Dysbiosis, Tolerance, and Development of Autoimmune Diseases

Abstract

The pathogenesis of autoimmune diseases (AIDS) is not only attributed to genetic susceptibility, but also to environmental factors, among which, those disturbing gut microbiota have attracted increasing attention lately. Healthy gut microbiota has beneficial effects on the development and activity of the immune system, playing a central role in peripheric tolerance. Compositional and functional changes in gut microbiota were reported in various AIDS, and increasing evidence suggests that disturbed gut microbiota contributes to their immunopathogenesis. Thyroid and intestinal diseases prevalently coexist—for instance, Hashimoto’s thyroiditis and Graves’ disease are the most common autoimmune thyroid diseases and often co-occur with celiac disease. This association can be at least explained by increased intestinal permeability, allowing antigens to cross the barrier more easily and activate the immune system. The passage of microbial antigens into the internal environment may break the self-tolerance, generating the production of autoantibodies and/or autoreactive T cells. In this chapter, we briefly present the roles of intestinal microbiota in human physiology, with a focus on the role of microbiota in immune tolerance.

1. Introduction

Immune tolerance is a physiological condition, characterized by the absence of an immune response to a specific antigen and the retention of the ability to develop an immune response to other different antigens. Tolerance to self-components develops both during embryonic development (i.e., central tolerance, which occurs in the primary lymphoid organs, along with the process of lymphocyte differentiation), and after birth (i.e., peripheral tolerance) [1].

Currently, the microbiota is considered an anatomically integrated meta-organ that performs functions through which it interferes with the host’s physiology [2]. Thus, microbiota eubiosis is a major parameter of physiological homeostasis. Human microbiota establishes three types of relationships with the host—symbiotic, commensal, and pathobiontic, respectively [3]. The terms “microbiota” and “microbiome” are equivalent, but not identical. The first refers to the population of microorganisms residing on the mucous membranes of the digestive, urogenital and respiratory tract, as well as on the skin, and the second designates the collective genome of the microbiota, called the metagenome [4]. The community microbiome was evaluated at 3.3 million redundant bacterial genes, about 150 times larger than the human gene complex [5]. The gut microbiota is influenced by various conditions, such as diet, health, mental stress, gender, or exercise, and conversely, it influences all body metabolism, immune reactivity, and behavior [6]. The microbiota contributes to the peripheral tolerance of the immune system toward autoantigens, with the retention of the immune reactivity against all antigens that do not cross-react with the tolerated antigen. Interruption of tolerance initiates an immune response to self-antigens characterized by the production of autoantibodies or autoreactive lymphocytes, which trigger an autoimmune conflict [7]. The purpose of this chapter is to highlight the role of the normal microbiota in the state of immune tolerance and to investigate the correlations of dysbiosis with endocrine AIDS.

2. The role of the intestinal microbiota in the physiology of the human body

The microbiota of the digestive tract consists of about 3 × 10¹³ to 40 × 10¹³ (3–40 trillion) bacterial cells, counting at least 10 times more than their host cells. The groups of Bacteroidetes and Firmicutes predominate in a numerical proportion of 90%, aside from lower density of Proteobacteria, Actinobacteria, Fusobacteria, and Verrucomicrobia and small populations of fungi, Archaea, and viruses, all exerting major functional effects on different organs. The bacterial microbiota belongs to 1000–1160 types of species [8]. The individual microbiota is evaluated in 150 to 160 species by the 16S RNA (rDNA analysis) ribotyping method [9]. The population composition of the intestinal microbiome stabilizes at the age of 3 years and is determined by various conditions, such as genetic factors, the maternal microbiota, the mode of birth (i.e., natural or by cesarean section), the antigenic exposure during early life, and is reconfigured mostly by diet [10, 11].

The microbiota is considered a virtual organ, whose functions must be integrated into general physiology. The host-microbiota interaction is primarily a symbiotic relationship, in which the host organism provides the ecological niche and nutrients for microbiota survival. The microbiota carries out fermentative and biosynthetic metabolic activities, thereby influencing systemic physiology [12]. The metabolism of the microbiota functions as a bridge between the diet with the human body. The intestinal microbiota increases the energy efficiency of the diet by fermenting the fibrous components, providing essential metabolites for organ systems, especially short-chain fatty acids (SCFA), such as acetic, propionic, and butyric acid. A proportion of 50% of the energy needs of epithelial cells is provided by SCFA [13, 14]. The modern diet is 7–10 times poorer in the fibrous component, compared to the traditional Mediterranean one. Microbiota synthesizes vitamin K and B, synthesizes amines through which it modifies endocrine function, stimulates the inflammatory process, has a protective role against the invasion of enteric pathogens (Shigella flexneri), metabolizes some drugs to their active form, ferments indigestible components of the diet (complex polysaccharides, amino acids, xenobiotics) [15], and modulates the lipid metabolism. The bile acids synthesized in the liver from cholesterol, facilitate the absorption of lipids and fat-soluble vitamins and maintain cholesterol balance. Also, the biliary acids have a signaling function through specific hepatocyte receptors [16, 17, 18, 19]. All these functions are impaired in patients with endocrine AIDS.

3. Role of the microbiota in the development of the mucosa-associated immune system

From an immunological perspective, the mucous membranes which cover a total area of about 400 m², represent both an anatomical and functional entity, because they are populated by a large number of immune cells.

