口腔鱗狀上皮細胞癌細胞株誘導人類調節性T細胞與Th17細胞分化與增生

Abstract

口腔鱗狀上皮細胞癌 (Oral squamous cell carcinoma)是全世界普遍的癌症，是經手術治療後還是伴隨低存活率的癌症。實驗室先前的研究發現調節性T細胞與Th17細胞在口腔麟狀上皮細胞癌病患的組織切片以及腫瘤浸潤細胞中都有高量比例之分佈，並且也發現一群特殊的細胞群同時表現IL-17+Foxp3+，但調節性T細胞、Th17細胞及IL-17+Foxp3 +細胞在腫瘤環境中所扮演的角色目前尚未明瞭。近期的研究指出，在體外培養下調節性T細胞有能力轉變為具有分泌IL-17能力之細胞，但在腫瘤環境中兩者之間的分化轉變關係尚未釐清。本研究利用CD4 T細胞與口腔麟狀上皮細胞癌細胞株共同培養系統，發現CD4 T細胞在腫瘤細胞株SAS的環境中，表現Foxp3 +、IL-17+、IL-17+Foxp3+的細胞比例有增加的現象，並且表現細胞激素IL-17，此外也表現對於Th17分化所需之細胞激素IL-6及IL-1β。實驗室先前的研究發現在腫瘤浸潤細胞中表現IL-17+Foxp3 +之細胞與之IL-17+細胞的分布具有非常高度的相關性。本研究發現在中和Th17分化所需之細胞激素IL-6及IL-1β後，CD4 T細胞與SAS共同培養系統中表現IL-17+Foxp3 +細胞之比例下降，顯示IL-17+Foxp3 +分化可能由Th17細胞轉變而來。文獻指出CCR6的表現與Th17細胞有關，本實驗進一步利用CD25及CCR6作為標記將調節性T細胞分為CD25highCCR6+、CD25high CCR6-以及CD25- CCR6+ CD25-CCR6-，四個細胞亞群分別與SAS共同培養後，CD25highCCR6+以及CD25- CCR6 +在共同培養後表現IL-17+Foxp3 +之細胞比例增加，顯示IL-17+Foxp3 +除了是由原本存在之IL-17+Foxp3 +增生而來也可由Th17分化而來。此外，由CD25- CCR6 +及CCR6 +CD25-與SAS細胞株共同培養的實驗發現CD25-在與SAS共同培養後能分化為Foxp3 +細胞。本研究由腫瘤細胞共同培養系統發現SAS細胞在Th17細胞及調節性T細胞之分化與增生扮演重要的角色。

Oral squamous-cell carcinoma (OSCC) is common worldwide with low survival rate after therapy. Previously, we found elevated Treg, Th17 and a subset of IL-17+ Foxp3+ tumor infiltrating lymphocytes (TILs) in OSCC by immunohistochemistry and flow cytometry. However, the origin of these subsets was unclear. In this study, a mixed CD4 T cells-tumor cells coculture system was established to delineate the induction and/or expansion of the IL-17+ Foxp3+ T cells. The percentage of either Foxp3+ or IL-17+Foxp3+ T cells was increased after co-cultured in the presence of an OSCC cell line. Neutralizing antibodies specific for IL-1β or IL-6 partially inhibited the induction/expansion of IL-17+Foxp3+, but direct cell-cell contact played a more important role as demonstrated by a transwell analysis. To further delineate the origin of the IL-17+Foxp3+ T cells, four subsets of CD4+CD25highCCR6+, CD4+CD25highCCR6-, CD4+CD25-CCR6+ and CD4+CD25-CCR6- cells were sorted and tested subsequently in the co-cultured system. The percentage of IL-17+ Foxp3+ T cells were increased in both CD4+CD25highCCR6+ and CD4+CD25-CCR6+ subsets, suggesting that the IL-17+ Foxp3+ cells maybe expanded from either IL-17+Foxp3+ or Th17 cells. The percentage of Foxp3+ cells was also increased in CD4+CD25-CCR6- and CD4+CD25-CCR6+ subsets, suggesting that CD4+CD25- cells could be differentiated into Foxp3+ cells. Therefore, our in vitro coculture assay suggested that OSCC may play a critical role in the induction and/or expansion both Th17 and Treg cells.