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1.
Ocul Immunol Inflamm ; : 1-8, 2019 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-31763950

RESUMO

Purpose: Anterior uveitis is the most common anatomic subset of uveitis. We developed a novel multi-parametric flow cytometry panel to identify immune dysregulation signatures in HLA B27-associated acute anterior uveitis (AAU) and axial spondyloarthritis (AxSpA).Methods: We used fluorescence activated cell sorting to characterize T cell cytokine expression in stimulated T cell subsets from patients with AAU (n = 4) compared to healthy controls (n = 14) or subjects with AxSpA (n = 6).Results: Positive findings among subjects with AAU included a statistically significant increase in stimulated granulocyte-macrophage colony stimulating factor (GM-CSF), IL-17, and IL-22 synthesized by CD8 cells, a trend for stimulated ILC (innate lymphoid cells)-3 cells to synthesize more IL-22 (p = .07), and stimulated MAIT (mucosa associated innate lymphoid cells)-like cells that express the T cell receptor V alpha 7.2 to express IL-17A, IL-17F, and IL-22 in a greater percentage of cells relative to controls. IL-17F, GM- CSF, and IL-22 represent potentially novel targets in AAU.Conclusion: Our report is arguably the first to implicate IL-17F or ILC-3 and MAIT cells in the pathogenesis of AAU.Abbreviations AAU: acute anterior uveitis; AxSpA: axial spondyloarthritis; BASDAI: Bath ankylosing spondylitis disease activity index; CCR: chemokine receptor; DMSO: dimethylsulfoxide; EULAR:European League Against Rheumatism; FACS: fluorescence activated cell sorter; FBS: fetal bovine serum; FSC: orward light scatter; GM-CSF: granulocyte-macrophage colony stimulating factor; HC: healthy control; ILC: innate lymphoid cell; KIR: killer immunoglobulin receptor; MAIT: mucosal associated immune T cell; ND: not detected; NK: natural killer cell; OHSU-Oregon Health & Science University; PBMC: peripheral blood mononuclear cell; SSC: side light scatter; TCR: T cell receptor.

2.
Arthritis Rheumatol ; 71(11): 1849-1857, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31216122

RESUMO

OBJECTIVE: To define inflammation-related host-microbe interactions in experimental spondyloarthritis (SpA) using novel inter-omic approaches. METHODS: The relative frequency of gut microbes was determined by 16S ribosomal RNA (rRNA) gene sequencing, and gene expression using RNA-Seq of host tissue. HLA-B27/human ß2 -microglobulin-transgenic (HLA-B27-transgenic) and wild-type rats from dark agouti, Lewis, and Fischer backgrounds were used. Inter-omic analyses using Cytoscape were employed to identify relevant relationships. PICRUSt was used to predict microbial functions based on known metagenomic profiles. RESULTS: Inter-omic analysis revealed several gut microbes that were strongly associated with dysregulated cytokines driving inflammatory response pathways, such as interleukin-17 (IL-17), IL-23, IL-17, IL-1, interferon-γ (IFNγ), and tumor necrosis factor (TNF). Many microbes were uniquely associated with inflammation in Lewis or Fischer rats, and one was relevant on both backgrounds. Several microbes that were strongly correlated with immune dysregulation were not differentially abundant in HLA-B27-transgenic compared to wild-type controls. A multi-omic network analysis revealed non-overlapping clusters of microbes in Lewis and Fischer rats that were strongly linked to overlapping dysregulated immune/inflammatory genes. Prevotella, Clostridiales, and Blautia were important in Lewis rats, while Akkermansia muciniphila and members of the Lachnospiraceae family dominated in Fischer rats. Inflammation-associated metabolic pathway perturbation (e.g., butanoate, propanoate, lipopolysaccharide, and steroid biosynthesis) was also predicted from both backgrounds. CONCLUSION: Inter-omic and network analysis of gut microbes and the host immune response in experimental SpA provides an unprecedented view of organisms strongly linked to dysregulated IL-23, IL-17, IL-1, IFNγ, and TNF. Functional similarities between these organisms may explain why animals of different genetic backgrounds exhibit common patterns of immune dysregulation, possibly through perturbation of similar metabolic pathways. These results highlight the power of linking analyses of gut microbiota with the host immune response to gain insights into the role of dysbiotic microbes in SpA beyond taxonomic profiling.

