IL-17 Positive Lymphocytes Predominantly Home At the Mucous Than in the Skin in cGvHD Patients

Abstract 3053

Chronic graft-versus-host-disease (cGvHD) is a distinct clinical and pathological entity as a late complication of hematopoietic stem cell transplantation (HSCT). In this study 23 biopsies taken from 12 patients with cGvHD who received PBPC from sibling (7 patients) or from matched unrelated donors (5 patients) were evaluated for immunpathomorphology including IL-17 producing cells, regulatory T-cells FOXP3 and CCR6 expression. Conditioning regimen (myeloablative - 5 pts or reduced intensity conditioning – 7 pts) included ATG - 9 pts and Campath – 1 patient. All patients were on cyclosporine (CsA) given to reach trough blood level of 200 ng/L as aGvHD prophylaxis. In 6 pts cGvHD followed acute manifestation but in 6 it was de novo cGvHD.

Biopsies were taken from the sites with evident cGvHD lesions of the skin (n=14), oral (n=3) or gastrointestinal mucosa (n=6). In addition to H+E staining the tissue infiltrations were analyzed for the presence of CD3, CD4, CD8, HLA-DR positive cells and for the sake of this study for the presence of IL-17 producing cells, FOXP3 and CCR6 expression. Immunoenzymatic double immunostaining (CD45 and IL-17 or cytokeratin and IL-17) were performed to describe more precisely lL-17 producing cells. Co-expression of CCR6 and IL-17 and that of IL-17 and CD138 epitopes were also verified with the use of double immunostaining.

Clinical manifestations were moderate and severe among patients suffering from oral mucosa lesions (10 out of 12 pts) and resulted in impairment of feeding and painful ulcerations (5 out of 10 pts).Gastric and intestinal mucosa lesions resulted in vomitus and diarrhea in 3 cases, in the skin poikiloderma and morphea restricted the function in 8 cases but without signs of an overt inflammation.

Histopathologic analysis, according to the NIH criteria, revealed that in our group all specimens met the criteria of cGvHD. In all oral mucous biopsies and in 8 out of 14 skin biopsies lymphocyte infiltration was rather heavy and the picture resembled lichen-planus eruptions. Notably, in the skin as well as in the mucosa lymphoplasmocytic-like cells were present within cellular infiltrates. In the skin and in the mucous CD8+ cells prevailed over CD4+cells. HLA-DR positive cells were only occasionally present. Strong HLA-DR expression was on endothelial cells of the vessel which were especially numerous in the upper area of the oral mucosa.

IL-17 producing cells were identified in 13 out of 14 skin biopsies and were localized predominantly around vessels and in the vicinity of eccrine coils. In oral mucosa IL-17 positive cells were scattered within the dense lymphocyte infiltration and in the gut they were rater in the interstitial tissue but not in the crypts. Double staining showed that IL-17+ cells were not seen in CD45 negative and in the cytokeratin positive populations.

FOXP3 expression differentiated tissue lesions of cGvHD by being present in the oral mucosa, scarce in the skin, but rather absent in the gut mucosa biopsies. The FOXP3 positive cells were seen mostly in cases with severe lymphocyte infiltrations.

CCR6 expression was especially strong expressed on the skin epithelium and not so evident in the mucous. Majority of the skin biopsies revealed CCR6 co-expression on IL-17 producing cells (13 out of 14 biopsies).

CD138+ cells were more abundant in the gut mucous than in the oral mucous or in the skin. CD138+ cells were frequently positive for IL-17 in the gut but not to the similar extent in the oral mucous or in the skin.

In conclusion, IL-17 producing cells were frequent in the tissues with an apparent local inflammation being frequent in the oral mucous (painful ulcerations) and in the intestine (gastritis, colitis). Therefore, due to their strong pro-inflammatory potential they contribute to the clinical picture of cGvHD affecting the organs known to be associated with Th17 guided immune-response.

Supported by the grant N N402 430039 from the Polish Ministry of Science & Higher Education