目的：研究涎腺腺样囊性癌（salivary adenoid cystic carcinoma ，SACC）来源的外泌体（exosome，EXO）对成纤维细胞PD-L1分子表达的影响。方法：通过EXO分离试剂盒提取SACC-83细胞系来源的外泌体并以电镜鉴定其颗粒大小、密度和表型；使用PKH67荧光标记后将EXO与成纤维细胞HPLF共孵育，以共聚焦显微镜观察EXO可否被HPLF细胞摄取；将EXO与HPLF细胞共孵育后，全转录组测序检测筛查该细胞中差异表达基因，GO分析结合KEGG富集分析阐明差异表达基因涉及的生物学功能和相关信号通路；使用qPCR、Western blotting 和流式细胞术检测肿瘤EXO对HPLF细胞中PD-L1、LAG3 和IDO1 在mRNA与蛋白水平表达的影响。结果：SACC-83细胞源EXO特异性表达CD63、CD81和TSG101分子，并可被HPLF细胞内吞摄取；EXO处理HPLF细胞后，细胞中PD-L1 分子显著上调表达（Fold change=10.19），差异基因显著富集于TNF、NF-κB、cGAS-String 等免疫反应信号通路；体外实验证明EXO可以从mRNA和蛋白水平显著促进HPLF细胞中PD-L1 的表达（24.7±4.75 vs 1.03±0.11，P<0.05）。结论：涎腺腺样囊性癌SACC-83 细胞来源EXO可以促进HPLF细胞中免疫检查点配体PD-L1的表达。

Objective: To investigate the effect of salivary adenoid cystic carcinoma (SACC) derived exosomes on PD-L1 expression in fibroblasts. Methods: Exosomes of SACC-83 cell line were extracted by exosome isolation kit, and its particle size, density and phenotypes were identified by electron microscope. After being labeled with PKH67 fluorescence, exosomes were co-incubated with HPLF to observe whether the exosomes could be ingested by fibroblasts under confocal microscope. After co-incubation with the exosomes,the differentially expressed genes (DEGs) in HPLF cells were detected by whole transcriptome sequencing. GO analysis together with KEGG enrichment analysis was used to clarify the biological functions and related signaling pathways related with the DEGs. qPCR,Western blotting and Flow cytometry were used to detect the effect of tumor exosomes on the mRNA and protein expressions of PD-L1,LAG3 and IDO1 in HPLF fibroblasts. Results: SACC exosomes specifically expressed CD63, CD81 and TSG101 molecules, and could be ingested by fibroblasts. After the treatment of fibroblasts by exosomes, the expression of PD-L1 molecule was significantly up-regulated (Fold change=10.19), and the DEGs were significantly enriched in the immune response signaling pathways, such as TNF, NF-κB and cGAS-String pathway, etc. In vitro experiments showed that exosomes could significantly promote the expression of PD-L1 in HPLF cells at both mRNA and protein levels (24.7±4.75 vs 1.03±0.11,P<0.05). Conclusion: Exosomes of SACC can promote the expression of immunocheckpoint ligand PD-L1 in fibroblasts.

国家自然科学基金资助项目（No.81602497）