Abstract 15253: Mitochondrial Trifunctional Protein HADHA Regulates Mitochondrial Cardiolipin Content in Right Ventricle in Pulmonary Hypertension

Abstract

Background: Pulmonary hypertension (PH) is prevalent in cardiopulmonary and metabolic diseases. Unmet energy needs due to mitochondrial (mito) dysfunction results in myocardial contractile failure, as seen in the dysfunctional right ventricles (RV). Cardiolipin (CL) is a critical lipid for mito structure and cristae organization that are essential for mito oxidative phosphorylation and ATP generation. The synthesis and maturation of CL involves many steps including a final step where acylation of monolysocardiolipin occurs and CL[18:1] 2 [18:2] 2 is converted to mature CL[18:2] 4 catalyzed by a mito trifunctional protein, HADHA. To date, the expression of mature CL and its regulation by HADHA in RV in PH remains unknown. Hypothesis: We hypothesize that HADHA regulates mito CL contents in RV in PH. Methods: Adult Sprague Dawley rats received 20 mg/kg sc SU5416 were exposed to 3 wks of hypoxia (10% FiO2), followed by 4 wks of normoxia. Control rats injected with vehicle were kept in normoxia. Hemodynamic and echocardiography were performed, followed by HADHA and CL analysis. We also generated in vitro models with H9c2 cells exposure in hypoxia and stable HADHA-knockdown H9c2 cells for mechanistic studies. Results: PH rats show RV dysfunction. Mito proteomics show a 70% decrease in HADHA expression in mito from RV of PH rats in comparison to the control rats, which is further validated by Western Blot and qPCR. Using mass spectrometry, mature CL (CL[18:2] 4 ) in RV of PH rats was found to decrease to 55% of that in RV of control rats (n=4-5/group, p <0.05). Exposing the H9c2 cells to hypoxia (1% O 2 ) for 72 hrs recapitulate the downregulation of HADHA observed in the dysfunctional RV of PH rats, which is associated with a 70% reduction in the account of mature CL (n=6 each, p <0.05). Importantly, adenoviral overexpression of HADHA almost brings the mature CL content in hypoxia to be comparable to cells in normoxia (n=6 each, p <0.05). Our data also show that HADHA-specific knockdown is sufficient to decrease mature CL in H9c2 cells (Control 100% vs. HADHA Knockdown 33%, n=3 each, p <0.05). Conclusions: Our data demonstrate that HADHA is critical in maintaining mature mito CL contents. Downregulation of HADHA contributes to reduced mito CL contents in RV in settings of PH.