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Title: miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine

Abstract

MicroRNAs (miRNAs) are short non-coding RNA and play crucial roles in a wide array of biological processes, including cell proliferation, differentiation and apoptosis. Our previous studies found that homocysteine(Hcy) can stimulate the proliferation of vascular smooth muscle cells (VSMCs), however, the underlying mechanisms were not fully elucidated. Here, we found proliferation of VSMCs induced by Hcy was of correspondence to the miR-125b expression reduced both in vitro and in the ApoE knockout mice, the hypermethylation of p53, its decreased expression, and DNA (cytosine-5)-methyltransferase 3b (DNMT3b) up-regulated. And, we found DNMT3b is a target of miR-125b, which was verified by the Dual-Luciferase reporter assay and western blotting. Besides, the siRNA interference for DNMT3b significantly decreased the methylation level of p53, which unveiled the causative role of DNMT3b in p53 hypermethylation. miR-125b transfection further confirmed its regulative roles on p53 gene methylation status and the VSMCs proliferation. Our data suggested that a miR-125b-DNMT3b-p53 signal pathway may exist in the VSMCs proliferation induced by Hcy.

Authors:
 [1];  [2];  [3];  [4]; ;  [1];  [5];  [3]; ;  [1];  [5];  [1];  [5]
  1. Key Laboratory of Basic Research in Cardio-Cerebral Vascular Diseases, Ningxia Medical University, Yinchuan (China)
  2. Department of Prenatal Diagnosis Center, General Hospital of Ningxia Medical University, Yinchuan (China)
  3. Department of Medical Laboratory, Ningxia Medical University, Yinchuan (China)
  4. Department of Basic Medicine, Ningxia Medical University, Yinchuan (China)
  5. (China)
Publication Date:
OSTI Identifier:
22649755
Resource Type:
Journal Article
Resource Relation:
Journal Name: Experimental Cell Research; Journal Volume: 347; Journal Issue: 1; Other Information: Copyright (c) 2016 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; APOPTOSIS; ARTERIOSCLEROSIS; CELL PROLIFERATION; CYTOSINE; DNA; FREE ENERGY; GENES; HOMOCYSTEINE; IN VITRO; KNOCK-OUT REACTIONS; LUCIFERASE; METHYL TRANSFERASES; METHYLATION; MICE; MUSCLES; POLYMERASE CHAIN REACTION; RNA

