- Email: [email protected]
A Hen in the Wolf Den: A Pathobiont Tale
Immunity
Previews of the mice previously treated with galunisertib rejected the tumor, despite not receiving ongoing TGF-b inhibitor therapy, suggesting that once anti-tumor T cell activity has been stimulated, immunological memory against prior antigens can persist. Whether transient dosing of TGF-b inhibitors in combination with immune checkpoint blockade can effectively induce long-term tumor responses in highly plastic and chromosomally unstable advanced human cancers, remains to be evaluated in prospective clinical trials. Beyond TGF-b, therapies targeting other oncogenic signaling pathways, including MAPK and AKT-PI3K-mTOR, have also shown promise in combination with immune checkpoint blockade, at least partly due to their effect on the tumor microenvironment (Ott et al., 2017). Future research dissecting the crosstalk among these pathways may highlight key signaling nodes for more effective therapeutic targeting.
ACKNOWLEDGMENTS K.G. is supported by American Cancer Society and Shulamit Katzman Endowed Fellowships and is an Investigator of the Stand Up to Cancer Colorectal Cancer Dream Team (SU2C-AACR-DT22-17). J.M. is supported by NIH grant CA34610. REFERENCES Calon, A., Lonardo, E., Berenguer-Llergo, A., Espinet, E., Hernando-Momblona, X., Iglesias, M., Sevillano, M., Palomo-Ponce, S., Tauriello, D.V., Byrom, D., et al. (2015). Stromal gene expression defines poor-prognosis subtypes in colorectal cancer. Nat. Genet. 47, 320–329. Gajewski, T.F. (2015). The Next Hurdle in Cancer Immunotherapy: Overcoming the Non-T-CellInflamed Tumor Microenvironment. Semin. Oncol. 42, 663–671. Gibney, G.T., Weiner, L.M., and Atkins, M.B. (2016). Predictive biomarkers for checkpoint inhibitor-based immunotherapy. Lancet Oncol. 17, e542–e551. Mariathasan, S., Turley, S.J., Nickles, D., Castiglioni, A., Yuen, K., Wang, Y., Kadel Iii, E.E., Koeppen, H., Astarita, J.L., Cubas, R., et al. (2018). TGFb attenuates tumour response to PD-L1
blockade by contributing to exclusion of T cells. Nature 554, 544–548. Massague´, J. (2012). TGFb signalling in context. Nat. Rev. Mol. Cell Biol. 13, 616–630. Neuzillet, C., Tijeras-Raballand, A., Cohen, R., Cros, J., Faivre, S., Raymond, E., and de Gramont, A. (2015). Targeting the TGFb pathway for cancer therapy. Pharmacol. Ther. 147, 22–31. Ott, P.A., Hodi, F.S., Kaufman, H.L., Wigginton, J.M., and Wolchok, J.D. (2017). Combination immunotherapy: a road map. J. Immunother. Cancer 5, 16. Pickup, M., Novitskiy, S., and Moses, H.L. (2013). The roles of TGFb in the tumour microenvironment. Nat. Rev. Cancer 13, 788–799. Ribas, A., and Wolchok, J.D. (2018). Cancer immunotherapy using checkpoint blockade. Science 359, 1350–1355. Tauriello, D.V.F., Palomo-Ponce, S., Stork, D., Berenguer-Llergo, A., Badia-Ramentol, J., Iglesias, M., Sevillano, M., Ibiza, S., Can˜ellas, A., Hernando-Momblona, X., et al. (2018). TGFb drives immune evasion in genetically reconstituted colon cancer metastasis. Nature 554, 538–543.
