血管紧张素Ⅱ-CaMK-CREB-CYP11B2和肾上腺素-PKA-CREB-CYP11B2途径在应激性高血压发病中的研究
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摘要
目的探讨血管紧张素Ⅱ-CaMK-CREB-CYP11B2和肾上腺素-PKA-CREB-CYP11B2途径在应激性高血压(SIH)发病中的作用。方法选取鼠龄在6～8w的SPF级雄性Wistar大鼠28只,适应性喂养7d后,按照完全随机法分为SIH模型组与正常组,每组14只。通过每天声光间断足底电、夹尾持续刺激12w构建SIH大鼠模型。采用放射免疫法检测SIH模型大鼠血浆血管紧张素Ⅱ含量,酶联免疫吸附(ELISA)方法检测SIH模型大鼠血清肾上腺素含量,实时荧光定量PCR(RT-qPCR)和免疫组织化学方法检测SIH模型大鼠肾上腺组织Creb、Nr4a1、Nr4a2、Cyp11b2基因mRNA表达和环磷酸腺苷反应元件结合蛋白(CREB)磷酸化水平及醛固酮合酶(CYP11B2)蛋白表达水平的变化。结果与正常组比较,SIH模型组大鼠体质量增长明显缓慢,收缩压与舒张压均明显升高(P <0.05),血浆血管紧张素Ⅱ含量无明显变化(P>0.05),血清肾上腺素含量明显升高(P <0.05),肾上腺组织Creb基因mRNA表达水平明显上调(P <0.05),Nr4a1与Nr4a2基因mRNA表达水平有下调趋势(P>0.05),Cyp11b2基因mRNA表达水平明显上调(P <0.05);与正常组比较,SIH模型组大鼠肾上腺CREB磷酸化水平及CYP11B2蛋白表达水平明显上调(P <0.05)。结论SIH模型组大鼠肾上腺组织肾上腺素-PKA-CREBCYP11B2途径的激活是血液醛固酮含量增多、应激性高血压发病的原因之一。
Objective To investigate the role of the angiotensin Ⅱ-CaMK-CREB-CYP11B2 and adrenergic-PKACREB-CYP11B2 pathways in the pathogenesis of stress induce hypertension(SIH).Methods Aged 6-8weeks of28 male SPF Wistar rats were randomly divided into SIH model group and normal group,with 14 rats in each group.The SIH rat model was constructed by daily sound and light interruption of the foot and continuous stimulation of the tail for 12 weeks.The content of plasma angiotensin Ⅱ in SIH model rats were detected by radioimmunoassay.The serum adrenaline contents in SIH model rats were examined by ELISA.The mRNAs of Creb,Nr4a1,Nr4a2 and Cyp11b2genes,the changes of CREB phosphorylation levels and CYP11B2 protein expression levels in adrenal tissue of SIH model rats were detected by RT-qPCR and immunohistochemistry,respectively.Results Compared with the normal group,the speed of body mass growth in SIH model group was significantly slower and the systolic blood pressure and diastolic blood pressure were significantly increased(P <0.05).Compared with the normal group,the content of plasma angiotensin Ⅱ in the SIH model group was not changed(P >0.05),but the content of serum adrenaline was significantly increased(P <0.05),and adrenal tissue Creb gene mRNA expression level and the level of Cyp11b2 mRNA were significantly up-regulated(P <0.05).Though Nr4a1 and Nr4a2gene mRNA expression levels were down-regulated,there was no statistical significance(P >0.05).Compared with the normal group,the phosphorylation level of CREB and the expression of CYP11B2 protein in the SIH model group were significantly up-regulated(P <0.05).Conclusion The activation of the adrenal-PKACREB-CYP 11B2 pathway in stress-induced hypertension group is the main cause of increased blood aldosterone content and onset of stress-induce hypertension.