3.
mSystems ; 4(4)2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164452

RESUMO

Microbial communities are commonly studied using culture-independent methods, such as 16S rRNA gene sequencing. However, one challenge in accurately characterizing microbial communities is exogenous bacterial DNA contamination, particularly in low-microbial-biomass niches. Computational approaches to identify contaminant sequences have been proposed, but their performance has not been independently evaluated. To identify the impact of decreasing microbial biomass on polymicrobial 16S rRNA gene sequencing experiments, we created a mock microbial community dilution series. We evaluated four computational approaches to identify and remove contaminants, as follows: (i) filtering sequences present in a negative control, (ii) filtering sequences based on relative abundance, (iii) identifying sequences that have an inverse correlation with DNA concentration implemented in Decontam, and (iv) predicting the sequence proportion arising from defined contaminant sources implemented in SourceTracker. As expected, the proportion of contaminant bacterial DNA increased with decreasing starting microbial biomass, with 80.1% of the most diluted sample arising from contaminant sequences. Inclusion of contaminant sequences led to overinflated diversity estimates and distorted microbiome composition. All methods for contaminant identification successfully identified some contaminant sequences, which varied depending on the method parameters used and contaminant prevalence. Notably, removing sequences present in a negative control erroneously removed >20% of expected sequences. SourceTracker successfully removed over 98% of contaminants when the experimental environments were well defined. However, SourceTracker misclassified expected sequences and performed poorly when the experimental environment was unknown, failing to remove >97% of contaminants. In contrast, the Decontam frequency method did not remove expected sequences and successfully removed 70 to 90% of the contaminants.IMPORTANCE The relative scarcity of microbes in low-microbial-biomass environments makes accurate determination of community composition challenging. Identifying and controlling for contaminant bacterial DNA are critical steps in understanding microbial communities from these low-biomass environments. Our study introduces the use of a mock community dilution series as a positive control and evaluates four computational strategies that can identify contaminants in 16S rRNA gene sequencing experiments in order to remove them from downstream analyses. The appropriate computational approach for removing contaminant sequences from an experiment depends on prior knowledge about the microbial environment under investigation and can be evaluated with a dilution series of a mock microbial community.

4.
Arthritis Rheumatol ; 71(10): 1642-1650, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31038287

RESUMO

OBJECTIVE: HLA alleles affect susceptibility to more than 100 diseases, but the mechanisms that account for these genotype-disease associations are largely unknown. HLA alleles strongly influence predisposition to ankylosing spondylitis (AS) and rheumatoid arthritis (RA). Both AS and RA patients have discrete intestinal and fecal microbiome signatures. Whether these changes are the cause or consequence of the diseases themselves is unclear. To distinguish these possibilities, we examined the effect of HLA-B27 and HLA-DRB1 RA risk alleles on the composition of the intestinal microbiome in healthy individuals. METHODS: Five hundred sixty-eight stool and biopsy samples from 6 intestinal sites were collected from 107 healthy unrelated subjects, and stool samples were collected from 696 twin pairs from the TwinsUK cohort. Microbiome profiling was performed using sequencing of the 16S ribosomal RNA bacterial marker gene. All subjects were genotyped using the Illumina CoreExome SNP microarray, and HLA genotypes were imputed from these data. RESULTS: Associations were observed between the overall microbial composition and both the HLA-B27 genotype and the HLA-DRB1 RA risk allele (P = 0.0002 and P = 0.00001, respectively). These associations were replicated using the stool samples from the TwinsUK cohort (P = 0.023 and P = 0.033, respectively). CONCLUSION: This study shows that the changes in intestinal microbiome composition seen in AS and RA are at least partially due to effects of HLA-B27 and HLA-DRB1 on the gut microbiome. These findings support the hypothesis that HLA alleles operate to cause or increase the risk of these diseases through interaction with the intestinal microbiome and suggest that therapies targeting the microbiome may be effective in preventing or treating these diseases.

5.
mSystems ; 4(1)2019.
Artigo em Inglês | MEDLINE | ID: mdl-30801029

RESUMO

Microbiome community composition plays an important role in human health, and while most research to date has focused on high-microbial-biomass communities, low-biomass communities are also important. However, contamination and technical noise make determining the true community signal difficult when biomass levels are low, and the influence of varying biomass on sequence processing methods has received little attention. Here, we benchmarked six methods that infer community composition from 16S rRNA sequence reads, using samples of varying biomass. We included two operational taxonomic unit (OTU) clustering algorithms, one entropy-based method, and three more-recent amplicon sequence variant (ASV) methods. We first compared inference results from high-biomass mock communities to assess baseline performance. We then benchmarked the methods on a dilution series made from a single mock community-samples that varied only in biomass. ASVs/OTUs inferred by each method were classified as representing expected community, technical noise, or contamination. With the high-biomass data, we found that the ASV methods had good sensitivity and precision, whereas the other methods suffered in one area or in both. Inferred contamination was present only in small proportions. With the dilution series, contamination represented an increasing proportion of the data from the inferred communities, regardless of the inference method used. However, correlation between inferred contaminants and sample biomass was strongest for the ASV methods and weakest for the OTU methods. Thus, no inference method on its own can distinguish true community sequences from contaminant sequences, but ASV methods provide the most accurate characterization of community and contaminants. IMPORTANCE Microbial communities have important ramifications for human health, but determining their impact requires accurate characterization. Current technology makes microbiome sequence data more accessible than ever. However, popular software methods for analyzing these data are based on algorithms developed alongside older sequencing technology and smaller data sets and thus may not be adequate for modern, high-throughput data sets. Additionally, samples from environments where microbes are scarce present additional challenges to community characterization relative to high-biomass environments, an issue that is often ignored. We found that a new class of microbiome sequence processing tools, called amplicon sequence variant (ASV) methods, outperformed conventional methods. In samples representing low-biomass communities, where sample contamination becomes a significant confounding factor, the improved accuracy of ASV methods may allow more-robust computational identification of contaminants.