A Hen in the Wolf Den: A Pathobiont Tale Matteo M. Guerrini,1 Alexis Vogelzang,1 and Sidonia Fagarasan1,* 1Laboratory for Mucosal Immunity, Center for Integrative Medical Sciences, RIKEN Yokohama Institute,1-7-22 Suehiro-cho Tsurumi-ku, Yokohama, 230-0045 Kanagawa, Japan *Correspondence: [email protected] https://doi.org/10.1016/j.immuni.2018.04.003
Disruption of the gut microbiota is thought to contribute to disease onset in individuals with a genetic predisposition to autoimmunity. In a recent issue of Science, Manfredo Vieira et al. (2018) identify translocation of the gut commensal Enterococcus gallinarum into the liver as a trigger for the autoimmune disease systemic lupus erythematous. Systemic lupus erythematous (SLE) is characterized by a type I interferon (IFN) gene signature and systemic autoimmunity. In lupus patients, autoantibodies target diverse antigens, including double-stranded DNA (dsDNA), phospholipids, cardiolipin, and b,2-glycoprotein. Genome-wide association studies have identified a component of genetic risk for SLE; however disease onset occurs in response to environmental insults of a biological, chemical, or physical nature. SLE is also accompanied by endothelial dysfunction that can lead to cardiovascu-
lar complications (Lisnevskaia et al., 2014). Manfredo Vieira et al. (2018) have now identified translocation of the gut commensal bacterium Enterococcus gallinarum as a trigger of SLE in the (NZW 3 BXSB) F1 strain of lupus-prone mice. The microbiota has been long thought to contribute to autoimmune diseases via mechanisms associated with molecular mimicry or bystander activation in which microbes stimulate the expansion of T and B cells that express antigen receptors capable of responding to self-antigens. In a more general sense, the microbiota af-
628 Immunity 48, April 17, 2018 ª 2018 Elsevier Inc.
fects the activation of the immune system. Several lines of evidence show that a loss of microbial diversity or microbial imbalance is associated with autoimmune disease. For example, segmented filamentous bacteria (SFB)—unculturable bacteria belonging to the Firmicutes phylum—expand in the small intestine of immunodeficient mice and sustain the differentiation of Th17 cells. This expanded T cell subset can contribute to the severity of autoimmune diseases, such as experimental autoimmune encephalomyelitis and collagen-induced arthritis, in mice.
Immunity
Previews
Figure 1. Translocation of the Gut Commensal Bacterium Enterococcus gallinarum Triggers the Onset of Systemic Lupus Erythematosus in the (NZW 3 BXSB) F1 Strain of Lupus-Prone Mice The gut of lupus-prone mice is characterized by compromised barrier function, reduced mucus expression, and an imbalance of microbial diversity, resulting in bacterial translocation. In the lamina propria of the small intestine, E. gallinarum induces an increase of plasmacytoid dendritic cells (pDCs) that produce IFNa. E. gallinarum produces AhR ligands, which enhance Th17 and T follicular helper (Tfh) cell activation and differentiation. In the liver, E. gallinarum induces expression of lupus-specific autoantigens and inflammatory factors, enhancing pathogenic deposition of immune complexes in multiple organs.
Gut dysbiosis has also been reported in patients with rheumatoid arthritis, type I diabetes, multiple sclerosis, and SLE (Chervonsky, 2013), but the underlying mechanisms linking the changes in bacterial diversity or function to autoimmunity remain largely elusive. Microbial imbalance might foster the expansion of autoimmunity-prone immune cells, which in turn drive further inflammation and dysbiosis, thus feeding a vicious cycle that appears to be common to many autoimmune diseases. Therefore, further work is required to identify the molecular mechanisms underlying the microbiota-driven expansion of autoimmune cells in specific autoimmune manifestations.
Manfredo Vieira et al. (2018) asked whether interventions targeting bacterial translocation from the gut could alleviate disease in animals predisposed to autoimmunity. The authors observed that treatment of the NZW x BXSB F1 strain of lupus-prone mice (lupus mice hereafter) with antibiotics, particularly vancomycin, extended their lifespan and lowered serum levels of antibodies against SLE autoantigens such as beta2 glycoprotein I (b2-GPI) and endogenous retrovirus glycoprotein 70 (Erv gp70). These observations led the authors to hypothesize that epithelial defects associated with lupus compromise the integrity of the gut epithelial barrier to allow bacterial translocation and thereby promote systemic
