AUTHOR=Wang Yulian , Huang Lisi , Huang Tian , Geng Suxia , Chen Xiaomei , Huang Xin , Lai Peilong , Du Xin , Weng Jianyu 

TITLE=The Gut Bacteria Dysbiosis Contributes to Chronic Graft-Versus-Host Disease Associated With a Treg/Th1 Ratio Imbalance

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.813576

DOI=10.3389/fmicb.2022.813576

ISSN=1664-302X

ABSTRACT=<sec><title>Introduction</title><p>Dysbiosis of gut bacteria has been discovered in a large number of autoimmune diseases. However, the influence of the gut bacteria in the mice model of chronic sclerodermatous graft-versus-host disease (Scl-GVHD), a disease that resembles an autoimmune disease characterized by chronic inflammation of multiple organs, such as skin, remains elusive. Here, we explore the role of gut bacteria in an Scl-cGVHD mice model.</p></sec><sec><title>Methods</title><p>We established a mouse model of Scl-cGVHD, collected fecal flora, analyzed the composition, and diversity of intestinal flora using 16S rDNA amplicon sequencing, and detected the proportion of Treg and Th1 cells in splenocytes of Scl-cGVHD mice. To verify the immunoregulatory effect of Scl-cGVHD intestinal flora, we prepared bacterial extracts, co-cultured with splenocytes <italic>in vitro</italic>, and used flow cytometry to detect T cell differentiation and cytokine secretion.</p></sec><sec><title>Results</title><p>By examining T-cell differentiation in splenocytes of cGVHD mice, we found that Treg cells were significantly reduced (15.27 ± 0.23 vs. 12.23 ± 0.47, <italic>p</italic> = 0.0045) and Th1 cells were increased (1.54 ± 0.18 vs. 6.68 ± 0.80, <italic>p</italic> = 0.0034) in cGVHD mice. Significant differences were observed in the composition and diversity of the gut bacteria in mice with Scl-cGVHD versus without GVHD. Analysis of mice fecal bacteria samples (<italic>n</italic> = 10, 5 Scl-cGVHD and 5 Non-GVHD) showed significant separation [<italic>R</italic> = 0.732, <italic>p</italic> = 0.015, non-parametric analysis (ANOSIM)] in Scl-cGVHD and non-GVHD mice. The abundance of the family and genus <italic>Ruminococcaceae</italic> bacteria decreased and the family <italic>Lachnospiraceae</italic> and limited to the species <italic>Lachnospiraceae_bacterium_DW17</italic> increased in Scl-cGVHD mice. <italic>In vitro</italic> results of the cellular level study suggest that the bacteria extracts of gut microbiota from Scl-cGVHD mice modulated the splenic T cells toward differentiation into CD4<sup>+</sup>IFN-γ<sup>+</sup> Th1 cells (14.37 ± 0.32 vs. 10.40 ± 2.19, <italic>p</italic> = 0.036), and the percentage of CD4<sup>+</sup>CD25<sup>+</sup>Foxp3<sup>+</sup> Tregs decreased (6.36 ± 0.39 vs. 8.66 ± 0.07, <italic>p</italic> = 0.001) compared with the non-GVHD mice. In addition, the secretion of proinflammatory interferon- γ (IFN-γ) cytokine in the supplement of cellular culture was increased (4,898.58 ± 235.82 vs. 4,347.87 ± 220.02 pg/ml, <italic>p</italic> = 0.042) in the mice model of the Scl-cGVHD group, but anti-inflammatory interleukin (IL)-10 decreased (7,636.57 ± 608.05 vs. 9,563.56 ± 603.34 pg/ml, <italic>p</italic> = 0.018).</p></sec><sec><title>Conclusion</title><p>Our data showed the different composition and diversity of gut bacteria in the Scl-cGVHD mice. The dysbiosis of gut bacteria may regulate the differentiation ratio of Treg and Th1 cells, which was associated with Scl-cGVHD.</p></sec>