6.
Invest Ophthalmol Vis Sci ; 60(1): 420-429, 2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30695094

RESUMO

Purpose: We determine the changes in intestinal microbiota and/or disruptions in intestinal homeostasis during uveitis. Methods: Experimental autoimmune uveitis (EAU) was induced in B10.RIII mice with coadministration of interphotoreceptor retinoid-binding protein peptide (IRBP) and killed mycobacterial antigen (MTB) as an adjuvant. Using 16S rRNA gene sequencing, we looked at intestinal microbial differences during the course of uveitis, as well as intestinal morphologic changes, changes in intestinal permeability by FITC-dextran leakage, antimicrobial peptide expression in the gastrointstinal tract, and T lymphocyte prevalence before and at peak intraocular inflammation. Results: We demonstrate that increased intestinal permeability and antimicrobial peptide expression in the intestinal tract coincide in timing with increased effector T cells in the mesenteric lymph nodes, during the early stages of uveitis, before peak inflammation. Morphologic changes in the intestine were most prominent during this phase, but also occurred with adjuvant MTB alone, whereas increased intestinal permeability was found only in IRBP-immunized mice that develop uveitis. We also demonstrate that the intestinal microbiota were altered during the course of uveitis, and that some of these changes are specific to uveitic animals, whereas others are influenced by adjuvant MTB alone. Intestinal permeability peaked at 2 weeks, coincident with an increase in intestinal bacterial strain differences, peak lipocalin production, and peak uveitis. Conclusions: An intestinal dysbiosis accompanies a disruption in intestinal homeostasis in autoimmune uveitis, although adjuvant MTB alone promotes intestinal disruption as well. This may indicate a novel axis for future therapeutic targeting experimentally or clinically.


Assuntos
Doenças Autoimunes/microbiologia , Disbiose/microbiologia , Microbioma Gastrointestinal/imunologia , Homeostase/fisiologia , Intestinos/fisiologia , Uveíte/microbiologia , Animais , Antígenos de Bactérias/imunologia , Doenças Autoimunes/imunologia , Ensaio de Imunoadsorção Enzimática , Proteínas do Olho , Citometria de Fluxo , Lipocalinas/metabolismo , Camundongos , Camundongos Mutantes , Modelos Animais , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/imunologia , RNA Ribossômico 16S/genética , Proteínas de Ligação ao Retinol , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Subpopulações de Linfócitos T/imunologia , Uveíte/imunologia , Proteína da Zônula de Oclusão-1/metabolismo
7.
Nat Rev Urol ; 15(12): 735-749, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30315209

RESUMO

Many studies have shown that the urinary tract harbours its own microbial community known as the urinary microbiota, which have been implicated in urinary tract disorders. This observation contradicts the long-held notion that urine is a sterile biofluid in the absence of acute infection of the urinary tract. In light of this new discovery, many basic questions that are crucial for understanding the role of the urinary microbiota in human health and disease remain unanswered. Given that the urinary microbiota is an emerging area of study, optimized techniques and protocols to identify microorganisms in the urinary tract are still being established. However, the low microbial biomass and close proximity to higher microbial biomass environments (for example, the vagina) present distinct methodological challenges for microbial community profiling of the urinary microbiota. A clear understanding of the unique technical considerations for obtaining and analysing low microbial biomass samples, as well the influence of key elements of experimental design and computational analysis on downstream interpretation, will improve our ability to interpret and compare results across methods and studies and is relevant for studies profiling the urinary microbiota and other sites of low microbial abundance.


Assuntos
Microbiota , Sistema Urinário/microbiologia , Coleta de Urina/métodos , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Microbiota/genética , Reação em Cadeia da Polimerase , Bexiga Urinária/microbiologia , Coleta de Urina/normas
8.
Nat Rev Rheumatol ; 14(12): 704-713, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30301938

RESUMO

Acute anterior uveitis (AAU) and the spondyloarthritis (SpA) subtypes ankylosing spondylitis, reactive arthritis and psoriatic arthritis are among the inflammatory diseases affected by the biology of the intestinal microbiome. In this Review, the relationship between AAU, SpA and the microbiome is discussed, with a focus on the major SpA risk gene HLA-B*27 and how it is associated with both intestinal tolerance and the loss of ocular immune privilege that can accompany AAU. We provide four potential mechanisms to account for how dysbiosis, barrier function and immune response contribute to the development of ocular inflammation and the pathogenesis of AAU. Finally, potential therapeutic avenues to target the microbiota for the clinical management of AAU and SpA are outlined.