inflammation. A gut bacterium from the phylum Firmicutes, E. gallinarum, could be cultured from the liver, mesenteric veins, and lymph nodes of lupus mice, but not from the tissues of wild-type mice or lupus mice receiving vancomycin. Interestingly, and perhaps somewhat ironically, E. gallinarum achieved notoriety for causing hospital-acquired infections due to its intrinsic resistance to vancomycin, which in the 1980s was used for treating enterococcal infections resistant to aminoglycosides (Lebreton et al., 2014). E. gallinarum is a pathobiont—a minor microbiota representative that can become infectious in different biological conditions—and can cause severe pathologies such as endocarditis and meningitis. Indeed, Manfredo Vieira et al. (2018) isolated antibiotic-resistant E. gallinarum from the liver of a vancomycin-treated mouse that developed a severe lupus phenotype. To assess the contribution of this bacterium to the autoimmune phenotype of lupus mice, the authors generated mice monocolonized with E. gallinarum. Mice monocolonized with Enterococcus fecalis, a bacterium belonging to the same genus, and Bacteroides Thetaiotaomicron, a commensal bacterium of the unrelated Bacteroidetes phylum, were generated as controls. Mice monocolonized with E. gallinarum alone showed decreased expression of adhesion molecules, antimicrobial peptides, and mucosal proteins in the ileum. Here too, the authors observed bacterial translocation to other organs, as well as induction of autoantibodies directed against RNA and dsDNA. Importantly, E. gallinarum induced the expansion of plasmacytoid dendritic cells (pDCs) in the lamina propria of the small intestine; pDCs are potent producers of type I IFNs, which are linked to SLE in both mice and humans. Bacterial screening suggested that although E. gallinarum was a relatively minor component of the gut microbiome of lupus mice, it was prominent in microbiological analyses of internal organs, including the liver. Therefore, hepatocytes from lupus mice were co-cultured with E. gallinarum, E. fecalis, and B. Thetaiotaomicron. Among these, E. gallinarum most efficiently induced the transcription of IFNa and the lupus autoantigens b,2-GPI and Erv gp70. As in the monocolonized ileum hepatocyte, Immunity 48, April 17, 2018 629
Immunity
Previews stimulation with E. gallinarum led to the induction of Ahr gene expression. Activation of the AhR pathway was confirmed by the increased transcription of the downstream genes Cyp1a1 and Cyp1a2, suggesting that this pathway drives autoimmunity in both organs. Consistent with the Th17-cell-promoting function of the AhR pathway, Th17 cell frequencies increased in the small-intestinal lamina propria and the mesenteric lymph nodes of E.-gallinarum-monocolonized mice. A specific antagonist blocking AhR signaling reduced the levels of serum anti-dsDNA autoantibodies in E.-gallinarum-monocolonized mice, supporting the role of AhR-Th17 axis in inducing autoimmune inflammation (Figure 1). How do these observations in mouse models relate to the pathology of SLE or autoimmune hepatitis? Longitudinal analysis of fecal samples from SLE patients revealed the presence of serum proteins, indicating gut leakiness. PCRbased analysis confirmed the presence of E. gallinarum in liver biopsies of patients with SLE or autoimmune hepatitis but not in the livers of healthy transplant donors or in patients with hepatic cirrhosis. Co-culture of human hepatocytes with E. gallinarum induced the expression of b,2-GPI type I IFNs and AhR, in line with the observations in mice. Lastly, patients with SLE and liver autoimmunity showed high titers of serum antibody directed against E. gallinarum and human RNA. No antibodies against E. fecalis or B. Thetaiotaomicron were found, suggesting that E. gallinarum is uniquely associated with SLE and liver autoimmunity in humans as well as in mice. It is surprising to see such a specific association with a single pathobiont in a disease with diverse clinical presentations. If further cohorts will confirm a specific E. gallinarum antibody signal in patients, this could be a useful diagnostic biomarker. As so often happens with interesting discoveries, the work by Kriegel and colleagues raises many questions for further study. For example, it is unclear why vancomycin effectively depletes E. gallinarum from the liver of the majority of lupusprone mice when it is known to have intrinsic resistance to this antibiotic. An intriguing hypothesis is that E. gallinarum physiology might be modulated by the surrounding environment (the healthy or 630 Immunity 48, April 17, 2018