The intestinal microbiota, epithelium and digestive, respiratory, genital, urinary mucosa-associated immune system form a functional triad whose components influence each other close interactions, with a rapid dynamic of change, induced by population changes of the microbiota, due to diet variation and/or administration of antibiotics. The modification of the functional parameters of a component of the triad has major influences on the physiology of the whole organism. The microbiota interacts directly with the epithelium of the adjacent mucosa and influences its permeability, and both local and systemic inflammatory responses [10] The interaction of the microbiota with the mucosal immune system (gut-associated lymphoid tissue—GALT) induces the synthesis of a wide set of cytokines, with local regulatory action of intestinal physiology [20, 21].

The microbiota has an essential role in the functional modulation (education), first of all of the GALT structures. Germ-free and gnotobiotic animal studies have made a decisive contribution to understanding the functional relationships of the microbiota-epithelium-immune system triad and provided new evidence for the role of the intestinal microbiota as a whole, but also of different groups of bacteria in the functional development and maturation of the systemic immune system, especially GALT. Germ-free mice have structural and functional defects of the immune system—decreased TCD4 lymphocyte count and Th-2 predominance in the spleen, altered Th-17 and T-reg differentiation in the lamina propria, and restoration of deficiencies after colonization with Bacteroides and segmented filamentous bacteria (SFB). The balance of effector T lymphocytes is disturbed in intestinal dysbiosis and accelerates or suppresses the autoimmune reactions [22, 23]. The constant interaction of the microbiota with the cells of adaptive immunity prevents bacterial invasion and pathogenesis, but also the systemic immune response with detrimental effects against the microbial antigens [24]. The structural but especially functional peculiarities of GALT tend to delimit it more and more from the systemic immune system.

M cells that cover the subepithelial immune structures engulf the luminal antigens, through the mechanism of pinocytosis and transfer them unaffected to the immune structures in the underlying follicles (i.e., macrophages, dendritic cells, T and B lymphocytes). Macrophages and dendritic cells respond to microbiota antigens in a nonspecific manner by TLR recognition followed by cytokines release (i.e., IFNα, IL-18, and IL-22), which stimulate the epithelial cells to synthesize antimicrobial peptides.

The microbiota, through the composition of bacterial phyla, has a major influence on the development of T lymphocyte subpopulations and in maintaining the numerical balance of Th-2/Th-1 lymphocyte populations in lymphoid organs. The differentiation of T lymphocyte sets is influenced by the antigenic specificity of the dominant bacterial population and its metabolic properties—(i) some bacteria stimulate the predominant differentiation of proinflammatory TCD4 lymphocytes that synthesize IFNγ and IL-17A [25]; and (ii) others stimulate the differentiation of regulatory CD25+ and Foxp3+ TCD4 lymphocytes (T-reg), the essential mediator of immune tolerance by decreasing Th-17 lymphocytes [26, 27]. The direct relationship between the concentration of butyric acid and the number of T-reg lymphocytes is well known. SCFA, particularly butyric acid harbor important roles, that is, stimulate gene transcription for mucin synthesis, strengthen the intestinal barrier and render it impermeable to toxins and bacterial cell translocation, thus preventing chronic systemic inflammation, inhibiting the synthesis of pro-inflammatory interleukins (IL) (TNFα and IL-6) induced by LPS and regulate the innate and adaptive immunity [13, 14]. Th-17 lymphocytes play an essential role in anti-bacterial and anti-fungal defense, but at the same time have an important role in the initiation of inflammatory diseases, through the synthesis of pro-inflammatory IL-17 and IL-22 and the recruitment of neutrophils. In germ-free animals, the lamina propria is populated by a very small number of Th-17 lymphocytes [9]. Th-17 lymphocytes also decrease after antibiotic treatment [27]. The group of Clostridium SFB, following the colonization of the epithelium, induces an increase in the number of Th-17 lymphocytes, whose proinflammatory IL can promote the onset of rheumatoid arthritis and multiple sclerosis in gnotobiotic animals. In patients suffering from inflammatory bowel disease, which manifests clinically similar to Crohn’s disease and ulcerative colitis, the number of T-reg lymphocytes with immunosuppressive function decreases in the lamina propria and the population of lymphocytes that have TCR for the bacterial microbiota antigens increases abnormally. The density of T-reg lymphocytes increases in gnotobiotic animals colonized by Clostridium SFB group, while the polysaccharide A of Bacteroides fragilis (which is attributed to symbiotic factor status) induces the differentiation of TCD4 lymphocytes to T-reg lymphocytes [9, 28]. TCD4 Foxp3 + lymphocytes secrete IL-10, the main anti-inflammatory cytokine, thus being involved in tolerance to microbial antigens. In germ-free animals, the dominance of Th-2 subpopulation in the spleen that favors allergic manifestations is restored by the polysaccharide A of Bacteroides fragilis.

The microbiota has also a profound influence on the development of B lymphocytes—it stimulates the synthesis of antibodies, especially of IgA type, targeted against thymus-dependent (Td), and thymus-independent (Ti) antigens. The Clostridium SFB and Alcaligenes group of bacteria are potential inducers of IgA synthesis specific for the intestinal microbiota antigens. In the absence of IgA, the Clostridium group is enriched, whereas the Alcaligenes group is diminished [9].