Citation Formats

Cao, ChengJian, Zhang, HuiPing, Zhao, Li, Zhou, Longxia, Zhang, Minghao, Xu, Hua, Department of Basic Medicine, Ningxia Medical University, Yinchuan, Han, Xuebo, Li, Guizhong, Yang, Xiaoling, Department of Basic Medicine, Ningxia Medical University, Yinchuan, Jiang, YiDeng, E-mail: jyjcyxy@yeah.net, and Department of Basic Medicine, Ningxia Medical University, Yinchuan. miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine. United States: N. p., 2016. Web. doi:10.1016/J.YEXCR.2016.07.007.
Cao, ChengJian, Zhang, HuiPing, Zhao, Li, Zhou, Longxia, Zhang, Minghao, Xu, Hua, Department of Basic Medicine, Ningxia Medical University, Yinchuan, Han, Xuebo, Li, Guizhong, Yang, Xiaoling, Department of Basic Medicine, Ningxia Medical University, Yinchuan, Jiang, YiDeng, E-mail: jyjcyxy@yeah.net, & Department of Basic Medicine, Ningxia Medical University, Yinchuan. miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine. United States. doi:10.1016/J.YEXCR.2016.07.007.
Cao, ChengJian, Zhang, HuiPing, Zhao, Li, Zhou, Longxia, Zhang, Minghao, Xu, Hua, Department of Basic Medicine, Ningxia Medical University, Yinchuan, Han, Xuebo, Li, Guizhong, Yang, Xiaoling, Department of Basic Medicine, Ningxia Medical University, Yinchuan, Jiang, YiDeng, E-mail: jyjcyxy@yeah.net, and Department of Basic Medicine, Ningxia Medical University, Yinchuan. 2016. "miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine". United States. doi:10.1016/J.YEXCR.2016.07.007.
@article{osti_22649755,
title = {miR-125b targets DNMT3b and mediates p53 DNA methylation involving in the vascular smooth muscle cells proliferation induced by homocysteine},
author = {Cao, ChengJian and Zhang, HuiPing and Zhao, Li and Zhou, Longxia and Zhang, Minghao and Xu, Hua and Department of Basic Medicine, Ningxia Medical University, Yinchuan and Han, Xuebo and Li, Guizhong and Yang, Xiaoling and Department of Basic Medicine, Ningxia Medical University, Yinchuan and Jiang, YiDeng, E-mail: jyjcyxy@yeah.net and Department of Basic Medicine, Ningxia Medical University, Yinchuan},
abstractNote = {MicroRNAs (miRNAs) are short non-coding RNA and play crucial roles in a wide array of biological processes, including cell proliferation, differentiation and apoptosis. Our previous studies found that homocysteine(Hcy) can stimulate the proliferation of vascular smooth muscle cells (VSMCs), however, the underlying mechanisms were not fully elucidated. Here, we found proliferation of VSMCs induced by Hcy was of correspondence to the miR-125b expression reduced both in vitro and in the ApoE knockout mice, the hypermethylation of p53, its decreased expression, and DNA (cytosine-5)-methyltransferase 3b (DNMT3b) up-regulated. And, we found DNMT3b is a target of miR-125b, which was verified by the Dual-Luciferase reporter assay and western blotting. Besides, the siRNA interference for DNMT3b significantly decreased the methylation level of p53, which unveiled the causative role of DNMT3b in p53 hypermethylation. miR-125b transfection further confirmed its regulative roles on p53 gene methylation status and the VSMCs proliferation. Our data suggested that a miR-125b-DNMT3b-p53 signal pathway may exist in the VSMCs proliferation induced by Hcy.},
doi = {10.1016/J.YEXCR.2016.07.007},
journal = {Experimental Cell Research},
number = 1,
volume = 347,
place = {United States},
year = 2016,
month = 9
}
  • Hyperhomocysteinemia induces the proliferation of vascular smooth muscle cells (VSMCs). Hydrogen sulfide (H{sub 2}S) inhibits the phenotype switch of VSMCs and calcium-sensing receptor (CaSR) regulated the production of endogenous H{sub 2}S. However, whether CaSR inhibits the proliferation of VSMCs by regulating the endogenous cystathionine-gamma-lyase (CSE, a major enzyme that produces H{sub 2}S) pathway in high homocysteine (HHcy) has not been previously investigated. The intracellular calcium concentration, the concentration of H{sub 2}S, the cell viability, the proliferation and the expression of proteins of cultured VSMCs from rat thoracic aortas were measured, respectively. The results showed that the [Ca{sup 2+}]{sub i} andmore » the expression of p-CaMK and CSE increased upon treatment with CaSR agonist. In HHcy, the H{sub 2}S concentration decrease, the proliferation and migration rate increased, the expression of Cyclin D1, PCNA, Osteopontin and p-Erk1/2 increased while the α-SM actin, P21{sup Cip/WAK−1} and Calponin decreased. The CaSR agonist or exogenous H{sub 2}S significantly reversed the changes of VSMCs caused by HHcy. In conclusion, our results demonstrated that CaSR regulate the endogenous CSE/H{sub 2}S is related to the PLC-IP{sub 3} receptor and CaM signal pathways which inhibit the proliferation of VSMCs, and the latter is involved in the Erk1/2 dependent signal pathway in high homocysteine. - Highlights: • CaSR activation increased the production of endogenous H{sub 2}S in high homocysteine VSMCs. • CaSR modulated the CSE/H{sub 2}S are related to the PLC-IP{sub 3}R and Ca{sup 2+}-CaM signal pathways. • Inhibition of H{sub 2}S on the proliferation of VSMCs is involved in the Erk1/2 pathway. • Explore the potential roles of CaSR in regulating VSMCs proliferation in high homocysteine.« less