dysbiotic gut, the liver), as has been documented in other bacteria. For example, Clostridium difficile is an ubiquitous pathobiont that modulates toxin production in response to differences in surrounding gut commensals after antibiotic administration (Theriot and Young, 2015). The authors observed that E. gallinarum has a singular capacity to induce autoantigen expression and autoantibodies against human nucleic acids. The mechanisms underlying these properties remain unknown. Does E. gallinarum induce expression of autoantigens in the liver through Toll-like receptors (TLRs)? In this respect, analysis of mice deficient for MyD88, a requisite component of TLR signaling, could be informative, as could the extension of these studies to the investigation of a link between E. gallinarum and gender susceptibly rates. SLE is disproportionally diagnosed in women (90%), and the TLR7 locus is among X-linked genes that might promote the disease in females. E. gallinarum is distinct from most other enterococci in its expression of a motilityenhancing flagellum, perhaps explaining its ability to translocate and colonize distant organs. Other bacteria with similar properties are likely to exist among the diversity of microbiota. Furthermore, other tryptophan-utilizing bacteria generate AhR ligands and are likely to have an impact on gut epithelium permeability, T cell expansion, and function; examples of such bacteria include those within the spore-forming Firmicutes phylum. Indeed, overrepresentation of Lachnospiraceae and Clostridiaceae is observed in lupus-prone mice, and the expansion of Lachnospiraceae is associated with an early onset and increased severity of lupus symptoms in females (Zhang et al., 2014) . The authors show that vancomycin treatment of lupus mice reduces autoimmunity. However, treatment of SLE patients with antibiotics, including vancomycin, could have detrimental effects. Targeting Gram-positive bacteria with vancomycin can select for highly resistant E. gallinarum, along with other species of Enterococci. Expansion of Enterococci in the gut often precedes bacterial invasion of the bloodstream (Ubeda et al., 2010), which could fuel autoimmunity by the mechanisms demonstrated by Manfredo Vieira et al. (2018). Broad-spectrum antibiotics targeting Gram-positive as
well as Gram-negative bacteria could pose additional problems. Antibiotic damage to microbial diversity can worsen the vulnerability of the gut epithelium, reducing the production of RegIIIg by Paneth cells, promoting outgrowth of antibiotic-resistant Enterococci (Brandl et al., 2008), or depleting beneficial bacterial species that attenuate immunopathology. Leaky gut and nephritis are ameliorated by colonization with Lactobacillus, by increasing local IL10 production, and by skewing the balance of Treg and Th17 cells toward a Treg-cell-dominant phenotype in another SLE model mouse (MRL/ Mp-Faslpr) (Mu et al., 2017). More sophisticated interventions targeting specific microorganisms on the basis of their functional properties are needed for controling the autoimmunity-driving properties of the microbiota without affecting its beneficial effects. Preliminary experiments investigating neonatal vaccination against E. gallinarum by Manfredo Vieira et al. (2018) suggest this might be a promising approach. Elegant studies of the fecal metabolites in SLE patients suggest that immune status regulates the intestinal bacterial functionality even in the absence of gross differences in the microbiota, further underlining the complexity of the system (Rojo et al., 2015). Combining microbiome transcriptome and metabolome phenotyping of fecal content has the potential to reveal how the functions of pathobionts change at a molecular level within the framework of the local microbial community. By revealing a link between a pathobiont and lupus, Manfredo Vieira et al. (2018) provide new insight into how the pathobionts can drive autoimmunity in the setting of gut dysbiosis. ACKNOWLEDGMENTS We thank N. Minato for comments. Supported by RIKEN Special Postdoctoral Researcher program (SPDR) (A.V.) and intramural grant (S.F. and M.M.G). REFERENCES Brandl, K., Plitas, G., Mihu, C.N., Ubeda, C., Jia, T., Fleisher, M., Schnabl, B., DeMatteo, R.P., and Pamer, E.G. (2008). Vancomycin-resistant enterococci exploit antibiotic-induced innate immune deficits. Nature 455, 804–807. Chervonsky, A.V. (2013). Microbiota and autoimmunity. Cold Spring Harb Perspect Biol 5, a,007294.
