EXERCISE, DIABETES, AND CORONARY SMOOTH MUSCLE CALCIUM

我们的长期目标是确定运动训练是否能减轻糖尿病患者的过量冠状动脉疾病（CAD）和所涉及的血管平滑肌（VSM） Ca （Cam）信号机制。在我们的猪糖尿病血脂异常模型中，冠状动脉对前列腺素f2 - α (PGF)的收缩增加，Cam对内皮素-1 （ET）的反应增加。正常猪的运动减少了肌浆网（SR）的钙释放，增加了钙内流，同时减少了et诱导的收缩和DNA合成。然而，et诱导的收缩不需要依赖于酪氨酸激酶的钙释放，酪氨酸激酶是细胞生长的信号。总体假设：运动训练后，增加的et诱导的Ca内流减弱收缩，而减少的SR Ca释放减弱平滑肌的生长。设计：低脂对照猪(C)、高脂喂养猪（HF）、四氧胺糖尿病猪和高脂喂养猪(D)分别维持20和30周，研究CAD的进展情况，并与D组猪（D + EX）进行比较。具体目的是验证糖尿病患者的假设：1)运动改善血糖和血脂状况。糖尿病将由血糖、糖化蛋白、胰岛素等指标来定义。HDL、LDL、VLDL、甘油三酯、载脂蛋白、糖化LDL和ET将定义糖尿病性血脂异常的特征。2)运动可减弱冠心病的增加。血管内超声将评估动脉粥样硬化和血管收缩对体内PGF和ET的反应。冠状动脉环对PGF和ET的组织学和收缩张力反应将提供CAD的体外测量。3)运动减弱了VSM从收缩表型（cVSM）向合成表型（sVSM）的转化。sVSM细胞表型将使用数字成像显微镜通过Cam对UTP、核周SR、DNA、平滑肌肌动蛋白、desmin和vimentin的反应来鉴定。4)运动通过增加ET受体依赖性钙内流而减弱cVSM的增加收缩。用锰和钡内流作为钙的替代物来评估钙内流。相对于ryanodine受体的肌上皮下Ca定位将被数字成像。5)运动通过减少sr中酪氨酸激酶依赖性钙的释放，减弱了ET诱导的Cam振幅和核钙定位的增加。成像将在单细胞中检测ET、ryanodine和IP3受体、酪氨酸磷酸化、DNA和Cam的相对含量和空间分布。意义：首次研究糖尿病血脂异常中过量CAD的钙定位机制及运动对CAD的治疗作用。

Our long-term goal is to determine if exercise training attenuates the excess coronary artery disease (CAD) in diabetes and the vascular smooth muscle (VSM) Ca (Cam) signaling mechanisms involved. In our porcine model of diabetic dyslipidemia coronary arteries show increases in contraction to prostaglandin F2alpha (PGF) and Cam responses to endothelin-1 (ET). Exercise of normal swine decreases Ca release from the sarcoplasmic reticulum (SR) and increases Ca influx, while decreasing ET-induced contraction and DNA synthesis. However, ET-induced contraction does not require Ca release that is dependent on tyrosine kinase, a signal for cell growth. Overall hypothesis: after exercise training increased ET-induced Ca influx attenuates contraction, while decreased SR Ca release attenuates growth of smooth muscle. Design: low fat control pigs (C), high fat fed (HF), and alloxan diabetic and high fat fed (D) pigs are maintained for 20 and 30 wk to study the progression of CAD and compared to D pigs exercise trained (D plus EX). Specific Aims are to test the hypotheses that in diabetes: 1) Exercise improves glycemic and lipidemic status. Diabetes will be defined by measures of blood glucose, glycated protein, insulin, etc. HDL, LDL, VLDL, triglycerides, apoproteins, glycated LDL, and ET will define features of diabetic dyslipidemia. 2) Exercise attenuates the increase in CAD. Intravascular ultrasound will assess atheroma and vasoconstrictor responses to PGF and ET in vivo. Histology and contractile tension responses of coronary rings to PGF and ET will provide in vitro measures of CAD. 3) Exercise attenuates the conversion of VSM from the contractile (cVSM) to synthetic (sVSM) phenotype. sVSM cell phenotype will be identified using digital imaging microscopy by the Cam response to UTP, perinuclear SR, DNA, smooth muscle actin, desmin, and vimentin. 4) Exercise attenuates the increased contraction of cVSM by increasing ET receptor-dependent Ca influx. Mn and Ba influx used as Ca surrogates will assess Ca influx. Subsarcolemmal Ca localization relative to ryanodine receptors will be digitally imaged. 5) Exercise attenuates the increased ET-induced Cam amplitude and nuclear Ca localization by decreasing tyrosine kinase-dependent Ca release from the SR. Imaging will assay in single cells the relative content and spatial distribution of ET, ryanodine, and IP3 receptors, tyrosine phosphorylation, DNA and Cam. Significance: the first study of Ca localization mechanisms involved in the excess CAD in diabetic dyslipidemia and the therapeutic effects of exercise on CAD.