NHE1 AND MITOCHONDRIAL DYSFUNCTION

DE FAZIO R; GODOY COTO J; CIANCIO MC; ORLOWSKI A,; AIELLO EA; JAQUENOD DE GUISTI C

Congreso; Reunión Anual SAFIS 2023; 2023

Introduction: Mitochondria play vital roles in both energy production and pathogenesis. Their dysfunction is linked to various diseases, such as heart failure and diabetes. NHE1, a key alkalinizing mechanism in the heart, is hyperactive in heart conditions like diabetic cardiomyopathy. In obese and diabetic mice (ob/ob), mitochondrial NHE1 expression increases despite similar cardiac levels, along with disrupted calcium handling, which can be restored with an NHE1 inhibitor.Objectives: understanding the consequences of increased mitochondrial NHE1 expression. Methods: Mitochondria were isolated from ob/ob mice. ∆ψm was measured with Rhodamine 123. PTP opening was assessed by monitoring Ca+2 release through the CRC assay with CsA as an inhibitor. Mitochondrial Ca+2 content was determined using Fluo-3 and a chemical assay. For the in vitro model HEK293T cells were transfected with pmitoNHE1, and ∆ψm was quantified as JC-1 aggregate/monomer fluorescence. PTP opening was measured by the release of calcein. ATP levels and mitochondrial ROS production were evaluated in both transfected cells and isolated mitochondria using a commercial ATP assay kit and H2DCFDA indicator, respectively. Data were analyzed with Student's t-test (means ± SEM). Results: In isolated mitochondria from ob/ob mice, membrane hyperpolarization and reduced calcium content were observed, accompanied by a decrease in ATP content and an increase in ROS. These changes came together with an increased sensitivity for PTP opening. In the in vitro model, mitochondrial NHE1 overexpression resulted in mitochondrial membrane hyperpolarization, accompanied with an increased ROS production and more sensibility for PTP opening. These cells also presented lower ATP levels. Conclusion: our results show a correlation between increased mitochondrial NHE1 expression with altered mitochondrial membrane potential, ROS production and ATP levels.