Immunity
Previews Lebreton, F., Willems, R.J.L., and Gilmore, M.S. (2014). Enterococcus Diversity Origins in Nature, and Gut Colonization. In Enterococci: From Commensals to Leading Causes of Drug Resistant Infection, M.S. Gilmore, D.B. Clewell, Y. Ike, and N. Shankar, eds. (Massachusetts Eye and Ear Infirmary). Lisnevskaia, L., Murphy, G., and Isenberg, D. (2014). Systemic lupus erythematosus. Lancet 384, 1878–1888. Manfredo Vieira, S., Hiltensperger, M., Kumar, V., Zegarra-Ruiz, D., Dehner, C., Khan, N., Costa, F.R.C., Tiniakou, E., Greiling, T., Ruff, W., et al. (2018). Translocation of a gut pathobiont drives
autoimmunity in mice and humans. Science 359, 1156–1161. Mu, Q., Zhang, H., Liao, X., Lin, K., Liu, H., Edwards, M.R., Ahmed, S.A., Yuan, R., Li, L., Cecere, T.E., et al. (2017). Control of lupus nephritis by changes of gut microbiota. Microbiome 5, 73. Rojo, D., Hevia, A., Bargiela, R., Lo´pez, P., Cuervo, A., Gonza´lez, S., Sua´rez, A., Sa´nchez, B., Martı´nez-Martı´nez, M., Milani, C., et al. (2015). Ranking the impact of human health disorders on gut metabolism: systemic lupus erythematosus and obesity as study cases. Sci. Rep. 5, 8310. Theriot, C.M., and Young, V.B. (2015). Interactions Between the Gastrointestinal Microbiome
and Clostridium difficile. Annu. Rev. Microbiol. 69, 445–461. Ubeda, C., Taur, Y., Jenq, R.R., Equinda, M.J., Son, T., Samstein, M., Viale, A., Socci, N.D., van den Brink, M.R., Kamboj, M., and Pamer, E.G. (2010). Vancomycin-resistant Enterococcus domination of intestinal microbiota is enabled by antibiotic treatment in mice and precedes bloodstream invasion in humans. J. Clin. Invest. 120, 4332–4341. Zhang, H., Liao, X., Sparks, J.B., and Luo, X.M. (2014). Dynamics of gut microbiota in autoimmune lupus. Appl. Environ. Microbiol. 80, 7551–7560.
Immunity 48, April 17, 2018 631
Previews of the mice previously treated with galunisertib rejected the tumor, despite not receiving ongoing TGF-b inhibitor therapy, suggesting that once anti-tumor T cell activity has been stimulated, immunological memory against prior antigens can persist. Whether transient dosing of TGF-b inhibitors in combination with immune checkpoint blockade can effectively induce long-term tumor responses in highly plastic and chromosomally unstable advanced human cancers, remains to be evaluated in prospective clinical trials. Beyond TGF-b, therapies targeting other oncogenic signaling pathways, including MAPK and AKT-PI3K-mTOR, have also shown promise in combination with immune checkpoint blockade, at least partly due to their effect on the tumor microenvironment (Ott et al., 2017). Future research dissecting the crosstalk among these pathways may highlight key signaling nodes for more effective therapeutic targeting.
ACKNOWLEDGMENTS K.G. is supported by American Cancer Society and Shulamit Katzman Endowed Fellowships and is an Investigator of the Stand Up to Cancer Colorectal Cancer Dream Team (SU2C-AACR-DT22-17). J.M. is supported by NIH grant CA34610. REFERENCES Calon, A., Lonardo, E., Berenguer-Llergo, A., Espinet, E., Hernando-Momblona, X., Iglesias, M., Sevillano, M., Palomo-Ponce, S., Tauriello, D.V., Byrom, D., et al. (2015). Stromal gene expression defines poor-prognosis subtypes in colorectal cancer. Nat. Genet. 47, 320–329. Gajewski, T.F. (2015). The Next Hurdle in Cancer Immunotherapy: Overcoming the Non-T-CellInflamed Tumor Microenvironment. Semin. Oncol. 42, 663–671. Gibney, G.T., Weiner, L.M., and Atkins, M.B. (2016). Predictive biomarkers for checkpoint inhibitor-based immunotherapy. Lancet Oncol. 17, e542–e551. Mariathasan, S., Turley, S.J., Nickles, D., Castiglioni, A., Yuen, K., Wang, Y., Kadel Iii, E.E., Koeppen, H., Astarita, J.L., Cubas, R., et al. (2018). TGFb attenuates tumour response to PD-L1
blockade by contributing to exclusion of T cells. Nature 554, 544–548. Massague´, J. (2012). TGFb signalling in context. Nat. Rev. Mol. Cell Biol. 13, 616–630. Neuzillet, C., Tijeras-Raballand, A., Cohen, R., Cros, J., Faivre, S., Raymond, E., and de Gramont, A. (2015). Targeting the TGFb pathway for cancer therapy. Pharmacol. Ther. 147, 22–31. Ott, P.A., Hodi, F.S., Kaufman, H.L., Wigginton, J.M., and Wolchok, J.D. (2017). Combination immunotherapy: a road map. J. Immunother. Cancer 5, 16. Pickup, M., Novitskiy, S., and Moses, H.L. (2013). The roles of TGFb in the tumour microenvironment. Nat. Rev. Cancer 13, 788–799. Ribas, A., and Wolchok, J.D. (2018). Cancer immunotherapy using checkpoint blockade. Science 359, 1350–1355. Tauriello, D.V.F., Palomo-Ponce, S., Stork, D., Berenguer-Llergo, A., Badia-Ramentol, J., Iglesias, M., Sevillano, M., Ibiza, S., Can˜ellas, A., Hernando-Momblona, X., et al. (2018). TGFb drives immune evasion in genetically reconstituted colon cancer metastasis. Nature 554, 538–543.
