Mechanisms of Muscle Microcirculatory Dysfunction in Heart Failure

描述（由申请人提供）：慢性心力衰竭（CHF）的运动不耐受有大量的骨骼肌成分。骨骼肌的反复收缩需要精确匹配氧气输送(QO2) -氧气利用（VO2）。然而，CHF会损害毛细血管血流动力学，从而降低QO2/VO2比率和微血管O2压，并损害心肌细胞O2通量。我们独特的活体显微镜模型（大鼠斜方肌-一种适合运动训练的肌肉）有助于直接观察肌肉微循环和高时间保真度-在收缩期间使用磷光猝灭来测定血液-肌细胞O2通量。基于观察到CHF降低肌肉一氧化氮（NO）的生物利用度，而NO的急性增加恢复了CHF中QO2-to-VO2的匹配，该提案将测试旨在提高NO生物利用度的多种策略，无论是直接（运动训练，增加四氢生物蝶呤（BH4，通过sepiapterin））还是通过调节活性氧（ROS，通过罗布宁，Gp91 ds-tat和Tempol降低O2O-）；减少OHO-通过去铁胺)将改善毛细血管血流动力学和心肌细胞氧通量。认识到CHF的炎症成分，我们将研究是否可以通过减少炎症介质IL-12（使用anakinra， IL-12受体拮抗剂）和TNF1（使用己酮茶碱）来改善CHF的微血管缺陷。该提案的显著优势包括：1。己酮茶碱，降低循环TNF-1，已被证明对人CHF有效，而TNF-1阻滞剂依那西普（重组TNF受体）和英夫利昔单抗（单克隆TNF-1抗体），例如，可能会加重病情。我们将检验己酮茶碱改善收缩期间毛细血管血流动力学的假设，并评估一氧化氮在这一过程中的作用。2. 2 .鼓励本科生和研究生参与有价值的前沿科学研究；通过定量确定一氧化氮生物利用度实际上促进毛细血管血流动力学的程度，避免了在收缩肌肉中进行精确微米级[NO]测量的技术上的不可能性。这将在每个实验条件下通过NOS阻断（L-NAME）和外源NO应用（硝普钠和/或NONOate）来完成。预计这些研究将确立一氧化氮在促进健康状态下毛细血管血流动力学和血细胞氧通量对肌肉收缩反应中的前哨作用，并揭示一氧化氮生物利用度的缺陷——由一氧化氮产生途径的多个步骤的功能障碍引起，包括bh4诱导的一氧化氮解偶联和ROS介导的一氧化氮破坏——是导致CHF诱导功能障碍的原因。拟议的研究将为心力衰竭患者肌肉毛细血管血流动力学功能障碍的机制提供新颖而重要的数据，并评估己酮茶碱治疗心力衰竭患者疗效的机制基础，同时履行AREA奖的任务，将真实的研究与本科生/研究生教育相结合。

DESCRIPTION (provided by applicant): The exercise intolerance of chronic heart failure (CHF) has a substantial skeletal muscle component. Repeated contractions of skeletal muscle require precise matching of O2 delivery (QO2) - to-O2 utilization (VO2). However, CHF impairs capillary hemodynamics thereby lowering the QO2/VO2 ratio and microvascular O2 pressure and compromising blood-myocyte O2 flux. Our unique intravital microscopy model (rat spinotrapezius - a muscle amenable to exercise training) facilitates direct observation of muscle microcirculation and high temporal fidelity-determination of blood-myocyte O2 flux using phosphorescence quenching during contractions. Building on the observation that CHF decreases muscle nitric oxide (NO) bioavailability and acute increases of NO restore QO2-to-VO2 matching in CHF this proposal will test the global hypothesis that multiple strategies designed to increase NO bioavailability either directly (exercise training, increased tetrahydrobiopterin (BH4, via sepiapterin)) or via modulation of reactive O2 species ( ROS, reduced O2O-, via apocynin, Gp91 ds-tat, and Tempol; reduced OHO- via deferoxamine) will improve capillary hemodynamics and blood-myocyte O2 flux. Recognizing the inflammatory component of CHF, we will investigate whether microvascular deficits in CHF can be ameliorated by reduction of inflammatory mediators IL-12 (using anakinra, IL-12 receptor antagonist) and TNF1 (using pentoxifylline). Significant strengths of this proposal include: 1. Pentoxifylline, which lowers circulating TNF-1, has proven efficacy in human CHF whereas TNF-1 blockers etanercept (recombinant TNF receptor) and infliximab (monoclonal TNF-1 antibody), for example, may worsen the condition. We will test the hypothesis that pentoxifylline improves capillary hemodynamics during contractions and assess the role of NO in this process. 2. Involvement of undergraduate and graduate students in meritorious cutting-edge scientific research, and 3. Avoiding the technical impossibility of making precise micron-level [NO] measurements in contracting muscle by determining quantitatively the extent to which NO bioavailability actually facilitates capillary hemodynamics. This will be accomplished using conditions of NOS blockade (L-NAME) and exogenous NO application (sodium nitroprusside and/or NONOate) under each experimental condition. It is anticipated that these investigations will establish a sentinel role for NO in facilitating the capillary hemodynamics and blood-myocyte O2 flux response to muscle contractions in health and reveal that deficits in NO bioavailability in CHF - evoked by dysfunction at multiple steps in the NO production pathway including BH4-induced NOS uncoupling and ROS- mediated NO destruction - are causative to CHF-induced dysfunction. The proposed studies will provide novel and important data addressing the mechanisms of muscle capillary hemodynamic dysfunction in CHF and assess the mechanistic bases for the efficacy of pentoxifylline in treating CHF patients whilst fulfilling the AREA award mandate to integrate authentic research with undergraduate/graduate student education. PUBLIC HEALTH RELEVANCE: The exercise intolerance of patients suffering from chronic heart failure (CHF) is based substantially in skeletal muscle and, irrespective of left-ventricular function per se, results from a mismatching of oxygen delivery to oxygen demands in contracting muscles. Recent evidence indicates that this mismatching is the consequence of impaired capillary blood flow secondary to decreased nitric oxide bioavailability. This proposal assesses the efficacy of strategies to increase nitric oxide bioavailability to improve capillary hemodynamics, oxygen delivery/utilization matching and muscle function in CHF. One key aspect is determination of the mechanistic bases for the efficacy of pentoxifylline to treat CHF patients. Under these auspices and the AREA Award mission this proposal integrates meritorious science with opportunities for development of undergraduate and graduate student scientists.