小型猪二尖瓣反流致心力衰竭模型构建及心肌硫化氢体系变化的研究
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摘要
目的:构建小型猪二尖瓣反流致心力衰竭模型,探究慢性心力衰竭过程中硫化氢(H_2S)体系的变化。方法:12只8月龄中华小型猪随机分为对照组(n=6)与二尖瓣反流组(n=6),采用小切口非体外循环下二尖瓣腱索拉伤造成二尖瓣反流建立慢性心力衰竭模型。6个月后采用超声心动图、在体血流动力学和病理学方法评价动物左心室结构和功能的变化,采用分子生物学方法检测心脏重构分子Ⅰ型胶原(CollagenⅠ)、Ⅲ型胶原(CollagenⅢ)和H_2S合成酶体系中胱硫醚-β-合成酶(CBS)、胱硫醚-γ-裂解酶(CSE)蛋白和mRNA表达。结果:术后6个月二尖瓣反流组动物与对照组相比,左心室收缩末期容积(LVESV)和左心室舒张末期容积(LVEDV)明显增加[(18.50±2.88)ml vs (10.50±0.99)ml,P<0.05;(87.50±12.12)ml vs (42.33±2.04)ml,P<0.01],左心室收缩末期压力明显降低[(86.66±5.60)mmHg vs (117.70±5.49)mmHg,1 mmHg=0.133 kPa,P<0.01],但左心室射血分数(LVEF)无明显变化[(78.67±1.87)%vs (75.33±1.87)%,P>0.05]。病理学可见二尖瓣反流组动物二尖瓣后瓣腱索断裂,瓣叶卷曲增厚,边缘不规则增生,石蜡切片染色可见左心室心肌细胞损伤、心肌间胶原纤维明显增多。二尖瓣反流组与对照组比较,血浆H_2S水平明显降低[(27.48±2.78)μmol/L vs (37.87±3.55)μmol/L,P=0.044],血清N末端B型利钠肽原(NT-proBNP)水平明显升高[(2 132.00±212.30)μg/L vs (456.70±40.79)μg/L,P<0.001]。分子生物学手段检测二尖瓣反流组心肌组织与心脏重构相关的CollagenⅠ、CollagenⅢ水平明显上调,H_2S合成相关的酶CSE、CBS明显下调。结论:H_2S合成体系下调可能参与二尖瓣反流所致心肌重构和心力衰竭的发展过程。
Objectives: To establish a mitral regurgitation model in swine and study the change of hydrogen sulfide(H_2 S) system in this chronic heart failure model.Methods: Miniature pigs were randomly divided into two groups, the control group(n=6) and mitral regurgitation group(n=6). Chronic heart failure models were established by pulling and destroying the mitral chordal through small incision off-pump surgery. After 6 months, echocardiography, in vivo hemodynamics and pathology examination were performed to evaluate the changes of left ventricular structure and function in this animal model. Molecular biology techniques were used to detect protein and mRNA level changes of type I collagen, type III collagen, cystathionine-β-synthase(CBS) and cystathionine-γ-lyase(CSE).Results: At 6 months after surgery, the LVESV and LVEDV were significantly increased([18.50 ± 2.88] ml vs [10.50±0.99] ml, P<0.05; [87.50 ± 12.12] ml vs [42.33 ± 2.04] ml, P<0.01), left ventricle diastolic pressure were significantly decreased([86.66 ± 5.60] mmHg vs [117.70 ± 5.49] mmHg, P<0.01) in mitral regurgitation group as compared to control group, while left ventricle ejection fraction was similar between the two groups([78.67 ± 1.87]% vs [75.33 ± 1.87]%,P>0.05). Pathologic evaluation showed disruption of mitral chordal of posterior mitral valve, thickened and curved valve with significant degenerative changes. HE stained myocardial slides revealed cardiomyocyte injury and increased extracellular collagen in left ventricle tissue in mitral value regurgitation group. Plasma H_2 S level was significantly decreased([27.48 ± 2.78]μmol/L vs [37.87 ± 3.55] μmol/L, P=0.044), serum NT-proBNP level was significantly increased([2 132.00 ± 212.30] μg/L vs [456.70 ± 40.79] μg/L, P<0.001) in mitral value regurgitation group as compared with control group. Protein and mRNA expressions of collagen type I and collagen type III were significantly upregulated, CSE and CBS were significantly downregulated in mitral value regurgitation group as compared with control group.Conclusions: The downregulation of H_2 S synthesis system may be involved in the development of myocardial remodeling and heart failure caused by mitral regurgitation in this swine model.