Assuntos
Microbioma Gastrointestinal/imunologia , Antígeno HLA-B27/metabolismo , Espondiloartropatias/microbiologia , Uveíte Anterior/microbiologia , Animais , Humanos , Imunidade Inata , Espondiloartropatias/imunologia , Uveíte Anterior/imunologia
9.
Front Med (Lausanne) ; 5: 80, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29675414

RESUMO

Spondyloarthritis is a common type of arthritis which affects mostly adults. It consists of idiopathic chronic inflammation of the spine, joints, eyes, skin, gut, and prostate. Inflammation is often asymptomatic, especially in the gut and prostate. The HLA-B*27 allele group, which presents intracellular peptides to CD8+ T cells, is by far the strongest risk factor for spondyloarthritis. The precise mechanisms and antigens remain unknown. In 1959, Catterall and King advanced a novel hypothesis explaining the etiology of spondyloarthritis: an as-yet-unrecognized sexually acquired microbe would be causing all spondyloarthritis types, including acute anterior uveitis. Recent studies suggest an unrecognized sexually acquired fungal infection may be involved in prostate cancer and perhaps multiple sclerosis. This warrants reanalyzing the Catterall-King hypothesis based on the current literature. In the last decade, many links between spondyloarthritis and fungal infections have been found. Antibodies against the fungal cell wall component mannan are elevated in spondyloarthritis. Functional polymorphisms in genes regulating the innate immune response against fungi have been associated with spondyloarthritis (CARD9 and IL23R). Psoriasis and inflammatory bowel disease, two common comorbidities of spondyloarthritis, are both strongly associated with fungi. Evidence reviewed here lends credence to the Catterall-King hypothesis and implicates a common fungal etiology in prostate cancer, benign prostatic hyperplasia, multiple sclerosis, psoriasis, inflammatory bowel disease, and spondyloarthritis. However, the evidence available at this time is insufficient to definitely confirm this hypothesis. Future studies investigating the microbiome in relation to these conditions should screen specimens for fungi in addition to bacteria. Future clinical studies of spondyloarthritis should consider antifungals which are effective in psoriasis and multiple sclerosis, such as dimethyl fumarate and nystatin.

10.
Curr Opin Rheumatol ; 30(4): 303-309, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29538010

RESUMO

PURPOSE OF REVIEW: The intestinal microbiome is thought to play a role in the pathogenesis of inflammatory bowel disease (IBD). There are many shared clinical manifestations between IBD and spondyloarthritis (SpA), of which the most common are peripheral arthritis and uveitis. Clinical overlap along with similar genetics between these diseases suggests a possible shared pathogenetic mechanism, which might center on the intestinal microbiota. In this review, we discuss the available evidence that SpA is a microbiome-driven disease and indicate how SpA-associated uveitis could be tied to gut dysbiosis. We conclude by discussing different treatment paradigms targeting the intestinal microbiome for SpA. RECENT FINDINGS: Recent studies support the growing evidence of the intestinal microbiome as a crucial player in SpA disease pathogenesis. There is emerging evidence that the gut microbiome may play a causative role in uveitis. SUMMARY: The field is beginning to discover a new level of understanding how the intestinal microbiome is involved in SpA. Treatment methods to alter intestinal microbiota to treat SpA-related diseases are still in its infancy.


Assuntos
Transplante de Microbiota Fecal , Espondilartrite/terapia , Uveíte/terapia , Animais , Microbioma Gastrointestinal , Humanos , Doenças Inflamatórias Intestinais/terapia , Espondilartrite/microbiologia , Uveíte/microbiologia
11.
Arthritis Rheumatol ; 70(4): 555-565, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29287307

RESUMO

OBJECTIVE: To investigate whether HLA-B27-mediated experimental spondyloarthritis (SpA) is associated with a common gut microbial signature, in order to identify potential drivers of pathogenesis. METHODS: The effects of HLA-B27 on 3 genetic backgrounds, dark agouti (DA), Lewis, and Fischer, were compared, using wild-type littermates and HLA-B7-transgenic Lewis rats as controls. Cecum and colon tissue specimens or contents were collected from the rats at 2, 3-4, and 6-8 months of age, and histologic analysis was performed to assess inflammation, RNA sequencing was used to determine gene expression differences, and 16S ribosomal RNA gene sequencing was used to determine microbiota differences. RESULTS: Both HLA-B27-transgenic Lewis rats and HLA-B27-transgenic Fischer rats developed gut inflammation, while DA rats were resistant to the effects of HLA-B27, and HLA-B7-transgenic rats were not affected. Immune dysregulation was similar in affected Lewis and Fischer rats and was dominated by activation of interleukin-23 (IL-23)/IL-17, interferon, tumor necrosis factor, and IL-1 cytokines and pathways in the colon and cecum, while DA rats exhibited low-level cytokine dysregulation without inflammation. Gut microbial changes in HLA-B27-transgenic rats were strikingly divergent on the 3 different host genetic backgrounds, including different patterns of dysbiosis in HLA-B27-transgenic Lewis and HLA-B27-transgenic Fischer rat strains, with some overlap. Interestingly, DA rats lacked segmented filamentous bacteria that promote CD4+ Th17 cell development, which may explain their resistance to disease. CONCLUSION: The effects of HLA-B27 on gut microbiota and dysbiosis in SpA are highly dependent on the host genetic background and/or environment, despite convergence of dysregulated immune pathways. These results implicate an ecological model of dysbiosis, with the effects of multiple microbes contributing to the aberrant immune response, rather than a single or small number of microbes driving pathogenesis.


