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Matrine inhibits vascular smooth muscle cell proliferation by modulating the expression of cell cycle regulatory genes.

Zhu P, Chen JM, Chen SZ, Zhang C, Zheng SY, Long G, Chen J, Zhou ZL, Fan RX, Fan XP, Chen YF, Zhuang J - Acta Pharmacol. Sin. (2010)

Bottom Line: Matrine did not affect p27 expression.Furthermore, the anti-proliferative effect of matrine was abolished by silencing of p21, but not by silencing of p27.Our data indicate that matrine has an inhibitory effect on VSMC proliferation via up-regulation of the p53/p21 signaling pathway and modulation of other cell cycle regulatory genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiovascular Surgery, Guangdong Cardiovascular Institute, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou, China.

ABSTRACT

Aim: To investigate the effect of matrine on proliferation of vascular smooth muscle cells (VSMCs) and elucidate the underlying mechanisms.

Methods: Rat aortic VSMCs were cultured in medium supplemented with 10% fetal bovine serum and treated with various concentrations (0, 5, 10, 15, and 20 mg/L) of matrine for 72 h. VSMCs proliferation and cell cycle profiling were assessed using a methylene blue incorporation assay and flow cytometry, respectively. The underlying protein signaling mechanisms were determined using Western blot analysis of the expression levels of cell cycle regulatory genes, including p53, p21, p27, cyclin D1, cyclin E, cyclin-dependent kinase 2 and 4 (cdk2, cdk4), and phosphorylated Rb. The involvement of p21 and p27 pathways was further determined using small interfering RNA (siRNA) knockdown.

Results: Matrine inhibited VSMC proliferation in a dose-dependent manner by promoting G(1) arrest. The G(1) arrest was accompanied by up-regulation of p53 and p21 protein levels, and down-regulation of cyclin D1/cdk4, cyclin E/cdk2 and phosphorylated Rb protein levels. Matrine did not affect p27 expression. Furthermore, the anti-proliferative effect of matrine was abolished by silencing of p21, but not by silencing of p27.

Conclusion: Our data indicate that matrine has an inhibitory effect on VSMC proliferation via up-regulation of the p53/p21 signaling pathway and modulation of other cell cycle regulatory genes.

Show MeSH
(A) Characterization of cultured rat aortic VSMCs (3th passage, 400×) by immunohistochemical staining with anti-α-smooth muscle actin monoclonal antibody; (B) Effect of matrine on VSMC proliferation in microwell plates. VSMCs were treated with various concentrations of matrine (0, 5, 10, 15, and 20 mg/L) for 72 h. Rapamycin (100 ng/L) was as a positive control. Cell growth was determined by colorimetric assay of methylene blue incorporation. The changes in percentage of control (0 mg/L matrine) were calculated. Data are expressed as mean±SEM. n=5/group, One-way ANOVA. bP<0.05, cP<0.01 vs control.
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fig1: (A) Characterization of cultured rat aortic VSMCs (3th passage, 400×) by immunohistochemical staining with anti-α-smooth muscle actin monoclonal antibody; (B) Effect of matrine on VSMC proliferation in microwell plates. VSMCs were treated with various concentrations of matrine (0, 5, 10, 15, and 20 mg/L) for 72 h. Rapamycin (100 ng/L) was as a positive control. Cell growth was determined by colorimetric assay of methylene blue incorporation. The changes in percentage of control (0 mg/L matrine) were calculated. Data are expressed as mean±SEM. n=5/group, One-way ANOVA. bP<0.05, cP<0.01 vs control.

Mentions: Rat aortic VSMCs were isolated from aortic explants. The purity of VSMCs was assessed via immunostaining for smooth muscle α-actin (Figure 1A). Primary cultures consisting of <95% VSMCs were discarded. Cellular viability was determined by trypan blue exclusion.


Matrine inhibits vascular smooth muscle cell proliferation by modulating the expression of cell cycle regulatory genes.

Zhu P, Chen JM, Chen SZ, Zhang C, Zheng SY, Long G, Chen J, Zhou ZL, Fan RX, Fan XP, Chen YF, Zhuang J - Acta Pharmacol. Sin. (2010)

(A) Characterization of cultured rat aortic VSMCs (3th passage, 400×) by immunohistochemical staining with anti-α-smooth muscle actin monoclonal antibody; (B) Effect of matrine on VSMC proliferation in microwell plates. VSMCs were treated with various concentrations of matrine (0, 5, 10, 15, and 20 mg/L) for 72 h. Rapamycin (100 ng/L) was as a positive control. Cell growth was determined by colorimetric assay of methylene blue incorporation. The changes in percentage of control (0 mg/L matrine) were calculated. Data are expressed as mean±SEM. n=5/group, One-way ANOVA. bP<0.05, cP<0.01 vs control.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4012916&req=5

fig1: (A) Characterization of cultured rat aortic VSMCs (3th passage, 400×) by immunohistochemical staining with anti-α-smooth muscle actin monoclonal antibody; (B) Effect of matrine on VSMC proliferation in microwell plates. VSMCs were treated with various concentrations of matrine (0, 5, 10, 15, and 20 mg/L) for 72 h. Rapamycin (100 ng/L) was as a positive control. Cell growth was determined by colorimetric assay of methylene blue incorporation. The changes in percentage of control (0 mg/L matrine) were calculated. Data are expressed as mean±SEM. n=5/group, One-way ANOVA. bP<0.05, cP<0.01 vs control.
Mentions: Rat aortic VSMCs were isolated from aortic explants. The purity of VSMCs was assessed via immunostaining for smooth muscle α-actin (Figure 1A). Primary cultures consisting of <95% VSMCs were discarded. Cellular viability was determined by trypan blue exclusion.

Bottom Line: Matrine did not affect p27 expression.Furthermore, the anti-proliferative effect of matrine was abolished by silencing of p21, but not by silencing of p27.Our data indicate that matrine has an inhibitory effect on VSMC proliferation via up-regulation of the p53/p21 signaling pathway and modulation of other cell cycle regulatory genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiovascular Surgery, Guangdong Cardiovascular Institute, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou, China.

ABSTRACT

Aim: To investigate the effect of matrine on proliferation of vascular smooth muscle cells (VSMCs) and elucidate the underlying mechanisms.

Methods: Rat aortic VSMCs were cultured in medium supplemented with 10% fetal bovine serum and treated with various concentrations (0, 5, 10, 15, and 20 mg/L) of matrine for 72 h. VSMCs proliferation and cell cycle profiling were assessed using a methylene blue incorporation assay and flow cytometry, respectively. The underlying protein signaling mechanisms were determined using Western blot analysis of the expression levels of cell cycle regulatory genes, including p53, p21, p27, cyclin D1, cyclin E, cyclin-dependent kinase 2 and 4 (cdk2, cdk4), and phosphorylated Rb. The involvement of p21 and p27 pathways was further determined using small interfering RNA (siRNA) knockdown.

Results: Matrine inhibited VSMC proliferation in a dose-dependent manner by promoting G(1) arrest. The G(1) arrest was accompanied by up-regulation of p53 and p21 protein levels, and down-regulation of cyclin D1/cdk4, cyclin E/cdk2 and phosphorylated Rb protein levels. Matrine did not affect p27 expression. Furthermore, the anti-proliferative effect of matrine was abolished by silencing of p21, but not by silencing of p27.

Conclusion: Our data indicate that matrine has an inhibitory effect on VSMC proliferation via up-regulation of the p53/p21 signaling pathway and modulation of other cell cycle regulatory genes.

Show MeSH