Objective To investigate the role of the angiotensin Ⅱ-CaMK-CREB-CYP11B2 and adrenergic-PKACREB-CYP11B2 pathways in the pathogenesis of stress induce hypertension(SIH).Methods Aged 6-8weeks of28 male SPF Wistar rats were randomly divided into SIH model group and normal group,with 14 rats in each group.The SIH rat model was constructed by daily sound and light interruption of the foot and continuous stimulation of the tail for 12 weeks.The content of plasma angiotensin Ⅱ in SIH model rats were detected by radioimmunoassay.The serum adrenaline contents in SIH model rats were examined by ELISA.The mRNAs of Creb,Nr4a1,Nr4a2 and Cyp11b2genes,the changes of CREB phosphorylation levels and CYP11B2 protein expression levels in adrenal tissue of SIH model rats were detected by RT-qPCR and immunohistochemistry,respectively.Results Compared with the normal group,the speed of body mass growth in SIH model group was significantly slower and the systolic blood pressure and diastolic blood pressure were significantly increased(P <0.05).Compared with the normal group,the content of plasma angiotensin Ⅱ in the SIH model group was not changed(P >0.05),but the content of serum adrenaline was significantly increased(P <0.05),and adrenal tissue Creb gene mRNA expression level and the level of Cyp11b2 mRNA were significantly up-regulated(P <0.05).Though Nr4a1 and Nr4a2gene mRNA expression levels were down-regulated,there was no statistical significance(P >0.05).Compared with the normal group,the phosphorylation level of CREB and the expression of CYP11B2 protein in the SIH model group were significantly up-regulated(P <0.05).Conclusion The activation of the adrenal-PKACREB-CYP 11B2 pathway in stress-induced hypertension group is the main cause of increased blood aldosterone content and onset of stress-induce hypertension.
引文
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[22]MCNUITY S E,BARRETT R M,VOGEL-CIERNIA A,et al.Differential roles for Nr4a1and Nr4a2in object location vs.object recognition long-term memory[J].Learn Mem,2012,19(12):588-592.
[23]潘志强,方肇勤,卢文丽,等.不同证型H22肝癌小鼠肾上腺皮质类固醇激素合成酶及其调节因子表达的特征[J].中国中西医结合杂志,2011,31(1):85-89.
[24]ERIKA N,ATSUSHI Y,MAKOTO K,et al.Endogenous purification of NR4A2(Nurr1)identified poly(ADP-Ribose)polymerase 1as a prime coregulator in human adrenocortical H295R Cells[J].Int J Mol Sci,2018,19(5):1406-1419.
[25]潘志强,方肇勤,卢文丽,等.不同程度邪毒证H22荷瘤小鼠肾上腺皮质酮合成酶及其转录因子表达的差异[J].中西医结合肝病杂志,2014,24(6):342-346.
[26]NOGUEIRA E F,RAINEY W E.Regulation of aldosterone synthase by activator transcription Factor/cAMP response element-binding protein family members[J].Endocrinology,2010,151(3):1060-1070.
[2]MCCUBBIN J A,NATHAN A,HIBDON M A,et al.Blood pressure,emotional dampening,and risk behav ior:implications for hypertension development[J].Psychosom Med,2018,80(6):544-550.
[3]孙瑞媛,吕秋凤.牛磺酸与应激性高血压[J].动物医学进展,2016,37(10):111-115.
[4]REDINA O E,MARKEL A L.Stress,genes,and hypertension.contribution of the ISIAH rat strain study[J].Curr Hypertens Rep,2018,20(8):66-76.
[5]RANHOTRA H S.The NR4Aorphan nuclear receptors:mediators in metabolism and diseases[J].J Recept Signal Transduct Res,2015,35(2):184-188.
[6]YASUHIRO N,YUTO Y,YUTA T,et al.Expression of CYP11B2in aldosterone-producing adrenocortical adenom a:regulatory mechanisms and clinical significance[J].Tohoku JExp Med,2016,240(3):183-190.
[7]WANG H M,D0NG J H,Li Q,et al.A stress response pathway in mice upregulates somatostatin level and transcription in pancreatic delta cells through Gs andβ-arrestin 1[J].Diabetologia,2014,57(9):1899-1910.
[8]SPAT A,HUNYADY L,SZANDA G.Signaling interactions in the adrenal cortex[J].Front Endocrinol(Lausanne),2016,7(2):17-26.
