CCL18通过与Nir1结合促进胶质瘤细胞侵袭的研究

Nir1 promotes invasion of glioma cells by binding to chemokine (C–C motif) ligand 18

背景与目的:Nir1是CCL18的跨膜受体,CCL18能与之特异性结合而促进乳腺癌细胞的侵袭与转移,但其在胶质瘤细胞中的作用尚不清楚。该文旨探讨Nir1在胶质瘤侵袭中的作用及分子机制。方法:应用蛋白[质]印迹法(Western blot)检测Nir1在不同胶质瘤细胞系中的表达;利用siRNA技术抑制U251细胞中Nir1的表达;采用Western blot检测转染后Nir1蛋白的表达和转染前后U251细胞中Akt磷酸化的情况;使用体外侵袭实验检测转染后细胞的运动和侵袭能力;采用F-actin聚合实验检测F-actin的聚合能力。结果:Nir1在各胶质瘤细胞中均呈高表达;小RNA干扰技术沉默Nir1基因后,U251细胞中Nir1的蛋白表达量明显下降,侵袭并穿透Matrigel胶的细胞数目明显比对照组少(P=0.00);在CCL18刺激后细胞内的F-actin聚合量比对照组减少;Akt磷酸化试验结果显示,对照组细胞在CCL18的刺激下Akt更易发生磷酸化,实验组细胞无论是否存在CCL18,Akt磷酸化都受到抑制。结论:在胶质瘤细胞中存在Nir1蛋白高表达,通过与细胞膜上CCL18特异性结合来调节胶质瘤细胞的F-actin聚合和Akt的磷酸化进而调控胶质瘤细胞的运动和侵袭能力。

Background and purpose： Nir1 is a transmembrane receptor for chemokine （C-C motif） ligand 18. CCL 18 specifically binds to Nir1 at the cellular membrane of breast cancer cells to exert its invasion and metastasis. However, the specific mechanism of Nir1 is not clear in glioma. This study probed the effect and mechanism of Nir1 in the invasion of glioma cells. Methods： Western blot was used to detect the expression of Nir1 in glioma cells. siRNA plasmid was used to transfect U251 cells. Western blot was used to analyze the expression of Nir1 and protein phosphorylation of Akt in the cells transfected by Nir1 plasmid. In vitro Matrigel invasion assay was used to detect the invasive ability in the cells that were transfected. F-actin polymerization assay was used to detect F-actin recognition ability in cells. Results： The expression of Nir1 was higher in all glioma cells. After transfection, the invasion of siNir1/ U251 was obviously decreased than the SCR/U251, F-actin content was reduced compared to the control group. Akt phosphorylation experiment result showed that the protein phosphorylation of Akt was enhanced in control group cells CCL18 following stimulation. However, the existence of CCL18 would affect the phosphorylation of Akt in siNir1/U251. Conclusion： Nir1 is high expression in glioma cells, and Nir1 binding to chemokine CCL18 promotes glioma cells invasion and metastasis through regulation the phosphorylation of Akt and F-actin polymerization.