A Hen in the Wolf Den: A Pathobiont Tale Matteo M. Guerrini,1 Alexis Vogelzang,1 and Sidonia Fagarasan1,* 1Laboratory for Mucosal Immunity, Center for Integrative Medical Sciences, RIKEN Yokohama Institute,1-7-22 Suehiro-cho Tsurumi-ku, Yokohama, 230-0045 Kanagawa, Japan *Correspondence: [email protected] https://doi.org/10.1016/j.immuni.2018.04.003
Disruption of the gut microbiota is thought to contribute to disease onset in individuals with a genetic predisposition to autoimmunity. In a recent issue of Science, Manfredo Vieira et al. (2018) identify translocation of the gut commensal Enterococcus gallinarum into the liver as a trigger for the autoimmune disease systemic lupus erythematous. Systemic lupus erythematous (SLE) is characterized by a type I interferon (IFN) gene signature and systemic autoimmunity. In lupus patients, autoantibodies target diverse antigens, including double-stranded DNA (dsDNA), phospholipids, cardiolipin, and b,2-glycoprotein. Genome-wide association studies have identified a component of genetic risk for SLE; however disease onset occurs in response to environmental insults of a biological, chemical, or physical nature. SLE is also accompanied by endothelial dysfunction that can lead to cardiovascu-
lar complications (Lisnevskaia et al., 2014). Manfredo Vieira et al. (2018) have now identified translocation of the gut commensal bacterium Enterococcus gallinarum as a trigger of SLE in the (NZW 3 BXSB) F1 strain of lupus-prone mice. The microbiota has been long thought to contribute to autoimmune diseases via mechanisms associated with molecular mimicry or bystander activation in which microbes stimulate the expansion of T and B cells that express antigen receptors capable of responding to self-antigens. In a more general sense, the microbiota af-
628 Immunity 48, April 17, 2018 ª 2018 Elsevier Inc.
fects the activation of the immune system. Several lines of evidence show that a loss of microbial diversity or microbial imbalance is associated with autoimmune disease. For example, segmented filamentous bacteria (SFB)—unculturable bacteria belonging to the Firmicutes phylum—expand in the small intestine of immunodeficient mice and sustain the differentiation of Th17 cells. This expanded T cell subset can contribute to the severity of autoimmune diseases, such as experimental autoimmune encephalomyelitis and collagen-induced arthritis, in mice.
Immunity
Previews
Figure 1. Translocation of the Gut Commensal Bacterium Enterococcus gallinarum Triggers the Onset of Systemic Lupus Erythematosus in the (NZW 3 BXSB) F1 Strain of Lupus-Prone Mice The gut of lupus-prone mice is characterized by compromised barrier function, reduced mucus expression, and an imbalance of microbial diversity, resulting in bacterial translocation. In the lamina propria of the small intestine, E. gallinarum induces an increase of plasmacytoid dendritic cells (pDCs) that produce IFNa. E. gallinarum produces AhR ligands, which enhance Th17 and T follicular helper (Tfh) cell activation and differentiation. In the liver, E. gallinarum induces expression of lupus-specific autoantigens and inflammatory factors, enhancing pathogenic deposition of immune complexes in multiple organs.
Gut dysbiosis has also been reported in patients with rheumatoid arthritis, type I diabetes, multiple sclerosis, and SLE (Chervonsky, 2013), but the underlying mechanisms linking the changes in bacterial diversity or function to autoimmunity remain largely elusive. Microbial imbalance might foster the expansion of autoimmunity-prone immune cells, which in turn drive further inflammation and dysbiosis, thus feeding a vicious cycle that appears to be common to many autoimmune diseases. Therefore, further work is required to identify the molecular mechanisms underlying the microbiota-driven expansion of autoimmune cells in specific autoimmune manifestations.
