不同浓度β_1-肾上腺素受体自身抗体对心肌细胞存活影响的差异性
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摘要
目的观察不同浓度的β_1-肾上腺素受体自身抗体(β_1-adrenergic receptor autoantibodies,β_1-AA)作用不同时间,对于心肌细胞产生的不同终末效应。方法分别使用10~(-10)、10~(-9)、10~(-8)、10~(-7)、10~(-6)mol/Lβ_1-AA作用于乳鼠心肌细胞(neonatal rat cardiomyocytes,NRCMs)和H9c2细胞检测细胞生存率(24、36、48 h),作用于H9c2细胞检测线粒体功能(ATP含量、线粒体膜电势)(48 h)。结果中、高浓度(10~(-7)、10~(-6)mol/L)β_1-AA刺激各个时间,NRCMs、H9c2细胞生存率明显降低;而低浓度(10~(-10)、10~(-9)、10~(-8)mol/L)使得H9c2细胞轻微增生。高浓度(10~(-6)mol/L)β_1-AA刺激后,H9c2细胞中ATP含量降低,而中低浓度(10~(-10)、10~(-9)、10~(-8)、10~(-7)mol/L)β_1-AA使H9c2细胞中ATP含量增加。高浓度β_1-AA(10~(-6)mol/L)刺激后,H9c2细胞线粒体膜电势降低,低浓度(10~(-10)、10~(-8)mol/L)β_1-AA则使H9c2细胞线粒体膜电势增加。结论高浓度β_1-AA刺激心肌细胞后表现为损伤作用,低浓度刺激表现为正性作用(即增强线粒体功能、促存活),与作用时间无明显相关,本研究为β_1-AA阳性人群中心功能差异的现象提供了病理生理学依据。
Objective To investigate the terminal effects on cardiomyocytes of β_1-adrenergic receptor autoantibodies( β_1-AA) at different concentrations for different time. Methods Cell survival was detected after 10~(-10),10~(-9),10~(-8),10~(-7),10~(-6) mol/L β_1-AA treating neonatal rat cardiomyocytes( NRCMs) and H9 c2 cells for 24,36,and 48 hours,while mitochondrial function( ATP content and mitochondrial membrane potential) of H9 c2 cells was detected after 48 hours. Results The cell survival of NRCMs and H9 c2 decreased significantly by β_1-AA in high-concentrations( 10~(-7),10~(-6) mol/L) at any time and the survival of H9 c2 proliferated slightly in low concentrations( 10~(-10),10~(-9),10~(-8) mol/L) at any time. ATP content of H9 c2 was decreased by β_1-AA in high concentration( 10~(-6) mol/L),while increased in medium and low concentrations( 10~(-10),10~(-9),10~(-8),10~(-7) mol/L). The mitochondrial membrane potential of H9 c2 was decreased by β_1-AA in high concentrations( 10~(-6) mol/L),while increased in low concentrations. Conclusion β_1-AA damage cardiomyocytes in high concentration while it showed the positive effects in low concentrations( ie,promoting ATP production and cell survival). And all the effects had no relation to time. In conclusion,our study provides a new foundation for the differences of cardiac function among β_1-AA positive population.
Objective To investigate the terminal effects on cardiomyocytes of β_1-adrenergic receptor autoantibodies( β_1-AA) at different concentrations for different time. Methods Cell survival was detected after 10~(-10),10~(-9),10~(-8),10~(-7),10~(-6) mol/L β_1-AA treating neonatal rat cardiomyocytes( NRCMs) and H9 c2 cells for 24,36,and 48 hours,while mitochondrial function( ATP content and mitochondrial membrane potential) of H9 c2 cells was detected after 48 hours. Results The cell survival of NRCMs and H9 c2 decreased significantly by β_1-AA in high-concentrations( 10~(-7),10~(-6) mol/L) at any time and the survival of H9 c2 proliferated slightly in low concentrations( 10~(-10),10~(-9),10~(-8) mol/L) at any time. ATP content of H9 c2 was decreased by β_1-AA in high concentration( 10~(-6) mol/L),while increased in medium and low concentrations( 10~(-10),10~(-9),10~(-8),10~(-7) mol/L). The mitochondrial membrane potential of H9 c2 was decreased by β_1-AA in high concentrations( 10~(-6) mol/L),while increased in low concentrations. Conclusion β_1-AA damage cardiomyocytes in high concentration while it showed the positive effects in low concentrations( ie,promoting ATP production and cell survival). And all the effects had no relation to time. In conclusion,our study provides a new foundation for the differences of cardiac function among β_1-AA positive population.
引文
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[20] Berezhnaya E,Neginskaya M,Uzdensky A B,et al. Photoinduced oxidative stress impairs mitochondrial metabolism in neurons and astrocytes[J]. Mol Neurobiol,2018,55(1):90-95.
[21] Ferreira G,Costa C,Bassaizteguy V,et al. Incubation of human sperm with micelles made from glycerophospholipid mixtures increases sperm motility and resistance to oxidative stress[J]. PLo S One,2018,13(6):e0197897.