In germ-free animals, GALT structures play a key role in inducing immune tolerance against auto-antigens from the intestinal mucosa, are less developed and indicators of immune response activation are lacking. In these animals, the number of TCD4 lymphocytes and IgA-secreting plasma cells decreases in Peyer’s patches, while in the spleen and lymph nodes, the number of B lymphocytes and germinal centers decreases.

In conclusion, the development, maturation, and function of the immune system are closely associated with the level of exposure to microbial antigens during early life, and as an opposite, insufficient exposure to various antigens increases the risk of autoimmune disorders occurrence [29].

4. Intestinal microbial antigens as inductors of central and peripheral tolerance

Despite its much-diversified antigen panel, the microbiota is tolerated by the immune system. Central tolerance is induced during fetal life, as immature lymphocytes are exposed to various antigenic peptides, and is essentially dependent on the specific process of antigenic peptide selection and presentation in association with the Human Leucocytes Antigen/ Major Histocompatibility Complex (HLA/MHC) molecules [30]. The occurrence of peripheral tolerance breaks results from a functional adaptation of the immune system to specific antigenic peptides that have not (sufficiently) been exposed to lymphocytes in the bone marrow or thymus during embryonic development. It is now considered that the T lymphocyte antigen receptor (TCR) is the major mediator of immune tolerance. That is why, from an evolutionary perspective, TCR recognizes both the genetic and microbial self [31].

The immune tolerance to commensal intestinal microbiota is peripheral and results from both an immediate neonate colonization of the digestive tract and a progressive co-evolution in which the interactions of gut-associated lymphoid tissue (i.e., GALT) with bacterial antigens have been modulating innate and adaptive primarily local immune reactivity. Commensal antigens, on contact with the intestinal mucosa, induce the state of tolerance, in which dendritic cells play an essential role, while the effectors are the epithelial cells with their covering molecular complex (i.e., antimicrobial peptides, mucin layer, surface immunoglobulin A—sIgA) [32]. Bacterial cells or their components (i.e., lipopolysaccharides, polysaccharides, peptidoglycans, teichoic acids, and DNA) that cross the intestinal barrier and reach the internal environment, activate the immune response [33].

4.1 Causes of losing immune tolerance to microbiota antigens

Interruption of immune tolerance to microbiota antigens is determined by several factors—genetic factors, the host’s immune system, disturbance of the diversity, and physiology of the microbiota—as triggering events [34].

Mechanisms that modulate immune tolerance loss to the intestinal microbiota include: (i) abnormal translocation of bacteria in the internal environment due to permeability of the intestinal barrier, (ii) antigenic similarity of some bacterial peptides with epithelial molecules. Immune cells are activated by bacterial peptides and become autoreactive; and (iii) disorder of local and systemic immunity under the stimulating action of some bacterial derivatives (nucleic acids, polysaccharides, metabolites, and toxins). Aberrant activation of the immune system leads to the excessive synthesis of proinflammatory IL (IFN type I, IL-12, IL-23) and a decreased rate of synthesis of anti-inflammatory cytokines (IL-10, TGF-β—transforming growth factor) (Figure 1) [35].

Figure 1.

The role of microbiota in mucosal homeostasis and immunological tolerance in healthy gut and activated inflammatory cascades in endocrine autoimmune disease. In germ-free animals, GALT structures are less developed and the microbiota has a major influence on the development of T lymphocyte subpopulations and in maintaining the numerical balance of Th-2/Th-1 lymphocyte populations in lymphoid organs. The healthy gut environment is characterized by high levels of antimicrobial peptides and metabolites (SCFAs), and the commensal-specific IgA is produced by plasma cells in the lamina propria, mediated by DCs in a T cell-independent mechanism. During homeostasis, gut microorganisms induce an immune tolerance phenotype in the host, whilst in inflammatory conditions, antigens from dysbiotic microorganisms activate Th1 and Th17 cells leading to decreased mucus layer, tissue injury, and microbial penetration and persistence in the intestinal tissues. This mucosal injury results in further uptake of microbial antigens that further perpetuate detrimental immune responses. Figure created with https://biorender.com/.

