Department of Otolaryngology, 74566The First Affiliated Hospital of Soochow University, Suzhou, China.
Department of Otolaryngology, Suzhou Ninth Hospital affiliated to Soochow University, Suzhou, China.
Technol Cancer Res Treat. 2021 Jan-Dec;20:15330338211046432. doi: 10.1177/15330338211046432.
This study aimed to explore the effect of programmed death-ligand 1 (PD-L1) in cancer-associated fibroblasts (CAFs) on advanced laryngeal squamous cell carcinoma (LSCC). The expression of PD-L1 in advanced LSCC tumor tissues was observed in 83 patients with LSCC by immunofluorescence microscopy and compared with that in normal laryngeal mucosa. The CAFs of LSCC and normal fibroblasts (NFs) were isolated, cultured, purified, and examined by fluorescence. The expression of PD-L1 in purified CAFs and NFs was measured by flow cytometry. The expression of PD-L1 in CAFs was downregulated through small interferring RNA (siRNA) transfection. The proliferation and migration capacities of CAFs were observed using proliferation and scratch tests, respectively. The proliferation of HEP-2 cells and T cells was measured after cocultured with CAFs. The secretion of interleukins IL-2 and IL-10 was detected using enzyme-linked immuno sorbent assay (ELISA). PD-L1 was expressed in 62 of 83 cases of the advanced LSCC tumor tissues. Also, CAFs expressed more PD-L1 compared with NFs. The proliferation and migration capacities of CAFs were significantly lower after transfection with PD-L1-siRNA. The proliferation rate of HEP-2 cells cocultured with CAFs decreased in PD-L1-siRNA-transfected cells. However, the proliferation rate of T cells increased in transfected cells. The ELISA results showed that the secretion of IL-2 increased and that of IL-10 decreased in PD-L1-siRNA-transfected cells. The expression of PD-L1 in CAFs of advanced LSCC was higher than that in NFs. The downregulation of PD-L1 reduced the proliferation and migration of CAFs and HEP-2 cells but enhanced the proliferation and pro-inflammatory function of T cells in the coculture experiment.
本研究旨在探讨肿瘤相关成纤维细胞（CAFs）中程序性死亡配体 1（PD-L1）对晚期喉鳞状细胞癌（LSCC）的影响。通过免疫荧光显微镜观察 83 例 LSCC 患者的晚期 LSCC 肿瘤组织中 PD-L1 的表达，并与正常喉黏膜进行比较。分离、培养、纯化 LSCC 和正常成纤维细胞（NFs），并用荧光法进行鉴定。通过流式细胞术检测纯化后的 CAFs 和 NFs 中 PD-L1 的表达。通过小干扰 RNA（siRNA）转染下调 CAFs 中 PD-L1 的表达。通过增殖和划痕试验分别观察 CAFs 的增殖和迁移能力。将 CAFs 与 HEP-2 细胞和 T 细胞共培养后，测量 HEP-2 细胞和 T 细胞的增殖情况。采用酶联免疫吸附试验（ELISA）检测白细胞介素（IL）-2 和 IL-10 的分泌。在 83 例晚期 LSCC 肿瘤组织中，有 62 例表达 PD-L1。此外，CAFs 表达的 PD-L1 明显多于 NFs。转染 PD-L1-siRNA 后，CAFs 的增殖和迁移能力显著降低。与 CAFs 共培养的 HEP-2 细胞的增殖率在转染 PD-L1-siRNA 的细胞中降低，但转染细胞中 T 细胞的增殖率增加。ELISA 结果显示，转染 PD-L1-siRNA 的细胞中 IL-2 的分泌增加，IL-10 的分泌减少。晚期 LSCC 中 CAFs 表达的 PD-L1 高于 NFs。下调 PD-L1 可降低 CAFs 和 HEP-2 细胞的增殖和迁移能力，但在共培养实验中增强了 T 细胞的增殖和促炎功能。
