Effect of rapamycin on cell proliferation and ORC1 expression on rat's vascular smooth muscle cells
- Author:
Qian WANG
1
Author Information
1. Department of cardiology
- Publication Type:Journal Article
- Keywords:
Cell proliferation;
Rapamycin;
Rat;
Vascular smooth muscle cells
- From:
Academic Journal of Second Military Medical University
2011;32(10):463-471
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the effect and mechanism of rapamycin on cell proliferation and ORC1's expression on rat's vascular smooth muscle cells (VSMCs) in vitro. Method VSMCs were divided randomly into two groups, three bottles/wells per group. Control group (CG) were cultured in DMEM with 10% FBS, and experimental group (EG) were cultured in DMEM with 10 μmol/L rapamycin plus 10% FBS. Synchronized cells were obtained by serum starvation method. Expression of PCNA protein were observed by immunocytochemistry, VSMCs ultramicrostructure were analyzed by electron microscope, and expressions of ORC1, P53, cyclinA and cyclinD1 protein at transcription and protein level were displayed by RT-PCR and Western Blot during VSMCs were cultured from 0 h to 48 h. Results Compared with CG, positive expression ratio of PCNA protein in EG decreased significantly (20±2.1% vs 80±3.0%, P<0.05), expression level of P53 protein increased gradually (P<0.05), Some chromatin assembled to the membrane of nucleus and forming massive or crescent. Expression level of cyclinD1 slightly increased, but expression level of cyclinA significantly decreased in EG. The expression level of ORC1mRNA in CG significantly increased from 12 h to 48 h, and its peak was at 12 h. But ORC1mRNA expression in EG was kept at high level. The expression change of ORC1 protein by western blotting in both EG and CG were similar to that of ORC1mRNA. Conclusion Rapamycin not only inhibits the proliferation of VSMCs, but also promotes their apoptosis; Rapamycin's inhibitive effect on VSMCs proliferation is probably through preventing transformation of G0/G1 phase to S phase and inducing cells keep quiescence phase to come true; Rapamycin induces high expression of ORC1 indicating that the effect of ORC1 in cell cycle occurs on the upstem of rapamycin, and further supported the view that ORC1 involves in the initiation process of DNA replication.