Assuntos
Disbiose/genética , Microbioma Gastrointestinal/genética , Antígeno HLA-B27/metabolismo , Espondiloartropatias/genética , Espondiloartropatias/microbiologia , Proteína Agouti Sinalizadora , Animais , Ceco/metabolismo , Ceco/microbiologia , Colo/metabolismo , Colo/microbiologia , Modelos Animais de Doenças , RNA Ribossômico 16S/metabolismo , Ratos , Ratos Endogâmicos F344 , Ratos Endogâmicos Lew , Ratos Transgênicos
12.
Sci Rep ; 7(1): 11745, 2017 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-28924192

RESUMO

Short chain fatty acids (SCFA) are metabolites of intestinal bacteria resulting from fermentation of dietary fiber. SCFA are protective in various animal models of inflammatory disease. We investigated the effects of exogenous administration of SFCAs, particularly propionate, on uveitis using an inducible model of experimental autoimmune uveitis (EAU). Oral SCFA administration attenuated uveitis severity in a mouse strain-dependent manner through regulatory T cell induction among lymphocytes in the intestinal lamina propria (LPL) and cervical lymph nodes (CLN). SCFA also suppressed effector T cell induction in the CLN and mesenteric lymph nodes (MLN). Alterations in intestinal morphology and gene expression demonstrated in the EAU model prior to the onset of uveitis were blunted by oral SCFA administration. Using a Kaede transgenic mouse, we demonstrated enhanced leukocyte trafficking between the intestine and the eye in EAU. Propionate suppressed T effector cell migration between the intestine and the spleen in EAU Kaede mice. In conclusion, our findings support exogenous administration of SCFAs as a potential treatment strategy for uveitis through the stabilization of subclinical intestinal alterations that occur in inflammatory diseases including uveitis, as well as prevention of trafficking of leukocytes between the gastrointestinal tract and extra-intestinal tissues.


Assuntos
Doenças Autoimunes/tratamento farmacológico , Movimento Celular/efeitos dos fármacos , Ácidos Graxos Voláteis/farmacologia , Intestinos , Linfócitos T Reguladores , Uveíte , Animais , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Movimento Celular/imunologia , Modelos Animais de Doenças , Feminino , Intestinos/imunologia , Intestinos/patologia , Camundongos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Uveíte/tratamento farmacológico , Uveíte/imunologia , Uveíte/patologia
13.
J Neuroimmunol ; 310: 51-59, 2017 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-28778445

RESUMO

Sex hormones promote immunoregulatory effects on multiple sclerosis. In the current study we evaluated the composition of the gut microbiota and the mucosal-associated regulatory cells in estrogen or sham treated female mice before and after autoimmune encephalomyelitis (EAE) induction. Treatment with pregnancy levels of estrogen induces changes in the composition and diversity of gut microbiota. Additionally, estrogen prevents EAE-associated changes in the gut microbiota and might promote the enrichment of bacteria that are associated with immune regulation. Our results point to a possible cross-talk between the sex hormones and the gut microbiota, which could promote neuroprotection.


Assuntos
Encefalomielite Autoimune Experimental/tratamento farmacológico , Estrogênios/uso terapêutico , Intestinos/microbiologia , Microbiota/efeitos dos fármacos , Membrana Mucosa/efeitos dos fármacos , Membrana Mucosa/patologia , Animais , Antígenos CD/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/induzido quimicamente , Encefalomielite Autoimune Experimental/patologia , Fezes/microbiologia , Feminino , Interleucina-10/genética , Interleucina-10/metabolismo , Intestinos/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Linfonodos/efeitos dos fármacos , Linfonodos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Glicoproteína Mielina-Oligodendrócito/toxicidade , Fragmentos de Peptídeos/toxicidade , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/metabolismo , Medula Espinal/patologia , Fatores de Tempo
14.
Arthritis Rheumatol ; 69(10): 1984-1995, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28622455