4.2 Consequences of losing immune tolerance

Although the autoimmune conflict occurs most of the time without clinical manifestations, it can generate under certain conditions, such as AIDS, that are characterized by the appearance of tissue lesions or disruption of physiological processes. AIDS have a multifactorial etiology involving genetic, epigenetic, and environmental factors. It is estimated that 70% of AIDS are due to environmental factors [36]. Among the multiple cellular and molecular mechanisms, yet not well established, by which the state of immune self-tolerance is disturbed, we can mention—(i) the genetic predisposition that may explain the familial character of AIDS, which, in general, have a polygenic determinism. The risk of a certain autoimmune disease for monozygotic twins is about 12 to 60%, and for dizygotic twins is 5%. The most important are certain specific polymorphisms generated by the change of a nucleotide, that is, SNP (single nucleotide polymorphism) in MHC genes [9]. For example, over 90% of Caucasians with ankylosing spondylitis express an allele of the HLA-B27 family, differing from that of normal individuals by two amino acids located in the peptide binding groove [37]; (ii) release of sequestered antigens after trauma, surgery, infectious processes, etc., become accessible to lymphocytes, triggering the autoimmune conflict and tissue damage (e.g., basic myelin protein in the central nervous system becomes the antigenic target in multiple sclerosis; crystalline proteins induce autoimmune ophthalmopathy; sperm proteins, in cases of sperm stasis, induce the synthesis of immobilizing or binder autoantibodies of sperm, leading to autoimmune infertility) [38, 39]; (iii) modification of the chemical structure of autoantigens (so-called altered self-theory), which occurs under the influence of some physical factors (such as burns or radiation), biological (i.e., bacteria, viruses, fungi), or chemical (i.e., drugs, alcohol) factors, with the exposure of some new antigenic determinants [40]; (iv) infectious agents, which may have an important role in triggering AIDS by various mechanisms, such as the antigenic resemblance of non-self to self-molecules and their cross-reactivity (e.g., protein M from Streptococcus pyogenes is antigenically similar to cardiomyocyte’s membrane proteins); (v) stimulation of the proinflammatory cytokines production that cause nonspecific activation of self-reactive immune cells; superantigens of infectious agents (i.e., Epstein-Barr virus, mycoplasmas, Staphylococcus aureus, Streptococcus pyogenes) that induce polyclonal activation of lymphocytes [41]; (vi) loss of peripheral immune tolerance, due to either mutation that generates the appearance of immunocompetent, self-reactive T or B lymphoid cell clones, or T-reg cell deficiency, or Th cell activation; (vii) disruption of the equilibrium state of the idiotypic network by the synthesis of anti-idiotypic antibodies, which may be autoantibodies [42]; and (viii) hormonal imbalances that may be involved in triggering AIDS, therefore explaining their increased frequency in women (8: 1 ratio) except for ankylosing spondylitis, or in men with higher levels of estrogen hormones. Moreover, pregnancy is associated with an improvement in the severity of AIDS, especially in rheumatoid arthritis cases.

AIDS resemble some general features—the pathological process has an individual intensity, dynamics, and evolution, may overlap with the same patient, and are rare in childhood, except for type 1 diabetes mellitus.

Regardless of the triggering mechanism, AIDS is characterized by the synthesis of autoantibodies (that are antibodies specific to self-tissue components) or by the generation of autoreactive T lymphocytes. Tissue injuries following the action of immune effectors occur in one of the above-mentioned scenarios—(i) autoantibodies recognize the tissue antigens and form immune complexes, the complement is activated, and the result is the cell lysis, or (ii) indirect action, in which case, the antigen-antibody—complement immune complexes are deposited in small vessels (arterioles, capillaries) from various organs and produces inflammatory reactions, with the consequence of tissue destruction; the AIDS that are mediated by various antibodies have a common feature, that is the target tissue is damaged by a chronic inflammatory reaction without a known infectious cause; and (iii) the lesions in the target tissue occur under the action of infiltrated Tc lymphocytes [43, 44].

Some AIDS are characterized by strictly localized pathological processes, that is, effectors (especially antibodies) have specific action against antigens specific to the target tissue (such is the case for autoantibodies specific only to B cells from Langerhans islands in type 1 diabetes mellitus, or autoantibodies specific to thyroid epithelial cell in Hashimoto’s thyroiditis) [45], sometimes the lesions are localized in a single organ, but autoantibodies do not have organ specificity (for instance, anti-mitochondrial antibodies in primary cirrhosis, or type IV anti-collagen autoantibodies in Goodpasture syndrome) [46, 47] while some AIDS are disseminated, characterized by the synthesis of autoantibodies to antigens with wide tissue distribution (e.g., antinuclear antibodies in systemic/disseminated lupus erythematosus) [48].

Often, in pathological cases, the body synthesizes auto-antibodies specific for components of the endocrine system, especially antibodies specific for a certain hormone receptor. The pathophysiological effects of these antibodies generated against hormone receptors are varied—they can stimulate the activity of the receptor, and the effect is to intensify the secretory activity of the gland (hormonal mimetic effect) or block the receptor, and the effect is to inhibit the secretory activity. Both antibodies can coexist in the same patient.

5. The role of microbiota in autoimmune-mediated endocrine diseases

The role of the microbiota in autoimmune pathology has been highlighted by experimental data collected from germ-free mice. The intestinal microbiota maintains the balance of protective reactions to pathogens and tolerance to commensals aimed at maintaining intestinal homeostasis [49, 50]. Alterations produced in the balance of the microbiota (that is dysbiosis) activate the proinflammatory immune response and favor the progression of autoimmune disorders, such as multiple sclerosis, inflammatory bowel disease, T1DM, rheumatoid arthritis, and other pathologies of the digestive tract and ancillary glands, including malignancies. However, the intimate mechanism of microbiota involvement in this pathogenesis remains unknown [51, 52, 53].

AIDS are caused primarily by predisposing genetic factors but also by other endogenous or environmental triggers. There is a permanent interaction of the local immune system with bacterial antigens in humans, and therefore dysbiosis of the microbiome is associated with autoimmune disorders and metabolic syndromes. Dysbiosis means, in fact, the numerical alteration, diversity, and physiology of the intestinal microbiota (the transcriptome, proteome, and metabolome change) [54].

