摘要
目的研究颗粒蛋白前体(progranulin,PGRN)对滋养层细胞侵袭、凋亡的影响,并探讨其在子痫前期发病机制中的作用。方法取子痫前期患者胎盘组织,测定其中PGRN mRNA和蛋白表达。选择子痫前期的3种滋养层细胞系JEG-3、BeWo、JAR以及人原代培养的滋养层细胞,通过定量聚合酶链反应(quantitative polymerase chain reaction,q PCR)和蛋白质免疫印迹(Western blot)实验确定PGRN在其中的表达情况。利用小干扰RNA(small interfering RNA,siRNA)干扰和过表达该细胞系中的PGRN,用qPCR、Western blot实验确定干扰或过表达有效后,分别通过Transwell侵袭实验和流式细胞术测定PGRN差异表达对滋养层细胞侵袭和凋亡的影响,同时在分子水平测定侵袭相关蛋白基质金属蛋白酶(matrix metalloproteinase,MMP)-2、-9和凋亡蛋白Caspase-3的表达及活性。结果子痫前期患者胎盘组织中PGRN在mRNA和蛋白水平的表达都显著高于正常组(P <0. 05)。JEG-3、BeWo、JAR细胞系中的PGRN蛋白水平显著高于原代滋养层细胞(P <0. 05)。在BeWo滋养层细胞中,干扰PGRN表达后,其侵袭能力减弱(P <0. 05),凋亡增加(P <0. 01)。分子水平上,MMP-2、MMP-9活性减弱(P <0. 05),Caspase-3裂解量增加(P <0. 01)。而过表达PGRN后,BeWo细胞侵袭能力(P <0. 05)和抗凋亡能力(P <0. 01)增加。结论 PGRN能够促进滋养层细胞侵袭和抑制细胞凋亡,通过上调PGRN可能减少子痫前期的发生和发展。
ObjectiveTo assess the effect of progranulin on the invasion and apoptosis of trophoblast cells and investigate its role in the pathogenesis of preeclampsia. MethodsProgranulin mRNA and protein levels were detected in the placenta tissues from 12 patients with preeclampsia and 12 women with normal pregnancy. The expression levels of progranulin in human trophoblast cells in primary cultures and 3 preeclamptic trophoblast cell lines （JEG-3, BeWo and JAR） were also detected using quantitative real-time PCR （qPCR） and Western blotting. The effects of progranulin silencing mediated by a specific siRNA and progranulin overexpression by an overexpression vector on the invasion and apoptosis of these cell lines were determined using Transwell assay and flow cytometry with Annexin V-PI staining; the expressions of invasion-related proteins MMP-2 and MMP-9 and the apoptotic protein Caspase-3 were also detected. ResultsThe mRNA and protein levels of progranulin were significantly higher in placental tissues of patients with preeclampsia than in normal placenta （P〈0.05）. The protein levels of progranulin were also significantly higher in JEG-3, BeWo and JAR cell lines than in normal human trophoblast cells （P〈0.05）. In BeWo trophoblast cells, progranulin silencing significantly reduced the cell invasion （P〈0.05）, increased cell apoptosis （P〈0.01）, reduced MMP-2 and MMP-9 expressions （P〈0.05）, and increased Caspase-3 activity （P〈0.01）, while the reverse changes occurred in cells with progranulin overexpression （P〈0.05）. ConclusionProgranulin is indispensable for the invasion and survival for trophoblast cells. The increased progranulin expression in the placenta may serve as a compensatory mechanism in preeclamptic patients, and progranulin upregulation may help to suppress the occurrence and progression of preeclampsia.
作者
吴秋雨
郇金亮
WU Qiuyu;HUAN Jinlian(College of Medicine,Jiangsu University,Zhenjiang,Jiangsu Province,212013;Department of Gynecology and Obstetrics,Huaqiao People＇s Hospital of Kunshan City,Kunshan,Jiangsu Province,215332;Department of General Surgery,Shanghai Eighth People＇s Hospital,Shanghai,200235,China)
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2018年第20期1858-1864,共7页
Journal of Third Military Medical University
