PHENOTYPE AND FUNCTION OF STROMAL CELLS CLONED FROM THE ILEAL PEYER PATCH OF SHEEP

The ileal Peyer's patch (PP) is the major site of B cell production and immunoglobulin diversification in lambs, but the factors which regulate these processes are poorly understood. As a first step toward identifying possible regulatory mechanisms, stable long-term cultures of ileal PP stromal cells were established at the clonal level. Four distinct cell types were identified by their phenotype and growth requirements. Immunohistochemical staining confirmed that all clones were mesenchymal (vimentin+; cytokeratin-) in origin and were negative for T cell, B cell, and macrophage markers. Three cell lines were negative for major histocompatibility complex (MHC) I and II molecules, but one cell line, SCN, expressed MHC I, MHC II and CD44 molecules, and a subpopulation of SCN cells expressed BAQ44A, a B cell differentiation molecule. The four cell lines produced different types and amounts of extracellular matrix proteins, and their growth was not influenced by exogenous human interleukin 1 (IL-1), IL-2, transforming growth factor beta1 (TGFbeta1), or bovine fibroblast growth factor (FGF) but was influenced by serum. When tested for their capacity to support lymphocyte growth, all clones produced a soluble factor(s) that was mitogenic for ileal and jejunal PP cells and thymocytes. Similar growth promoting activity was observed with culture supernatants of murine, human and bovine fibroblasts but could not be reproduced using recombinant human cytokines. Furthermore, coculture of stromal cells with ileal PP follicular B cells elicited a proliferative response unique to each stromal cell line. Coculture with increasing numbers of SCN cells inhibited B cell proliferative responses, whereas coculture with SCG2 and SCF32 cells enhanced B cell proliferative response at both low and high stromal cell densities. Ileal PP follicular B cells rapidly bound to the surface of all stromal cell clones, and this interaction was specific when compared with thymocytes or peripheral blood lymphocytes. These results suggest that ileal PP stromal cells are a phenotypically and functionally heterogeneous population that may enhance or inhibit B lymphopoiesis in the ileal PP.