Effects of extracts from Panax ginseng, Panax notoginseng and Ligusticum chuanxiong on expression of β-galactosidase and signal pathway p16-cyclin D/CDK-Rb in vascular smooth muscle cells

Abstract

To observe the pathological changes of vascular aging in spontaneously hypertensive rats (SHRs), and the effects of extracts from Panax ginseng, Panax notoginseng (Burk.) and Ligusticum chuanxiong on vascular aging. Vascular smooth muscle cells (VSMCs) isolated from thoracic aorta of Wistar-Kyoto (WKY) rats and SHRs were primarily cultured and divided into WKY group, SHR group, valsartan group, low-dose extract group and high-dose extract group. Aging situation of VSMCs was detected by senescence-associated β-galactosidase (SA-β-gal) staining; the cell cycle was analyzed by flow cytometry; mRNA and protein expressions of p16, cyclin D1, cyclin-dependent kinase 4 (CDK4) and retinoblastoma protein (Rb) were detected by real-time fluorescence quantitative polymerase chain reaction (PCR) and Western blotting. SA-β-gal stain showed that compared with the WKY group, number of SA-β-gal-positive cells in the SHR group was increased significantly (P<0.01); extracts from Panax ginseng, Panax notoginseng (Burk.) and Ligusticum chuanxiong reduced the number of SA-β-gal-positive cells in the SHR group (P<0.01). Flow cytometry analysis showed that the number of VSMCs in G1 phase was reduced and in S phase was increased significantly (P<0.05) in the SHR group compared with the WKY group, and the number of VSMCs in G(1) phase was increased and in S phase was reduced significantly in the extracts groups (P<0.05). PCR result showed that compared with the WKY group, the mRNA expressions of p16 and Rb in the SHR group were reduced and those of cyclin D1 and CDK4 were increased significantly, the mRNA expressions of p16 and Rb in the extracts group were increased and mRNA expressions of cyclin D1 and CDK4 were reduced significantly. Western blotting result showed that compared with the WKY group, the protein expression of p16 in the SHR group was reduced and the protein expressions of cyclin D1, CDK4 and phospho-Rb were increased significantly (P<0.05), while the protein expression of p16 in the extracts group was increased and the protein expressions of cyclin D1, CDK4 and phospho-Rb were reduced significantly (P<0.05). The extracts from Panax ginseng, Panax notoginseng (Burk.) and Ligusticum chuanxiong can delay vascular aging of SHRs, which works by p16-cyclin D/CDK-RB pathways to inhibit VSMC proliferation.