CD4CD8αα Lymphocytes, A Novel Human Regulatory T Cell Subset Induced by Colonic Bacteria and Deficient in Patients with Inflammatory Bowel Disease

It has become evident that bacteria in our gut affect health and disease, but less is known about how they do this. Recent studies in mice showed that gut Clostridium bacteria and their metabolites can activate regulatory T cells (Treg) that in turn mediate tolerance to signals that would ordinarily cause inflammation. In this study we identify a subset of human T lymphocytes, designated CD4CD8αα T cells that are present in the surface lining of the colon and in the blood. We demonstrate Treg activity and show these cells to be activated by microbiota; we identify F. prausnitzii, a core Clostridium strain of the human gut microbiota, as a major inducer of these Treg cells. Interestingly, there are fewer F. prausnitzii in individuals suffering from inflammatory bowel disease (IBD), and accordingly the CD4CD8αα T cells are decreased in the blood and gut of patients with IBD. We argue that CD4CD8αα colonic Treg probably help control or prevent IBD. These data open the road to new diagnostic and therapeutic strategies for the management of IBD and provide new tools to address the impact of the intestinal microbiota on the human immune system

DP8α are frequently found in the <i>lamina propria</i> of healthy colon mucosa.

<p>Freshly dissociated CD3 LPL and cell lines were analyzed by flow cytometry for the co-expression of CD4 and either CD8α or CD8β. (<b>a</b>) Representative dot plots and frequencies of CD4 T cells co-expressing CD8α or CD8β among the CD3 LPL from 12 donors; ***<i>p</i><0.001 (paired <i>t</i>-test). (<b>b</b>) Frequencies of CD4CD8αα cells among the CD3 or CD3CD4 LPL (<i>n</i> = 18). (<b>c</b>) Stable co-expression of CD4 and CD8α but not CD8β, by a cell line (representative of four) obtained from FACS-sorted DP8α colonic LPL, after several transfers in culture.</p

Specificity of DP8α-LPL lines and-PBL lines for <i>F. prausnitzii</i>.

<p>DP8α LPL lines (<i>n</i> = 3, C114, C139, C140) and one PBL line (DTC4) were stimulated for 6 h by monocytes, loaded overnight by different Clostridium species (1∶5).</p>a<p>Percent IL-10 or IFN-γ positive cells.</p

Presence of F-reactive DP8α T lymphocytes in the blood of healthy individuals.

<p>(a) Flow cytometry analysis of the frequencies of DP8α T lymphocytes in the blood of 18 donors: representative dot plot of PBMC co-labelled with anti-CD3, anti-CD4, and anti-CD8α antibodies, representative histogram of CD8β expression by gated CD8, CD4, and DP8α PBMC, and the percentages of DP8α lymphocytes among the CD3 and the CD3CD4 PBMC. (b) Proliferative responses of DP8α peripheral blood T cells to F in the presence or absence of an anti-MHC class-II antibody or an irrelevant antibody, and to B, L, and E. PBMC were cultured for 5 d with the antibody and/or indicated bacteria at a bacterium∶PBMC ratio 1∶1: representative FlowJo analysis of the VPD dilution and percentage of F-specific divided DP8α T cells in the PBMC from several donors (<i>n</i> = 18); ***<i>p</i><0.001 and **<i>p</i><0.01 (paired <i>t</i>-test). (c) Percent VPD dilution in paired DP8α and CD4 T cells among PBMC co-cultured with F for 5 d; ***<i>p</i><0.001 (paired <i>t</i>-test). (d) CTLA4 and LAG3 expression in proliferative DP8α and CD4 PBL after 5 d of co-culture with allogeneic monocytes loaded overnight with F (<i>n</i> = 2). (e) Flow cytometry analysis of the proliferative response of freshly isolated DP8α or CD4 PBL after 5 d of co-culture with autologous monocytes alone or loaded overnight with F or E: representative cytometry data of a freshly sorted DP8α PBL population and mean percentage of F-specific divided cells (calculated on FlowJo software) (experiments performed with five freshly isolated DP8α PBL (black circles) and two CD4 PBL (white circles).</p