Experimental results in germ-free or induced dysbiotic animals support either the microbiota’s direct or indirect involvement in the pathogenesis of some AIDS. Hence, in patients with type 1 diabetes mellitus, rheumatoid arthritis, multiple sclerosis, or lupus, as in those suffering from inflammatory bowel disease (both Chron’s disease and ulcerative colitis), Sjögren’s syndrome, Behcet’s disease, autoimmune skin diseases (such as vitiligo, psoriasis, atopic dermatitis), the digestive microbiota is altered in terms of diversity and numerical representation of some species [9, 18, 35]. Kriegel et al. consider that dysbiosis is an essential trigger of autoimmunity both at the mucosal and systemic levels [9]. The spread of autoimmune response seems to be generated either by disseminating bacterial antigens but mostly by cross-immune reactivity under homeostasis conditions [55]. Such a mechanism is supported by a rheumatic fever induced by M and SLO antigens of Streptococcus pyogenes, or Guillain-Barre syndrome induced by Campylobacter jejuni infection, both as transient autoimmune syndromes. Cross-reactivity of lipopolysaccharides, bacterial polysaccharides, or D amino acid polymers would be an important mechanism for initiating the autoimmune conflict. Patients with autoimmune disorders often have vitamin D deficiency; its administration in experimental settings to animals improves the course of the disease. Vitamin D deficiency is also associated with an increased risk of infectious diseases. Inflammatory cells convert vitamin D to its active form, which is calcitriol. Vitamin D is an essential factor for the activation and proliferation of inflammatory cells (macrophages, neutrophils) [56]. The probiotics could also reverse the chronic systemic inflammation associated with AIDS.

5.1 Type I diabetes mellitus

Type 1 diabetes mellitus (T1DM) has a well-defined autoimmune component, characterized by selective immune aggression against β-cells that secrete insulin [57]. The genetic predisposition for T1DM is unanimously accepted, but the interaction of genetic factors with environmental ones explains the sudden increase in incidence in Western countries [58]. More than 50% of monozygotic twins who have a sibling with T1DM remain healthy, showing that environmental factors (such as infectious agents, consumption of cow’s milk in early childhood, or ingestion of contaminated food) play a major role in triggering the disease. Hence, out of 50 individuals suffering from congenital rubella virus infection [59, 60], nine developed diabetes at an average age of 28 years. However, some infections (i.e., M. tuberculosis, viruses, or parasites) exert a nonspecific inhibition on the onset of T1DM, probably by stimulating regulatory T cells [61, 62, 63, 64].

The pathological mechanisms leading to the autoimmune destruction of pancreatic beta-cells in T1DM are very complex and incompletely elucidated. The pancreatic beta-cells express MHC II and co-stimulatory molecules, suggesting their role as antigen-presenting cells to TCD4 cells. Auto-antigens that stimulate the specific immune reactivity against pancreatic beta-cell are represented by insulin, glutamic acid decarboxylase—isoform 2 of 65 kD from beta-cell cytoplasm, a Zn transporter protein (ZnT8) involved in active secretion of insulin from islet granules, insulinoma-associated antigen 2 (alpha and beta), and a membrane protein acting as tyrosine phosphatase. The presence of humoral autoimmunity defines the risk of T1DM; antibodies against insulin were identified in 40% of children with the overt disease [65].

In patients with T1DM, it has been shown by immunohistochemical staining that the islets are infiltrated with macrophages, dendritic cells, TCD4, TCD8, NK, and fewer B lymphocytes, which can act as antigen-presenting cells for TCD4 cells. The immune response against islet antigens is associated with an inflammatory one in which IL-1, TNFα, and IFNγ are released [66]. The immune and inflammatory process destroys the beta cells. When about 80% of the beta-cell mass has been destroyed, the disease overt. This silent period may last for several years, sometimes decades. Along with the progressive destruction of β cells, the humoral antibody response and decreased glucose tolerance are documented until the clinical onset of the disease. Immune effectors selectively lyse insular β cells, leaving the other cell types intact. After the onset of hyperglycemia, the degree of mononuclear infiltration decreases [67].

The inflammatory diseases of the pancreas (such as chronic pancreatitis, neoplasia) are characterized by mast cells infiltrates into the acinar parenchyma, which releases various proteases (chymase, tryptase), acting as direct destroyers on islet’s beta cells. The B4 type of leukotrienes, which derives from mast cells, exerts a chemoattractant effect on T lymphocytes [68].

Loss of pancreatic beta cells leads to insulin secretion deficiency, while the glucagon secretion becomes excessive and disrupts metabolism, resulting (in the absence of insulin) in diabetic ketoacidosis [69].

5.1.1 Experimental studies

The experimental results argue for the interference of the microbiota and T1DM pathological mechanisms—the incidence of diabetes is higher in mice raised in aseptic conditions, and the antibiotics administered to conventional animals accelerate the evolution of diabetic pathology. The NOD (non-obese diabetes) mice have a distinct microbiota from other resistant lines, and the incidence of type 1 diabetes mellitus is higher in specific pathogen-free animals [70].

5.1.2 Analytical results

Dysbiosis is shaped by host-related individual factors and early-life exposure to certain microorganisms, and its alterations undergo extensive changes with the change in diet. The permeability of the intestinal barrier plays an important role in the initiation and evolution of autoimmune conflict, aside from the background of genetic predisposition. The intercellular tight junctions control the permeability of the epithelium, allowing the absorption of nutrients, but preventing the passage of various environmental antigens (i.e., food, bacterial, viral, and fungal). Dysbiosis decreases intestinal permeability and facilitates the translocation of bacterial antigens [52].