RESUMO

OBJECTIVE: HLA-B27-associated spondyloarthritides are associated with an altered intestinal microbiota and bowel inflammation. We undertook this study to identify HLA-B27-dependent changes in both host and microbial metabolites in the HLA-B27/ß2 -microglobulin (ß2 m)-transgenic rat and to determine whether microbiota-derived metabolites could impact disease in this major model of spondyloarthritis. METHODS: Cecal contents were collected from Fischer 344 33-3 HLA-B27/ß2 m-transgenic rats and wild-type controls at 6 weeks (before disease) and 16 weeks (with active bowel inflammation). Metabolomic profiling was performed by high-throughput gas and liquid chromatography-based mass spectrometry. HLA-B27/ß2 m-transgenic rats were treated with the microbial metabolites propionate or butyrate in drinking water for 10 weeks, and disease activity was subsequently assessed. RESULTS: Our screen identified 582 metabolites, of which more than half were significantly altered by HLA-B27 expression at 16 weeks. Both microbial and host metabolites were altered, with multiple pathways affected, including those for amino acid, carbohydrate, xenobiotic, and medium-chain fatty acid metabolism. Differences were even observed at 6 weeks, with up-regulation of histidine, tyrosine, spermidine, N-acetylmuramate, and glycerate in HLA-B27/ß2 m-transgenic rats. Administration of the short-chain fatty acid propionate significantly attenuated HLA-B27-associated inflammatory disease, although this was not associated with increased FoxP3+ T cell induction or with altered expression of the immunomodulatory cytokines interleukin-10 (IL-10) or IL-33 or of the tight junction protein zonula occludens 1. HLA-B27 expression was also associated with altered host expression of messenger RNA for the microbial metabolite receptors free fatty acid receptor 2 (FFAR2), FFAR3, and niacin receptor 1. CONCLUSION: HLA-B27 expression profoundly impacts the intestinal metabolome, with changes evident in rats even at age 6 weeks. Critically, we demonstrate that a microbial metabolite, propionate, attenuates development of HLA-B27-associated inflammatory disease. These and other microbiota-derived bioactive mediators may provide novel treatment modalities in HLA-B27-associated spondyloarthritides.


Assuntos
Ceco/metabolismo , Microbioma Gastrointestinal , Antígeno HLA-B27/genética , Espondiloartropatias/metabolismo , Animais , Ácido Butírico/farmacologia , Ceco/microbiologia , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Ácidos Graxos Voláteis/metabolismo , Citometria de Fluxo , Cromatografia Gasosa-Espectrometria de Massas , Perfilação da Expressão Gênica , Ácidos Glicéricos/metabolismo , Histidina/metabolismo , Interleucina-10/imunologia , Interleucina-33/imunologia , Linfonodos/citologia , Espectrometria de Massas , Mesentério , Metabolômica , Ácidos Murâmicos/metabolismo , Propionatos/farmacologia , Ratos , Ratos Endogâmicos F344 , Ratos Transgênicos , Espermidina/metabolismo , Baço/citologia , Espondiloartropatias/genética , Espondiloartropatias/imunologia , Linfócitos T/imunologia , Tirosina/metabolismo , Regulação para Cima , Microglobulina beta-2/genética
15.
Curr Rheumatol Rep ; 18(10): 62, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27641915

RESUMO

PURPOSE OF REVIEW: The microbiome is the term that describes the microbial ecosystem that cohabits an organism such as humans. The microbiome has been implicated in a long list of immune-mediated diseases which include rheumatoid arthritis, ankylosing spondylitis, and even gout. The mechanisms to account for this effect are multiple. The clinical implications from observations on the microbiome and disease are broad. RECENT FINDINGS: A growing number of microbiota constituents such as Prevotella copri, Porphyromonas gingivalis, and Collinsella have been correlated or causally related to rheumatic disease. The microbiome has a marked effect on the immune system. Our understanding of immune pathways modulated by the microbiota such as the induction of T helper 17 (Th17) cells and secretory immunoglobulin A (IgA) responses to segmented filamentous bacteria continues to expand. In addition to the gut microbiome, bacterial communities of other sites such as the mouth, lung, and skin have also been associated with the pathogenesis of rheumatic diseases. Strategies to alter the microbiome or to alter the immune activation from the microbiome might play a role in the future therapy for rheumatic diseases.


