摘要
目的探讨组织蛋白酶S(Cat S)对人骨肉瘤细胞增殖、迁移和侵袭的影响及其潜在调控机制。方法以正常成骨细胞(hFOB)和人骨肉瘤细胞(SAOS2和MG63)作为研究对象,将Cat S小干扰RNA(si-Cat S)和阴性对照序列(si-NC)转染至SAOS2和MG63细胞中以改变Cat S在人骨肉瘤细胞中的表达。将实验细胞分为4组:hFOB组(hFOB细胞)、Control组(未转染的SAOS2或MG63细胞)、si-NC组(转染了si-NC的SAOS2或MG63细胞)和si-Cat S组(转染了si-Cat S的SAOS2或MG63细胞)。RT-PCR和Western blot法检测Cat S在SAOS2和MG63细胞中的表达;CCK-8和克隆形成实验检测敲低Cat S对SAOS2和MG63细胞增殖活性的影响;划痕和Transwell实验分别检测敲低Cat S对SAOS2和MG63细胞迁移和侵袭能力的影响;Western blot实验检测敲低SAOS2对SAOS2细胞凋亡相关蛋白(Bcl-2、Bax和Caspase-3)和Wnt/β-catenin通路相关蛋白(LRP5、β-catenin、C-myc和Cyclin D1)的影响。结果与hFOB组比较,Cat S在SAOS2组和MG63组中的表达上调(P<0.001)。此外,与si-NC组比较,si-Cat S组细胞的增殖、迁移和侵袭能力均受到抑制(P<0.001)。Western blot结果显示:与si-NC组比较,si-Cat S组细胞的Bcl-2表达下调,而Bax和Caspase-3表达上调(P<0.001)。同时,与si-NC组比较,si-Cat S组LRP5、β-catenin、C-myc和Cyclin D1的表达均下调(P<0.001)。结论siRNA敲低Cat S可以通过调控Wnt/β-catenin通路来抑制人骨肉瘤细胞的增殖、迁移和侵袭并诱导细胞凋亡,预示了Cat S可能是骨肉瘤治疗的潜在靶点之一。
Objective To investigate the effects of cathepsin S(Cat S)on proliferation,migration and invasion of osteosarcoma cells and its potential regulatory mechanism.Methods Normal osteoblasts(hFOB)and osteosarcoma cells(SAOS2 and MG63)were selected as the subjects of this study.Cat S small interfering(si)RNA(si-Cat S)and negative control sequence(si-NC)were transfected into SAOS2 and MG63 cells to modulate the expression of Cat S in osteosarcoma cells.The experimental cells were randomly divided into four groups:hFOB group(hFOB cells),Control group(untransfected SAOS2 or MG63 cells),si-NC group(SAOS2 or MG63 cells transfected with si-NC)and si-Cat S group(SAOS2 or MG63 cells transfected with si-Cat S).The expression of Cat S in SAOS2 and MG63 cells was detected by RT-PCR and Western blot.Effect of Cat S knockdown on the proliferation of SAOS2 and MG63 cells was assessed by CCK-8 and clone formation assays.And effects of Cat S knockdown on the migration and invasion of SAOS2 and MG63 cells were determined by wound-healing and Transwell assays,respectively.Western blot assay was performed to measure the effects of SAOS2 knockdown on the expressions of apoptosis-related proteins(Bcl-2,Bax and Caspase-3)and Wnt/β-catenin pathway related proteins(LRP5,β-catenin,C-myc and Cyclin D1)in SAOS2 cells.Results Compared with hFOB group,the expression of Cat S in SAOS2 group and MG63 group was upregulated(P<0.001).In addition,compared with si-NC group,the proliferation,migration and invasion of cells in si-Cat S group were reduced(P<0.001).Results of Western blot showed that compared with si-NC group,the expression of Bcl-2 in si-Cat S group was downregulated,while the expression of Bax and Caspase-3 were upregulated(P<0.001).Meanwhile,compared with si-NC group,the expression of LRP5,β-catenin,C-myc and Cyclin D1 in si-Cat S group was downregulated(P<0.001).Conclusion Cat S siRNA knockdown can inhibit the proliferation,migration and invasion of osteosarcoma cells and induce apoptosis by regulating Wnt/β-catenin pathway,indicating that Cat S may be one of the potential targets for the treatment of osteosarcoma.
作者
纪海茹
孔令伟
曹胜
吕家兴
李佳欣
刘春雨
金宇
Ji Hairu;Kong Lingwei;Cao Sheng;LüJiaxing;Li Jiaxin;Liu Chunyu;Jin Yu(Dept of Pathology,Chengde Medical College,Chengde 067000;Dept of Trauma Surgery,Hospital of Chengde Medical College,Chengde 067000)
出处
《安徽医科大学学报》
CAS
北大核心
2022年第9期1459-1465,共7页
Acta Universitatis Medicinalis Anhui
基金
河北省高等学校自然科学研究青年基金项目(编号:QN2020107)
河北省医学科学研究课题计划项目(编号:20200378、20200352)。