Microbiota derangements have been implicated in the evolution of both T1DM and T2DM [71]. Dysbiosis occurs very early in subjects with a genetic predisposition for T1DM, probably since the neonatal period [51]. It is unknown whether the genetic predisposition to T1DM shapes the microbiota of high-risk individuals or whether the microbiota is the cause or effect of the disease [71].

As stated above, the human microbiota stabilizes during the first 3 years of life, while three parallel phenomena occur—(i) development of the immune system, (ii) maturation of the microbiota, and (iii) seroconversion to T1DM-associated autoantibodies. The possible conditioning of the two (i.e., seroconversion and T1DM occurrence) events is unknown. In a longitudinal study, Kostic et al. showed a decrease in the bacterial diversity of the microbiota that occurs before the development of the clinical disease in children positive for anti-insulin antibodies [70]. The Clostridium, Veillonella Bacteroides increase in abundance, while Lactobacillus, Bifidobacterium, Prevotella genera decrease compared to healthy subjects, suggesting the correlation between microbiota disturbance and T1DM [18, 19]. Different authors reported other changes associated with T1DM. Increases of Bacteroidetes (Gram-negative) and decreases of Prevotella and Firmicutes (Gram-positive producing SCFA) observed in children with T1DM when compared to healthy subjects suggest an increased intestinal barrier permeability and decreased SCFA production [70]. The microbiota of children with T1DM is unstable and has a smaller population of butyrate-producing bacteria, which correlates with an increased barrier permeability. Healthy children have higher levels of Lactobacillus rhamnosus and Bifidobacterium dentium, while the group of Streptococcus mitis/oralis/pneumoniae is abundant in subjects with T1DM.

Furthermore, the microbiota changes evolve with disease progression [65].

The fungal microbiome of the human population is evaluated in 267 species, with the most commonly represented by g. Candida, Saccharomyces, Penicillium, and Aspergillus. The individual mycobiome rarely contains more than one genus, but this panel is enough to influence the entire composition of the microbiota population, either directly by interactions with bacterial cells or indirectly by immune modulation. In patients with type 1 diabetes and those with inflammatory bowel disease, there was an overgrowth of Candida [70].

Despite the abundance of experimental and clinical results suggesting a bidirectional relationship between dysbiosis and T1DM onset and progress, there are questions that still need an answer—(i) is their relationship causal or simultaneous? and (ii) the condition of causality is that the change of one variable leads to the change of another repeatedly and generally? [65].

5.2 Autoimmune thyroid diseases

Thyroid AIDS are conditioned as other auto-immunities by a genetic predisposition, but other factors play an important role in triggering and evolving the autoimmune pathological process [72]. They occur with a frequency of about 4% in the human population and express by either hyper- or hypothyroidism. In both cases, the thyroid may increase in volume (goiter), while ophthalmopathy may develop in hyperthyroidism only [73]. Autoimmune thyroid disease affects especially women and from an immunological point of view, it is characterized by the presence of circulating autoantibodies, activated T cells against thyroid antigens, and by lymphocytic infiltration of the organ. Three specificities of anti-thyroid autoantibodies have been described—anti-thyroid peroxidase (microsomal antigen); anti-thyroglobulin; anti-TSH receptor of thyroid acinar cells [74, 75].

AIDS that cause thyroid failure, generically called thyroiditis, are characterized by lymphocytic infiltration. Depending on the clinical aspects there are two pathological conditions—Hashimoto’s thyroiditis and atrophic thyroiditis (primary myxedema). In both cases, the thyroid tissue is lysed. Autoimmune thyroid disease is influenced by various factors, such as age, sex, race, and hormonal status [76, 77].

Autoimmune thyroid diseases (Graves and Hashimoto’s thyroiditis) often coexist with intestinal diseases, especially celiac disease. The composition of the microbiota population is influenced by diet, affects the thyroid function, mostly by providing the micronutrients essential for the synthesis of thyroid hormones—iodine, iron, and copper. Selenium and zinc are essential for the conversion of T4 to T3, and vitamin D has an immune regulatory effect. Probiotic supplementation favorably influences the secretion of thyroid hormones [26].

Autoimmune thyroiditis is the most common thyroid disorder, with a prevalence of 10–12%. It is triggered by genetic and environmental factors (viral infections) and has an increased prevalence in patients with celiac disease. The commensal microbiota activates the proinflammatory response through innate immunity receptors from the toll-like receptor family and disrupts the intestinal permeability, which may be a triggering factor for Hashimoto’s thyroiditis [78].

Hashimoto’s thyroiditis is the most common endocrine AIDS (i.e., 10–12% of total autoimmune endocrinopathies), which is characterized by autoimmune destruction of thyroid follicles. The incidence increases with age and is 10 times higher in women. In the serum of patients with Hashimoto’s thyroiditis are detected various specific autoantibodies, such as anti-thyroglobulin and/or anti-TPO (thyroid-peroxidase), anti-TSH receptor. Definitive for Hashimoto’s disease is the replacement of thyroid tissue with lymphoid tissue. An impressive increase in thyroid volume may be observed, but no hormones are synthesized instead (dry goiter). The symptoms of Hashimoto’s thyroiditis and celiac disease often overlap and share epidemiological, clinical, serological, pathological, hormonal, genetic, and immune similarities. Microbiome analysis performed on patients with this ailment revealed that abundance levels of Blautia, Roseburia, Ruminococcus torques groups, Dorea, Fusicatenibacter, and Eubacterium hallii group genera were significantly higher whereas Faecalibacterium, Prevotella, and Bacteroides genera were decreased [79, 80, 81, 82].

