xie\nand Xifu Zheng",authors:[{id:"190985",title:"Dr.",name:"Xifu",middleName:null,surname:"Zheng",slug:"xifu-zheng",fullName:"Xifu Zheng"},{id:"194981",title:"BSc.",name:"Yonghui",middleName:null,surname:"Zhu",slug:"yonghui-zhu",fullName:"Yonghui Zhu"},{id:"194982",title:"MSc.",name:"Wenqing",middleName:null,surname:"Wang",slug:"wenqing-wang",fullName:"Wenqing Wang"},{id:"194985",title:"Dr.",name:"Jingchu",middleName:null,surname:"Hu",slug:"jingchu-hu",fullName:"Jingchu Hu"},{id:"194986",title:"MSc.",name:"Biao",middleName:null,surname:"Feng",slug:"biao-feng",fullName:"Biao Feng"},{id:"194987",title:"Ph.D. Student",name:"Jiawei",middleName:null,surname:"Xie",slug:"jiawei-xie",fullName:"Jiawei Xie"}]},{id:"52472",title:"Gender and Health",slug:"gender-and-health",totalDownloads:3430,totalCrossrefCites:5,totalDimensionsCites:11,abstract:"Research has found differences between women and men in some health indicators. Women’s life expectancy is higher than men’s, but research on differences in morbidity has proved less consistent than on the differences in mortality. These differences vary in terms of the type of health indicator used, the life cycle period analyzed, and even the country where research is conducted. Generally, men have more life-threatening chronic diseases at younger ages, including coronary heart disease, as well as more externalizing mental health problems and substance use disorders. Women present higher rates of chronic debilitating conditions such as arthritis, frequent or severe headaches, gallbladder conditions, and also more internalizing mental problems such as affective and anxiety disorders. Results of research on the differences between women and men in self-rated health have also highlighted the complexity of gender differences in health. Although several studies have shown that women have poorer self-rated health than men, this is not the case in all countries. Also, differences in self-rated health vary depending on other psychosocial and demographic variables. The present study reviews the main differences in women’s and men’s health as well as the most relevant factors that may account for them.",book:{id:"5472",slug:"gender-differences-in-different-contexts",title:"Gender Differences in Different Contexts",fullTitle:"Gender Differences in Different Contexts"},signatures:"María Pilar Matud",authors:[{id:"189729",title:"Prof.",name:"M. Pilar",middleName:null,surname:"Matud",slug:"m.-pilar-matud",fullName:"M. Pilar Matud"}]},{id:"53212",title:"Broken Dreams—Balancing Self and Family Well-Being: The Experiences of Women Immigrants to Hamilton, ON",slug:"broken-dreams-balancing-self-and-family-well-being-the-experiences-of-women-immigrants-to-hamilton-o",totalDownloads:1512,totalCrossrefCites:0,totalDimensionsCites:0,abstract:"This chapter presents the preliminary analysis of a study conducted in Hamilton, ON. It explores the intersection of women’s immigration, integration and mental health. Their perceptions of what is needed from them in relation to the various challenges/changes that moving to a new country entails is a particular focus of this research. To begin with, the term “women immigrant” (WI) is used, rather than immigrant women as commonly used—as the participants were women long before they became immigrants. 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In the process of validating a scale of consumer boycott in Brazil, the 13 items of the original scale were kept, but were grouped in different factors. The emerged factors were perception of guilt, influence from others, boycott efficiency, and purchase frequency. Testing relationship among demographic variables and these factors, only gender was significant on perception of guilt. In this sense, we seek in psychology, psychoanalysis (also briefly in anthropology and history), features that could explain the reasons why women feel guiltier than men, and thus are more likely to boycott.",book:{id:"5472",slug:"gender-differences-in-different-contexts",title:"Gender Differences in Different Contexts",fullTitle:"Gender Differences in Different Contexts"},signatures:"Breno de P.A. Cruz, Ricardo José Marques Pires-Jr. and Steven D. 