Department of Otolaryngology Head and Neck Surgery, The First Affiliated Hospital of Liaoning Medical University, Jinzhou, Liaoning 121000, P.R. China.
Department of Otolaryngology Head and Neck Surgery, Liaoning Medical University, Jinzhou, Liaoning 121000, P.R. China.
Oncol Rep. 2018 Mar;39(3):1306-1312. doi: 10.3892/or.2017.6160. Epub 2017 Dec 15.
The aim of the present study was to observe the effects of cytokine signaling suppressor 1 (SOCS1)-silenced dendritic cells (DCs) pulsed with epidermal growth factor receptor (EGFR) fusion protein on the activation of T lymphocyte and cytotoxic T-lymphocyte (CTL) activity against Hep-2 cells. DCs were derived from the medullary cells of mice and authenticated by flow cytometry (FCM). Recombinant glutathione-S-transferase (GST)-EGFR fusion protein was produced and purified. After being pulsed with it, DCs were modified by recombinant SOCS1-siRNA adenoviral to silence SOCS1 gene expression. The maturation of DCs was evaluated by FCM. The effects of modified DCs on T-cell proliferation were assessed by MTT assay. The killing effects against Hep-2 cells of CTL were assessed by lactate dehydrogenase (LDH) release assay. High-purity DCs from the medullary cells of mice were obtained. Compared with the control, EGFR-pulsed DCs displayed higher expression of cell surface molecules, including CD83, CD860 and HLA-DR. The MTT assay revealed that all of the EGFR-pulsed, SOCS1‑silenced and EGFR-pulsed plus SOCS1-silenced DCs had an enhanced capacity to stimulate T-lymphocyte proliferation. As expected, EGFR-pulsed plus SOCS1-silenced DCs had the strongest effects on T-cell proliferation. The splenic T cells isolated from both EGFR-pulsed DC-immunized mice and EGFR-pulsed plus SOCS1-silenced DC-immunized mice enhanced the cytotoxicity against Hep-2 cells, while T cells from EGFR‑pulsed plus SOCS1-silenced DC-immunized mice exhibited significantly higher cytotoxicity than those from EGFR-DC-immunized mice. The EGFR-pulsed SOCS1‑siRNA-silenced DCs had the strongest effects on activation of T-cell proliferation and the CTL activity against Hep-2 cells.
本研究旨在观察表皮生长因子受体（EGFR）融合蛋白脉冲致敏的细胞因子信号转导抑制因子 1（SOCS1）沉默树突状细胞（DC）对 T 淋巴细胞激活和细胞毒性 T 淋巴细胞（CTL）对 Hep-2 细胞活性的影响。DC 由小鼠髓细胞衍生，并通过流式细胞术（FCM）鉴定。产生并纯化重组谷胱甘肽-S-转移酶（GST）-EGFR 融合蛋白。用它脉冲后，用重组 SOCS1-siRNA 腺病毒修饰 DC 以沉默 SOCS1 基因表达。通过 FCM 评估 DC 的成熟。通过 MTT 测定评估修饰后的 DC 对 T 细胞增殖的影响。通过乳酸脱氢酶（LDH）释放测定评估 CTL 对 Hep-2 细胞的杀伤作用。从小鼠骨髓细胞中获得高纯度的 DC。与对照组相比，EGFR 脉冲 DC 表面分子（包括 CD83、CD860 和 HLA-DR）表达更高。MTT 测定显示，所有 EGFR 脉冲、SOCS1 沉默和 EGFR 脉冲加 SOCS1 沉默的 DC 均增强了刺激 T 淋巴细胞增殖的能力。不出所料，EGFR 脉冲加 SOCS1 沉默的 DC 对 T 细胞增殖的影响最强。从 EGFR 脉冲 DC 免疫小鼠和 EGFR 脉冲加 SOCS1 沉默 DC 免疫小鼠分离的脾 T 细胞增强了对 Hep-2 细胞的细胞毒性，而来自 EGFR 脉冲加 SOCS1 沉默 DC 免疫小鼠的 T 细胞比来自 EGFR-DC 免疫小鼠的 T 细胞具有显著更高的细胞毒性。EGFR 脉冲 SOCS1-siRNA 沉默的 DC 对 T 细胞增殖的激活和对 Hep-2 细胞的 CTL 活性具有最强的作用。
