PDE5-Herp通路诱导26S蛋白酶体降解SERCA2a促进心力衰竭的作用和机制

Downregulation of sarcoplasmic reticulum Ca2+-ATPase2a (SERCA2a) is attributed to the reduced contractility in heart failure (HF). But the factors modulating the expression of SERCA2a remains unknown. Our previous study showed that the phosphodiesterase type 5 (PDE5) inhibitor is involved in upregulating SERCA2a protein expression. Recent studies demonstrated that Human homocysteine-induced endoplasmic reticulum(ER) protein (Herp) plays a critical role in recruiting the 26S proteasome to the ER and the 26S proteasome is involved in the process of protein degradation. Our previous study found Herp expression and 26S proteasome activity is increased in HF, PDE5 inhibition can downregulate Herp expression, and that Herp overexpression decreases SERCA2a expression. Based on the findings above, we hypothesize that increased PDE5 expression in HF will upregulate Herp expression, resulting in 26S proteasome-dependent protein degradation of SERCA2a, and reduced cardiac contractility during the development of HF. Therefore, we will use molecular and functional study, both in vivo and in vitro, to verify the regulatory role of PDE5-Herp pathway in modulating SERCA2a expression. The present study may be helpful in elucidating the mechanisms of cardiac contractility modulation, and provide a novel mechanism for preventing HF.

肌浆网Ca2+-ATP酶2a(SERCA2a)表达下调是心力衰竭（心衰）心脏收缩力降低的重要机制，但SERCA2a表达改变的调控机制尚不清楚。我们前期首次发现心衰时磷酸二酯酶5（PDE5）抑制剂能显著缓解SERCA2a表达下调。同型半胱氨酸诱导的内质网蛋白（Herp）可通过召集26S蛋白酶体至内质网，引起多种蛋白降解。我们发现心衰时Herp表达显著增加，26S蛋白酶体活性增强，抑制PDE5可明显降低Herp表达。心室肌细胞过表达Herp后SERCA2a蛋白表达下调。因此，本课题组在前期工作基础上提出假设：心衰时PDE5表达上调并促进Herp表达，后者可通过召集26S蛋白酶体降解SERCA2a，导致心脏收缩力下降，促进心衰发展。我们将应用分子生物学和功能学技术，在体和细胞水平验证上述假设，深入阐明上述信号通路调控SERCA2a的分子机制和在心衰发生及进展中的确切作用，为心衰防治提供新靶点。

研究背景：心力衰竭（心衰）是各种心脏疾病晚期的严重表现，我国心衰患者数量庞大，发病率不断增长，死亡率和再住院率居高不下，是21世纪主要的心血管疾病之一。肾素-血管紧张素系统（RAAS）激活可导致心力衰竭恶化，抑制血管紧张素-2(Ang Ⅱ) 活性能延缓心衰的进展。然而近年大量临床证据表明，作用于RAAS（ACEI和ARB）的药物对心衰的改善程度有限，因为在病理状态下，Ang-(1-12)-Chymase途径是产生AngII的主要方式，而非经典的ACE途径。然而，目前Ang-(1-12)对心脏功能的直接作用和机制尚不清楚。主要研究内容：本研究将探索外源性Ang-(1-12)对正常和心衰大鼠左心室心肌收缩力的影响，Ang-(1-12)对单个心肌细胞收缩和舒张功能影响，细胞水平验证Ang-(1-12)-Chymase通路对SERCA2a表达的调控作用。重要结果和关键数据：我们首次发现外源性Ang-(1-12) 能增加正常对照大鼠左心室收缩末期压力，对心率、动脉弹性、左室舒张末期压力，左室舒张时间和射血分数无影响，静脉注射Ang-(1-12)能显著增加左室心肌收缩力。然而，在心衰大鼠外源性Ang-(1-12)确表现出截然相反的效果，和心衰给药前基线比较，Ang-(1-12)显著延长了左室舒张时间，增加动脉僵硬度和舒张末期压力，降低射血分数，使左室压力容积环向右上偏移，Ang-(1-12)显著降低心衰大鼠左室射血分数，每搏输出量和收缩力（EES 和MSW显著降低），使左室-动脉耦合指数下降43%。心衰静息时Ang-(1-12)使左心室压力-容积曲线右移，并使左室收缩末期压力-容积关系斜率[Ang-(1-12): 4.4 vs 基线： 5.6 mmHg/ml]和SW- VED关系斜率[Ang-(1-12): 60.2 vs 基线：68.5mmHg]降低。以上结果说明Ang-(1-12)对心衰清醒狗左室收缩功能产生负性肌力作用。左室-动脉偶联EES/EA也降低68%（Ang-(1-12)：0.38 vs 基线：0.64）。科学意义：本试验的重要性在于它证实了Ang-(1-12)-Chymase通路能促进心力衰竭发展，为治疗心力衰竭提供新靶点。