Manfredo Vieira et al. (2018) asked whether interventions targeting bacterial translocation from the gut could alleviate disease in animals predisposed to autoimmunity. The authors observed that treatment of the NZW x BXSB F1 strain of lupus-prone mice (lupus mice hereafter) with antibiotics, particularly vancomycin, extended their lifespan and lowered serum levels of antibodies against SLE autoantigens such as beta2 glycoprotein I (b2-GPI) and endogenous retrovirus glycoprotein 70 (Erv gp70). These observations led the authors to hypothesize that epithelial defects associated with lupus compromise the integrity of the gut epithelial barrier to allow bacterial translocation and thereby promote systemic
inflammation. A gut bacterium from the phylum Firmicutes, E. gallinarum, could be cultured from the liver, mesenteric veins, and lymph nodes of lupus mice, but not from the tissues of wild-type mice or lupus mice receiving vancomycin. Interestingly, and perhaps somewhat ironically, E. gallinarum achieved notoriety for causing hospital-acquired infections due to its intrinsic resistance to vancomycin, which in the 1980s was used for treating enterococcal infections resistant to aminoglycosides (Lebreton et al., 2014). E. gallinarum is a pathobiont—a minor microbiota representative that can become infectious in different biological conditions—and can cause severe pathologies such as endocarditis and meningitis. Indeed, Manfredo Vieira et al. (2018) isolated antibiotic-resistant E. gallinarum from the liver of a vancomycin-treated mouse that developed a severe lupus phenotype. To assess the contribution of this bacterium to the autoimmune phenotype of lupus mice, the authors generated mice monocolonized with E. gallinarum. Mice monocolonized with Enterococcus fecalis, a bacterium belonging to the same genus, and Bacteroides Thetaiotaomicron, a commensal bacterium of the unrelated Bacteroidetes phylum, were generated as controls. Mice monocolonized with E. gallinarum alone showed decreased expression of adhesion molecules, antimicrobial peptides, and mucosal proteins in the ileum. Here too, the authors observed bacterial translocation to other organs, as well as induction of autoantibodies directed against RNA and dsDNA. Importantly, E. gallinarum induced the expansion of plasmacytoid dendritic cells (pDCs) in the lamina propria of the small intestine; pDCs are potent producers of type I IFNs, which are linked to SLE in both mice and humans. Bacterial screening suggested that although E. gallinarum was a relatively minor component of the gut microbiome of lupus mice, it was prominent in microbiological analyses of internal organs, including the liver. Therefore, hepatocytes from lupus mice were co-cultured with E. gallinarum, E. fecalis, and B. Thetaiotaomicron. Among these, E. gallinarum most efficiently induced the transcription of IFNa and the lupus autoantigens b,2-GPI and Erv gp70. As in the monocolonized ileum hepatocyte, Immunity 48, April 17, 2018 629
Immunity
Previews stimulation with E. gallinarum led to the induction of Ahr gene expression. Activation of the AhR pathway was confirmed by the increased transcription of the downstream genes Cyp1a1 and Cyp1a2, suggesting that this pathway drives autoimmunity in both organs. Consistent with the Th17-cell-promoting function of the AhR pathway, Th17 cell frequencies increased in the small-intestinal lamina propria and the mesenteric lymph nodes of E.-gallinarum-monocolonized mice. A specific antagonist blocking AhR signaling reduced the levels of serum anti-dsDNA autoantibodies in E.-gallinarum-monocolonized mice, supporting the role of AhR-Th17 axis in inducing autoimmune inflammation (Figure 1). How do these observations in mouse models relate to the pathology of SLE or autoimmune hepatitis? Longitudinal analysis of fecal samples from SLE patients revealed the presence of serum proteins, indicating gut leakiness. PCRbased analysis confirmed the presence of E. gallinarum in liver biopsies of patients with SLE or autoimmune hepatitis but not in the livers of healthy transplant donors or in patients with hepatic cirrhosis. Co-culture of human hepatocytes with E. gallinarum induced the expression of b,2-GPI type I IFNs and AhR, in line with the observations in mice. Lastly, patients with SLE and liver autoimmunity showed high titers of serum antibody directed against E. gallinarum and human RNA. No antibodies against E. fecalis or B. Thetaiotaomicron were found, suggesting that E. gallinarum is uniquely associated with SLE and liver autoimmunity in humans as well as in mice. It is surprising to see such a specific association with a single pathobiont in a disease with diverse clinical presentations. If further cohorts will confirm a specific E. gallinarum antibody signal in patients, this could be a useful diagnostic biomarker. As so often happens with interesting discoveries, the work by Kriegel and colleagues raises many questions for further study. For example, it is unclear why vancomycin effectively depletes E. gallinarum from the liver of the majority of lupusprone mice when it is known to have intrinsic resistance to this antibiotic. An intriguing hypothesis is that E. gallinarum physiology might be modulated by the surrounding environment (the healthy or 630 Immunity 48, April 17, 2018
