Cataract is a main cause of blindness worldwide. Results Manifestation of RAC1 was increased after transfection with RAC1 overexpression plasmid To explore the effect of RAC1, a RAC1 overexpression plasmid was constructed and transfected into SRA01/04 cells using lentivirus. The effect of RAC1 overexpression plasmid transfection was detected by qRT-PCR and western blot. Result of qRT-PCR showed that the mRNA level of RAC1 was increased to 3.320.34-fold after transfection with RAC1 overexpression plasmid, but no significant difference in RAC1 expression level after transfection with Vector (Figure 1A). The protein level of RAC1 was increased to 3.080.45-fold after transfection with RAC1 overexpression plasmid, leaving no significant changes in that of cells transfected with Vector (Figure 1B and ?and1C).1C). These results exhibited that the manifestation of RAC1 was enhanced in cells transfected with RAC1 overexpression plasmid, both in mRNA level and in protein level. Physique 1 RAC1 level was increased after transfection with RAC1 overexpression plasmid. A. The mRNA level of RAC1 was detected by quantitative real time PCR. The comparative mRNA level was calculated using 2-??Ct method. W, C. The protein level of … RAC1 overexpression promoted the proliferation of lens epithelial cells To explore the effect of RAC1 overexpression on the proliferation of lens epithelial cells, a MTT assay was carried out. As shown in Physique 2A, cells transfected with RAC1 overexpression Rabbit polyclonal to CCNA2 plasmid showed a quick growth compared with that of cells transfected with Vector. Then PCNA, which is usually a marker of proliferation, was also detected by western blot. Result showed that the protein level of PCNA was increased to 2.370.38-fold by 213261-59-7 RAC1 overexpression compared with Vector (Figure 2B and ?and2C).2C). These results suggested that RAC1 overexpression promoted the proliferation of lens epithelial cells. Physique 2 Overexpression of RAC1 accelerated the proliferation of lens epithelial cells. A. Cell viability was decided by MTT assay. W, C. Protein level of proliferating cell nuclear antigen (PCNA) was detected using western blot. The comparative protein level … RAC1 overexpression promoted the migration and invasion of lens epithelial cells Wound-healing assay was performed to detect the impact of RAC1 overexpression on the migration capability of lens epithelial cells. As shown in Physique 213261-59-7 3A, cells transfected with RAC1 overexpression plasmid showed a 213261-59-7 rapid migration at 12 h and 24 h compared with Vector. Transwell assay was then performed to detect the effect of RAC1 overexpression on the invasion of lens epithelial cells. Results of transwell assay showed that presently there were 107.411.7 cells passing 213261-59-7 through the micropore membrane in the RAC1 overexpression group, significantly more than that of Vector (72.67.92) (Physique 3B, < 0.001). These results indicated that RAC1 overexpression promoted the migration and invasion of lens epithelial cells. Physique 3 RAC1 overexpression promoted the migration and invasion of lens epithelial cells. A. Wound-healing assay was performed to evaluate the migration capability of cells. The comparative migration rate at 12 h and 24 h was calculated. Scale bar = 100 m. ... RAC1 overexpression promoted EMT of lens epithelial cells EMT is usually a key process in the metastasis of cells and the effect of RAC1 overexpression on EMT of lens epithelial cells was examined in this study. The morphological analysis of cells was first carried out. Cells transfected with Vector showed round or diamond appearance, but cells transfected with RAC1 overexpression plasmid showed long spindle appearance (Physique 4A). Then markers of EMT were detected by western blot. Results of western blot showed that 213261-59-7 after transfection with RAC1 overexpression plasmid, the protein level of E-cadherin was decreased to 618%, and the protein levels of N-cadherin, vimentin, fibronectin and -SMA was.