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That is exactly what he does, diving into Machine Learning algorithms and technologies to help TECNALIA to decide whether something is great in theory or will actually impact on the product or processes of its projects. So, he is expert at framing experiments, developing hypotheses, and proving whether they’re true or not, in order to investigate fundamental problems with a longer time horizon. He is also able to design and develop PoCs and system prototypes in simulation. He has participated in several national and internacional R&D projects.\n\nAs another relevant part of his everyday research work, he usually publishes his findings in reputed scientific refereed journals and international conferences, occasionally acting as reviewer and Programme Commitee member. Concretely, since 2018 he has published 9 JCR (8 Q1) journal papers, 9 conference papers (e.g. ECML PKDD 2021), and he has co-edited a book. He is also active in popular science writing data science stories for reputed blogs (KDNuggets, TowardsDataScience, Naukas). Besides, he has recently embarked on mentoring programmes as mentor, and has also worked as data science trainer.",institutionString:"TECNALIA Research & Innovation",institution:{name:"Tecnalia",country:{name:"Spain"}}},{id:"103779",title:"Prof.",name:"Yalcin",middleName:null,surname:"Isler",slug:"yalcin-isler",fullName:"Yalcin Isler",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRyQ8QAK/Profile_Picture_1628834958734",biography:"Yalcin Isler (1971 - Burdur / Turkey) received the B.Sc. degree in the Department of Electrical and Electronics Engineering from Anadolu University, Eskisehir, Turkey, in 1993, the M.Sc. degree from the Department of Electronics and Communication Engineering, Suleyman Demirel University, Isparta, Turkey, in 1996, the Ph.D. degree from the Department of Electrical and Electronics Engineering, Dokuz Eylul University, Izmir, Turkey, in 2009, and the Competence of Associate Professorship from the Turkish Interuniversity Council in 2019.\n\nHe was Lecturer at Burdur Vocational School in Suleyman Demirel University (1993-2000, Burdur / Turkey), Software Engineer (2000-2002, Izmir / Turkey), Research Assistant in Bulent Ecevit University (2002-2003, Zonguldak / Turkey), Research Assistant in Dokuz Eylul University (2003-2010, Izmir / Turkey), Assistant Professor at the Department of Electrical and Electronics Engineering in Bulent Ecevit University (2010-2012, Zonguldak / Turkey), Assistant Professor at the Department of Biomedical Engineering in Izmir Katip Celebi University (2012-2019, Izmir / Turkey). He is an Associate Professor at the Department of Biomedical Engineering at Izmir Katip Celebi University, Izmir / Turkey, since 2019. In addition to academics, he has also founded Islerya Medical and Information Technologies Company, Izmir / Turkey, since 2017.\n\nHis main research interests cover biomedical signal processing, pattern recognition, medical device design, programming, and embedded systems. He has many scientific papers and participated in several projects in these study fields. He was an IEEE Student Member (2009-2011) and IEEE Member (2011-2014) and has been IEEE Senior Member since 2014.",institutionString:null,institution:{name:"Izmir Kâtip Çelebi University",country:{name:"Turkey"}}},{id:"339677",title:"Dr.",name:"Mrinmoy",middleName:null,surname:"Roy",slug:"mrinmoy-roy",fullName:"Mrinmoy Roy",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/339677/images/16768_n.jpg",biography:"An accomplished Sales & Marketing professional with 12 years of cross-functional experience in well-known organisations such as CIPLA, LUPIN, GLENMARK, ASTRAZENECA across different segment of Sales & Marketing, International Business, Institutional Business, Product Management, Strategic Marketing of HIV, Oncology, Derma, Respiratory, Anti-Diabetic, Nutraceutical & Stomatological Product Portfolio and Generic as well as Chronic Critical Care Portfolio. A First Class MBA in International Business & Strategic Marketing, B.Pharm, D.Pharm, Google Certified Digital Marketing Professional. Qualified PhD Candidate in Operations and Management with special focus on Artificial Intelligence and Machine Learning adoption, analysis and use in Healthcare, Hospital & Pharma Domain. Seasoned with diverse therapy area of Pharmaceutical Sales & Marketing ranging from generating revenue through generating prescriptions, launching new products, and making them big brands with continuous strategy execution at the Physician and Patients level. Moved from Sales to Marketing and Business Development for 3.5 years in South East Asian Market operating from Manila, Philippines. Came back