Metformin protects against diclofenac-induced toxicity in primary rat hepatocytes by preserving mitochondrial integrity via a pathway involving EPAC

ABSTRACT: Background and purpose: It has been shown that the antidiabetic drug metformin protects hepatocytes against toxicity by various stressors. Chronic or excessive consumption of diclofenac (DF) - a pain-relieving drug, leads to drug-induced liver injury via a mechanism involving mitochondrial damage and ultimately apoptotic death of hepatocytes. However, whether metformin protects against DF-induced toxicity is unknown. Recently, it was also shown that cAMP elevation is protective against DF-induced apoptotic death in hepatocytes, a protective effect primarily involving the downstream cAMP effector EPAC and preservation of mitochondrial function. This study therefore aimed at investigating whether metformin protects against DF-induced toxicity via cAMP-EPACs. Experimental approach: Primary rat hepatocytes were exposed to 400 µmol/L DF. CE3F4 or ESI-O5 were used as EPAC-1 or 2 inhibitors respectively. Apoptosis was measured by caspase-3 activity and necrosis by Sytox green staining. Seahorse X96 assay was used to determine mitochondrial function. Mitochondrial reactive oxygen species (ROS) production was measured using MitoSox, mitochondrial MnSOD expression was determined by immunostaining and mitochondrial morphology (fusion and fission ratio) by 3D refractive index imaging. Key results: Metformin (1 mmol/L) was protective against DF-induced apoptosis in hepatocytes. This protective effect was EPAC-dependent (mainly EPAC-2). Metformin restored mitochondrial morphology in an EPAC-independent manner. DF-induced mitochondrial dysfunction which was demonstrated by decreased oxygen consumption rate, an increased ROS production and a reduced MnSOD level, were all reversed by metformin in an EPAC-dependent manner. Conclusion and implications: Metformin protects hepatocytes against DF-induced toxicity via cAMP-dependent EPAC-2. Keywords: Apoptosis; CAMP; Diclofenac; EPAC; Hepatocyte; Metformin; Mitochondria.

• Autores: Arroyave Ospina, Johanna Carolina; Aguilar Mora, Fabio Alejandro; Musheshe, Nshunge; Geng, Yana; Soto, Juan M.; Rodrigo, José A.; Alieva, Tatiana; Buist Homan, Manon; Lezoualc'h, Frank; Cheng, Xiaodong; Schmidt, Martina; Moshage, Han
• Tipo de recurso: Article of investigation
• Fecha de publicación: 2021
• Institución: Universidad de Antioquia
• Repositorio: Repositorio UdeA
• Idioma: eng
• OAI Identifier: oai:bibliotecadigital.udea.edu.co:10495/40154
• Acceso en línea: https://hdl.handle.net/10495/40154
• Palabra clave: Antioxidantes; Antioxidants; Apoptosis; Apoptosis; Caspasa 3; Caspase 3; Células Cultivadas; Cells, Cultured; Enfermedad Hepática Inducida por Sustancias y Drogas; Chemical and Drug Induced Liver Injury; AMP Cíclico; Cyclic AMP; Inhibidores de la Ciclooxigenasa; Cyclooxygenase Inhibitors; Diclofenaco; Diclofenac; Factores de Intercambio de Guanina Nucleótido; Guanine Nucleotide Exchange Factors; Hepatocitos; Hepatocytes; Metformina; Metformin; Mitocondrias Hepáticas; Mitochondria, Liver; Estrés Oxidativo; Oxidative Stress; Cultivo Primario de Células; Primary Cell Culture; Ratas Wistar; Rats, Wistar; https://id.nlm.nih.gov/mesh/D022781; https://id.nlm.nih.gov/mesh/D000975; https://id.nlm.nih.gov/mesh/D017209; https://id.nlm.nih.gov/mesh/D053148; https://id.nlm.nih.gov/mesh/D002478; https://id.nlm.nih.gov/mesh/D056486; https://id.nlm.nih.gov/mesh/D000242; https://id.nlm.nih.gov/mesh/D016861; https://id.nlm.nih.gov/mesh/D004008; https://id.nlm.nih.gov/mesh/D020662; https://id.nlm.nih.gov/mesh/D008687; https://id.nlm.nih.gov/mesh/D008930; https://id.nlm.nih.gov/mesh/D018384; https://id.nlm.nih.gov/mesh/D061251; https://id.nlm.nih.gov/mesh/D017208
• Rights: openAccess
• License: https://creativecommons.org/licenses/by/4.0/