Regulation of Vascular Smooth Muscle Calcium by NADPH Redox

NADPH 氧化还原对血管平滑肌钙的调节

• 批准号: 8204769
• 负责人: SACHIN A GUPTE
• 金额: $ 36.75万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-01-01 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8204769
• 关键词: 6-Aminonicotinamide; 6-phosphogluconate; Address; Angiotensin II; Aorta; Binding; Biochemical; Biochemical Reaction; Biological Assay; Blood Vessels; Calcium; Calcium Channel; Cell Survival; Cell membrane; Cell physiology; Cells; Co-Immunoprecipitations; Commit; Complex; Confusion; Contracts; Coronary; Coronary artery; Critiques; Cysteine; Cytosol; DNA; Data; Deletion Mutation; Diabetes Mellitus; Disease; Drug Delivery Systems; Electrophoresis; Enzymes; Epiandrosterone; Functional disorder; Future; Genes; Glucose; Glucose-6-Phosphate; Glucosephosphate Dehydrogenase; Glutathione; Glutathione Disulfide; Glycolysis; Goals; Grant; Heart; Heart failure; Human; Hydrogen Peroxide; Hypertension; In Vitro; Influentials; Ion Channel; Ion Channel Protein; L-Type Calcium Channels; L-type calcium channel alpha(1C); Label; Laboratories; Link; Lung; Measures; Mediating; Membrane Potentials; Metabolic; Metabolic Diseases; Metabolic Pathway; Methods; Modification; Mus; Muscle function; Myocardial; NADP; Oxidation-Reduction; Oxidoreductase; PMCA1 protein; Pathway interactions; Pentosephosphate Pathway; Perfusion; Phosphorylation; Play; Principal Investigator; Process; Protein Kinase C; Proteins; Protocols documentation; Publishing; Pulmonary Hypertension; Pulmonary artery structure; RNA chemical synthesis; Radio; Radioisotopes; Regulation; Relaxation; Rest; Role; Sarcoplasmic Reticulum; Signal Pathway; Signal Transduction; Signaling Protein; Site; Site-Directed Mutagenesis; Small Interfering RNA; Smooth Muscle; Smooth Muscle Myocytes; Sodium-Calcium Exchanger; Techniques; Testing; Thromboxane A2; Tracer; Transfection; Vascular Smooth Muscle; Vasomotor; analog; attenuation; base; channel blockers; dehydroepiandrosterone; glucose metabolism; inhibitor/antagonist; innovation; mutant; mutant mouse model; novel; novel therapeutic intervention; oxidation; patch clamp; programs; research study; response; ribose-5-phosphate; thioredoxin glutathione reductase; voltage

evidence that the pentose phosphate pathway (PPP)/glucose-6-phosphate dehydrogenase (G6PD) and NADPH redox is involved in modulating contractile function of the coronary (CA) artery. However, the machanism(s) by which G6PD and NADPH modulates contractile function of CA are obscure. Therefore, the primary focus of this proposal will be, to elucidate the signaling pathways involved in mediating the effects of G6PD and NADPH redox on smooth muscle cell L-type Ca2+ currents and CA function. To achieve these goals, we will, in Aim #1 determine if G6PD is active in the sub- cellular fractions of resting and contracting CA, by estimating the rate of glucose oxidation, and the G6PD activity levels by biochemical and radioisotope tracer assays. Furthermore, we will identify mechanism(s) involved in contractile agents-induced-G6PD activation, by investigating the role of PKC and metabolic pathways. In Aim #2, we will determine whether G6PD mediates L-type Ca2+ channel activity, intracellular Ca2+, and vasomotor tone in resting and contracting CA, by examining L-type Ca2+ function, measure intracellular Ca2+ changes and vasomotor function after inhibiting G6PD with pharmacological agents and siRNA transfection, and in G6PD deficient mouse aorta. In Aim #3, we will determine whether glucose-6-phosphate dehydrogenase modulates the L-type Ca2+ channel function, intracellular Ca2+, contraction and redox changes, in CA via direct physical interaction with the ion channel proteins (alpha subunit of CaV1.2), by co-immunoprecipitation, co-localization and in-vitro binding assays. Additionally, we will determine whether direct binding of NADP+ or NADPH to the L-type Ca2+ channel protein inactivates the channel and whether changes in the levels of reduced/oxidized glutathione (GSH) or hydrogen peroxide (H2O2), induced by decrease in NADPH levels (due to the inhibition of G6PD activity), modulates L-type Ca2+ channel function, in smooth muscle cells isolated from coronary and aorta of G6PD deficient mouse. The PPP/G6PD and NADPH redox is up-regulated in diabetes, pulmonary hypertension and heart failure, thereby suggesting a potential role for G6PD and NADPH redox in profoundly impairing the contractile function of blood vessels in these diseases. This study, on completion as anticipated, will prove to be useful in developing novel therapies for the treatment of vascular dysfunction in pulmonary hypertension, diabetes, and heart failure. Project Description Page 6

