Role of Mitochondrial NAD(P)+ Transhydrogenase (NNT) in the Viability and Proliferation of Astrocytes and Glioblastoma Cells in Culture

• Grant number: 25/09703-1
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: August 01, 2025
• End date: July 31, 2027
• Field of knowledge: Biological Sciences - Biochemistry - Metabolism and Bioenergetics
• Principal Investigator: Roger Frigério Castilho
• Grantee: Stephanie Maia Acuna
• Host Institution: Faculdade de Ciências Médicas (FCM). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil
• Associated research grant: 23/00229-0 - Pathophysiological roles of mitochondrial NAD(P)+ transhydrogenase, AP.TEM

Abstract

The mitochondrial transmembrane protein NAD(P)+ transhydrogenase (NNT) catalyzes the transfer of a hydride between NAD(H) and NADP(H), coupled to the translocation of a proton from the intermembrane space into the mitochondrial matrix. This process can occur in the direction of either NADPH or NADH generation (i.e., forward and reverse reactions). Although both reactions are physiologically relevant, the NADPH generated by NNT activity stands out for providing reducing power to peroxide detoxification systems and participating in anabolic reactions, such as the mitochondrial steps of steroidogenesis. NADPH provided by NNT is also relevant to the metabolism and proliferation of tumor cells, suggesting its potential as a therapeutic target.This project aims to elucidate the role of NNT in the proliferation and viability of cultured astrocytes and glioblastoma cells. To this end, comparative studies will be conducted using astrocytes obtained from C57BL/6 mice carrying a spontaneous mutation in the Nnt gene (C57Unib.B6-Nnt-/-), which results in the absence of protein expression, and from congenic control mice expressing this protein (C57Unib.B6-Nnt+/+). Regarding human glioblastoma cells, the cell lines A172, T98G, and U-87MG will be used, employing cell culture techniques, gene silencing by siRNA, and biochemical and molecular analyses.Specifically, we aim to:1) Characterize the importance of NNT in redox signaling and proliferation of cultured astrocytes, as well as the effects of inhibitors of mitochondrial respiratory chain and ATP synthase in this model;2) Investigate the effects of different metabolic states on NNT activity in astrocytes, including the analysis of post-translational modifications;3) Evaluate the effect of NNT silencing on redox signaling, proliferation, and viability of glioblastoma cells and astrocytes in culture, determining the expression of genes related to NADPH metabolism and antioxidant response.The development of this project is expected to contribute to a better understanding of the importance of NNT in the redox metabolism of astrocytes and glioblastoma cells, as well as provide evidence for the potential development of therapeutic strategies directed at modulating NNT activity in glioblastomas. (AU)