to India and handled and developed Brands such as Gluconorm, Lupisulin, Supracal, Absolut Woman, Hemozink, Fabiflu (For COVID 19), and many more. In my previous assignment I used to develop and execute strategies on Sales & Marketing, Commercialization & Business Development for Institution and Corporate Hospital Business portfolio of Oncology Therapy Area for AstraZeneca Pharma India Ltd. Being a Research Scholar and Student of ‘Operations Research & Management: Artificial Intelligence’ I published several pioneer research papers and book chapters on the same in Internationally reputed journals and Books indexed in Scopus, Springer and Ei Compendex, Google Scholar etc. Currently, I am launching PGDM Pharmaceutical Management Program in IIHMR Bangalore and spearheading the course curriculum and structure of the same. I am interested in Collaboration for Healthcare Innovation, Pharma AI Innovation, Future trend in Marketing and Management with incubation on Healthcare, Healthcare IT startups, AI-ML Modelling and Healthcare Algorithm based training module development. I am also an affiliated member of the Institute of Management Consultant of India, looking forward to Healthcare, Healthcare IT and Innovation, Pharma and Hospital Management Consulting works.",institutionString:null,institution:{name:"Lovely Professional University",country:{name:"India"}}},{id:"310576",title:"Prof.",name:"Erick Giovani",middleName:null,surname:"Sperandio Nascimento",slug:"erick-giovani-sperandio-nascimento",fullName:"Erick Giovani Sperandio Nascimento",position:null,profilePictureURL:"https://intech-files.s3.amazonaws.com/0033Y00002pDKxDQAW/ProfilePicture%202022-06-20%2019%3A57%3A24.788",biography:"Prof. Erick Sperandio is the Lead Researcher and professor of Artificial Intelligence (AI) at SENAI CIMATEC, Bahia, Brazil, also working with Computational Modeling (CM) and HPC. He holds a PhD in Environmental Engineering in the area of Atmospheric Computational Modeling, a Master in Informatics in the field of Computational Intelligence and Graduated in Computer Science from UFES. He currently coordinates, leads and participates in R&D projects in the areas of AI, computational modeling and supercomputing applied to different areas such as Oil and Gas, Health, Advanced Manufacturing, Renewable Energies and Atmospheric Sciences, advising undergraduate, master's and doctoral students. He is the Lead Researcher at SENAI CIMATEC's Reference Center on Artificial Intelligence. In addition, he is a Certified Instructor and University Ambassador of the NVIDIA Deep Learning Institute (DLI) in the areas of Deep Learning, Computer Vision, Natural Language Processing and Recommender Systems, and Principal Investigator of the NVIDIA/CIMATEC AI Joint Lab, the first in Latin America within the NVIDIA AI Technology Center (NVAITC) worldwide program. He also works as a researcher at the Supercomputing Center for Industrial Innovation (CS2i) and at the SENAI Institute of Innovation for Automation (ISI Automação), both from SENAI CIMATEC. He is a member and vice-coordinator of the Basic Board of Scientific-Technological Advice and Evaluation, in the area of Innovation, of the Foundation for Research Support of the State of Bahia (FAPESB). He serves as Technology Transfer Coordinator and one of the Principal Investigators at the National Applied Research Center in Artificial Intelligence (CPA-IA) of SENAI CIMATEC, focusing on Industry, being one of the six CPA-IA in Brazil approved by MCTI / FAPESP / CGI.br. He also participates as one of the representatives of Brazil in the BRICS Innovation Collaboration Working Group on HPC, ICT and AI. He is the coordinator of the Work Group of the Axis 5 - Workforce and Training - of the Brazilian Strategy for Artificial Intelligence (EBIA), and member of the MCTI/EMBRAPII AI Innovation Network Training Committee. He is the coordinator, by SENAI CIMATEC, of the Artificial Intelligence Reference Network of the State of Bahia (REDE BAH.IA). He leads the working group of experts representing Brazil in the Global Partnership on Artificial Intelligence (GPAI), on the theme \"AI and the Pandemic Response\".",institutionString:"Manufacturing and Technology Integrated Campus – SENAI CIMATEC",institution:null},{id:"1063",title:"Prof.",name:"Constantin",middleName:null,surname:"Volosencu",slug:"constantin-volosencu",fullName:"Constantin Volosencu",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/1063/images/system/1063.png",biography:"Prof. Dr. Constantin