Celiac disease (CD) is an autoimmune condition characterized by a specific serological and histological profile triggered by gluten ingestion in genetically predisposed individuals [83]. CD is the only AID known to be triggered by an exogenous antigen, that is, wheat gluten. Gluten is a mixture of proteins grouped in the fraction of gliadin and glutenin, which is the source of carbon and nitrogen for germinating seedlings. Gliadin triggers specific auto-antibody synthesis, the clinical feature being strictly dependent on dietary exposure to gluten and homologous proteins from other cereals. CD is one of the most common autoimmune disorders, with a reported prevalence of 0.5–1% of the general population, except in areas showing a low frequency of CD-predisposing genes and low gluten consumption [84]. Studies have shown that most CD cases remain undetected in the absence of serological screening due to heterogeneous symptoms and/or poor disease awareness. CD has a strong hereditary component confirmed by its high familial recurrence (~10–15%) and the high concordance of the disease among monozygotic twins (75–80%) [85]. Also common to other AIDS, the HLA class II heterodimers, specifically DQ2 and DQ8, have a relevant role, in the heritability of CD. HLA-DQ2 homozygosis confers a much higher risk (25–30%) of developing early-onset CD in infants with a first-degree family member affected by the disease [86].

Dysbiosis is considered an important factor in the interaction of intestinal and thyroid AIDS. The mechanisms that mediate the interaction of microbiota imbalance and thyroid auto-immunities include: (i) intestinal dysbiosis, which interrupts self-tolerance and tolerance to non-pathogenic bacteria, by post-translational modification of proteins. The bacterial enzymatic apparatus can transform the self or nonself peptide into initiators of the autoimmune reaction, (ii) lipopolysaccharides-induced TLR activation, which is associated with thyroiditis and synthesis of anti-thyroglobulin antibodies, (iii) induction of Th-2 lymphocyte differentiation, inhibition of Th-17 lymphocyte differentiation and induction of oral acid tolerance to retinoic acid, which can activate an immune response of tolerance at intestinal level, (iv) permeabilization of the intestinal barrier through injuries of the integrity of tight junctions, deficiency of butyric acid produced by the fermented components in the microbiota or excess of ingested proteins that are metabolized by the microbiota with an increase of putrefaction components; all these factors increase the permeability of the intestinal barrier, facilitating the passage of gliadin and activation of the immune response [26]; (v) changes in the transcriptome, proteome and metabolome of the microbiota [34].

Hashimoto’s thyroiditis and CD share common antibodies, that is anti-tissular transglutaminase (anti-tTg). In patients with CD, tTg binds to the thyroid follicles and the extracellular matrix of the follicles, therefore amplifying the interactions of the microbiota with the thyroid tissue. There is a direct correlation between serum titers of anti-tTg anti-TPO antibodies. DR3-DQ2 and DR4-DQ8 alleles, involved in CD, are reported as common genes that predispose to endocrine AIDS [80].

6. The microbiota interference with other autoimmune-mediated diseases

Rheumatoid arthritis is characterized by a severe and chronic inflammatory condition of the joints. The clinical course of the disease underlines the potential role of dysbiosis in triggering an inflammatory process that involves autoimmune components [87]. Germ-free animals are protected from rheumatoid arthritis in experimental settings. However, the disease is induced in mice exposed to Clostridium SFB, which may act as pathobiont or symbiont, depending on conditions that are host-dependent or independent. Clostridial antigens stimulate Th17 (proinflammatory) lymphocytes that contribute to the progressive evolution of rheumatoid arthritis. Conversely, neutralization of Th17 lymphocytes halts the evolution of the disease. The microbiome of patients with rheumatoid arthritis is altered, with the abundance of Prevotella copri. Citrullinated peptides and specific anti-citrullinated proteins antibodies (ACPA) have been identified in patients suffering from rheumatoid arthritis. Citrullinated peptides result from the peptidyl-Arg-deiminase (PAD)-catalyzed deamination reaction. The enzyme is mainly released after the lysis of granulocytes, monocytes, and macrophages that accumulate in the inflammatory spreads. However, it is also produced by Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, which citrulinate human fibrinogen, synovial fluid proteins [88]. Citrullination is a common physiological process, especially associated with inflammatory processes. Citrullinated proteins, identified in the inflamed synovial membrane of the arthritic joint, exhibit new epitopes and induce the synthesis of ACPA. Circulating ACPAs incorporate into immune complexes aside from citrullinated peptides originating in the joints.

Periodontitis that is caused by oral microbiota bacteria progresses similarly to rheumatoid arthritis—leukocyte infiltration and the progressive destruction of alveolar bone. Leukocytes release the set of proinflammatory interleukins (such as TNF, Il-1, IL-6, IL-12, IL-17, IL-18, and IL-33), growth factors (such as colony-stimulating factors—i.e., GM-CSF, monocyte-CSF), activator receptor of nuclear factor kappa-β ligand (RANKL), metalloproteases, nitric oxide, and PG E2 [89].

