沉默N-cadherin对人牙髓干细胞增殖和迁移能力的影响  

Effects of N-cadherin silencing on the proliferation and migration of human dental pulp stem cells

作　　者：邓子龙[1,2] 闫文娟[1,2] 赵望泓[1,2] 吴补领[2,3] DENG Zilong;YAN Wenjuan;ZHAO Wanghong;WU Buling(Department of Stomatology,Nanfang Hospital,Southern Medical University,Guangzhou 510515,China;College of Stomatology,Southern Medical University,Guangzhou 510515,China;Department of Operative Dentistry and Endodontics,Shenzhen Stomatology Hospital(Pingshan),Southern Medical University,Shenzhen 518118,China)

机构地区：[1]南方医科大学南方医院口腔科,广东广州510515 [2]南方医科大学口腔医学院,广东广州510515 [3]南方医科大学深圳口腔医院(坪山)牙体牙髓病科,广东深圳518118

出　　处：《口腔疾病防治》2022年第11期779-784,共6页Journal of Prevention and Treatment for Stomatological Diseases

基　　金：国家自然科学基金项目(82100997)。

摘　　要：目的探讨沉默N-cadherin对人牙髓干细胞(dental pulp stem cells,DPSCs)增殖和迁移能力的影响,为基于DPSCs的牙髓再生提供实验依据。方法采用N-cadherin shRNA慢病毒转染DPSCs,qRT-PCR、Western blot检测N-cadherin的表达水平,验证沉默效率。实验分为阴性对照组(shRNA-NC)和N-cadherin shRNA沉默组。采用CCK-8法检测细胞增殖能力,流式细胞术检测细胞周期和细胞凋亡情况,Transwell法检测细胞迁移能力。结果N-cadherin shRNA可显著降低DPSCs的N-cadherin mRNA和蛋白的表达水平(P<0.001)。N-cadherin shRNA组细胞增殖活性分别在细胞接种的第3、4天高于shRNA-NC组,第6~8天低于shRNA-NC组,差异均具有统计学意义(P<0.05);在第3天时,N-cadherin shRNA组处于S期和G2期的细胞比例大于shRNANC组(P<0.05);在第6天时,N-cadherin shRNA组细胞处于S期和G2期的比例小于shRNA-NC组(P<0.05),同时N-cadherin shRNA组凋亡细胞比例大于shRNA-NC组(P<0.01);分别以低细胞密度和高细胞密度接种于Transwell上室培养20 h,N-cadherin shRNA组穿过Transwell上室小孔的细胞数量均多于shRNA-NC组(P<0.001)。结论沉默N-cadherin表达可促进DPSCs早期的增殖和迁移能力。Objective To investigate the effects of N-cadherin silencing on the proliferation and migration of human dental pulp stem cells(DPSCs)and to provide experimental evidence for DPSCs-based dental pulp regeneration.Methods DPSCs were transfected with N-cadherin shRNA lentivirus,and the knockdown efficiency of N-cadherin at both the mRNA and protein levels was confirmed by qRT-PCR and Western blot.The experiment included a negative control group(shRNA-NC)and an N-cadherin shRNA silencing group.Cell proliferation was detected by the CCK-8 method.Cell cycle and apoptosis were assessed by flow cytometry,and cell migration was detected using the Transwell method.Results N-cadherin shRNA significantly reduced the expression levels of N-cadherin mRNA and protein in DPSCs(P<0.001).The proliferation activity of the N-cadherin shRNA group was significantly greater than that of the shRNA-NC group on the 3rd and 4th days after cell inoculation and lower than that of the shRNA-NC group from the 6th to 8th days(P<0.05).On the 3rd day after cell inoculation,the proportion of cells in S phase and G2 phase in the N-cadherin shRNA group was greater than that in the shRNA-NC group(P<0.05).On the 6th day after cell inoculation,the proportion of cells in S phase and G2 phase in the N-cadherin shRNA group was lower than that in the shRNA-NC group(P<0.05),and the proportion of apoptotic cells in the N-cadherin shRNA group was greater than that in the shRNA-NC group(P<0.01).Low densities cells and high densities cells were inoculated into Transwell upper chamber for 20 h,the number of cells passing through the membrane pores of upper chamber in the N-cadherin shRNA group was greater than that in the shRNA-NC group(P<0.001).Conclusion Silencing N-cadherin expression can promote the early proliferation and migration of DPSCs.

关 键 词：牙髓干细胞 增殖 迁移 N-CADHERIN 牙髓再生 细胞周期 细胞凋亡 慢病毒 微环境 

分 类 号：R78[医药卫生—口腔医学]