线粒体ATP敏感性钾离子通道开放剂改善冠心病大鼠模型心肌氧化应激损伤的分子机制
|
摘要
目的探索线粒体ATP敏感性钾离子通道(mito KATP)开放剂改善冠心病大鼠模型心肌氧化应激损伤的分子机制。方法选取40只SD大鼠随机分为4组:正常对照组(不造模且不给予二氮嗪药物干预)、假手术组(仅切皮不进行造模手术)、模型组(制作冠心病大鼠模型,但不给予二氮嗪药物干预)和药物组(制作冠心病大鼠模型,给予二氮嗪药物3 mg/kg干预),每组10只。利用实时荧光定量聚合酶链式反应及Western blotting实验测定各组血管生成因子[成纤维细胞生长因子2(FGF-2)和血管内皮生长因子(VEGF)]m RNA及相关蛋白的表达量,同时比较各组大鼠细胞内的乳酸脱氢酶、线粒体膜电位(MMP)和细胞死亡率。采用SPSS 22.0软件进行数据处理。组间比较采用方差分析或者t检验。结果与模型组相比,药物组大鼠FGF-2[(100. 21±12. 33)×103vs (120. 43±10. 33)×103IU]与VEGF [(163. 31±9. 33)×10~3vs (181. 33±11. 13)×10~3IU]m RNA转录水平、FGF-2 [(0. 69±0. 33) vs (1. 32±0. 33)与VEGF [(0. 68±0. 33) vs (1. 20±0. 13)]表达水平、乳酸脱氢酶[(49. 32±3. 51) vs (156. 12±10. 18) U/L]表达均显著降低(P <0. 05)。与模型组相比,药物组细胞死亡率显著降低[(30. 32±3. 48)%vs (66. 12±3. 23)%],而荧光强度显著增加[(780. 12±9. 20) vs (220. 24±6. 15),P<0. 05]。结论 mito KATP通道开放剂可通过促进冠心病大鼠血管FGF-2和VEGF增加,增强乳酸脱氢酶活性,降低MMP、细胞死亡率和冠心病大鼠心肌氧化应激损伤。
Objective To investigate the molecular mechanism of mitochondrial mitochondrial ATP-sensitive potassium channels( mito KATP) opener in improving myocardial oxidative stress/injury in rates with coronary heart disease( CHD). Methods A total of40 SD rats were randomly divided into 4 groups with 10 in each: control group( without modeling and intervention of diazoxide drug),sham operation group( with skin cutting but without modeling),and model group( modeling of coronary heart disease but without inter-vention of diazoxide) and drug group( with modeling of coronary heart disease and intervention of diazoxide at 3 mg/kg). m RNA ex-pression of angiogenic factors[fibroblast growth factor 2( FGF-2) and vascular endothelial growth factor( VEGF) ] and related proteinsfor each group were measured using real-time fluorescence quantitative polymerase chain reaction and Western blotting. The groups werecompared in lactate dehydrogenase,mitochondrial membrane potential( MMP) and cell death rate. SPSS statistics 22.0 was used of da-ta processing,and analysis of variance( ANOVA) or t test for comparison between groups. Results Compared with the model group,drug group had significantly lower m RNA transcription of FGF-2 [( 100. 21 ± 12. 33) × 10~3 vs( 120. 43 ± 10. 33) × 10~3 IU] and VEGF[( 163. 31±9. 33) ×10~3 vs( 181. 33±11. 13) ×10~3 IU] and expression of FGF-2 [( 0. 69±0. 33) vs( 1. 32±0. 33) ],VEGF [( 0. 68±0. 33) vs( 1. 20±0. 13) ],and lactate dehydrogenase [( 49. 32±3. 51) vs( 156. 12±10. 18) U/L]. Compared with the model group,the cell death rate in the drug group decreased [( 30. 32 ± 3. 48) % vs( 66. 12 ± 3. 23) %],but the fluorescence intensityincreased [( 780. 12± 9. 20) vs( 220. 24 ± 6. 15) ]( P<0. 05). Conclusion mito KATP channel opener can decrease MMP,celldeath rate and myocardial oxidative stress/injury in rats with coronary heart disease by increasing vascular FGF-2 and VEGF andenhancing activity of lactate dehydrogenase.
