骨髓间充质干细胞分泌的外泌体诱导肿瘤相关巨噬细胞极化调控胰腺癌细胞实验研究

The treatment of pancreatic cancer cells by exosome derived from bone marrow mesenchymal stem cells through regulation of tumor associated macrophages polarization in vitro

目的探讨骨髓间充质干细胞(BMSC)分泌的外泌体调控肿瘤相关巨噬细胞(TAM)极化进而发挥对胰腺癌细胞的调控作用。方法将小鼠胰腺癌细胞株(Pan02)分为3组:单纯细胞组:Pan02单独培养不做特殊处理;共培养组:Pan02与诱导后M0型巨噬细胞共培养;外泌体组:Pan02与诱导后M0型巨噬细胞共培养后,加入外泌体。随后检测M1型以及M2型巨噬细胞活化标志物的基因表达、胰腺癌细胞标志物B7-H4,CA199的含量、细胞增殖能力以及细胞的侵袭和迁移能力。结果共培养细胞加入外泌体后,M1型活化标志物iNOS(6.5±2.1比13.6±4.2,P<0.05),CD68(4.5±1.6比15.3±4.9,P<0.05)表达增高,而M2型活化标志物Arginase(11.4±3.2比4.3±1.4,P<0.05),CD206(7.9±2.6比3.1±0.9,P<0.05)表达降低;同时,加入外泌体后,胰腺癌细胞标志物B7-H4(10.9±3.4比4.6±1.5,P<0.05),CA199(21.6±7.3比8.2±2.9,P<0.05)表达下降,癌细胞侵袭力(103.4±34.5比54.6±19.1,P<0.05)及迁移能力(104.6±34.9比59.3±19.8,P<0.05)减弱,而凋亡率增高(3.26%比24.3%,P<0.05)。结论BMSC分泌的外泌体可抑制TAM向M2表型转化,并诱导其向M1表型转化,进而抑制胰腺癌细胞的增殖、侵袭及迁移能力,达到抑癌的作用。

Objective To investigate the effect of exosomes derived from bone marrow mesenchymal stem cells(BMSCs)on pancreatic cancer through the polarization of tumor associated macrophages(TAM).Methods The mouse pancreatic cancer cell line Pan02 was divided into 3 groups:Pan02 group(Pan02 cells without any special treatment);co-cultured group(Pan02 cells co-cultured with M0 macrophages);exosome group(Pan02 cells co-cultured with M0 macrophages,and then exosomes were added).The mRNA expression of M1 biomarkers and M2 biomarkers,the content of pancreatic cancer biomarkers,the proliferation and apoptosis of Pan02 cell line,and the invasion and migration ability were detected.Results After exosomes were added,the expression of iNOS(6.5±2.1 vs.13.6±4.2,P<0.05)and CD68(4.5±1.6 vs.15.3±4.9,P<0.05)increased while the expression of Arginase(11.4±3.2 vs.4.3±1.4,P<0.05)and CD206(7.9±2.6 vs.3.1±0.9,P<0.05)decreased with significant difference.Also,the exosomes treatment could significantly decrease the expression of B7-H4(10.9±3.4 vs.4.6±1.5,P<0.05)and CA199(21.6±7.3 vs.8.2±2.9,P<0.05),meanwhile significantly inhibit the invasion(103.4±34.5 vs.54.6±19.1,P<0.05),migration rate(104.6±34.9 vs.59.3±19.8,P<0.05)while significantly promoting apoptosis rate(3.26%vs.24.3%,P<0.05).Conclusion Exosomes derived from BMSCs can inhibit the polarization of TAM to the M2 phenotype and induce their polarization to the M1 phenotype,thereby suppressing the development of pancreatic cancer.