[22] Agnihotri S K,Agrawal A K,Hakim B A,et al. Mitochondrial membrane potential(MMP)regulates sperm motility[J]. In Vitro Cell Dev Biol Anim,2016,52(9):953-960.
[2] Turcani M,Fizel A,Maasova D,et al. The pathophysiology of heart failure[J]. Bratisl Lek Listy,1988,89(3):211-219.
[3] Verschure D O,Van Eck-Smit B L,Somsen G A,et al.Cardiac sympathetic activity in chronic heart failure:cardiac123I-mIBG scintigraphy to improve patient selection for ICD implantation[J]. Neth Heart J,2016,24(12):701-708.
[4] Andersen S,Andersen A,De Man F S,et al. Sympathetic nervous system activation and beta-adrenoceptor blockade in right heart failure[J]. Eur J Heart Fail,2015,17(4):358-366.
[5] Holthoff H P,Zeibig S,Jahns-Boivin V,et al. Detection of anti-beta1-AR autoantibodies in heart failure by a cell-based competition ELISA[J]. Circ Res, 2012, 111(6):675-684.
[6] Becker N P,Goettel P,Mueller J,et al. Functional autoantibody diseases:basics and treatment related to cardiomyopathies[J]. Front Biosci(Landmark Ed),2019,24:48-95.
[7] Bovo E,Mazurek S R,De Tombe P P,et al. Increased energy demand during adrenergic receptor stimulation contributes to Ca2+wave generation[J]. Biophys J,2015,109(8):1583-1591.
[8] Wang L,Li Y,Ning N,et al. Decreased autophagy induced by beta1-adrenoceptor autoantibodies contributes to cardiomyocyte apoptosis[J]. Cell Death Dis,2018,9(3):406.
[9] Liu H R,Zhao R R,Zhi J M,et al. Screening of serum autoantibodies to cardiac beta1-adrenoceptors and M2-muscarinic acetylcholine receptors in 408 healthy subjects of varying ages[J]. Autoimmunity,1999,29(1):43-51.
[10]杜芸辉,李笑,于海存,等.一种新型β1肾上腺素受体单克隆抗体的制备及其临床应用[J].心肺血管病杂志,2017,36(8):696-700.
[11] Lohse M J,Engelhardt S,Eschenhagen T. What is the role of beta-adrenergic signaling in heart failure?[J]. Circ Res,2003,93(10):896-906.
[12] He J Q,Balijepalli R C,Haworth R A,et al. Crosstalk of beta-adrenergic receptor subtypes through Gi blunts beta-adrenergic stimulation of L-type Ca2+channels in canine heart failure[J]. Circ Res,2005,97(6):566-573.
[13] Reina S,Ganzinelli S,Sterin-Borda L,et al. Pro-apoptotic effect of anti-beta1-adrenergic receptor antibodies in periodontitis patients[J]. Int Immunopharmacol,2012,14(4):710-721.
[14] Jahns R,Boivin V,Siegmund C,et al. Autoantibodies activating human beta1-adrenergic receptors are associated with reduced cardiac function in chronic heart failure[J].Circulation,1999,99(5):649-654.
[15] Jahns R,Boivin V,Krapf T,et al. Modulation of beta1-adrenoceptor activity by domain-specific antibodies and heart failure-associated autoantibodies[J]. J Am Coll Cardiol,2000,36(4):1280-1287.
[16] Wallukat G,Morwinski M,Kowal K,et al. Autoantibodies against the beta-adrenergic receptor in human myocarditis and dilated cardiomyopathy:beta-adrenergic agonism without desensitization[J]. Eur Heart J,1991,12(Suppl D):178-181.
[17] Liang C S,Mao W,Liu J. Pro-apoptotic effects of anti-beta1-adrenergic receptor antibodies in cultured rat cardiomyocytes:actions on endoplasmic reticulum and the prosurvival PI3K-Akt pathway[J]. Autoimmunity,2008,41(6):434-441.
[18] Teerlink J R,Pfeffer J M,Pfeffer M A. Progressive ventricular remodeling in response to diffuse isoproterenol-induced myocardial necrosis in rats[J]. Circ Res,1994,75(1):105-113.
[19] Banoth B,Cassel S L. Mitochondria in innate immune signaling[J]. Transl Res,2018,202:52-68.
[20] Berezhnaya E,Neginskaya M,Uzdensky A B,et al. Photoinduced oxidative stress impairs mitochondrial metabolism in neurons and astrocytes[J]. Mol Neurobiol,2018,55(1):90-95.
[21] Ferreira G,Costa C,Bassaizteguy V,et al. Incubation of human sperm with micelles made from glycerophospholipid mixtures increases sperm motility and resistance to oxidative stress[J]. PLo S One,2018,13(6):e0197897.
[22] Agnihotri S K,Agrawal A K,Hakim B A,et al. Mitochondrial membrane potential(MMP)regulates sperm motility[J]. In Vitro Cell Dev Biol Anim,2016,52(9):953-960.