Objectives: To establish a mitral regurgitation model in swine and study the change of hydrogen sulfide(H_2 S) system in this chronic heart failure model.Methods: Miniature pigs were randomly divided into two groups, the control group(n=6) and mitral regurgitation group(n=6). Chronic heart failure models were established by pulling and destroying the mitral chordal through small incision off-pump surgery. After 6 months, echocardiography, in vivo hemodynamics and pathology examination were performed to evaluate the changes of left ventricular structure and function in this animal model. Molecular biology techniques were used to detect protein and mRNA level changes of type I collagen, type III collagen, cystathionine-β-synthase(CBS) and cystathionine-γ-lyase(CSE).Results: At 6 months after surgery, the LVESV and LVEDV were significantly increased([18.50 ± 2.88] ml vs [10.50±0.99] ml, P<0.05; [87.50 ± 12.12] ml vs [42.33 ± 2.04] ml, P<0.01), left ventricle diastolic pressure were significantly decreased([86.66 ± 5.60] mmHg vs [117.70 ± 5.49] mmHg, P<0.01) in mitral regurgitation group as compared to control group, while left ventricle ejection fraction was similar between the two groups([78.67 ± 1.87]% vs [75.33 ± 1.87]%,P>0.05). Pathologic evaluation showed disruption of mitral chordal of posterior mitral valve, thickened and curved valve with significant degenerative changes. HE stained myocardial slides revealed cardiomyocyte injury and increased extracellular collagen in left ventricle tissue in mitral value regurgitation group. Plasma H_2 S level was significantly decreased([27.48 ± 2.78]μmol/L vs [37.87 ± 3.55] μmol/L, P=0.044), serum NT-proBNP level was significantly increased([2 132.00 ± 212.30] μg/L vs [456.70 ± 40.79] μg/L, P<0.001) in mitral value regurgitation group as compared with control group. Protein and mRNA expressions of collagen type I and collagen type III were significantly upregulated, CSE and CBS were significantly downregulated in mitral value regurgitation group as compared with control group.Conclusions: The downregulation of H_2 S synthesis system may be involved in the development of myocardial remodeling and heart failure caused by mitral regurgitation in this swine model.
引文
[1]潘文志,周达新,葛均波.二尖瓣反流机制的新认识及其应用[J].中国医学前沿杂志(电子版),2017,9(10):43-46.DOI:10.12037/YXQY.2017.10-07.
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[11]颜艳.超声心动图在诊断二尖瓣脱垂中的应用价值[J].医学信息,2018,31(8):149-151.DOI:10.3969/j.issn.1006-1959.2018.08.052.
[12]Abe K,Kimura H.The possible role of hydrogen sulfide as an endogenous neuromodulator[J].J Neurosci,1996,16(3):1066-1071.DOI:10.1523/JNEUROSCI.16-03-01066.1996.
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[14]Zhang HH,Hu J,Zhou YL,et al.Promoted interaction of nuclear factor-kappa B with demethylated cystathionine-beta-synthetase gene contributes to gastric hypersensitivity in diabetic rats[J].J Neurosci,2013,33(21):9028-9038.DOI:10.1523/jneurosci.1068-13.2013.
[15]Mishra PK,Tyagi N,Sen U,et al.H2S ameliorates oxidative and proteolytic stresses and protects the heart against adverse remodeling in chronic heart failure[J].Am J Physiol Heart Circ Physiol,2010,298(2):451-456.DOI:10.1152/ajpheart.00682.2009.
[16]郭蓉.硫化氢对肾血管性高血压大鼠颈动脉窦压力感受器活动的易化作用[D].石家庄:河北医科大学,2013.
