The role of plasminogen activator inhibitor-1 (PAI-1) in mediating age-related fi

描述（由申请方提供）：心血管老化导致心肌纤维化、舒张功能障碍和心肌梗死（MI）后伤口愈合减弱。虽然许多研究已经将纤溶酶原激活物抑制剂-1（派-1）的增加与衰老相关联，但是没有一项研究直接研究派-1介导的胶原蛋白周转抑制的可能机制。这项研究旨在确定派-1在调节年龄相关性心脏纤维化中的作用。派-1依赖性升高与基质金属蛋白酶（MMP）活性降低、胶原沉积增加和炎症反应减弱相关。作为受损的ECM降解和成纤维细胞迁移的结果，MMP活性的降低可能通过增加胶原沉积和减少MI后的炎症反应而引起心室纤维化。 将使用分离的大鼠心脏成纤维细胞（CF）来研究派-1释放与对胶原沉积和CF迁移的潜在影响之间的关系。本研究的第一个目的是询问由CFs产生和释放的派-1是否通过抑制MMP活化来增加胶原沉积。将通过蛋白质印迹法、酶谱法和ELISA测定CF中的总MMP和活性MMP。第二个目的是比较从年轻和老年大鼠分离的CFs中派-1的表达，以检查表达是否随年龄增加而增加，并负责增加胶原蛋白的积累。假设派-1随年龄增加而减弱MMP活性。这反过来又减少了胶原降解，也可能抑制CF迁移。最后，本研究的第三个目的将使用离体大鼠缺血-再灌注模型。这提供了一个系统，以确定是否派-1的表达是增加的基础上，在老年心脏和派-1是否上调，在生理相关的浓度缺血应激。派-1对MMP活性的急性抑制可以解释老年人MI后炎症和伤口愈合反应减弱的原因。总之，这些发现可能确定了一种新的派-1依赖性机制，其中老年心脏更容易在缺血性损伤后发生纤维化和伤口愈合减弱。 公共卫生相关性：心脏纤维化导致心脏硬化、肺部积液和心脏效率降低，并可能干扰损伤后的适当伤口愈合。年龄是心脏纤维化发展的一个重要因素，因为所有老年心脏都经历胶原沉积并遭受减弱的愈合反应。本研究旨在研究纤溶酶原激活物抑制剂-1，一种随年龄增加的蛋白质，可通过增加纤维蛋白沉积量和降低心脏损伤后的愈合反应对心脏产生负面影响。

DESCRIPTION (provided by applicant): Cardiovascular aging results in cardiac fibrosis, diastolic dysfunction, and attenuated wound healing post-myocardial infarction (MI). While numerous studies have correlated increases in plasminogen activator inhibitor-1 (PAI-1) with aging, none have directly examined possible mechanisms of PAI-1-mediated inhibition of collagen turnover. The proposed study seeks to identify a role of PAI-1 in regulating age-related cardiac fibrosis. Age-dependent increases of PAI-1 correlate with decreased matrix metalloprotease (MMP) activity, increased collagen deposition, and blunted inflammatory response. As a consequence of impaired ECM degradation and fibroblast migration, the reduction in MMP activity may cause ventricular fibrosis by increasing collagen deposition and diminished inflammatory responses post-MI. Isolated rat cardiac fibroblasts (CFs) will be used to investigate the relationship between PAI-1 release and potential effects on collagen deposition and CF migration. The first aim of this study asks whether PAI-1 produced and released by CFs increases collagen deposition by inhibiting MMP activation. Total and active MMPs will be assayed in CFs by Western blotting, zymography, and ELISA. The second aim compares PAI-1 expression in CFs isolated from young and aged rats to examine whether expression increases with age and is responsible for increased collagen accumulation. The increase of PAI-1 with age is hypothesized to attenuate MMP activity. This in turn decreases collagen degradation and may also inhibit CF migration. Finally, the third aim of the study will use an ex vivo rat ischemia-reperfusion model. This provides a system to ascertain whether PAI-1 expression is increased basally in aged hearts and whether PAI-1 is up-regulated in physiologically relevant concentrations following ischemic stress. Acute inhibition of MMP activity by PAI-1 may explain the attenuated inflammatory and wound healing responses post-MI seen in aged individuals. Together, these findings may identify a novel PAI-1 dependent mechanism in which aged hearts are more prone to both fibrosis and attenuated wound healing following ischemic damage. PUBLIC HEALTH RELEVANCE: Cardiac fibrosis leads to stiffening of the heart, fluid buildup in the lungs, and decreased cardiac efficiency and can interfere with proper wound healing after injury. Age is an important factor in the development of cardiac fibrosis as all aged hearts undergo collagen deposition and suffer from attenuated healing responses. This study seeks to investigate plasminogen activator inhibitor-1, a protein that increases with age and can negatively affect the heart by increasing the amount of fibrous protein deposition and decreasing the healing response after cardiac injury.