Mitochondrial dysfunction dependent on deficiency of Complex I (CI, NADH: ubiquinone oxidoreductase) contributes to cardiac hypertrophy in animal models. NDUFC2 subunit deletion causes CI disassembly and mitochondrial dysfunction. In this project, the impact of NDUFC2 subunit deletion on the cardiac phenotype associated with hypertension will be investigated to understand the molecular mechanism of CI dysfunction underlying impaired mitochondrial function and the hypertrophic phenotype. Two rat models will be used for in-vitro and ex-vivo studies: the spontaneously hypertensive rat (SHR/wt) and the heterozygous Ndufc2-KO SHR (SHR/ndufc2+/-). On cardiac mitochondria from the two models, we will investigate the coupling between the electron transport chain and the ATP synthesis machinery to provide information on the mitochondrial oxidative phosphorylation (OXPHOS) function/dysfunction. The link between the ROS level and the mitochondrial membrane potential (mmp) hormesis, which are related to the respiratory supercomplex (SC) organization and the event of permeability transition pore (PTP) opening, will be considered. The mitochondrial bioenergetic parameters, OXPHOS, mmp, ROS production and PTP activity, the proteomics of SCs and transcriptomics of SCs assembly factors, will be compared in SHR/wt and SHR/ndufc2+/- primary cardiomyocytes. The information obtained at the molecular level will be related to i) the single-cell phenotype analysis upon Ndufc2 silencing; ii) the in-vivo echocardiographic analysis of the two rat strains; iii) the ex-vivo evaluation of protein and gene expression analysis in rat hearts. The gene expression analysis of sirtuin 3 (SIRT3), a mitochondrial NAD+-dependent protein deacetylase, as well as the SIRT3-dependent signalling, will be assessed. A NAD+ booster by nicotinamide riboside (NR) supplementation will be examined. In-vitro, ex-vivo and in-vivo experiments will be translated into the human hypertensive disease by studying the impact of the T allele at the rs11237379/NDUFC2, which is associated with reduced gene expression and CI-dependent mitochondrial dysfunction, on the cardiac phenotype of affected patients. Luciferase reporter assays will be performed to explore the impact of this allele on NDFUC2 transcriptional regulation. Furthermore, peripheral blood mononuclear cells (PBMCs), isolated from venous blood samples of hypertensive patients carrying the T allele, will be used to obtain a large-scale analysis (RNA-Seq and miRNA-Seq) to explain the link between NDUFC2 downregulation and mitochondrial disorder by impaired CI function. The PBMCs bioenergetic parameters, the mitochondrial function and the NR impact to overcome the deleterious NDUFC2 action deficiency on mitochondrial function will be evaluated. This study will provide key mechanistic information on CI dependent deficiency contributing to hypertensive cardiac hypertrophy and will attempt a translation of experimental findings to the human disease.
Codice identificativo 2022E75TWB – CUP B53D23016150006
ENTE FINANZIATORE: Ministero dell’Istruzione, dell’Università e della Ricerca (MIUR)
ANNO INIZIO: 2023
ANNO FINE: 2025
RESPONSABILE SCIENTIFICO: Dott. Flaviana Marzano