Objective To investigate the molecular mechanism of mitochondrial mitochondrial ATP-sensitive potassium channels( mito KATP) opener in improving myocardial oxidative stress/injury in rates with coronary heart disease( CHD). Methods A total of40 SD rats were randomly divided into 4 groups with 10 in each: control group( without modeling and intervention of diazoxide drug),sham operation group( with skin cutting but without modeling),and model group( modeling of coronary heart disease but without inter-vention of diazoxide) and drug group( with modeling of coronary heart disease and intervention of diazoxide at 3 mg/kg). m RNA ex-pression of angiogenic factors[fibroblast growth factor 2( FGF-2) and vascular endothelial growth factor( VEGF) ] and related proteinsfor each group were measured using real-time fluorescence quantitative polymerase chain reaction and Western blotting. The groups werecompared in lactate dehydrogenase,mitochondrial membrane potential( MMP) and cell death rate. SPSS statistics 22.0 was used of da-ta processing,and analysis of variance( ANOVA) or t test for comparison between groups. Results Compared with the model group,drug group had significantly lower m RNA transcription of FGF-2 [( 100. 21 ± 12. 33) × 10~3 vs( 120. 43 ± 10. 33) × 10~3 IU] and VEGF[( 163. 31±9. 33) ×10~3 vs( 181. 33±11. 13) ×10~3 IU] and expression of FGF-2 [( 0. 69±0. 33) vs( 1. 32±0. 33) ],VEGF [( 0. 68±0. 33) vs( 1. 20±0. 13) ],and lactate dehydrogenase [( 49. 32±3. 51) vs( 156. 12±10. 18) U/L]. Compared with the model group,the cell death rate in the drug group decreased [( 30. 32 ± 3. 48) % vs( 66. 12 ± 3. 23) %],but the fluorescence intensityincreased [( 780. 12± 9. 20) vs( 220. 24 ± 6. 15) ]( P<0. 05). Conclusion mito KATP channel opener can decrease MMP,celldeath rate and myocardial oxidative stress/injury in rats with coronary heart disease by increasing vascular FGF-2 and VEGF andenhancing activity of lactate dehydrogenase.
引文
[1]胡文超,曹丰.无创血流储备分数测定对冠心病患者心肌缺血功能评估的临床进展[J].中华老年多器官疾病杂志,2018,17(2):145-149. DOI:10.11915/j.issn.1671-5403.2018.02.032.Hu WC,Cao F. Clinical application of noninvasive fractional flowreserve in assessment of myocardial ischemic function in patientswith coronary artery disease[J]. Chin J Mult Organ Dis Elderly,2018,17(2):145-149. DOI:10.11915/j.issn.1671-5403.2018.02.032.
[2]张琳,严松彪,陈晖,等.99mTc-MIBI SPECT负荷心肌灌注显像对怀疑心肌缺血糖尿病患者的预后价值[J].中华老年多器官疾病杂志,2017,16(9):672-676. DOI:10.119155/j.issn.1671-5403.2017.09.158.Zhang L,Yan SB,Chen H,et al. Prognostic value of stressmyocardial perfusion imaging with99mTc-MIBI SPECT for diabeticpatients with suspected myocardial ischemia[J]. Chin J MultOrgan Dis Elderly,2017,16(9):672-676. DOI:10.119155/j.issn.1671-5403.2017.09.158.
[3] Liu G,Wang ZK,Wang ZY,et al. Mitochondrial permeabilitytransition and its regulatory components are implicated in apoptosisof primary cultures of rat proximal tubular cells exposed to lead[J].Arch Toxicol, 2016, 90(5):1193-1209. DOI:10. 1007/s00204-015-1547-0.
[4] Zhou W,Marinelli F,Nief C,et al. Atomistic simulations indicatethe c-subunit ring of the F1FoATP synthase is not themitochondrial permeability transition pore[J]. Elife,2017,6(2):1-10. DOI:10.7554/e Life.23781.
[5] Arabian M,Aboutaleb N,Soleimani M,et al. Activation of mito-chondrial KATP channels mediates neuroprotection induced bychronic morphine preconditioning in hippocampal CA-1 neuronsfollowing cerebral ischemia[J]. Adv Med Sci,2018,63(2):213-219. DOI:10.1016/j.advms.2017.11.003.
[6] Onukwufor JO,Stevens D,Kamunde C. Bioenergetic and volumeregulatory effects of mito KATP channel modulators protect againsthypoxia-reoxygenation induced mitochondrial dysfunction[J]. JExp Biol, 2016, 219(17):2743-2751. DOI:10. 1242/jeb.140186.
[7] Pecoraro M,Ciccarelili M,Fiordelisi A,et al. Diazoxide improvesmitochondrial connexin 43 expression in a mouse model of doxoru-bicin-induced cardiotoxicity[J]. Int J Mol Sci,2018,19(3). pii:E757. DOI:10.3390/ijms19030757.
[8] Pohjoismki JL,Goffart S. The role of mitochondria in cardiacdevelopment and protection[J].Free Radic Biol Med,2017,106(5):345-354. DOI:10.1016/j.freeradbiomed.2017.02.032.
[9] Best B,Moran P,Ren B. VEGF/PKD-1 signaling mediates arte-riogenic gene expression and angiogenic responses in reversiblehuman microvascular endothelial cells with extended lifespan[J].Mol Cell Biochem,2018,446(1-2):199-207. DOI:10.1007/s11010-018-3286-z.
