| The proliferation of vascular smooth muscle cell (VSMC) is the main pathological change of the atherosclerosis (AS) , blood vessel restenosis of percutaneous coronary intervention (PCI)and other diseases. AngiotensinII (AngII) is the important biologically active peptide in renin angiotensin system and it can induce cell to proliferate. The proliferation of VSMC is regulated by all kinds of gene, such as proto-oncogene c-myc and c-fos. The proto-oncogene c-myc, c-fos and c-jun can transcripte and rise their expression level under the stimulation of AngII, growth factor and et.al. And then transcripte all kinds of regulation factors to synthesize and proliferate. Panax Quinquefolium Diolsaponins (PQDS) belongs to panax plant, PQDS can resist myocardial ischemia, obviously deflate myocardial infarction size. At present, there haven't been and report about the effects of PQDS on proliferation of VSMC. This article will study of the effect of PQDS on VSMC proliferation.OBJECTIVETo investigate the effect of PQDS on VSMC proliferation and the expression of the proto-oncogene c-myc, c-fos and c-jun mRNA. To discuss the effect of PQDS inhibiting VSMC proliferation and mechanisms.METHODSRats aorta thoracalis vascular smooth muscle cell be cultured by tissue-sticking method in the experiment. Cells were random divided to: control group,AngII model group,AngII+PQDS groups, among the model+PQDS groups, PQDS divided to 25,50,100mg/L different dosage. In order to make model, 10-7mol/L AngII was used to induce VSMC to proliferate. To determine the cell proliferation ability with MTT, to determine the cell generation cycle and prolifation index with Flow cytometer,to evaluate the expression of c-myc,c-fos and c-jun mRNA with reverse transcription-polymerase chain reaction method.Results1. Compared with blank control group: OD value of AngII model group obviously increased (P<0.05). Compared with AngII model group: OD values of PQDS high and middle dosage groups obviously decreased. PQDS low dosage groups had tendency of decrease (P>0.05). With dosage of PQDS increased, OD value of them decreased. The OD value of PQDS low and middle dosage groups were higher than OD value of blank control group, while the OD value of PQDS high group was lower than it.2. Compared with blank control group: the VSMC percentage in G0/G1 period of AngII model group obviously decreased (P<0.05), it in S period and G2/M period and proliferation index of it obviously increased (P<0.05). Compared with model group: the VSMC percentage in G0/G1 period of all groups of PQDS obviously increased (P<0.05), it in S period and proliferation index of them obviously decreased (P<0.05).The VSMC percentage in G2/M period of PQDS high and middle dosage groups obviously decreased (P<0.05), it in G2/M period of PQDS low dosage group had tendency of decrease (P>0.05). The VSMC percentage in G0/G1 period of PQDS low and middle dosage groups were lower than blank control group and it in G0/G1 period of PQDS high dosage group was higher than blank control group, while the result of S, G2/M period and proliferation index were opposite. Among the groups of PQDS: with dosage of PQDS increased, the VSMC percentage in G0/G1 period of obviously increased (P<0.05), it in S period and proliferation index of them obviously decreased (P<0.05).3. Compared with blank control group: the mRNA expression of c-myc, c-fos and c-jun obviously increased (P<0.05)in AngII group. Compared with model group,the mRNA expression of c-myc and c-jun in all groups of PQDS obviously discreased (P<0.05).The expression of c-fos in PQDS high and middle dosage groups obviously discreased (P<0.05), it in PQDS low dosage group had tendency of decrease (P>0.05). The expression of c-myc and c-fos in the PQDS low and middle group were higher and in the PQDS high group was lower than them in the blank control group. The expression of c-jun in the PQDS low group was higher and in the PQDS high and middle dosage groups were lower than them in the blank control group. With dosage of PQDS increased, the mRNA expression of c-myc, c-fos and c-jun increased gradually decreased. There was statistical significance in the PQDS high group compared with middle and low dosage group in the expression of c-myc and c-jun(P<0.05). There was statistical significance in the PQDS high group compared with low dosage group in the expression of c-fos(P<0.05).DiscussionThe operation of tissue-sticking method is simple, the result is stable, the purity is high, reproducibility is good, it is a commonly used, convenient and high achievement ratio primary culture method.The rat aorta pectoralis vascular smooth muscle cells were cultured by tissue-sticking method in the experiment, After observation under inversion microscop, dyeing of actin, the endochylema were buffy, which confirmed that cultured cells were VSMCs.AngII was used to induce VSMC to proliferate in the experiment. AngII is the important biologically active peptide in renin angiotensin system. The study founds AngII can induce VSMC to proliferate at home and abroad. The results of MTT and FCM indicated that AngII can induce VSMC to proliferate, it differentiate VSMC from G0/G1 period to S and G2/M period, which indicated that the model was succeed made.We found that AngII can up regulate the expression of c-myc, c-fos and c-jun. That means the mechanisms of AngII induced VSMC to proliferate can relate to up-regulating the expression of c-myc, c-fos and c-jun. Proto-oncogene can regulate growth. The mechanisms of c-myc induce cell to proliferate are:â‘ The carboxyl terminus of c-myc can combine with special DNA sequence and open the gene which relate to cell proliferation.â‘¡The amino terminus of c-myc combine with inhibiting gene and relief inhibitory action and induce cell to proliferate.â‘¢Inducing the production of serin-threonine kinase P34cdc2 and make cell to migrate. The proteinum expression product of c-fos and c-jun can form leucine zipper, it can made activator protein-1 (AP-1) with Fos /Jun heterozygosis dimeric format. After combine with special DNA sequence, AP-1 can regulate the expression downstream gene under transcriptional level, regulate cell protein to synthesis and effect cell proliferation and differentiation.PQDS is panax plant, It can resist myocardial ischemia. Compared with AngII model group, OD values of PQDS obviously decreased and all groups of PQDS can prevent cell proliferating from G0/G1 to S period and made proliferation index decreased.the effect of inhibition has a positive correlation with the amount of dosage,all groups of PQDS can down regulate the mRNA expression of c-myc, c-fos and c-jun. it means that the effect of PQDS inhibiting VSMC proliferation implement with down regulate the expression of c-myc, c-fos and c-jun. The study indicate the c-Myc proteinum can synergise with cyclinD1,cyclinE and cyclinA. It make the cells in G1 period pass the restriction point, enter into S period and initiated cell period. The expression of c-fos make cell from G1 period to S period, precipitate cell period to work. PQDS can inhibit VSMC at G0/G1 period that may be relate to down regulating the expression of c-myc and c-fos.Conclusions1. The expression of proto-oncogene c-myc, c-fos and c-jun increse under the effect of AngII, which induce VSMC to proliferate.2. PQDS can inhibit VSMC proliferating, it can inhibit VSMC at G0/G1 period .The effect of inhibition has a positive correlation with the amount of dosage.3. PQDS can down regulate the mRNA expression of c-myc,c-fos and c-jun.4. The meschansims of PQDS inhibiting VSMC proliferation concerned with down regulate the expression of c-myc,c-fos and c-jun.5. PQDS can inhibit VSMC at G0/G1 period which may be relate to down regulating the expression of c-myc and c-fos.
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