Voloşencu graduated as an engineer from\nPolitehnica University of Timișoara, Romania, where he also\nobtained a doctorate degree. He is currently a full professor in\nthe Department of Automation and Applied Informatics at the\nsame university. Dr. Voloşencu is the author of ten books, seven\nbook chapters, and more than 160 papers published in journals\nand conference proceedings. He has also edited twelve books and\nhas twenty-seven patents to his name. He is a manager of research grants, editor in\nchief and member of international journal editorial boards, a former plenary speaker, a member of scientific committees, and chair at international conferences. His\nresearch is in the fields of control systems, control of electric drives, fuzzy control\nsystems, neural network applications, fault detection and diagnosis, sensor network\napplications, monitoring of distributed parameter systems, and power ultrasound\napplications. He has developed automation equipment for machine tools, spooling\nmachines, high-power ultrasound processes, and more.",institutionString:'"Politechnica" University Timişoara',institution:null},{id:"221364",title:"Dr.",name:"Eneko",middleName:null,surname:"Osaba",slug:"eneko-osaba",fullName:"Eneko Osaba",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/221364/images/system/221364.jpg",biography:"Dr. Eneko Osaba works at TECNALIA as a senior researcher. He obtained his Ph.D. in Artificial Intelligence in 2015. He has participated in more than twenty-five local and European research projects, and in the publication of more than 130 papers. He has performed several stays at universities in the United Kingdom, Italy, and Malta. Dr. Osaba has served as a program committee member in more than forty international conferences and participated in organizing activities in more than ten international conferences. He is a member of the editorial board of the International Journal of Artificial Intelligence, Data in Brief, and Journal of Advanced Transportation. He is also a guest editor for the Journal of Computational Science, Neurocomputing, Swarm, and Evolutionary Computation and IEEE ITS Magazine.",institutionString:"TECNALIA Research & Innovation",institution:{name:"Tecnalia",country:{name:"Spain"}}},{id:"275829",title:"Dr.",name:"Esther",middleName:null,surname:"Villar-Rodriguez",slug:"esther-villar-rodriguez",fullName:"Esther Villar-Rodriguez",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/275829/images/system/275829.jpg",biography:"Dr. Esther Villar obtained a Ph.D. in Information and Communication Technologies from the University of Alcalá, Spain, in 2015. She obtained a degree in Computer Science from the University of Deusto, Spain, in 2010, and an MSc in Computer Languages and Systems from the National University of Distance Education, Spain, in 2012. Her areas of interest and knowledge include natural language processing (NLP), detection of impersonation in social networks, semantic web, and machine learning. Dr. Esther Villar made several contributions at conferences and publishing in various journals in those fields. Currently, she is working within the OPTIMA (Optimization Modeling & Analytics) business of TECNALIA’s ICT Division as a data scientist in projects related to the prediction and optimization of management and industrial processes (resource planning, energy efficiency, etc).",institutionString:"TECNALIA Research & Innovation",institution:{name:"Tecnalia",country:{name:"Spain"}}},{id:"49813",title:"Dr.",name:"Javier",middleName:null,surname:"Del Ser",slug:"javier-del-ser",fullName:"Javier Del Ser",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/49813/images/system/49813.png",biography:"Prof. Dr. Javier Del Ser received his first PhD in Telecommunication Engineering (Cum Laude) from the University of Navarra, Spain, in 2006, and a second PhD in Computational Intelligence (Summa Cum Laude) from the University of Alcala, Spain, in 2013. He is currently a principal researcher in data analytics and optimisation at TECNALIA (Spain), a visiting fellow at the Basque Center for Applied Mathematics (BCAM) and a part-time lecturer at the University of the Basque Country (UPV/EHU). His research interests gravitate on the use of descriptive, prescriptive and predictive algorithms for data mining and optimization in a diverse range of application fields such as Energy, Transport, Telecommunications, Health