Protective effects of γ-aminobutyric acid against H2O2-induced oxidative stress in RIN-m5F pancreatic cells

Xue Tang; Renqiang Yu; Qin Zhou; Shanyu Jiang; Guowei Le

doi:10.1186/s12986-018-0299-2

Protective effects of γ-aminobutyric acid against H₂O₂-induced oxidative stress in RIN-m5F pancreatic cells

Nutr Metab (Lond). 2018 Sep 3:15:60. doi: 10.1186/s12986-018-0299-2. eCollection 2018.

Authors

Xue Tang^{1

2}, Renqiang Yu³, Qin Zhou³, Shanyu Jiang³, Guowei Le^{1

2}

Affiliations

¹ 1State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, 214122 Jiangsu China.
² 3School of Food Science and Technology, Jiangnan University, Wuxi, 214122 Jiangsu China.
³ 2The Affiliated Wuxi Maternity and Child Health Care Hospital of Nanjing Medical University, Wuxi, 214002 Jiangsu China.

Abstract

Background: γ-Aminobutyric acid (GABA) is a major inhibitory neurotransmitter in the central nervous system and reported to maintain the redox homeostasis and insulin secretion function of pancreatic β cells. This study tested the hypothesis that GABA maintains cellular redox status, and modulates glycogen synthase kinase (GSK)-3β and antioxidant-related nuclear factor erythroid 2-related factor 2 (NRF2) nuclear mass ratio in the H₂O₂-injured RINm5F cells.

Methods: RINm5F cells were treated with/without GABA (50, 100 and 200 μmol/L) for 48 h and then exposed to 100 μmol/L H₂O₂ for 30 min. Viable cells were harvested, and dichloro-dihydro-fluorescein diacetate (DCFH-DA) was used to detect reactive oxygen species (ROS) level; cellular redox status and insulin secretion were measured; cell viability was determined by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay; mitochondrial membrane potential (MMP) was detected by flow cytometry; relative genes levels were analyzed by reverse transcriptase polymerase chain reaction (RT-PCR); western blotting was used to determine protein expression of GSK-3β and p-GSK-3β (Ser9), and nuclear and cytoplasmic NRF2.

Results: H₂O₂ increased ROS production, and induced adverse affects in relation to antioxidant defense systems and insulin secretion. These changes were restored by treatment with 100 and 200 μmol/L GABA. In addition, 100 or 200 μmol/L GABA induced membrane depolarization and increased cell viability. These effects were mediated by Caspase-3, Bcl-2 associated X protein (Bax) and B-cell lymphoma-2 (Bcl-2) expression. Western blotting indicated that GABA inhibited GSK-3β by increasing p-GSK-3β (Ser9) level, and directed the transcription factor NRF2 to the nucleus.

Conclusion: In rat insulin-producing RINm5F cells, GABA exerts its protective effect by regulating GSK-3β and NRF2, which governs redox homeostasis by inhibiting apoptosis and abnormal insulin secretion by exposure to H₂O_2.

Keywords: Anti-apoptosis; Anti-oxidation; GABA; Insulin secretion; RINm5f cells.