Effects Of Insulin Like Growth Factor 1 On Proliferation And Apoptosis Of Vascular Smooth Muscle Cells

Objective:Objective to investigate the effects of insulin like growth factor-1 on proliferation and apoptosis of vascular smooth muscle cells which is in inflammatory conditions.Methods:1.Male SD rats were broken the neck before separation of aorta,removing the intima and adventitia in clean bench,and obtained the primary vascular smooth muscle cells by tissue explant method and enzyme digestion method.4-8 generation cell were quality to the next experiment,and positive expression of alpha actin was identified as smooth muscle cells.2.RAW264.7 cell induced by different concentrations of lipopolysaccharide(LPS)preparated inflammation model and ELISA method was used to detect the level of IL-6in the supernatant of each group,compared with the control group,the difference was statistically significant is successfully made named conditioned medium(CM)and without LPS group as the control group named unconditioned culture medium(nCM).3.Vascular smooth muscle cell co-cultured with nCM、CM、CM+IGF-1 60ng/ml、CM+IGF-1 90ng/ml 、 CM+IGF-1 120ng/ml in vitro and the OD value reflecting the number of VSMC was detected by MTT method,Annex in V-FITC/PI flow cytometry was used to determine the rate of smooth muscle cell apoptosis.4.SPSS17.0 software for statistical analysis,measurement data symbolized as the mean ± standard deviation.T-test was token to compare between the two groups.Analysis of variance was used in multiple groups,P < 0.05 was statistically significant.Results:1.Vascular smooth muscle cells can be successfully obtained by tissue explant method and enzyme digestion method.2.The expression of IL-6 in the supernatant of RAW264.7 cells was significantly increased induced by a certain concentration of LPS.RAW264.7 cell stimulated by0ng/ml、10ng/ml、100ng/ml、1ug/ml、10ug/ml LPS,the concentrations of IL-6 in the supernatant were(6.75±0.12)pg/ml、(7.82±1.53)pg/ml、(44.09±1.58)pg/ml、(155.71±23.93)pg/ml 、(436.59±3.15)pg/ml.Compared with the control group,there was no significant difference in the 10ng/ml group(P=0.916);however there was significant difference between the latter three groups and control group(P=0.009,P=0.000,P=0.000).3.CM could inhibit the proliferation of vascular smooth muscle cells,which could be reverted by addition of IGF-1 at optimal concentration.Vascular smooth muscle cell co-cultured with five different interventions,the OD value were(0.53±0.07)、(0.26±0.06)、(0.30±0.10)、(0.62±0.09)、(0.55±0.05).Compared with nCM group,the OD value of CM group were significantly decreased(P=0.000),the difference between the two groups was statistically significant.Compared with CM group,the OD value of CM+IGF-160ng/ml group,CM+IGF-190ng/ml group and CM+IGF-1120ng/ml group was increased(P=0.993,P=0.001,P=0.000),the latter two groups have statistical significance difference group 90ng/ml had the best effect.4.CM can promote the apoptosis of vascular smooth muscle cells,which could be reverted by addition of IGF-1 at optimal concentration.Vascular smooth muscle cell co-cultured with five different interventions,the apoptosis rate were(4.89±0.09)%;(10.86±0.15)%;(9.64±0.65)%;(3.85±0.51)%;(4.82±0.08)%.Compared with nCM group,the apoptosis of CM group were significantly increased(P=0.000),the difference between the two groups was statistically significant.Compared with CM group,the apoptosis of CM+IGF-160ng/ml group,CM+IGF-190ng/ml group and CM+IGF-1120ng/ml group was decreased(P=0.045,P=0.000,P=0.000),the difference have statistical sense.Group 90ng/ml had the best effect.Conclusion:LPS could induce RAW264.7 cell polarization into type M1 macrophages.The inflammatory state induced by type M1 macrophages could promote the apoptosis of vascular smooth muscle cells and inhibit the proliferation of vascular smooth muscle cells.Given a certain concentration of IGF-1,the apoptosis was decreased and the proliferation was increased.