BMP7过表达慢病毒载体构建及其对小鼠主动脉平滑肌细胞钙化的影响

付仕林; 易雪娇; 潘文旭; 尹纯; 康华利; 钱德慧

doi:10.11659/jjssx.10E023055

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BMP7过表达慢病毒载体构建及其对小鼠主动脉平滑肌细胞钙化的影响

基础研究 | 更新时间：2024-02-02

- BMP7过表达慢病毒载体构建及其对小鼠主动脉平滑肌细胞钙化的影响
- BMP7 overexpression lentiviral vector construction and its effect on calcification of mouse aortic smooth muscle cells
- 局解手术学杂志 2024年33卷第2期页码：95-99
- 作者机构：
  
  陆军军医大学第二附属医院心血管内科，重庆 400037
- 作者简介：
  
  钱德慧，E-mail：qiandehui@163.com
- 基金信息：
  
  重庆市自然科学基金面上项目(CSTB2023NSCQ-MSX0686);陆军军医大学医院青年博士人才孵化计划项目(2022YQB029)
- DOI：10.11659/jjssx.10E023055
  中图分类号： R34;R329.2
- 纸质出版日期：2024-02-25，
  
  收稿日期：2023-10-13，
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付仕林,易雪娇,潘文旭等.BMP7过表达慢病毒载体构建及其对小鼠主动脉平滑肌细胞钙化的影响[J].局解手术学杂志,2024,33(02):95-99.

FU Shi-lin,YI Xue-jiao,PAN Wen-xu,et al.BMP7 overexpression lentiviral vector construction and its effect on calcification of mouse aortic smooth muscle cells[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2024,33(02):95-99.
付仕林,易雪娇,潘文旭等.BMP7过表达慢病毒载体构建及其对小鼠主动脉平滑肌细胞钙化的影响[J].局解手术学杂志,2024,33(02):95-99. DOI： 10.11659/jjssx.10E023055.

FU Shi-lin,YI Xue-jiao,PAN Wen-xu,et al.BMP7 overexpression lentiviral vector construction and its effect on calcification of mouse aortic smooth muscle cells[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2024,33(02):95-99. DOI： 10.11659/jjssx.10E023055.

摘要

目的

构建骨形态发生蛋白7（BMP7）过表达慢病毒载体，分析BMP7过表达对小鼠主动脉内皮细胞Jagged1表达的影响及其对共培养血管平滑肌细胞（VSMCs）钙化的影响。

方法

根据目的基因信息Mouse-BMP7（NM_007557.3）和质粒信息pLVX-zsGreen-C1进行基因序列合成，构建BMP7过表达慢病毒。qPCR检测BMP7过表达慢病毒感染效率；Western blot检测其感染的小鼠主动脉内皮细胞Jagged1蛋白表达。将过表达BMP7慢病毒感染的内皮细胞与VSMCs共培养，茜素红染色观察共培养后VSMCs钙化情况。

结果

成功构建BMP7过表达慢病毒载体并转染至小鼠主动脉内皮细胞。qPCR检测结果提示，与正常对照组比较，BMP7过表达组的BMP7 mRNA相对表达量显著升高（

0.01），空载体对照组BMP7 mRNA无显著变化（

0.05）。Western blot结果显示，BMP7过表达组内皮细胞Jagged1蛋白表达水平较正常对照组显著降低（

0.01），而空载体对照组内皮细胞Jagged1蛋白水平较正常对照组无显著改变（

0.05）。茜素红染色结果提示，与感染过表达BMP7慢病毒的内皮细胞共培养后，VSMCs的钙化程度明显增加。

结论

成功构建小鼠BMP7过表达慢病毒载体，并发现过表达BMP7可以降低小鼠主动脉内皮细胞Jagged1表达，并促进共培养的VSMCs钙化。

Abstract

Objective

To construct a lentiviral vector for overexpression of bone morphogenetic protein 7 （BMP7） in mice，and the effect of BMP7 overexpression on the expression of Jagged1 in mouse aortic endothelial cells and the calcification of the co-cultured vascular smooth muscle cells（VSMCs） were analyzed.

Methods

According to the target gene information Mouse-BMP7（NM_007557.3） and plasmid information pLVX-zsGreen-C1，gene sequence synthesis was carried out to construct BMP7 overexpression lentivirus. The efficiency of BMP7 overexpression lentivirus infection was detected by qPCR；the expression of Jagged1 protein in aortic endothelial cells from infected mice was detected by Western blot. The endothelial cells with lentivirus overexpressing BMP7 were co-cultured with VSMCs，and the calcification of VSMCs was observed by alizarin red staining.

Results

BMP7 overexpression lentiviral vector was successfully constructed and transfected into aortic endothelial cells.qPCR test results showed that the expression level of BMP7 mRNA was significantly increased in the BMP7 overexpression group than that in the normal control group （

0.01），while there was no significant difference in the expression of BMP7 mRNA between the empty vector control group and the normal control group （

0.05）. Western blot results showed that the expression level of Jagged1 protein in endothelial cells of mouse in the BMP7 overexpression group was significantly lower than that in the normal control group （

0.01），while there was no significant difference in the expression level of Jagged1 protein in endothelial cells between the empty vector control group and the normal control group （

0.05）. The results of alizarin red staining showed that the calcification of VSMCs was significantly increased after co-cultured with endothelial cells infected with BMP7 lentivirus.

Conclusion

Mouse BMP7 overexpression lentiviral vector was successfully constructed，and overexpression of BMP7 can reduce the expression of Jagged1 in mouse aortic endothelial cells and promote the calcification of co-cultured VSMCs.