[9]FERRON L,RUCHON Y,RENAUD J F,et al.T-type Ca2+signalling regulates aldosterone-induced CREB activation and cell death through PP2Aactivation in neonatal cardiomyocytes[J].Cardiovasc Res,2011,90(1):105-112.
[10]阿地拉·阿不都艾尼,吴桂霞,卡思木江·阿西木江,等.应激性高血压大鼠血浆中醛固酮水平及肾上腺超微结构的变化[J].新疆医科大学学报,2017,40(6):797-799.
[11]钟航海,冯英凯.慢性复合应激诱发高血压动物模型的制备[J].现代医药卫生,2010,26(23):3523-3525.
[12]RAZZAGHY-AZAR M,YAU M,KHATTAB A,et al.Apparent mineralocorticoid excess and the long term treatment of genetic hypertension[J].J Steroid Biochem Mol Biol,2016,165(Pt A):145-150.
[13]BARBARO N R,KIRABO A,HARRISON D G.A new role of mister(MR)T in hypertension:mineralocorticoid receptor,immune system,and hypertension[J].Circ Res,2017,120(10):1527-1529.
[14]FUNDER J W.Sensitivity to aldosterone:plasma levels are not the full story[J].Hypertension,2014,63(6):1168-1170.
[15]SUZUKI D,SAITOHAKODA A,Ito R,et al.Suppressive effects of RXR agonist PA024on adrenal CYP11B2expression,aldosterone secretion and blood pressure[J].PloS One,2017,12(8):1-22.
[16]SZEKERES M,TURU G,ORIENT A,et al.Mechanisms of angiotensin Ⅱ-mediated regulation of aldosterone synthase expression in H295R human adrenocortical and rat adrenal glomerulosa cells[J].Mol Cell Endocrinol,2009,302(2):244-253.
[17]ZHOU X,CHEN K,WANG Y,et al.Antiaging gene klotho regulates adrenal CYP11B2expression and aldosterone synthesis[J].J Am Soc Nephrol,2016,27(6):1765-1776.
[18]OLALA L O,CHOUDHARY V,JOHNSON M H,et al.Angiotensin Ⅱ-induced protein kinase D activates the ATF/CREB family of transcription factors and promotes StARmRNA expression[J].Endocrinology,2014,155(7):2524-2533.
[19]LI W C,NGUYEN Q D,GIGANTELLI G,et al.Regulation of CREB functions by phosphorylation and sumoylation in nervous and visual systems[J].Curr Mol Med,2016,16(10):885-892.
[20]MATSUDA K,URUNO A,KOGURE N,et al.Angiotensin Ⅱreceptor blockers differentially affect CYP11B2expression in human adrenal H295Rcells.[J].Mol Cell Endocrinol,2014,383(1-2):60-68.
[21]张池,胡吉,张朝云.NR4A孤儿核受体亚家族在代谢综合征中的作用[J].医学综述,2014,20(18):3273-3276.
[22]MCNUITY S E,BARRETT R M,VOGEL-CIERNIA A,et al.Differential roles for Nr4a1and Nr4a2in object location vs.object recognition long-term memory[J].Learn Mem,2012,19(12):588-592.
[23]潘志强,方肇勤,卢文丽,等.不同证型H22肝癌小鼠肾上腺皮质类固醇激素合成酶及其调节因子表达的特征[J].中国中西医结合杂志,2011,31(1):85-89.
[24]ERIKA N,ATSUSHI Y,MAKOTO K,et al.Endogenous purification of NR4A2(Nurr1)identified poly(ADP-Ribose)polymerase 1as a prime coregulator in human adrenocortical H295R Cells[J].Int J Mol Sci,2018,19(5):1406-1419.
[25]潘志强,方肇勤,卢文丽,等.不同程度邪毒证H22荷瘤小鼠肾上腺皮质酮合成酶及其转录因子表达的差异[J].中西医结合肝病杂志,2014,24(6):342-346.
[26]NOGUEIRA E F,RAINEY W E.Regulation of aldosterone synthase by activator transcription Factor/cAMP response element-binding protein family members[J].Endocrinology,2010,151(3):1060-1070.