  • Platelet-derived growth factor (PDGF) is known to induce phenotypic switching of vascular smooth muscle cells (VSMCs) from contractile to a pathological synthetic state, which played an essential role in proliferation of VSMCs. Sonic hedgehog (Shh) contributes to the proliferation of VSMCs when induced by PDGF. Here, we investigated the probable role of Shh in PDGF-induced VSMC dedifferentiation and its underlying mechanisms. We found that PDGF stimulated Shh expression in VSMCs, which was mediated by activation of PDGFRβ/ERK1/2 cell signaling pathway. Further, we found PDGF-induced VSMC phenotypic modulation was accompanied by up-regulation of Shh/Gli family zinc finger 2 (Gli2) signaling andmore » Krüppel-like factor 4 (KLF4). When inhibited Shh in the presence of PDGF, the expressions of KLF4 and VSMC dedifferentiation markers were down-regulated and the effect of PDGF in inducing VSMC dedifferentiation was blocked. In the absence of PDGF, Shh signaling activation increased the expression of KLF4 and promoted VSMC dedifferentiation. The results indicate Shh participated in the regulation of PDGF-induced VSMC dedifferentiation. Finally, we found that KLF4 was closely involved in this process. On inhibition of KLF4, PDGF induced VSMC dedifferentiation was abrogated, even in the presence of Shh. Taken together, the results provide critical insights into the newly discovered role of Shh in phenotypic modulation of VSMCs which depends on KLF4. - Highlights: • Shh as a downstream effector of PDGF participates in PDGF-induced VSMC phenotypic modulation. • Shh can promote VSMC phenotypic switching from contractile to synthetic state. • Shh mediates VSMC phenotypic modulation through regulation of KLF4.« less
  • Highlights: •The mechanism underlying the effects of Apelin-13 on VSMC was investigated. •Apelin-13 induced VSMC migration, proliferation and Egr-1 and OPN upregulation. •These effects were inhibited by the Egr-1 specific deoxyribozyme, ED5. •The effects of Apelin-13 on VSMC are mediated via Egr-1 upregulation. •These data will help in attempts to prevent and treat vascular remodeling diseases. -- Abstract: Apelin-13 plays an important role in the migration and proliferation of vascular smooth muscle cells (VSMCs); however, the underlying mechanisms are still unclear. Egr-1 is a nuclear transcription factor, which is considered to be the critical initiating factor of the processes ofmore » VSMC proliferation and migration. Egr-1 is known to regulate the expression of osteopontin (OPN), which is a marker of the phenotypic modulation that is a necessary condition of VSMC proliferation and migration. We hypothesized that the role of Apelin-13 is mediated via upregulation of Egr-1. To test this hypothesis, we analyzed the effects of Apelin-13 treatment on Egr-1 mRNA and protein expression in A10 rat aortic VSMCs by RT-PCR and Western blotting, respectively. Results showed that, Apelin-13 upregulated the expression of Egr-1. Furthermore, treatment with the extracellular-regulated protein kinase (ERK) inhibitor, PD98059, inhibited the upregulation of Egr-1 by Apelin-13. In addition, this upregulation was inhibited by treatment of VSMCs with the Egr-1 specific deoxyribozyme ED5 (DNAenzyme/10-23 DRz). Furthermore, ED5 treatment was found to significantly inhibit Apelin-13-induced migration and proliferation of VSMCs using transwell and MTT assays, respectively. The evaluation of OPN mRNA and protein expression levels by RT-PCR and Western blot analyses revealed that ED5 treatment also inhibited Apelin-13-induced OPN upregulation. The results of this study indicated that Apelin-13 upregulates Egr-1 via ERK. Furthermore, Apelin-13 induced the proliferation and migration of VSMCs as well as the upregulation of OPN via the upregulation of Egr-1. These results will provide an important theoretical and experimental basis for the control of inappropriate remodeling of vessel walls, and will hopefully lead to the prevention and treatment of vascular remodeling diseases.« less
  • AngiotensinII (AngII) induces vascular smooth muscle cell (VSMC) proliferation, which plays an important role in the development and progression of hypertension. AngII-induced cellular events have been implicated, in part, in the activation of protein kinase C (PKC) and extracellular signal-regulated kinases 1/2 (ERK1/2). In the present study, we investigated the effect of Ib, a novel nonpeptide AngII receptor type 1 (AT{sub 1}) antagonist, on the activation of PKC and ERK1/2 in VSMC proliferation induced by AngII. MTT, and [{sup 3}H]thymidine incorporation assay showed that AngII-induced VSMC proliferation was inhibited significantly by Ib. The specific binding of [{sup 125}I]AngII to AT{submore » 1} receptors was blocked by Ib in a concentration-dependent manner with IC{sub 50} value of 0.96 nM. PKC activity assay and Western blot analysis demonstrated that Ib significantly inhibited the activation of PKC and phosphorylation of ERK1/2 induced by AngII, respectively. Furthermore, AngII-induced ERK1/2 activation was obviously blocked by GF109203X, a PKC inhibitor. These findings suggest that the suppression of Ib on AngII-induced VSMC proliferation may be attributed to its inhibitory effect on PKC-dependent ERK1/2 pathway.« less
  • Proliferation of vascular smooth muscle cells (VSMCs) contributes to the development of various cardiovascular diseases. Curcumin, extracted from Curcumae longae, has been shown a variety of beneficial effects on human health, including anti-atherosclerosis by mechanisms poorly understood. In the present study, we attempted to investigate whether curcumin has any effect on VSMCs proliferation and the potential mechanisms involved. Our data showed curcumin concentration-dependently abrogated the proliferation of primary rat VSMCs induced by Chol:M{beta}CD. To explore the underlying cellular and molecular mechanisms, we found that curcumin was capable of restoring caveolin-1 expression which was reduced by Chol:M{beta}CD treatment. Moreover, curcumin abrogatedmore » the increment of phospho-ERK1/2 and nuclear accumulation of ERK1/2 in primary rat VSMCs induced by Chol:M{beta}CD, which led to a suppression of AP-1 promoter activity stimulated by Chol:M{beta}CD. In addition, curcumin was able to reverse cell cycle progression induced by Chol:M{beta}CD, which was further supported by its down-regulation of cyclinD1 and E2F promoter activities in the presence of Chol:M{beta}CD. Taking together, our data suggest curcumin inhibits Chol:M{beta}CD-induced VSMCs proliferation via restoring caveolin-1 expression that leads to the suppression of over-activated ERK signaling and causes cell cycle arrest at G1/S phase. These novel findings support the beneficial potential of curcumin in cardiovascular disease.« less