and Industry, among others. In these fields he has published more than 240 articles, co-supervised 8 Ph.D. theses, edited 6 books, coauthored 7 patents and participated/led more than 40 research projects. He is a Senior Member of the IEEE, and a recipient of the Biscay Talent prize for his academic career.",institutionString:"Tecnalia Research & Innovation",institution:{name:"Tecnalia",country:{name:"Spain"}}},{id:"278948",title:"Dr.",name:"Carlos Pedro",middleName:null,surname:"Gonçalves",slug:"carlos-pedro-goncalves",fullName:"Carlos Pedro Gonçalves",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRcmyQAC/Profile_Picture_1564224512145",biography:'Carlos Pedro Gonçalves (PhD) is an Associate Professor at Lusophone University of Humanities and Technologies and a researcher on Complexity Sciences, Quantum Technologies, Artificial Intelligence, Strategic Studies, Studies in Intelligence and Security, FinTech and Financial Risk Modeling. He is also a progammer with programming experience in:\n\nA) Quantum Computing using Qiskit Python module and IBM Quantum Experience Platform, with software developed on the simulation of Quantum Artificial Neural Networks and Quantum Cybersecurity;\n\nB) Artificial Intelligence and Machine learning programming in Python;\n\nC) Artificial Intelligence, Multiagent Systems Modeling and System Dynamics Modeling in Netlogo, with models developed in the areas of Chaos Theory, Econophysics, Artificial Intelligence, Classical and Quantum Complex Systems Science, with the Econophysics models having been cited worldwide and incorporated in PhD programs by different Universities.\n\nReceived an Arctic Code Vault Contributor status by GitHub, due to having developed open source software preserved in the \\"Arctic Code Vault\\" for future generations (https://archiveprogram.github.com/arctic-vault/), with the Strategy Analyzer A.I. module for decision making support (based on his PhD thesis, used in his Classes on Decision Making and in Strategic Intelligence Consulting Activities) and QNeural Python Quantum Neural Network simulator also preserved in the \\"Arctic Code Vault\\", for access to these software modules see: https://github.com/cpgoncalves. He is also a peer reviewer with outsanding review status from Elsevier journals, including Physica A, Neurocomputing and Engineering Applications of Artificial Intelligence. Science CV available at: https://www.cienciavitae.pt//pt/8E1C-A8B3-78C5 and ORCID: https://orcid.org/0000-0002-0298-3974',institutionString:"University of Lisbon",institution:{name:"Universidade Lusófona",country:{name:"Portugal"}}},{id:"241400",title:"Prof.",name:"Mohammed",middleName:null,surname:"Bsiss",slug:"mohammed-bsiss",fullName:"Mohammed Bsiss",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/241400/images/8062_n.jpg",biography:null,institutionString:null,institution:null},{id:"276128",title:"Dr.",name:"Hira",middleName:null,surname:"Fatima",slug:"hira-fatima",fullName:"Hira Fatima",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/276128/images/14420_n.jpg",biography:"Dr. Hira Fatima\nAssistant Professor\nDepartment of Mathematics\nInstitute of Applied Science\nMangalayatan University, Aligarh\nMobile: no : 8532041179\nhirafatima2014@gmal.com\n\nDr. Hira Fatima has received his Ph.D. degree in pure Mathematics from Aligarh Muslim University, Aligarh India. Currently working as an Assistant Professor in the Department of Mathematics, Institute of Applied Science, Mangalayatan University, Aligarh. She taught so many courses of Mathematics of UG and PG level. Her research Area of Expertise is Functional Analysis & Sequence Spaces. She has been working on Ideal Convergence of double sequence. She has published 17 research papers in National and International Journals including Cogent Mathematics, Filomat, Journal of Intelligent and Fuzzy Systems, Advances in Difference Equations, Journal of Mathematical Analysis, Journal of Mathematical & Computer Science etc. She has also reviewed few research papers for the and international journals. She is a member of Indian Mathematical Society.",institutionString:null,institution:null},{id:"414880",title:"Dr.",name:"Maryam",middleName:null,surname:"Vatankhah",slug:"maryam-vatankhah",fullName:"Maryam Vatankhah",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Borough of Manhattan Community College",country:{name:"United States of America"}}},{id:"414879",title:"Prof.",name:"Mohammad-Reza",middleName:null,surname:"Akbarzadeh-Totonchi",slug:"mohammad-reza-akbarzadeh-totonchi",fullName:"Mohammad-Reza Akbarzadeh-Totonchi",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Ferdowsi University of Mashhad",country:{name:"Iran"}}},{id:"414878",title:"Prof.",name:"Reza",middleName:null,surname:"Fazel-Rezai",slug:"reza-fazel-rezai",fullName:"Reza Fazel-Rezai",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"American Public University System",country:{name:"United States of America"}}},{id:"426586",title:"Dr.",name:"Oladunni A.",middleName:null,surname:"Daramola",slug:"oladunni-a.