[10] Jiang Z,Wang W,Guo C. Tetrahydroxy stilbene glucoside amelio-rates H_2O_2-induced human brain microvascular endothelial celldysfunction in vitro by inhibiting oxidative stress and inflammatoryresponses[J]. Mol Med Rep,2017,16(4):5219-5224. DOI:10.3892/mmr.2017.7225.
[11] Filippi I,Carrarelli P,Luisi S,et al. Different expression ofhypoxic and angiogenic factors in human endometriotic lesions[J].Reprod Sci, 2016, 23(4):492-497. DOI:10. 1177/19337191-15607978.
[12] Szubert S,Moszynski R,Michalak S,et al. The associationsbetween serum VEGF,bFGF and endoglin levels with microvesseldensity and expression of proangiogenic factors in malignant andbenign ovarian tumors[J]. Microvasc Res,2016,107(9):91-96.DOI:10.1016/j.mvr.2016.06.002.
[13] Telford W,Tamul K,Bradford J. Measurement and characterizationof apoptosis by flow cytometry[J]. 2016,77(1):9.49.1-9.49.28.DOI:10.1002/cpcy.1.
[2]张琳,严松彪,陈晖,等.99mTc-MIBI SPECT负荷心肌灌注显像对怀疑心肌缺血糖尿病患者的预后价值[J].中华老年多器官疾病杂志,2017,16(9):672-676. DOI:10.119155/j.issn.1671-5403.2017.09.158.Zhang L,Yan SB,Chen H,et al. Prognostic value of stressmyocardial perfusion imaging with99mTc-MIBI SPECT for diabeticpatients with suspected myocardial ischemia[J]. Chin J MultOrgan Dis Elderly,2017,16(9):672-676. DOI:10.119155/j.issn.1671-5403.2017.09.158.
[3] Liu G,Wang ZK,Wang ZY,et al. Mitochondrial permeabilitytransition and its regulatory components are implicated in apoptosisof primary cultures of rat proximal tubular cells exposed to lead[J].Arch Toxicol, 2016, 90(5):1193-1209. DOI:10. 1007/s00204-015-1547-0.
[4] Zhou W,Marinelli F,Nief C,et al. Atomistic simulations indicatethe c-subunit ring of the F1FoATP synthase is not themitochondrial permeability transition pore[J]. Elife,2017,6(2):1-10. DOI:10.7554/e Life.23781.
[5] Arabian M,Aboutaleb N,Soleimani M,et al. Activation of mito-chondrial KATP channels mediates neuroprotection induced bychronic morphine preconditioning in hippocampal CA-1 neuronsfollowing cerebral ischemia[J]. Adv Med Sci,2018,63(2):213-219. DOI:10.1016/j.advms.2017.11.003.
[6] Onukwufor JO,Stevens D,Kamunde C. Bioenergetic and volumeregulatory effects of mito KATP channel modulators protect againsthypoxia-reoxygenation induced mitochondrial dysfunction[J]. JExp Biol, 2016, 219(17):2743-2751. DOI:10. 1242/jeb.140186.
[7] Pecoraro M,Ciccarelili M,Fiordelisi A,et al. Diazoxide improvesmitochondrial connexin 43 expression in a mouse model of doxoru-bicin-induced cardiotoxicity[J]. Int J Mol Sci,2018,19(3). pii:E757. DOI:10.3390/ijms19030757.
[8] Pohjoismki JL,Goffart S. The role of mitochondria in cardiacdevelopment and protection[J].Free Radic Biol Med,2017,106(5):345-354. DOI:10.1016/j.freeradbiomed.2017.02.032.
[9] Best B,Moran P,Ren B. VEGF/PKD-1 signaling mediates arte-riogenic gene expression and angiogenic responses in reversiblehuman microvascular endothelial cells with extended lifespan[J].Mol Cell Biochem,2018,446(1-2):199-207. DOI:10.1007/s11010-018-3286-z.
[10] Jiang Z,Wang W,Guo C. Tetrahydroxy stilbene glucoside amelio-rates H_2O_2-induced human brain microvascular endothelial celldysfunction in vitro by inhibiting oxidative stress and inflammatoryresponses[J]. Mol Med Rep,2017,16(4):5219-5224. DOI:10.3892/mmr.2017.7225.
[11] Filippi I,Carrarelli P,Luisi S,et al. Different expression ofhypoxic and angiogenic factors in human endometriotic lesions[J].Reprod Sci, 2016, 23(4):492-497. DOI:10. 1177/19337191-15607978.
[12] Szubert S,Moszynski R,Michalak S,et al. The associationsbetween serum VEGF,bFGF and endoglin levels with microvesseldensity and expression of proangiogenic factors in malignant andbenign ovarian tumors[J]. Microvasc Res,2016,107(9):91-96.DOI:10.1016/j.mvr.2016.06.002.
[13] Telford W,Tamul K,Bradford J. Measurement and characterizationof apoptosis by flow cytometry[J]. 2016,77(1):9.49.1-9.49.28.DOI:10.1002/cpcy.1.