Abstract
Aim:
To explore the effect of neferine on angiotensin II (Ang II)-induced vascular smooth muscle cell (VSMC) proliferation.
Methods:
Human umbilical vein smooth muscle cells (HUVSMCs) were used. Cell proliferation was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry analysis. Heme oxygenase (HO)-1 protein expression was tested by Western blot analysis. Extracellular signal-regulated protein kinase 1/2 (ERK1/2) activation was determined by using immunoblotting.
Results:
Pre-incubation of HUVSMCs with neferine (0.1, 0.5, 1.0, and 5.0 μmol/L) significantly inhibited Ang II-induced cell proliferation in a concentration-dependent manner and neferine 5.0 μmol/L increased HO-1 expression by 259% compared with control. The antiproliferative effect of neferine was significantly attenuated by coapplication of zinc protoporphyrin IX (ZnPP IX, an HO-1 inhibitor) with neferine. Ang II-enhanced ERK1/2 phosphorylation was markedly reversed by neferine. By inhibiting HO-1 activity with ZnPP IX, the inhibitive effect of neferine on ERK1/2 phosphorylation was significantly attenuated. Cobalt-protoporphyrin (CoPP), an HO-1 inducer, significantly decreased Ang II-induced ERK1/2 phosphorylation and inhibited Ang II-induced cell proliferation. The ERK1/2 pathway inhibitor PD98059 significantly blocked Ang II-enhanced ERK1/2 phosphorylation and inhibited cell proliferation.
Conclusion:
These findings suggest that neferine can inhibit Ang II-induced HUVSMC proliferation by upregulating HO-1, leading to the at least partial downregulation of ERK1/2 phosphorylation.
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Abbreviations
- MTT:
-
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
- CoPP:
-
cobalt-protoporphyrin
- ERK:
-
extracellular signal-regulated kinase
- HO:
-
heme oxygenase
- PD98059:
-
2'-amino-3'-methoxyflavone
- MAPK:
-
mitogen-activated protein kinase
- MEK:
-
MAP kinase
- ZnPP:
-
zinc protoporphyrin
- GAPDH:
-
glyceraldehyde 3-phosphate dehydrogenase
- Ang:
-
angiotensin
- HUVSMC:
-
human umbilical vascular smooth muscle cell
- DMEM:
-
Dulbecco's modified Eagle's medium
- ROS:
-
reactive oxygen species
- Nef:
-
neferine
- JNK:
-
Jun N-terminal kinase
- SnPP:
-
tin protoporphyrin
- EGFR:
-
epidermal growth factor receptor
- PDGF:
-
platelet-derived growth factor
- cdk-2:
-
cyclin-dependent kinase-2
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Acknowledgements
This study was supported by the Administration of Traditional Chinese Medicine of Zhejiang Province (No 2007CA112).
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Li, Xc., Tong, Gx., Zhang, Y. et al. Neferine inhibits angiotensin II-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1. Acta Pharmacol Sin 31, 679–686 (2010). https://doi.org/10.1038/aps.2010.57
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DOI: https://doi.org/10.1038/aps.2010.57
