沉默PGAM1基因对食管癌细胞生物学功能的影响

目的   探讨沉默磷酸甘油酸变位酶1（PGAM1）基因对食管癌细胞生物学功能的影响及可能机制。方法   收集2016年7月至2017年6月间在我院行手术切除的67例食管癌及对应癌旁正常组织，采用免疫组化SP法检测PGAM1表达并分析其表达与临床病理特征的关系。PGAM1shRNA（shPGAM1组）或shRNA-NC（NC组）转染至食管癌Eca-109细胞，分析PGAM1基因沉默对Eca-109细胞凋亡和增殖的影响。采用实时荧光定量PCR（QPCR）和Western blotting法检测PGAM1对Akt／mTOR信号通路关键分子的影响。结果   食管癌组织中PGAM1高表达率为58.2％（39／67），高于癌旁组织的31.3％（21／67），差异有统计学意义（P＜0.05）。PGAM1表达与临床分期、分化程度、浸润深度有关（P＜0.05），而与年龄、性别、淋巴结转移等无关（P＞0.05）。shPGAM1组和NC组中PGAM1 mRNA表达量分别为0.48±0.10和1.01±0.14，差异有统计学意义（P＜0.05）。MTT法检测结果 显示，培养24、48、72、96 h后，shPGAM1组Eca-109细胞增殖率分别为（87.65±7.42）％、（79.34±9.11）％、（70.17±6.84）％、（60.36±7.95）％，明显低于NC组，差异有统计学意义（P＜0.05）。48 h后shPGAM1组和NC组的Eca-109细胞凋亡率分别为（45.36±5.38）％和（24.81±4.67）％，差异有统计学意义（P＜0.05）；shPGAM1组和NC组Eca-109细胞培养上清的葡萄糖消耗量分别为（56.84±11.35）％和（99.87±10.48）％，shPGAM1组和NC组Eca-109细胞培养上清的乳酸含量分别为（48.02±10.18）％和（99.00±12.35）％，差异均有统计学意义（P＜0.05）。shPGAM1组细胞的PTEN表达量高于NC组（P＜0.05），而p-Akt、p-mTOR表达量均低于NC组（P＜0.05）。结论   PGAM1高表达与食管癌的发生、发展有关，通过激活Akt／mTOR信号通路诱导的Warburg效应，促使肿瘤细胞增殖，抑制其凋亡。

Experimental study on the biological function and mechanism ofPGAM1 on esophageal cancer

Objective  To investigate the effect of silencingphosphoglycerate mutase 1 （PGAM1） gene on thebiological function of esophageal carcinoma cells and its possible mechanism. Methods  The expression of PGAM1 was detected byimmunohistochemical SP method in 67 fresh esophageal cancer tissues and theiradjacent normal tissues from July 2016 to June 2017. In order to analyze theeffect of PGAM1 gene silencing on the apoptosis and proliferation of Eca-109cells， PGAM1 shRNA （shPGM1 group） or shRNA-NC （NC group） was transfected intoEca-109 cells. QPCR and Western blotting were used to detect the effect ofPGAM1 on the key molecules in Akt／mTOR signalingpathway. Results The high expression rate of PGAM1 in esophageal carcinoma was58.2％ （39／67）， which was higher than that in adjacent tissues （31.3％）， and the differencewas statistically significant （P＜0.05）. The expression ofPGAM1 was correlated with clinical stage， differentiationdegree and depth of invasion （P＜0.05）， but not with age， sex and lymph nodemetastasis （P＞0.05）. The expression of PGAM1 mRNA in shPGAM1 group and NC group were 0.48±0.10 and 1.01±0.14 respectively， and the difference was statistically significant （P＜0.05）. MTT assay showed that after 24， 48， 72 and 96 hours of culture， the proliferation rates Eca-109 cells in shPGAM1 group were （87.65±7.42）％， （79.34±9.11）％， （70.17± 6.84）％ and （60.36±7.95）％ respectively， which were significantly lower than those of NC group （P＜0.05）. The apoptosis rates of Eca-109 cells inshPGAM1 group and NC group were （45.36±5.38）％ and （24.81±4.67）％ respectively after 48 hours （P＜0.05）. The content of glucose in culture supernatantof Eca-109 cells in shPGAM1 group and NC group was （56.84±11.35）％ and （99.87±10.48）％， respectively. Thecontent of lactic acid in culture supernatant of Eca-109 cells in shPGAM1 groupand NC group was （48.02±10.18）％ and （99.00±12.35）％， respectively. The difference was statisticallysignificant （P＜0.05）. The expression ofPTEN in shPGAM1 group was higher than that in NC group （P＜0.05）， but the expression of p-Akt and p-mTOR waslower than that in NC group （P＜0.05）. Conclusion Thereare significant evidences that PGAM1 up-regulation would be related withesophageal cancer， which promote theproliferation of Eca-109 cells by activating the Akt／mTOR signaling pathway and Warburg effect.