Increased Th17/Treg Ratio In Liver Chronic Graft-Versus-Host-Disease
Authors: Malard Florent , Bossard Céline , Brissot Eolia , Chevallier Patrice , Guillaume Thierry , Delaunay Jacques , Mosnier jean François , Moreau Philippe , Gregoire Marc , Gaugler Béatrice , Mohty Mohamad 
Published: October 2019

Abstract

Background
Chronic GVHD is the main cause of late non-relapse mortality and mortality after allogeneic stem cell transplantation (allo-SCT). In contrast to acute GVHD, pathophysiology of chronic GVHD remains poorly understood. Chronic GVHD presents clinical features that mimic autoimmune diseases. The identification of proinflammatory Th17 cells which contribute to autoimmune diseases pathophysiology, raised the issue of the role of Th17 cells in human chronic GVHD. Indeed, the contribution of Th17 cells in chronic GVHD was assessed in GVHD mouse models. Furthermore, theses studies indicates that the expansion of Th1 and Th17 cells is favored by the progressive loss CD4+CD25+Foxp3+ regulatory T cells (Treg) leading to chronic GVHD. This report investigated the role of Th17 cells and Treg in liver biopsies taken from patients with or without chronic GVHD.

Patients and Methods
Studies described in this report were performed in a single centre series of 17 patients who underwent allo-SCT for different hematological malignancies (cGVHD group). The median age of patients was 54 years (range, 27-71). The stem cell source was PBSCs in 9 cases (53%) and BM in 8 cases (47%). 13 patients received transplant from a matched-related donor, and 4 patients from an unrelated donor. A reduced-intensity conditioning regimen was used 8 patients (47%) and a myeloablative conditioning in 9 patients (53%). The control group included 8 patients without hematological malignancies who underwent liver biopsy for sleeve gastrectomy for morbid obesity (n=2) or adjacent tumor surgery (n=6). Immunohistochemistry was performed on deparaffinized tissues sections using an indirect immunoperoxydase method. A quantitative evaluation of antigens expression was performed by counting the number of positive cells in the whole biopsy at 200 magnifications for each sample.

Results
In the cGVHD group, based on standard pathology criteria, all 17 patients had a histologically proven liver chronic GVHD. Biopsies were taken at time of first hepatic symptoms declaration or during their reappearance and prior to corticosteroid treatment initiation or resumption. In the control group, all patients present histologically normal liver biopsy. In order to identify the Th17 cell population, biopsies were tested for expression of the CD161 and CCR6 markers, and ROR-gamma-t, the key transcription factor that orchestrates the differentiation of Th17 cells. Significantly higher numbers of ROR-gamma-t+, CD161+ and CCR6+ cells were counted in the liver of patients with chronic GVHD compared with liver of patients in the control group, mainly found in the portal space (p=0.0001, p=0.03 and p=0.03 for CD161, CCR6 and ROR-gamma-t, expression respectively). We also assessed the presence of T cells expressing Tbet (the transcription factor characterizing Th1 cells) and Foxp3 (the master regulator gene of Treg cells) in the liver biopsies. There was no difference in the number of Th1 and Treg cells between the 2 groups (p=0.88 and p=0.12 respectively). Finally we look at the Th17/Th1 and Th17/Treg ratios, considering Th17 cells as ROR- gamma-t positive cells as ROR- gamma-t is the more specific hallmark of Th17 cells. Both Th17/Th1 and Th17/Treg ratios were significantly increased in the liver of patients with liver cGVHD (p=0.005 and p=0.002 respectively).

Conclusion
The current study shed some light on the role of Th17 cells in the context of liver chronic GVHD. Using well-established specific markers, we show that Th17 cells infiltrate liver biopsies from patients with acute GVHD. In addition, Th17/Treg ratio was significantly increased in the liver of patients with liver chronic GVHD, suggesting a regulatory defect in liver chronic GVHD. These data raise the prospect of future innovative approaches to optimize immunosuppression regimens for the treatment or prophylaxis of chronic GVHD by targeting the Th17 response.

Disclosures:
Moreau: CELGENE: Honoraria, Speakers Bureau; JANSSEN: Honoraria, Speakers Bureau.