Assuntos
Microbiota/imunologia , Doenças Reumáticas/terapia , Humanos , Doenças Reumáticas/imunologia , Doenças Reumáticas/microbiologia
16.
Artigo em Inglês | MEDLINE | ID: mdl-27512653

RESUMO

OBJECTIVES: Traditionally, the urinary tract has been thought to be sterile in the absence of a clinically identifiable infection. However, recent evidence suggests that the urinary tract harbors a variety of bacterial species, known collectively as the urinary microbiome, even when clinical cultures are negative. Whether these bacteria promote urinary health or contribute to urinary tract disease remains unknown. Emerging evidence indicates that a shift in the urinary microbiome may play an important role in urgency urinary incontinence (UUI). The goal of this prospective pilot study was to determine how the urinary microbiome is different between women with and without UUI. We also sought to identify if characteristics of the urinary microbiome are associated with UUI severity. METHODS: We collected urine from clinically well-characterized women with UUI (n = 10) and normal bladder function (n = 10) using a transurethral catheter to avoid bacterial contamination from external tissue. To characterize the resident microbial community, we amplified the bacterial 16S rRNA gene by PCR and performed sequencing using Illumina MiSeq. Sequences were processed using the workflow package QIIME. We identified bacteria that had differential relative abundance between UUI and controls using DESeq2 to fit generalized linear models based on the negative binomial distribution. We also identified relationships between the diversity of the urinary microbiome and severity of UUI symptoms with Pearson's correlation coefficient. RESULTS: We successfully extracted and sequenced bacterial DNA from 95% of the urine samples and identified that there is a polymicrobial community in the female bladder in both healthy controls and women with UUI. We found the relative abundance of 14 bacteria significantly differed between control and UUI samples. Furthermore, we established that an increase in UUI symptom severity is associated with a decrease in microbial diversity in women with UUI. CONCLUSIONS: Our study provides further characterization of the urinary microbiome in both healthy controls and extensively phenotyped women with UUI. Our results also suggest that the urinary microbiome may play an important role in the pathophysiology of UUI and that the loss of microbial diversity may be associated with clinical severity.


Assuntos
Bactérias/isolamento & purificação , Microbiota/fisiologia , Incontinência Urinária/microbiologia , Adulto , Idoso , Bactérias/classificação , Bactérias/genética , Sequência de Bases , Biodiversidade , Estudos de Casos e Controles , DNA Bacteriano/genética , Feminino , Humanos , Microbiota/genética , Pessoa de Meia-Idade , Filogenia , Projetos Piloto , Estudos Prospectivos , Índice de Gravidade de Doença , Doenças da Bexiga Urinária/microbiologia , Doenças da Bexiga Urinária/urina , Incontinência Urinária/fisiopatologia , Incontinência Urinária/urina , Sistema Urinário/microbiologia
17.
Invest Ophthalmol Vis Sci ; 57(8): 3747-58, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-27415793

RESUMO

PURPOSE: To investigate the contribution of the gut microbiota to the pathogenesis of uveitis. METHODS: Experimental autoimmune uveitis (EAU) in B10.RIII mice was induced using interphotoreceptor binding protein peptide. Mice were treated with oral or intraperitoneal (IP) antibiotics. Effector (Teff) and regulatory (Treg) T lymphocytes were identified using flow cytometry; 16S rRNA gene sequencing and qPCR were performed on gastrointestinal (GI) contents. RESULTS: Broad-spectrum (four antibiotics given simultaneously) oral, but not IP, antibiotics reduced mean uveitis clinical scores significantly compared with water-treated animals (0.5 vs. 3.0, P < 0.0001 for oral; 3.4 vs. 3.4, P > 0.99 for IP). Both oral metronidazole (P = 0.02) and vancomycin (P < 0.0001) alone decreased inflammation, whereas neomycin (P = 0.7) and ampicillin (P = 0.4) did not change mean uveitis scores. Oral broad-spectrum antibiotics increased Tregs in the GI lamina propria of EAU animals at 1 week, and in extraintestinal lymphoid tissues later, whereas Teff and inflammatory cytokines were reduced. 16S sequencing of GI contents revealed altered microbiota in immunized mice compared with nonimmunized mice, and microbial diversity clustering in EAU mice treated with uveitis-protective antibiotics. Experimental autoimmune uveitis mice also demonstrated gut microbial diversity clustering associated with clinical score severity. CONCLUSIONS: Oral antibiotics modulate the severity of inducible EAU by increasing Tregs in the gut and extraintestinal tissues, as well as decreasing effector T cells and cytokines. 16S sequencing suggests that there may be protective and, conversely, potentially uveitogenic, gut microbiota. These findings may lead to a better understanding of how uveitis can be treated or prevented by modulating the gut microbiome.


Assuntos
Doenças Autoimunes/microbiologia , Microbioma Gastrointestinal/imunologia , Uveíte/microbiologia , Administração Oral , Animais , Antibacterianos/administração & dosagem , Antibacterianos/farmacologia , Doenças Autoimunes/prevenção & controle , Biomarcadores/metabolismo , Células Cultivadas , Citocinas/metabolismo , Combinação de Medicamentos , Proteínas do Olho/toxicidade , Feminino , Microbioma Gastrointestinal/efeitos dos fármacos , Injeções Intraperitoneais , Camundongos Endogâmicos , Retina/imunologia , Retina/microbiologia , Proteínas de Ligação ao Retinol/toxicidade , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Uveíte/prevenção & controle
18.
Curr Opin Rheumatol ; 28(4): 405-12, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27152700

