Effect of senescence marker protein 30 on the proliferation and apoptosis of human lens epithelial cells SRA01/04.

AIM

To study the effect of senescence marker protein 30 (SMP30) on the proliferation and apoptosis of human lens epithelial cell (HLEC) SRA01/04.

METHODS

SMP30 overexpression (OE) and knock down (KD) type cell lines were cultivated by using two groups regucalcin (RGN; SMP30) lentiviral vectors (LV-RGN, LV-RGN-RNAi) and the respective negative control virus infect SRA01/04 cells. Western blot and real-time quantitative polymerase chain reaction (q-PCR) analysis were used to determine RGN overexpression and knock down efficiency. We use cell counting kit-8 (CCK8) assay to measure cell viability and 5-bromodeoxyuridine (BrdU) assay to test cell proliferation. Cell cycle was measured by PI FACS assay and cell apoptosis was tested by Annexin V-APC assay through flow cytometry. We use Western blot to measure the content of caspase-3 in SRA01/04.

RESULTS

We used PCR and Western blot techniques to determine the successful transfection of SMP30 OE and KD SRA01/04 cell lines. By CCK8, Brdu and PI FACS cell cycle assay, it was found that the SMP30 OE group promoted cell proliferation (<0.05) compared with the control group, and the KD group inhibited cell proliferation (<0.05). The results of Annexin V-APC signal staining detection indicated that compared with respective control group, the cell apoptosis rate was higher in KD group (<0.05) but lower in OE group (<0.01). The expression of caspase-3 was down-regulated in OE group through Western blot assay and up-regulated in KD group compared with respective control group.

CONCLUSION

Proliferation of SRA01/04 was promoted by SMP30 OE and apoptosis was suppressed. Increasing the expression of SMP30 may protect HLEC SRA01/04 against apoptosis in cataract.