| Objective: Cysteine-rich acidic secretory protein like 1(Sparcl1)is an extracellular matrix protein,involved in the regulation of intercellular adhesion,cell migration and invasion.In the last 30 years,a lot of discussion about the role of Sparcl1 genes in different tumors,including ovarian cancer.Sparcl1 can limit cell migration and invasion in tumors by regulating the expression of adhesion-related molecules and migration-related molecules,and reduce tumor metastasis,but its detailed mechanism is unclear.By detecting the expression of Sparcl1 in ovarian cancer,preliminary analysis of its effect on ovarian cancer;then the effect of transfection of Sparcl1 gene on biological behavior of human ovarian cancer SKOV3 cell line and its possible mechanism were discussed.Methods:(1)The expression of Sparcl1 protein was examined by IHC staining in paraffin sections of 58 cases of ovarian cancer and 25 cases of normal ovarian tissue.(2)Transfection of Sparcl1 plasmid and empty plasmid vector into SKOV3 cell lines;a group of transfection strains with high Sparcl1 expression and a group of empty plasmids were obtained.(3)qRT-PCR methods were used to identify the successful transfection of Sparcl1 genes in SKOV3 cells.The expression of Sparcl1 protein,ERK protein and p-ERK protein was determined by Western blotting(Western Blot,WB)in Control group,OE-NC group,OE-Sparcl 1 group.(4)The Optical Density(OD)values of Control,OE-NC and OE-Sparcl1 groups at 1d,2d,3d,4d were detected by the method CCK-8,which reflected the inhibition of cell proliferation.(5)Transwell experiments were performed to detect the number of cells entering the lower chamber in the Control group,the OE-NC group and the OE-Sparcl1 group at1 d and 2d,to evaluate the invasion ability of the cells by examining the number of cells.(6)The scratch widths of Control group,OE-NC group and OE-Sparcl1 group at 1d,2d were measured by scratch test,and the mobility was calculated.(7)The apoptosis or survival of 1 cell in Control group,OE-NC group and OE-Sparcl1 group were detected by 7-AAD/p I-Annexin V flow cytometry;Results:(1)The IHC staining intensity of ovarian sections was lower than that of normal ovarian sections(p<0.05).There was no significant difference in the expression level of Sparcl1 protein among mass size,histological classification and age in 58 ovarian cancer tissues((p>0.05),but significant difference with histological grade,clinical stage,regional lymph node metastasis(p<0.05).(2)The expression level of Sparcl1 protein in OE-Sparcl1 group was the highest and significantly higher than that in Control group and OE-NC group(p<0.05).There was no difference in ERK expression level of OE-Sparcl 1 compared with OE-NC group and Control group(p>0.05);The expression level of p-ERK in OE-Sparcl1 group was the lowest,also significantly lower than that in Control group and OE-NC group(p<0.05).(3)CCK-8 results showed that the inhibition rate of proliferation was significantly higher in group 2d、3d、4d than that in group Control and OE-NC(p<0.05).(4)Transwell experimental results showed that OE-Sparcl1 group had less invasive ability than group Control and group OE-NC(p<0.05).(5)Scratch test showed that the mobility of OE-Sparcl1 group was significantly lower than that of Control group and OE-NC group(p<0.05).(6)The apoptosis rate of SPARCl1 group by 7-AAD/p E-Annexin V staining flow cytometry was significantly higher than that of Control group and NC group(p<0.05).Conclusion:(1)Sparcl1 protein is low or absent in ovarian cancer tissue and high in normal ovarian tissue;suggests that Sparcl1 gene on the occurrence and progress of ovarian cancer may have certain inhibition.(2)Sparcl1 gene can inhibit the proliferation,migration and invasion of SKOV3 cells and promote the apoptosis of SKOV3 cells.(3)The effect of Sparcl1 on the biological behavior of SKOV3 cells may be related to the down regulation of p-ERK. |
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