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Early-Life Gut Dysbiosis: A Driver of Later-Life Fibrosis?

      Using a novel mouse model of scleroderma induced by immunization with topoisomerase-I peptide-loaded dendritic cells, Mehta et al. found that early-life antibiotic exposure resulted in increased later-life fibrosis in the skin and lungs. These observations advance the novel concept that gut microbiome alterations caused by early-life exposures may contribute to scleroderma pathogenesis, and warrant in-depth characterization and validation in complementary disease models.
      • Skin and internal organ fibrosis in scleroderma is associated with anti- topoisomerase-I autoimmunity and vascular and fibrotic damage driven by poorly defined environmental exposures.
      • Alterations in the gut microbiome are implicated in chronic inflammatory and autoimmune disorders, including inflammatory bowel disease, type 1 diabetes, rheumatoid arthritis, multiple sclerosis, psoriasis, and possibly scleroderma.
      • In a novel mouse model of scleroderma associated with anti-topoisomerase-I immunity, skin and lung fibrosis were exacerbated by perinatal antibiotic treatment, suggesting that early-life changes in the gut microbial community have durable and potentially pathological effects on shaping later-life fibrotic responses.
      Systemic sclerosis (SSc) is a sporadic autoimmune disease of unknown cause. Although genome-wide association studies have identified a number of genetic risk loci shared with autoimmune and inflammatory diseases such as lupus and psoriasis, SSc shows only modest familial heritability, and environmental factors are therefore thought to have a crucial role. Recent studies have started to unravel alterations in gut microbial composition and diversity, that is, dysbiosis, associated with SSc, suggesting a potential pathogenic role (
      • Volkmann E.R.
      Intestinal microbiome in scleroderma: recent progress.
      ).

      Systemic sclerosis: complex tripartite pathogenesis

      Skin induration in SSc is invariably accompanied by internal organ fibrosis, most prominent in the lungs, that may cause considerable morbidity and mortality. Despite encouraging recent clinical trials with mycophenolate mofetil, autologous hematopoietic stem cell transplantation, and drugs targeting transforming growth factor-β, IL-6, CD20/B cells, cannabinoid receptors, and others, effective disease-modifying therapy for SSc (and other forms of fibrosis) remains elusive. A major impediment relates to the complexity of SSc pathogenesis, with overlapping autoimmune, vascular, and fibrotic processes, each contributing to protean disease manifestations (
      • Gabrielli A.
      • Avvedimento E.V.
      • Krieg T.
      Scleroderma.
      ). Furthermore, SSc demonstrates patient-to-patient heterogeneity, variable natural history, and unpredictable outcomes. In view of these considerations, preclinical disease models of SSc assume a pivotal role for revealing key pathogenic cell types, pathways and mediators, and for evaluating the efficacy and mechanism of action of novel therapies.
      In its earliest stage, SSc is dominated by autoimmune and inflammatory features, with tissue infiltration with activated T and B cells, monocytes, macrophages, innate lymphoid cells, and dendritic cells (
      • Allanore Y.
      • Simms R.
      • Distler O.
      • Trojanowska M.
      • Pope J.
      • Denton C.P.
      • et al.
      Systemic sclerosis.
      ). Additionally, mutually exclusive autoantibodies against topoisomerase-I, centromere, or RNA polymerase III can be detected in most patients. Although the origins and pathogenic roles of these disease-specific antibodies remain unknown, they are exceptionally useful as clinical tools in SSc to stratify patients and anticipate particular disease complications. For instance, anti-RNA polymerase III antibodies identify patients with SSc at increased risk for scleroderma renal crisis, whereas anti-topoisomerase-I positivity predicts extensive skin and lung involvement and a poor outcome. In addition to cellular and humoral autoimmunity, vascular damage is prevalent, and chronic fibrosis affects the skin and many organs.

      Preclinical disease models for SSc: an innovative cellular immunization approach

      Despite considerable effort, there are currently no preclinical models of SSc that recapitulate the three pathomechanistic hallmarks of the human disease (
      • Tsujino K.
      • Sheppard D.
      Critical appraisal of the utility and limitations of animal models of scleroderma.
      ). In particular, widely used rodent models of skin and lung fibrosis induced by bleomycin or other noxious agents typically cause only localized and self-limited fibrosis, do not generate disease-specific autoantibodies, and lack the microvascular features of SSc. Similarly, modeling SSc in mutant or genetically engineered mouse strains has utility for probing the fibrosis-inflammation link, but not other features of SSc. To maximize the value of preclinical SSc models, whether for understanding pathogenesis or evaluating the efficacy and mode of action of novel therapies, a combination strategy using multiple complementary disease models is recommended (
      • Del Galdo F.
      • Matucci-Cerinic M.
      The search for the perfect animal model discloses the importance of biological targets for the treatment of systemic sclerosis.
      ).
      • Mehta H.
      • Goulet P.O.
      • Mashiko S.
      • Desjardins J.
      • Pérez G.
      • Koenig M.
      • et al.
      Early-life antibiotic exposure causes intestinal dysbiosis and exacerbates skin and lung pathology in experimental systemic sclerosis.
      present an interesting and potentially informative novel approach to model SSc predicated on the putative roles of dendritic cells and topoisomerase-I-specific autoimmunity. The authors performed immunization with dendritic cells loaded with the topoisomerase-I peptide to induce immunity, inflammation, and skin and lung fibrosis in BALB/c mice. Remarkably, unlike other inducible models of SSc, topoisomerase-I peptide-immunized mice develop durable fibrosis detectable as late as 10 weeks after the final immunization (
      • Mehta H.
      • Goulet P.O.
      • Nguyen V.
      • Perez G.
      • Koenig M.
      • Senecal J.L.
      • et al.
      Topoisomerase I peptide-loaded dendritic cells induce autoantibody response as well as skin and lung fibrosis.
      ). Although the mice mount an anti-topoisomerase-I antibody response similar to patients with SSc, fibrosis preceded the appearance of these antibodies, excluding their direct role in fibrosis initiation, though perhaps not in its persistence. Microangiopathy does not appear to develop in the immunized mice, underlying the inability of preclinical disease models to fully recapitulate the pathomechanistic spectrum of human SSc.

