沉默LncRNA H19通过调节神经生长因子抑制心肌梗死后交感神经重构

doi:10.6040/j.issn.1671-7554.0.2020.1534

摘要/Abstract

摘要： 目的探讨H19与神经生长因子(NGF)在心肌梗死(MI)后室性快速性心律失常(VAs)发生过程中的作用与调控关系。方法采用左冠状动脉结扎法建立大鼠MI模型。(1)将14只健康SD大鼠分为MI组和假手术组(sham组),每组7只,采用qRT-PCR法检测MI后7d长链非编码RNA H19(LncRNA H19)的表达;(2)将32只大鼠随机分为假手术+对照病毒组(sham+NC组)、假手术+沉默病毒组(sham+siH19组)、MI+对照病毒组(MI+NC组)、MI+沉默病毒组(MI+siH19组),每组8只,在MI手术当天,将H19沉默慢病毒或对照病毒注射到左心室心肌,在MI后7 d处死之前,进行电生理检查,取出心脏,分别采用Western blotting、qRT-PCR和免疫荧光等方法检测H19的表达及其对NGF、神经重构指标酪氨酸羟化酶(TH)、生长相关蛋白43(GAP43)及心律失常易感性的影响;(3)将24只大鼠随机分为MI+NC组、MI+siH19组和MI+siH19+LV-NGF组,每组8只,进行回复实验,采用Western blotting、免疫荧光法检测NGF、TH和GAP43的表达。结果大鼠MI 7 d后,H19表达上调(P<0.01);沉默H19后,免疫荧光结果显示,大鼠MI后TH、GAP43的阳染面积下降(P<0.001);程序性电刺激显示, MI后VAs的易感性降低(P<0.001);沉默H19可以有效抑制NGF的表达(Z=-2.402 P=0.016),而且沉默H19对交感神经重构的抑制作用也可被过表达的NGF改善( P_TH<0.001,P_GAP43=0.001)。结论 MI后沉默H19可通过调节NGF的表达而抑制交感神经重构,从而降低MI后心律失常的发生率。

关键词: 心肌梗死, 交感神经重构, 室性心律失常, LncRNA H19, 神经生长因子

Abstract: Objective To explore the roles of H19 and nerve growth factor(NGF)in the development of ventricular arrhythmias(VAs)after myocardial infarction(MI). Methods MI rat models were established by left coronary artery occlusion. (1)A total of 14 healthy Sprague Dawley(SD)rats were randomly divided into the MI group and sham group, with 7 rats in each group. Seven days after MI was induced, the rats were sacrificed to determine the H19 expression with qRT-PCR. (2)A total of 32 rats were randomly divided into 4 groups: sham+NC group, sham+siH19 group, MI+NC group and MI+siH19 group, with 8 rats in each group. H19-siRNA or the control virus was injected into the left ventricular myocardium of the rats. Programmed electrical stimulation was performed 7 days after MI was induced to observe the susceptibility to VAs. The effects of H19 on NGF, tyrosine hydroxylase(TH)and growth-associated protein 43(GAP43)were detected with qRT-PCR, Western blotting and immunofluorescence staining. (3)A total of 24 rats were randomly divided into the MI+NC group, MI+siH19 group and MI+siH19+LV-NGF group for rescue test, with 8 rats in each group. The expressions of NGF, TH and GAP43 were detected with Western blotting and immunofluorescence staining. Results The expression of H19 were up-regulated 7 days after MI was induced(P<0.001). H19 knockdown suppressed the positive staining area of TH and GAP43(P<0.001), decreased the susceptibility to VAs(P<0.01), and inhibited the expression of NGF(Z=-2.402, P=0.016). The inhibitory effect of H19 knockdown on sympathetic remodeling could be saved by overexpressed NGF(P_TH<0.001, P_GAP43=0.001). Conclusion H19 knockdown after myocardial infarction can inhibit sympathetic nerve remodeling by regulating the expressiou of NGF, thus reducing the incidence of arrhythmia.

Key words: Myocardial infarction, Sympathetic remodeling, Ventricular arrhythmias, LncRNA H19, Nerve growth factor

中图分类号:

R541

张学丽, 郑璐, 王瑜, 王康, 闫素华. 沉默LncRNA H19通过调节神经生长因子抑制心肌梗死后交感神经重构[J]. 山东大学学报 (医学版), 2021, 59(5): 73-81.