证明戊糖磷酸途径（PPP）/葡萄糖-6-磷酸 脱氢酶（G6 PD）和NADPH氧化还原参与调节冠状动脉的收缩功能 (CA)动脉然而，G6 PD和NADPH调节心肌收缩功能的机制是不确定的。 CA是模糊的。因此，本提案的主要重点将是阐明信号通路 参与介导G6 PD和NADPH氧化还原对平滑肌细胞L型钙电流的影响 CA功能。为了实现这些目标，我们将在目标#1中确定G6 PD是否在亚细胞中活跃， 静息和收缩CA的细胞分数，通过估计葡萄糖氧化速率，和G6 PD 通过生物化学和放射性同位素示踪分析确定放射性水平。此外，我们将确定机制， 参与收缩剂诱导的G6 PD激活，通过研究PKC和代谢的作用， 途径。在目标#2中，我们将确定G6 PD是否介导L型Ca 2+通道活性， 细胞内Ca 2+和血管紧张素在静息和收缩CA，通过检查L型Ca 2+功能， 用药物抑制G6 PD后测定细胞内Ca ~（2+）变化和血管功能 试剂和siRNA转染，并在G6 PD缺陷型小鼠主动脉中。在目标#3中，我们将确定 葡萄糖-6-磷酸脱氢酶是否调节L-型Ca 2+通道功能，细胞内 CA中的Ca 2+、收缩和氧化还原变化通过与离子通道蛋白的直接物理相互作用 （CaV1.2的α亚基），通过免疫共沉淀、共定位和体外结合测定。 此外，我们将确定NADP+或NADPH是否直接结合到L型Ca 2+通道， 蛋白质失活通道，以及还原型/氧化型谷胱甘肽（GSH）水平的变化 或过氧化氢（H2 O2），由NADPH水平降低诱导（由于G6 PD抑制 活动），调节L-型钙通道功能，在平滑肌细胞分离的冠状动脉和 G6 PD缺陷小鼠的主动脉。PPP/G6 PD和NADPH氧化还原在糖尿病中上调， 肺动脉高压和心力衰竭，从而表明G6 PD和NADPH的潜在作用 氧化还原在这些疾病中严重损害血管的收缩功能。这项研究，在 如预期的那样完成，将被证明在开发用于治疗以下疾病的新疗法中是有用的： 肺动脉高压、糖尿病和心力衰竭中的血管功能障碍。 项目描述第6页

项目成果

Glucose-6-phosphate dehydrogenase: a novel therapeutic target in cardiovascular diseases.

• 发表时间: 2008-09
• 期刊: Current opinion in investigational drugs
• 作者: S. Gupte

Effects of laparoscopic Roux-en-Y gastric bypass on glucose-6 phosphate dehydrogenase activity in obese type 2 diabetics.

腹腔镜 Roux-en-Y 胃绕道手术对肥胖 2 型糖尿病患者葡萄糖-6 磷酸脱氢酶活性的影响。

• DOI: 10.1007/s00464-011-1959-8
• 发表时间: 2012
• 期刊: Surgical endoscopy
• 作者: Schneider,AndrewM;Rawat,Dhwajbahadur;Weinstein,LSteve;Gupte,SachinA;Richards,WilliamO
• 通讯作者: Richards,WilliamO

Glucose-6-phosphate dehydrogenase and NADPH redox regulates cardiac myocyte L-type calcium channel activity and myocardial contractile function.

• DOI: 10.1371/journal.pone.0045365
• 发表时间: 2012
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Rawat DK;Hecker P;Watanabe M;Chettimada S;Levy RJ;Okada T;Edwards JG;Gupte SA
• 通讯作者: Gupte SA

Cyp2c44 gene disruption is associated with increased hematopoietic stem cells: implication in chronic hypoxia-induced pulmonary hypertension.

Cyp2c44 基因破坏与造血干细胞增加有关：对慢性缺氧诱导的肺动脉高压的影响。

• DOI: 10.1152/ajpheart.00785.2016
• 发表时间: 2017
• 期刊: American journal of physiology. Heart and circulatory physiology
• 作者: Hashimoto,Ryota;Joshi,SachindraRaj;Jiang,Houli;Capdevila,JorgeH;McMurtry,IvanF;LaniadoSchwartzman,Michal;Gupte,SachinA
• 通讯作者: Gupte,SachinA

Cholesterol depletion modulates basal L-type Ca2+ current and abolishes its -adrenergic enhancement in ventricular myocytes.

• DOI: 10.1152/ajpheart.00824.2007
• 发表时间: 2008
• 期刊: American journal of physiology. Heart and circulatory physiology
• 作者: H. Tsujikawa;Y. Song;Makino Watanabe;H. Masumiya;S. Gupte;R. Ochi;T. Okada