-daramola",fullName:"Oladunni A. Daramola",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Federal University of Technology",country:{name:"Nigeria"}}},{id:"357014",title:"Prof.",name:"Leon",middleName:null,surname:"Bobrowski",slug:"leon-bobrowski",fullName:"Leon Bobrowski",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Bialystok University of Technology",country:{name:"Poland"}}},{id:"302698",title:"Dr.",name:"Yao",middleName:null,surname:"Shan",slug:"yao-shan",fullName:"Yao Shan",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Dalian University of Technology",country:{name:"China"}}},{id:"354126",title:"Dr.",name:"Setiawan",middleName:null,surname:"Hadi",slug:"setiawan-hadi",fullName:"Setiawan Hadi",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Padjadjaran University",country:{name:"Indonesia"}}},{id:"125911",title:"Prof.",name:"Jia-Ching",middleName:null,surname:"Wang",slug:"jia-ching-wang",fullName:"Jia-Ching Wang",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"National Central University",country:{name:"Taiwan"}}},{id:"332603",title:"Prof.",name:"Kumar S.",middleName:null,surname:"Ray",slug:"kumar-s.-ray",fullName:"Kumar S. Ray",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Indian Statistical Institute",country:{name:"India"}}},{id:"415409",title:"Prof.",name:"Maghsoud",middleName:null,surname:"Amiri",slug:"maghsoud-amiri",fullName:"Maghsoud Amiri",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Allameh Tabataba'i University",country:{name:"Iran"}}},{id:"357085",title:"Mr.",name:"P. Mohan",middleName:null,surname:"Anand",slug:"p.-mohan-anand",fullName:"P. Mohan Anand",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Indian Institute of Technology Kanpur",country:{name:"India"}}},{id:"356696",title:"Ph.D. Student",name:"P.V.",middleName:null,surname:"Sai Charan",slug:"p.v.-sai-charan",fullName:"P.V. Sai Charan",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Indian Institute of Technology Kanpur",country:{name:"India"}}},{id:"357086",title:"Prof.",name:"Sandeep K.",middleName:null,surname:"Shukla",slug:"sandeep-k.-shukla",fullName:"Sandeep K. Shukla",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Indian Institute of Technology Kanpur",country:{name:"India"}}}]}},subseries:{item:{id:"18",type:"subseries",title:"Proteomics",keywords:"Mono- and Two-Dimensional Gel Electrophoresis (1-and 2-DE), Liquid Chromatography (LC), Mass Spectrometry/Tandem Mass Spectrometry (MS; MS/MS), Proteins",scope:"With the recognition that the human genome cannot provide answers to the etiology of a disorder, changes in the proteins expressed by a genome became a focus in research. Thus proteomics, an area of research that detects all protein forms expressed in an organism, including splice isoforms and post-translational modifications, is more suitable than genomics for a comprehensive understanding of the biochemical processes that govern life. The most common proteomics applications are currently in the clinical field for the identification, in a variety of biological matrices, of biomarkers for diagnosis and therapeutic intervention of disorders. From the comparison of proteomic profiles of control and disease or different physiological states, which may emerge, changes in protein expression can provide new insights into the roles played by some proteins in human pathologies. Understanding how proteins function and interact with each other is another goal of proteomics that makes this approach even more intriguing. Specialized technology and expertise are required to assess the proteome of any biological sample. Currently, proteomics relies mainly on mass spectrometry (MS) combined with electrophoretic (1 or 2-DE-MS) and/or chromatographic techniques (LC-MS/MS). MS is an excellent tool that has gained popularity in proteomics because of its ability to gather a