Effect Of Ginsenoside Compound K On Biological Behavior Of Pulmonary Arterial Smooth Muscle Cells

Objective: To explore the effects and the related mechanisms of ginsenoside compound K(CK)on the proliferation,apoptosis,migration,phenotypic conversion,and collagen deposition of platelet-derived growth factor-BB(PDGF-BB)-induced pulmonary arterial smooth muscle cells(PASMCs).This study provides a new insight and theoretical basis for the treatment of pulmonary arterial hypertension(PAH).Methods: 1.PASMCs of SD rats were primary cultured by tissue block attachment method and were identified by immunofluorescence smooth muscleα-actin(α-SMA),and then cell passage was performed for subsequent experimental study.2.In this study,PDGF-BB-induced PASMCs in vitro simulate the pathological state of PASMCs in PAH;CK was used to intervene the biological behavior of PDGF-BB-induced PASMCs.The experiment was divided into three groups: control group(no serum culture medium),model group(PDGF-BB for induction)and intervention group(PDGF-BB+CK for treatment);3.Direct blood count method and CCK-8 method was used to detect the proliferation ability of PASMCs in each group;4.Flow cytometry was used to detect cell cycle and apoptosis of PASMCs;5.Cell scratch test was used to detect the migration ability of PASMCs;6.RT-q PCR and Western blot were used to detect m RNA and protein levels of differentiation marker α-SMA and smooth muscle 22α in PASMCs,respectively;7.ELISA,RT-q PCR,and Western blot were used to detect m RNA and protein expression level and secretion of Collagen type Ⅰ(COL Ⅰ)and Collagen type Ⅲ(COL Ⅲ)of PASMCs.8.Western blot was used to detect the relative protein levels of β-catenin,PGSK-3β,and GSK-3β proteins related to Wnt/β-catenin signaling pathway in PASMCs in each group.Results: 1.A large number of cells could be observed radiating from the edge of the tissue mass at the 7th to 10 th day,with long fusiform cell morphology.Cells close to the tissue mass fuse into sheets that exhibit a typical "peak-valley" pattern.The positive rate of α-SMA staining by immunofluorescence assay was more than 95%,which was suitable for subsequent experimental study.2.Compared with the control group,PDGF-BB significantly induced the proliferation of PASMCs(P<0.05).Compared with model group,CK inhibited the proliferation of PASMCs induced by PDGF-BB,in a concentrationdependent manner(P<0.05),and this inhibition was non-cytotoxic.3.Compared with model group,the cell cycle of PDGF-BB-induced PASMCs was significantly arrested at G0/G1 phase by CK(P<0.05),and CK significantly promoted apoptosis of PDGF-BB-induced PASMCs(P<0.05);4.Compared with the control group,PDGF-BB significantly induced migration of PASMCs(P<0.05);Compared with model group,the migration ability of PASMCs was significantly decreased under CK intervention(P<0.05);5.Compared with the control group,the m RNA and protein expression levels of α-SMA and SM22α in PASMCs were significantly decreased by PDGF-BB(P<0.05);Compared with model group,the m RNA and protein expression levels of α-SMA and SM22α in PASMCs were significantly up-regulated by CK(P<0.05).6.Compared with the control group,the protein level of β-catenin in PDGF-BB-induced PASMCs was significantly increased,and the protein levels of p GSK-3β/GSK-3β were significantly decreased(P<0.05).Compared with model group,the protein level of β-catenin in PASMCs were significantly decreased,and the protein levels of PGSK-3β/GSK-3β were significantly increased by CK treatment(P<0.05).7.Compared with control groups,PDGF-BB significantly promoted the protein synthesis and secretion of COL Ⅰ and COL Ⅲ in the PASMCs(P<0.05).Compared with model groups,CK significantly inhibited the protein synthesis and secretion of COL Ⅰ and COL Ⅲ in the PASMCs(P<0.05).Conclusion: 1.CK inhibited the proliferation,migration,and phenotypic conversion of PDGF-BB-induced PASMCs,and promoted the apoptosis of PDGF-BB-induced PASMCs;CK inhibited the collagen deposition induced by PDGF-BB;3.CK inhibited Wnt/β-catenin signaling,which may be related to its inhibition of proliferation,migration,phenotypic conversion,and collagen deposition.4.CK has the ability to inhibit the biological behavior of PASMCs,suggesting that CK may have the potential ability to inhibit vessel remodeling of PAH.