[17]Liu W,Xu C,You X,et al.Hydrogen sulfide delays LPS-induced preterm birth in mice via anti-inflammatory pathways[J].PLoS One,2016,11(4):e0152838.DOI:10.1371/journal.pone.0152838.
[2]Olas B.Hydrogen sulfide in signaling pathways[J].Clinica Chimica Acta,2015,439:212-218.DOI:10.1016/j.cca.2014.10.037.
[3]Dunn WR,Alexander SP,Ralevic V,et al.Effects of hydrogen sulphide in smooth muscle[J].Pharmacol Ther,2016,158:101-113.DOI:10.1016/j.pharmthera.2015.12.007.
[4]Hu L F,Lu M,Hon Wong P T,et al.Hydrogen sulfide:neurophysiology and neuropathology[J].Antioxid Redox Signal,2011,15(2):405-419.DOI:10.1089/ars.2010.3517.
[5]Zhao X,Zhang LK,Zhang CY,et al.Regulatory effect of hydrogen sulfide on vascular collagen content in spontaneously hypertensive rats[J].Hypertens Res,2008,31(8):1619-1630.DOI:10.1291/hypres.31.1619.
[6]Calvert JW,Elston M,Nicholson CK,et al.Genetic and pharmacologic hydrogen sulfide therapy attenuates ischemia-induced heart failure in mice[J].Circulation,2010,122(1):11-19.DOI:10.1161/circulationaha.109.920991.
[7]田毅,张宏,孙嘉康,等.小型猪二尖瓣反流联合球囊封堵序贯法建立慢性心力衰竭模型[J].中国循环杂志,2014,29(z1):40-41.
[8]李凯,罗富良,田毅,等.比格犬二尖瓣反流慢性心力衰竭模型的建立[J].中国循环杂志,2014,29(z1):37.
[9]Liu SY,Duan XC,Jin S,et al.Hydrogen sulfide improves myocardial remodeling via downregulated angiotensin/AT1R pathway in renovascular hypertensive rats[J].Am J Hypertens,2017,30(1):67-74.DOI:10.1093/ajh/hpw104.
[10]沈明,郭志福,鲍礼智,等.大鼠心梗后心力衰竭模型的制备及心功能评价[J].实验动物科学,2017,34(2):66-70.DOI:10.3969/j.issn.1006-6179.2017.02.015.
[11]颜艳.超声心动图在诊断二尖瓣脱垂中的应用价值[J].医学信息,2018,31(8):149-151.DOI:10.3969/j.issn.1006-1959.2018.08.052.
[12]Abe K,Kimura H.The possible role of hydrogen sulfide as an endogenous neuromodulator[J].J Neurosci,1996,16(3):1066-1071.DOI:10.1523/JNEUROSCI.16-03-01066.1996.
[13]Shibuya N,Koike S,Tanaka M,et al.A novel pathway for the production of hydrogen sulfide from D-cysteine in mammalian cells[J].Nat Commun,2013,1366(4):1-7.DOI:10.1038/ncomms2371.
[14]Zhang HH,Hu J,Zhou YL,et al.Promoted interaction of nuclear factor-kappa B with demethylated cystathionine-beta-synthetase gene contributes to gastric hypersensitivity in diabetic rats[J].J Neurosci,2013,33(21):9028-9038.DOI:10.1523/jneurosci.1068-13.2013.
[15]Mishra PK,Tyagi N,Sen U,et al.H2S ameliorates oxidative and proteolytic stresses and protects the heart against adverse remodeling in chronic heart failure[J].Am J Physiol Heart Circ Physiol,2010,298(2):451-456.DOI:10.1152/ajpheart.00682.2009.
[16]郭蓉.硫化氢对肾血管性高血压大鼠颈动脉窦压力感受器活动的易化作用[D].石家庄:河北医科大学,2013.
[17]Liu W,Xu C,You X,et al.Hydrogen sulfide delays LPS-induced preterm birth in mice via anti-inflammatory pathways[J].PLoS One,2016,11(4):e0152838.DOI:10.1371/journal.pone.0152838.
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