In 2013, Rinaldi identified auto-antibodies against the cellular wall of Saccharomyces cerevisiae in rheumatoid arthritis, lupus erythematosus, and antiphospholipid syndrome. These antibodies are also observed in the sera of 32% of patients with celiac disease before its clinical occurrence, and they are considered as a specific serological marker of the disease [90].

Behcet’s disease is a chronic, multisystemic inflammation that is characterized by uveitis, which is a major cause of blindness, and recurrent ulcerative lesions involving the mouth and genital mucosa. There have been reported changes in Th-1, Th-17, and T-reg lymphocytes, whose activity is regulated by the microbiome [91], as well as the diversification of potentially pathogenic bacteria and the decrease of those that produce butyrate (Clostridium).

The pathological change in ulcerative colitis consists of diffuse inflammation, with limited ulcers in the chorion of the colonic mucosa. The pathological process is extended over the entire mucosa of the intestinal epithelium [92].

In Crohn’s disease, the inflammatory infiltrates often generate extensive granulomas in the submucosa and even in the muscular layer of the colon and small intestine. The pathological process of Crohn’s disease is localized, with the damaged areas of the intestine alternating with the healthy ones [93].

Crohn’s disease and ulcerative colitis are not AIDS in the strict sense, because triggering antigens appear to be components of the intestinal microbiota translocated into the chorion, but are the consequence of a large immune response in non-pathogenic antigens, which occurs in people with a genetic predisposition. The inflammatory condition increases the permeability of the colonic epithelium, and the microbiome is modified—the method of 16S rDNA sequencing has shown a decrease in bacterial diversity, especially of the non-pathogenic population, in favor of potentially pathogenic ones [94].

Lupus erythematosus is the prototype of systemic autoimmune disease—an autoimmune response characterized by hyper-reactivity of B lymphocytes and the presence of a large spectrum of serum antibodies [95]. As its name, the disease involves many organs and systems and has various clinical manifestations. Lupus erythematosus affects especially women (female/male ratio = 9/1), with the highest risk during pregnancy [96]. The intestinal microbiota is altered—depletion of lactobacilli, increased Lachnospiraceae density and general diversity, compared to healthy individuals. A large proportion (over 65%) of patients have periodontitis [97], which is always associated with extensive changes in the oral microbiota, in which species with potential pathogens predominate—Fusobacterium nucleatum, Actinomyces naeslundii, Ps. anaerobius, Bacteroides intermedius, and Porphyromonas gingivalis [98].

Multiple sclerosis is a chronic demyelinating inflammatory disease of the central nervous system, characterized by destruction of the integrity of the haemato-encephalic barrier, T lymphocyte infiltrates, and autoimmune reaction against myelin proteins [99]. The immune response in experimental autoimmune encephalitis is mediated by Th-1 and Th-17 cells. The causative agent is not known, but the modification of the microbiota may be important in the onset and/or progression of autoimmune disease. The autoimmune encephalitis diminishes to extinction in germ-free mice, and colonization with Clostridium SFB restores the severity of the disease, as it stimulates the growth of the population of Th-17 (proinflammatory) lymphocytes [100]. Conversely, the administration of Bacteroides protects against demyelination and expansion of tissue-specific inflammation induced by Treg Foxp3 + [55].

The liver autoimmune disease appears to have a direct connection to the microbial load (cells, lipopolysaccharides, peptidoglycans, flagellin, DNA, RNA, toxins, and metabolites) that reaches the Kupffer cells and sinusoidal capillaries by passaging the portal vein. The immune response to these antigens can initiate liver damage and fibrosis [55, 101].

Vitiligo is a systemic autoimmune disease, which is characterized by areas of skin depigmentation, as a result of melanocyte lysis under the action of TCD8 lymphocytes. Melanocytes are located at the border between the epidermis and the dermis, but the disease is systemic because melanocytes are also found in other tissues. The number of melanocytes is the same in different individuals, but differences in pigmentation result from the number, distribution, and size of melanosomes in keratinocytes. The intestinal microbiota in patients with vitiligo is altered and is characterized by decreased taxonomic diversity [18, 102].

Atopic dermatitis is an inflammatory skin disease, clinically characterized by pruritus and xerosis (dry skin). The underlying cause is delayed hypersensitivity mediated by T lymphocytes. The local trigger is the colonization of the skin with Staphylococcus aureus. The toxins released by S. aureus exert a cytotoxic effect [103, 104].

7. Conclusions

Intestinal dysbiosis alters the permeability of the intestinal barrier. The passage of the microbiota antigens into the internal environment may induce the loss of self-tolerance with the generation of autoantibodies and/or autoreactive T cells, leading to the occurrence of cross-reactions. The microbiota alterations lead to an increase in enteric barrier permeability and the occurrence of lymphocyte infiltrates into the epithelial layer, augmenting the risk of cell-mediated auto-immune response. Many questions still need an answer about the role of the microbiota in triggering AIDS, such as—what are the roles of sex hormones and the role of X-linked genes expression in correlation with the microbiome in the polarization of gender-dependent AIDS. Do the changes in the microbiota, which are reported by many authors, contribute to the onset of AIDS by breaking the peripheric tolerance or they are the consequence of AIDS?

Acknowledgments

This research was funded by projects PD224/2021 (PD-2019-0499) and PFE-CDI.2021-587.

Conflict of interest

The authors declare no conflict of interest.