RESUMO

PURPOSE OF REVIEW: The intestinal microbiome is increasingly implicated in the pathogenesis of ankylosing spondylitis, reactive arthritis, and other diseases collectively known as the spondyloarthropathies (SpAs). In common with other complex inflammatory diseases, SpAs have both a strong genetic and environmental component. Recent genetic studies have highlighted host pathways that may intersect the host-microbiota interaction and offer novel paradigms to understand the pathophysiology of these diseases. RECENT FINDINGS: Genetic association studies have identified genes such as RUNX3, PTPEN2, and IL-33 as susceptibility loci for SpAs. Functional studies in humans have extended knowledge of established genetic risk factors for ankylosing spondylitis that include ERAP1, ERAP2, and interleukin-23R. Recent basic research has identified new mechanisms that regulate host immune responses to the microbiota that conceivably may be dysregulated in SpA. SUMMARY: Intestinal barrier function, deletional tolerance, Th17 signature response, and endoplasmic reticulum stress pathways have been recently linked to SpA. Dysregulated immune responses to the gut microbiota and an altered microbial community structure are shared features of SpA. Although the cause-effect dynamic of this relationship remains equivocal, it nonetheless has major implications for both intestinal and extra-intestinal pathology observed in SpA.


Assuntos
Microbioma Gastrointestinal , Espondiloartropatias/genética , Espondiloartropatias/microbiologia , Aminopeptidases/genética , Microbioma Gastrointestinal/imunologia , Predisposição Genética para Doença , Antígeno HLA-B27/genética , Humanos , Antígenos de Histocompatibilidade Menor/genética , Espondiloartropatias/imunologia , Espondilite Anquilosante/genética , Espondilite Anquilosante/imunologia , Espondilite Anquilosante/microbiologia
19.
Ocul Immunol Inflamm ; 24(4): 440-4, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27002532

RESUMO

The microbiome is strongly implicated in a broad spectrum of immune-mediated diseases. Data support the concept that HLA molecules shape the microbiome. We provide hypotheses to reconcile how HLA-B27 might affect the microbiome and in turn predispose to acute anterior uveitis. These theories include bacterial translocation, antigenic mimicry, and dysbiosis leading to alterations in regulatory and effector T-cell subsets. Received 31 October 2015; revised 7 January 2016; accepted 8 January 2016; published online 22 March 2016.


Assuntos
Microbioma Gastrointestinal/fisiologia , Antígeno HLA-B27/sangue , Uveíte Anterior/microbiologia , Doença Aguda , Animais , Humanos , Uveíte Anterior/imunologia
20.
Arthritis Rheumatol ; 68(9): 2151-62, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-26992013

RESUMO

OBJECTIVE: The HLA-B27/ß2 -microglobulin (ß2 m)-transgenic (Tg) rat is a leading model of B27-associated spondyloarthritis (SpA), and the disease is dependent on the presence of intestinal bacteria. Previous studies have shown that adult HLA-B27/ß2 m-Tg rats have an altered intestinal microbiota. This study sought to better define the age-dependent changes to both mucosal immune function and dysbiosis in this rat model of SpA. METHODS: Intestinal contents were collected from wild-type and HLA-B27/ß2 m-Tg rats postweaning (ages 3 and 6 weeks), at disease onset (age 10 weeks), and after the establishment of disease (ages ≥16 weeks). The microbial community structure was determined by 16S ribosomal RNA sequencing and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Mucosal and systemic Th1, Th17, and Treg cell responses were analyzed by flow cytometry, as was the frequency of IgA-coated intestinal bacteria. Intestinal expression of inflammatory cytokines and antimicrobial peptides (AMPs) was determined by qRT-PCR. RESULTS: An inflammatory cytokine signature and elevated AMP expression during the postweaning period preceded the development of clinical bowel inflammation and dysbiosis in HLA-B27/ß2 m-Tg rats. An early and sustained expansion of the Th17 cell pool was specifically observed in the cecal and colonic mucosa of HLA-B27/ß2 m-Tg rats. Strongly elevated intestinal colonization of Akkermansia muciniphila and an increased frequency of IgA-coated fecal bacteria were significantly associated with expression of HLA-B27 and arthritis development. CONCLUSION: HLA-B27/ß2 m expression in this rat model renders the host hyperresponsive to microbial antigens from infancy. Early activation of innate immunity and expansion of a mucosal Th17 signature are soon followed by dysbiosis in HLA-B27/ß2 m-Tg animals. The pathologic processes of perturbed mucosal immunity and dysbiosis strongly merit further study in both prediseased and diseased populations of patients with SpA.


Assuntos
Disbiose/imunologia , Imunidade nas Mucosas , Espondilartrite/complicações , Espondilartrite/imunologia , Animais , Modelos Animais de Doenças , Antígeno HLA-B27/imunologia , Ratos , Ratos Transgênicos
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