ABSTRACT
Objective To study the effect of Numb gene overexpression on invasion and migration in human renal cell carcinoma cells and its related mechanism. Methods The Numb-ORF plasmid was transfected into renal cell carcinoma cells 786-O, set the negative control and blank control. The expression of Numb and matrix metalloproteinase-9 (MMP-9) was detected by real-time PCR and western blot. Then abilities of cells invasion and migration were measured respectively by transwell assay and scratch-wound-healing assay. Results Compared to negative control (2.51±0.27) and blank control (2.87±0.21), Numb expression in Numb-ORF group (36.13 ±8.33) was significantly higher (P= 0.00). Meanwhile, MMP-9 level in Numb-ORF group was decreased: Numb-ORF group was 2.36±0.29, negative control was 8.73±0.91, blank control was 8.99±0.78 (P=0.00). In scratch-wound-healing assay,the repair ratio of 12 h was as follow: Numb-ORF group was 0.53 ±0.06, negative control was 0.73 ±0.09, blank control was 0.75 ±0.08 (P= 0.02). In transwell assay, the number of invasion cells in Numb-ORF group was 31.40±5.96, less than negative control (126.93±13.61) and blank control (131.87 ±15.42) (P= 0.00). Conclusion Overexpression of Numb significantly decreases abilities of invasion and migration in 786-O cells, and the suppressive effect may be due to the down-regulation of MMP-9 expression in human renal cell carcinoma.
ABSTRACT
Objective To study the effect of Numb overexpression on invasion and migration in human bladder cancer cell line T24,and its related mechanism.Methods In June 2013,the plasmids including Numb-ORF plasmid and its blank vector pCMV6-Entry were transfected into T24 cells,and set the blank control as usual.The expressions of Numb and MMP-9 (Matrix metalloproteinase-9) were detected by Real-Time PCR and Western Blot.Then T24 cell invasion and migration were measured respectively by Transwell assay and Scratch Wound Healing Assay.Results Compared to negative control group,which transfected with blank-vector plasmid and untransfected group,Numb expression level in Numb-ORF group was significantly higher (P＜0.01).Meanwhile,MMP-9 level in Numb-ORF group was decreased,△ CT-value as follows:Numb-ORF group was 8.423±0.202,negative control group was 7.294±0.138,untransfected group was 7.220±0.118 (P＜0.01).In Scratch Wound Healing Assay,the repair ratio of Numb-ORF group was less than negative control group and untransfected group.The repair ratio in 12 hours as follow:Numb-ORF group was 0.525±0.037,negative control group was 0.693±0.034,untransfected group was 0.701 ±0.038(P=0.004).In Transwell assay,the number of invasion cells in Numb-ORF group was 12.6±3.2,less than in control group (130.8±9.2) and untransfected group (132.2± 10.6) (P＜0.05).Conclusions Overexpression of Numb by transfection significantly decreases the capability of T24 cell invasion and migration.This suppressed effect may be due to the down-regulation of MMP-9 expression in human bladder cancer.