Deficiency Of Claudin-5 Leads To Myocardial Atrophy And Dilation In Mouse Heart With Severe Mitochondrial Dysfunction And Electrical Dissociation

Background: Cardiac overexpression of Claudin-5 (Cldn5) exerts protection against myocardial ischemia and reperfusion injury. However, whether cardiac deficiency of Cldn5 affects cardiac function is still unclear. Here we investigated the role of Cldn5 in mouse heart by means of cardiac Cldn5 knockdown to better understand the intracellular function of Cldn5 in cardio-protection. Methods and Results: Cldn5 shRNA adeno-associated virus (AAV) was used to knockdown Cldn5 in the left ventricle of male C57BL/6J mice by intramyocardial injection. Four weeks post AAV injection, echocardiography and electrocardiography were performed to measure cardiac function. We found 25% of mice died in Cldn5 shRNA AAV injection group. The residual mice were terminated followed by left ventricles isolation for proteomics analysis, cytokine analysis, WGA staining, Masson-trichrome staining and transmission electron microscope (TEM) observation. The signal pathway markers of apoptosis, autophagy and lipid peroxidation were also detected by western blot. The protein level of Cldn5 in left ventricles was decreased by 75% in shRNA AAV injection group accompanied by changes of bcl-2, bax, LC3II, p62, Cox2 and GPX4. The fraction shortening of the heart was decreased by 23.5% with ventricular dilation and ST elevation. We also observed the increase of IL-1beta, TNF alpha, BNP, CT and cTnI in myocardium. WGA staining showed the area of cross section of single cardiomyocyte was decreased and Masson staining showed the collagen deposition was increased in myocardium. The TEM indicated Cldn5 knockdown induced mitochondrial fission and depletion. Finally, the proteomics analysis demonstrated that Cldn5 knockdown severely disturbed ATP metabolic regulation, mitochondrial permeability transition pore opening, calcium ion transport, cardiac contraction, endoplasmic reticulum stress response and antioxidant activity in the AAV injected myocardium. Conclusion: We demonstrated for the first time the deficiency of Cldn5 in myocardium caused myocardial atrophy and dilation with deteriorative heart function. The mechanism of Cldn5 deficiency-induced cardiac dysfunction is associated with severe mitochondrial dysfunction and excitation-contraction coupling.