行政院國家科學委員會專題研究計畫成果報告:犬傳染性花柳性腫瘤細胞所分泌之毒殺樹枝狀細胞物質之分離及特性分析

For therapeutic purposes, large numbers of dendritic cells (DC) are essential. However, the isolation efficiency of canine DC was low. In this study, we established a modified adherence step to reliably enrich CD14+ monocytes from peripheral blood mononuclear cells (PBMC). Canine DC were generated from the adherent monocytes. Adherent monocytes were cultured for 6 days in human GM-CSF, Flt3L, and canine IL-4. This improved procedure yielded 2–4 times more canine DC than other published methods. Canine DC characteristics, including surface phenotype and biological functions during maturation, were also investigated in this study. In canine immature DC (iDC), DLA class II, CD1a, CD11c, CD40, and CD86 expression was high, based on flow cytometry and RT-PCR assays. During maturation, which was stimulated by LPS, CD80 and CD83 were up-regulated in mDC. However, DLA class II, CD1a, CD11c, and CD40 did not increase in mature DC (mDC). Functional maturation was assessed by antigen uptake ability and the allogenic MLR. Our data showed that incubating canine iDC with LPS decreased antigen uptake and increased the immunostimulatory capacity, indicating that LPS accelerates DC maturation. The TNF-a gene expression was decreased during DC differentiation from monocytes confirm the functional difference between monocytes and DC. This new protocol markedly increased the cell number with in vitro culture of DC precursors and thus may facilitate the use of DC in clinical cellular immunotherapy