ZHANG Xueli, ZHENG Lu, WANG Yu, WANG Kang, YAN Suhua. Knockdowa of LncRNA H19 inhibits sympathetic nerve remodeling after myocardial infarction by regulating nerve growth factor[J]. Journal of Shandong University (Health Sciences), 2021, 59(5): 73-81.

参考文献

[1] Pokorny J, Staněk V, Vrána M. Sudden cardiac death thirty years ago and at present. The role of autonomic disturbances in acute myocardial infarction revisited [J]. Physiol Res, 2011, 60(5): 715-728.
[2] Chen PS, Chen LS, Cao JM, et al. Sympathetic nerve sprouting, electrical remodeling and the mechanisms of sudden cardiac death [J]. Cardiovasc Res, 2001,50(2): 409-416.
[3] Wang Y, Xuan YL, Hu HS, et al. Risk of ventricular arrhythmias after myocardial infarction with diabetes associated with sympathetic neural remodeling in rabbits [J]. Cardiology, 2012, 121(1): 1-9.
[4] 马金, 陈秋雄, 吕渭辉. 神经重构在心肌梗死后心律失常中的作用及机制[J]. 心血管病学进展, 2019, 40(2): 260-263. MA Jin, CHEN Qiuxiong, LYU Weihui. Nerve remodeling in arrhythmia after myocardial infarction [J]. Advances in Cardiovascular Diseases, 2019, 40(2): 260-263.
[5] Hasan W, Jama A, Donohue T, et al. Sympathetic hyperinnervation and inflammatory cell NGF synthesis following myocardial infarction in rats [J]. Brain Res, 2006, 1124(1): 142-154.
[6] Hu H, Xuan Y, Wang Y, et al. Targeted NGF siRNA delivery attenuates sympathetic nerve sprouting and deteriorates cardiac dysfunction in rats with myocardial infarction [J]. PLoS One, 2014, 9(4): e95106. doi: 10.1371/journal.pone.0095106.
[7] Jathar S, Kumar V, Srivastava J, et al. Technological developments in lncRNA biology [J]. Adv Exp Med Biol, 2017, 1008:283-323. doi: 10.1007/978-981-10-5203-3_10.
[8] Wang C, Wang L, Ding Y, et al. LncRNA structural characteristics in epigenetic regulation [J]. Int J Mol Sci, 2017, 18(12): 2659.
[9] Wieczorek E, Reszka E. mRNA, microRNA and lncRNA as novel bladder tumor markers [J]. Clin Chim Acta, 2018, 477:141-153. doi:10.1016/j.cca.2017.12.009.
[10] Li H, Li J, Jia S, et al. miR675 upregulates long noncoding RNA H19 through activating EGR1 in human liver cance [J]. Oncotarget, 2015, 6(31): 31958-31984.
[11] Marquardt JU, Fischer K, Baus K, et al. Sirtuin-6-dependent genetic and epigenetic alterations are associated with poor clinical outcome in hepatocellular carcinoma patients [J]. Hepatology, 2013, 58(3): 1054-1064.
[12] Tao SC, Rui BY, Wang QY, et al. Extracellular vesicle-mimetic nanovesicles transport LncRNA-H19 as competing endogenous RNA for the treatment of diabetic wounds [J]. Drug Deliv, 2018, 25(1): 241-255.
[13] Wang JX, Zhang XJ, Li Q, et al. MicroRNA-103/107 regulate programmed necrosis and myocardial ischemia/reperfusion injury through targeting FADD [J]. Circ Res, 2015, 117(4): 352-363.
[14] Liu L, An X, Li Z, et al. The H19 long noncoding RNA is a novel negative regulator of cardiomyocyte hypertrophy [J]. Cardiovasc Res, 2016, 111(1): 56-65.
[15] Greco S, Zaccagnini G, Perfetti A, et al. Long noncoding RNA dysregulation in ischemic heart failure [J]. J Transl Med, 2016, 14(1): 183.
[16] Yin J, Hu HS, Li XL, et al. Inhibition of Notch signaling pathway attenuates sympathetic hyperinnervation together with the augmentation of M2 macrophages in rats post-myocardial infarction [J]. Am J Physiol Cell Physiol, 2016, 310(1): C41-C53.