complex body of information such as cataloging protein expression, identifying protein modification sites, and defining protein interactions. The Proteomics topic aims to attract contributions on all aspects of MS-based proteomics that, by pushing the boundaries of MS capabilities, may address biological problems that have not been resolved yet.",coverUrl:"https://cdn.intechopen.com/series_topics/covers/18.jpg",hasOnlineFirst:!0,hasPublishedBooks:!0,annualVolume:11414,editor:{id:"200689",title:"Prof.",name:"Paolo",middleName:null,surname:"Iadarola",slug:"paolo-iadarola",fullName:"Paolo Iadarola",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bSCl8QAG/Profile_Picture_1623568118342",biography:"Paolo Iadarola graduated with a degree in Chemistry from the University of Pavia (Italy) in July 1972. He then worked as an Assistant Professor at the Faculty of Science of the same University until 1984. In 1985, Prof. Iadarola became Associate Professor at the Department of Biology and Biotechnologies of the University of Pavia and retired in October 2017. Since then, he has been working as an Adjunct Professor in the same Department at the University of Pavia. His research activity during the first years was primarily focused on the purification and structural characterization of enzymes from animal and plant sources. During this period, Prof. Iadarola familiarized himself with the conventional techniques used in column chromatography, spectrophotometry, manual Edman degradation, and electrophoresis). Since 1995, he has been working on: i) the determination in biological fluids (serum, urine, bronchoalveolar lavage, sputum) of proteolytic activities involved in the degradation processes of connective tissue matrix, and ii) on the identification of biological markers of lung diseases. In this context, he has developed and validated new methodologies (e.g., Capillary Electrophoresis coupled to Laser-Induced Fluorescence, CE-LIF) whose application enabled him to determine both the amounts of biochemical markers (Desmosines) in urine/serum of patients affected by Chronic Obstructive Pulmonary Disease (COPD) and the activity of proteolytic enzymes (Human Neutrophil Elastase, Cathepsin G, Pseudomonas aeruginosa elastase) in sputa of these patients. More recently, Prof. Iadarola was involved in developing techniques such as two-dimensional electrophoresis coupled to liquid chromatography/mass spectrometry (2DE-LC/MS) for the proteomic analysis of biological fluids aimed at the identification of potential biomarkers of different lung diseases. He is the author of about 150 publications (According to Scopus: H-Index: 23; Total citations: 1568- According to WOS: H-Index: 20; Total Citations: 1296) of peer-reviewed international journals. He is a Consultant Reviewer for several journals, including the Journal of Chromatography A, Journal of Chromatography B, Plos ONE, Proteomes, International Journal of Molecular Science, Biotech, Electrophoresis, and others. He is also Associate Editor of Biotech.",institutionString:null,institution:{name:"University of Pavia",institutionURL:null,country:{name:"Italy"}}},editorTwo:{id:"201414",title:"Dr.",name:"Simona",middleName:null,surname:"Viglio",slug:"simona-viglio",fullName:"Simona Viglio",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRKDHQA4/Profile_Picture_1630402531487",biography:"Simona Viglio is an Associate Professor of Biochemistry at the Department of Molecular Medicine at the University of Pavia. She has been working since 1995 on the determination of proteolytic enzymes involved in the degradation process of connective tissue matrix and on the identification of biological markers of lung diseases. She gained considerable experience in developing and validating new methodologies whose applications allowed her to determine both the amount of biomarkers (Desmosine and Isodesmosine) in the urine of patients affected by COPD, and the activity of proteolytic enzymes (HNE, Cathepsin G, Pseudomonas aeruginosa elastase) in the sputa of these patients. Simona Viglio was also involved in research dealing with the supplementation of amino acids in patients with brain injury and chronic heart failure. She is presently engaged in the development of 2-DE and LC-MS techniques for the study of proteomics in biological fluids. The aim of this research is the identification of potential biomarkers of lung diseases. 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