      Early-life gut dysbiosis influences fibrosis propensity in the mouse

      Employing their innovative immunization-induced disease model,
      • Mehta H.
      • Goulet P.O.
      • Mashiko S.
      • Desjardins J.
      • Pérez G.
      • Koenig M.
      • et al.
      Early-life antibiotic exposure causes intestinal dysbiosis and exacerbates skin and lung pathology in experimental systemic sclerosis.
      advance a relatively novel concept for SSc pathogenesis based on gut dysbiosis. The gut microbiome represents a dynamic ecosystem that is relatively stable in adults, but is shaped by early-life exposures. These include diet composition and complexity, maturation of the immune system, host genetics, lifestyle, social interactions, environmental exposures, and antibiotic therapy (
      • Yatsunenko T.
      • Rey F.E.
      • Manary M.J.
      • Trehan I.
      • Dominguez-Bello M.G.
      • Contreras M.
      • et al.
      Human gut microbiome viewed across age and geography.
      ). In the Mehta et al. study, immunized mice were treated with streptomycin, an oral antibiotic that is only minimally absorbed through the gastrointestinal tract. Antibiotic treatment has been shown to induce long-lasting changes in gut microbiome (
      • Langdon A.
      • Crook N.
      • Dantas G.
      The effects of antibiotics on the microbiome throughout development and alternative approaches for therapeutic modulation.
      ). Mehta et al. demonstrate that streptomycin treatment during early life, that is, in utero and during the perinatal period, resulted in increased later-life skin and lung fibrosis in topoisomerase-I peptide-immunized mice. Persistent taxonomic changes at the phylum, family, and species levels were evident in the gut microbiome. Early-life antibiotic exposure was associated with altered expression of fibrosis-related genes and evidence of skin and pulmonary fibrosis, and dysregulated T-cell responses in the lungs. The mechanistic links between the microbial alterations and increased fibrosis, and potential direct connections between the gut microbiome and the skin (
      • Zanvit P.
      • Konkel J.E.
      • Jiao X.
      • Kasagi S.
      • Zhang D.
      • Wu R.
      • et al.
      Antibiotics in neonatal life increase murine susceptibility to experimental psoriasis.
      ) or the lung (
      • Barfod K.K.
      • Roggenbuck M.
      • Hansen L.H.
      • Schjorring S.
      • Larsen S.T.
      • Sorensen S.J.
      • et al.
      The murine lung microbiome in relation to the intestinal and vaginal bacterial communities.
      ), or the effect of antibiotic by itself on lung and skin fibrosis, were not explored. Validation of these antibiotic exposure-induced fibrotic responses in alternate murine models of scleroderma, in addition to the present topoisomerase-1 autoimmunity-specific model, will be eagerly awaited.

      Summary

      These concerns do not detract from the novelty of the conceptual model implicating early-life gut dysbiosis in subsequent fibrosis propensity. Precedents for this concept are increasingly abundant. Early-life dysbiosis in antibiotic-treated or germ-free mice has been linked to adult-onset obesity, diabetes mellitus, eosinophilic esophagitis, inflammatory bowel disease, and asthma (reviewed in
      • Gensollen T.
      • Blumberg R.S.
      Correlation between early-life regulation of the immune system by microbiota and allergy development.
      ). The mechanism may involve imprinting of poorly defined immune responses during an early-life “window of opportunity” by interactions between commensal organisms and the host that have profound implications for subsequent disease propensity. For example, microbial colonization during pregnancy in mice has powerful effects on innate immunity in the offspring (
      • Gomez de Aguero M.
      • Ganal-Vonarburg S.C.
      • Fuhrer T.
      • Rupp S.
      • Uchimura Y.
      • Li H.
      • et al.
      The maternal microbiota drives early postnatal innate immune development.
      ).
      The intriguing study by Mehta et al. using a novel SSc-relevant immunization approach to model scleroderma provides evidence that early-life antibiotic exposure causes durable gut dysbiosis associated with exacerbated later-life fibrosis. Whether the profibrotic effects of gut dysbiosis are specific to the novel model induced by anti-topoisomerase-I immunity, or are reproducible in more commonly used and extensively characterized disease models, is an important question to be addressed. Additionally, the cellular mechanisms underlying this phenomenon remain to be elucidated. For instance, is increased later-life fibrosis propensity in the immunized mice due to antibiotic-induced early-life alterations shaping the immune system, or due to persistence of the microbiome alterations? Although the clinical implications of the findings by Mehta et al. are not yet clear, the findings are in line with accumulating evidence that early-life dysbiosis causes durable alterations in the intestinal microbiota that are associated with long-lasting health effects and implicated in a variety of autoimmune, atopic, and inflammatory diseases in adulthood. The present findings add fibrotic conditions to this list, suggest new directions in contemplating the role of environment in SSc pathogenesis, and should inspire further investigations. Ultimately, delineating the nature and contribution of gut dysbiosis to SSc pathogenesis might present new therapeutic opportunities.

      Conflict of Interest

      The authors state no conflict of interest.

      Acknowledgments

      Per journal regulations, we are restricted to a limited number of references. However, many original papers are cited in the listed review articles. We acknowledge partial support by grants from National Institutes of Health (K08 HL130601 to KJH and R56 AG054207 to JV).

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