dysbiotic gut, the liver), as has been documented in other bacteria. For example, Clostridium difficile is an ubiquitous pathobiont that modulates toxin production in response to differences in surrounding gut commensals after antibiotic administration (Theriot and Young, 2015). The authors observed that E. gallinarum has a singular capacity to induce autoantigen expression and autoantibodies against human nucleic acids. The mechanisms underlying these properties remain unknown. Does E. gallinarum induce expression of autoantigens in the liver through Toll-like receptors (TLRs)? In this respect, analysis of mice deficient for MyD88, a requisite component of TLR signaling, could be informative, as could the extension of these studies to the investigation of a link between E. gallinarum and gender susceptibly rates. SLE is disproportionally diagnosed in women (90%), and the TLR7 locus is among X-linked genes that might promote the disease in females. E. gallinarum is distinct from most other enterococci in its expression of a motilityenhancing flagellum, perhaps explaining its ability to translocate and colonize distant organs. Other bacteria with similar properties are likely to exist among the diversity of microbiota. Furthermore, other tryptophan-utilizing bacteria generate AhR ligands and are likely to have an impact on gut epithelium permeability, T cell expansion, and function; examples of such bacteria include those within the spore-forming Firmicutes phylum. Indeed, overrepresentation of Lachnospiraceae and Clostridiaceae is observed in lupus-prone mice, and the expansion of Lachnospiraceae is associated with an early onset and increased severity of lupus symptoms in females (Zhang et al., 2014) . The authors show that vancomycin treatment of lupus mice reduces autoimmunity. However, treatment of SLE patients with antibiotics, including vancomycin, could have detrimental effects. Targeting Gram-positive bacteria with vancomycin can select for highly resistant E. gallinarum, along with other species of Enterococci. Expansion of Enterococci in the gut often precedes bacterial invasion of the bloodstream (Ubeda et al., 2010), which could fuel autoimmunity by the mechanisms demonstrated by Manfredo Vieira et al. (2018). Broad-spectrum antibiotics targeting Gram-positive as
well as Gram-negative bacteria could pose additional problems. Antibiotic damage to microbial diversity can worsen the vulnerability of the gut epithelium, reducing the production of RegIIIg by Paneth cells, promoting outgrowth of antibiotic-resistant Enterococci (Brandl et al., 2008), or depleting beneficial bacterial species that attenuate immunopathology. Leaky gut and nephritis are ameliorated by colonization with Lactobacillus, by increasing local IL10 production, and by skewing the balance of Treg and Th17 cells toward a Treg-cell-dominant phenotype in another SLE model mouse (MRL/ Mp-Faslpr) (Mu et al., 2017). More sophisticated interventions targeting specific microorganisms on the basis of their functional properties are needed for controling the autoimmunity-driving properties of the microbiota without affecting its beneficial effects. Preliminary experiments investigating neonatal vaccination against E. gallinarum by Manfredo Vieira et al. (2018) suggest this might be a promising approach. Elegant studies of the fecal metabolites in SLE patients suggest that immune status regulates the intestinal bacterial functionality even in the absence of gross differences in the microbiota, further underlining the complexity of the system (Rojo et al., 2015). Combining microbiome transcriptome and metabolome phenotyping of fecal content has the potential to reveal how the functions of pathobionts change at a molecular level within the framework of the local microbial community. By revealing a link between a pathobiont and lupus, Manfredo Vieira et al. (2018) provide new insight into how the pathobionts can drive autoimmunity in the setting of gut dysbiosis. ACKNOWLEDGMENTS We thank N. Minato for comments. Supported by RIKEN Special Postdoctoral Researcher program (SPDR) (A.V.) and intramural grant (S.F. and M.M.G). REFERENCES Brandl, K., Plitas, G., Mihu, C.N., Ubeda, C., Jia, T., Fleisher, M., Schnabl, B., DeMatteo, R.P., and Pamer, E.G. (2008). Vancomycin-resistant enterococci exploit antibiotic-induced innate immune deficits. Nature 455, 804–807. Chervonsky, A.V. (2013). Microbiota and autoimmunity. Cold Spring Harb Perspect Biol 5, a,007294.
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