关键词

骨形态发生蛋白7慢病毒载体构建Jagged1血管钙化

Keywords

bone morphological protein 7lentiviral vector constructionJagged1vascular calcification

references

Hill JM，Kereiakes DJ，Shlofmitz RA，et al.Intravascular lithotripsy for treatment of severely calcified coronary artery disease［J］.J Am Coll Cardiol，2020，76（22）：2635-2646.doi：10.1016/j.jacc. 2020. 09.603http://dx.doi.org/10.1016/j.jacc.2020.09.603.

张玉霞，汤日宁，刘必成. 慢性肾脏病患者血管钙化发生机制研究进展［J］.中华肾脏病杂志，2021，37（9）：779-784.doi：10.3760/cma.j.cn441217-20200817-00115http://dx.doi.org/10.3760/cma.j.cn441217-20200817-00115.

Furmanik M，Chatrou M，van Gorp R，et al.Reactive oxygen-forming Nox5 links vascular smooth musclecell phenotypic switching and extracellular vesicle-mediated vascular calcification［J］.Circ Res，2020，127（7）：911-927.doi：10.1161/CIRCRESAHA.119.316159http://dx.doi.org/10.1161/CIRCRESAHA.119.316159.

Wu X，Zou Y，Zhou Q，et al. Role of Jagged1 in arterial lesions after vascular injury［J］. Arterioscler Thromb Vasc Biol，2011，31：2000-2006.doi：10.1161/ATVBAHA.111.225144http://dx.doi.org/10.1161/ATVBAHA.111.225144.

Cao G，Xuan X，Hu J，et al. How vascular smooth muscle cell phenotype switching contributes to vascular disease［J］. Cell Commun Signal，2022，20（1）：180. doi：10.1186/s12964-022-00993-2http://dx.doi.org/10.1186/s12964-022-00993-2.

Li X，Li K，Yu G，et al. miR-342-5p inhibits expression of Bmp7 to regulate proliferation，differentiation and migration of osteoblasts［J］. Mol Immunol，2019，114：251-259.doi：10.1016/j.molimm. 2019.07.027http://dx.doi.org/10.1016/j.molimm.2019.07.027.

Kobayashi M，Inoue K，Warabi E，et al. A simple method of isolating mouse aortic endothelial cells［J］.J Atheroscler Thromb，2005，12（3）：138-142. doi：10.5551/jat.12.138http://dx.doi.org/10.5551/jat.12.138.

郝思雨，田江天，刘洋，等.小鼠主动脉平滑肌细胞的分离培养及鉴定［J］.现代生物医学进展，2016，16（27）：5237-5240. doi：10.13241/j.cnki.pmb.2016.27.009http://dx.doi.org/10.13241/j.cnki.pmb.2016.27.009.

Fillinger MF，Sampson LN，Cronenwett JL，et al. Coculture of endothelial cells and smooth muscle cells in bilayer and conditioned media models［J］.J Surg Res，1997，67（2）：169-178. doi：10.1006/jsre. 1996.4978http://dx.doi.org/10.1006/jsre.1996.4978.

Leopold JA. Vascular calcifification： mechanisms of vascular smooth muscle cell calcifification［J］. Trends Cardiovasc Med，2015，25（4）： 267-274.doi：10.1016/j.tcm.2014.10.021http://dx.doi.org/10.1016/j.tcm.2014.10.021.

Shi LL，Hao M，Jin ZY，et al. Liraglutide alleviates diabetic atherosclerosis through regulating calcification of vascular smooth muscle cells［J］. Dis Markers，2022，25：5013622.doi：10.1155/2022/5013622http://dx.doi.org/10.1155/2022/5013622.

Yang P，Troncone L，Augur ZM，et al.The role of bone morphogenetic protein signaling in vascular calcification［J］. Bone，2020，141：115542.doi：10.1016/j.bone.2020.115542http://dx.doi.org/10.1016/j.bone.2020.115542.

Oštrić M，Kukuljan M，Markić D，et al. Expression of bone-related proteins in vascular calcification and its serum correlations with coronary artery calcification score［J］. J Biol Regul Homeost Agents，2019，33（1）：29-38.

Liu H，Wang H，Yang S，et al.Downregulation of miR-542-3p promotes osteogenic transition of vascular smooth muscle cells in the aging rat by targeting BMP7［J］. Human Genomics，2019，13（1）：67.doi：10.1186/s40246-019-0245-zhttp://dx.doi.org/10.1186/s40246-019-0245-z.

Baeten JT，Lilly B. Notch signaling in vascular smooth muscle cells［J］. Adv Pharmacol，2017，78：351-382. doi：10.1016/bs.apha. 2016.07.002http://dx.doi.org/10.1016/bs.apha.2016.07.002.

张清，向明，陈杭，等.Notch信号通路在骨关节炎软骨细胞中的表达及意义［J］.局解手术学杂志，2018，27（4）：240-245.doi：10.11659/jjssx.08E017010http://dx.doi.org/10.11659/jjssx.08E017010.

Mao C，Ma Z，Jia Y，et al. Nidogen-2 maintains the contractile phenotype of vascular smooth muscle cells and prevents neointima formation via bridging Jagged1-Notch3 signaling［J］. Circulation，2021，144（15）：1244-1261.doi：10.1161/CIRCULATIONAHA. 120.053361http://dx.doi.org/10.1161/CIRCULATIONAHA.120.053361.

Zhang Y，Zhang J，Navrazhina K，et al.TGF beta1 induces Jagged1 expression in astrocytes via ALK5 and Smad3 and regulates the balance between oligodendrocyte progenitor proliferation and differentiation［J］. Glia，2010，58：964-974.doi：10.1002/glia.20978http://dx.doi.org/10.1002/glia.20978.

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