[17] Lee TM, Lai PY, Chang NC. Effect of N-acetylcysteine on sympathetic hyperinnervation in post-infarcted rat hearts [J]. Cardiovasc Res, 2010, 85(1): 137-146.
[18] Yin J, Wang Y, Hu H, et al. P2X7 receptor inhibition attenuated sympathetic nerve sprouting after myocardial infarction via the NLRP3/IL-1β pathway [J]. J Cell Mol Med, 2017, 21(11): 2695-2710.
[19] Shi Y, Li Y, Yin J, et al. A novel sympathetic neuronal GABAergic signalling system regulates NE release to prevent ventricular arrhythmias after acute myocardial infarction [J]. Acta Physiol(Oxf), 2019, 227(2): e13315.
[20] Wang Y, Yin J, Wang C, et al. Microglial Mincle receptor in the PVN contributes to sympathetic hyperactivity in acute myocardial infarction rat [J]. J Cell Mol Med, 2019, 23(1): 112-125.
[21] Tang J, Cui X, Caranasos TG, et al. Heart repair using nanogel-encapsulated human cardiac stem cells in mice and pigs with myocardial infarction [J]. ACS nano, 2017, 11(10): 9738-9749.
[22] Voroshilovsky O, Qu Z, Lee MH, et al. Mechanisms of ventricular fibrillation induction by 60-Hz alternating current in isolated swine right ventricle [J]. Circulation, 2000, 102(13): 1569-1574.
[23] Reichardt LF. Neurotrophin-regulated signalling pathways [J]. Philos Trans R Soc Lond B Biol Sci, 2006, 361(1473): 1545-1564.
[24] Zhou S, Chen LS, Miyauchi Y, et al. Mechanisms of cardiac nerve sprouting after myocardial infarction in dogs [J]. Circ Research, 2004, 95(1): 76-83.
[25] Waterston RH, Lindblad-Toh K, Birney E, et al. Initial sequencing and comparative analysis of the mouse genome [J]. Nature, 2002, 420(6915): 520-562.
[26] Wahlestedt C. Targeting long non-coding RNA to therapeutically upregulate gene expression [J]. Nat Rev Drug Discov, 2013, 12(6): 433-446.
[27] Schmitz SU, Grote P, Herrmann BG. Mechanisms of long noncoding RNA function in development and disease [J]. Cell Mol Life Sci, 2016, 73(13): 2491-2509.
[28] Batista PJ, Chang HY. Long noncoding RNAs: cellular address codes in development and disease [J]. Cell, 2013, 152(6): 1298-1307.
[29] Ballantyne MD, McDonald RA, Baker AH. lncRNA/MicroRNA interactions in the vasculature [J]. Clin Pharmacol Ther, 2016, 99(5): 494-501.
[30] García-Padilla C, Domínguez JN, Aránega AE, et al. Differential chamber-specific expression and regulation of long non-coding RNAs during cardiac development [J]. Biochim Biophys Acta Gene Regul Mech, 2019, 1862(10): 194435.
[31] Viereck J, Bührke A, Foinquinos A, et al. Targeting muscle-enriched long non-coding RNA H19 reverses pathological cardiac hypertrophy [J]. Eur Heart J, 2020, 41(36): 3462-3474.
[32] Omura J, Habbout K, Shimauchi T, et al. Identification of Long Noncoding RNA H19 as a new biomarker and therapeutic target in right ventricular failure in pulmonary arterial hypertension [J]. Circulation, 2020, 142(15): 1464-1484.
[33] Zhang X, Cheng L, Xu L, et al. The lncRNA, H19 mediates the protective effect of hypoxia postconditioning against hypoxia-reoxygenation injury to senescent cardiomyocytes by targeting microRNA-29b-3p [J]. Shock, 2019, 52(2): 249-256.
[34] Hadji F, Boulanger MC, Guay SP, et al. Altered DNA methylation of Long Noncoding RNA H19 in calcific aortic valve disease promotes mineralization by silencing NOTCH1 [J]. Circulation, 2016, 134(23): 1848-1862.

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