Protective Effect of GIP against Monosodium Glutamate-Induced Ferroptosis in Mouse Hippocampal HT-22 Cells through the MAPK Signaling Pathway

Abstract

The effect of glucose-dependent insulinotropic polypeptide (GIP) on cells under oxidative stress induced by glutamate, a neurotransmitter, and the underlying molecular mechanisms were assessed in the present study. We found that in the pre-treatment of HT-22 cells with glutamate in a dose-dependent manner, intracellular ROS were excessively generated, and additional cell damage occurred in the form of lipid peroxidation. The neurotoxicity caused by excessive glutamate was found to be ferroptosis and not apoptosis. Other factors (GPx-4, Nrf2, Nox1 and Hspb1) involved in ferroptosis were also identified. In other words, it was confirmed that GIP increased the activity of sub-signalling molecules in the process of suppressing ferroptosis as an antioxidant and maintained a stable cell cycle even under glutamate-induced neurotoxicity. At the same time, in HT-22 cells exposed to ferroptosis as a result of excessive glutamate accumulation, GIP sustained cell viability by activating the mitogen-activated protein kinase (MAPK) signalling pathway. These results suggest that the overexpression of the GIP gene increases cell viability by regulating mechanisms related to cytotoxicity and reactive oxygen species production in hippocampal neuronal cell lines.

Rhinacanthin-C but Not -D Extracted from Rhinacanthus nasutus (L.) Kurz Offers Neuroprotection via ERK, CHOP, and LC3B Pathways

“…In previous studies, glutamate activated autophagy in HT-22 cells [ 12 , 21 ], whereas CHOP expression had been shown to prevent autophagy [ 32 , 33 , 34 , 35 ] and ERK phosphorylation was regulated by autophagy proteins [ 36 ]; therefore, we investigated the effects of Rn-C and NpQ on autophagy induction in HT-22 cells subjected to 5 mM glutamate. LC3I conversion to LC3II is an indicator of autophagy induction, and thus, we measured the ratio of LC3II to LC3I via western blot ( Figure 8 A).…”

“…In contrast, some others find them ineffective. Previous studies from this laboratory have reported caspase-12 cleavage, annexin V detection in glutamate treated HT-22 cells [ 21 ], and apoptosis-inducing factor (AIF) activation and caspase-independent cell death [ 43 ]. This all suggests that oxidative stress, caused by ER stress and mitochondrial leakage, induces a caspase-dependent or independent form of apoptosis; moreover, other conditions of cell death have been linked to glutamate toxicity in HT-22 cells, including the calpain, cathepsin, and the ubiquitin-proteasome system [ 44 , 45 ].…”

“…Another form of cell death other than necrotic and apoptotic resulting from mitochondrial and ER stress seen in HT-22 cells treated with glutamate is autophagic cell death; previous studies have shown that autophagy plays a significant component of the cell death seen in HT-22 cells subjected to glutamate treatment [ 12 , 21 , 40 , 47 , 48 ]. In this study, we show that autophagy is activated by glutamate treatment shown by LC3BI conversion to LC3BII and LC3B localization with P63.…”

“…Glutamate has been shown in previous studies to induce the production of reactive oxygen species (ROS) in HT-22 cells [ 16 , 20 , 21 ]. ROS was measured using H 2 DCFDA following the previously described protocol [ 17 ].…”

Role of NADPH Oxidases in Blood–Brain Barrier Disruption and Ischemic Stroke

“…In Zn 2+ -mediated neurodegeneration, protein kinase C (PKC)-induced Nox1 activation mediates transient receptor potential melastatin 2 (TRPM2)-dependent intercellular Ca 2+ overload via adenosine diphosphate ribose (ADPR) production, resulting in apoptosis [ 20 ]. A recent study also demonstrated that glutamate-induced ROS production, lipid peroxidation and hippocampal cell death is mediated by glucose-dependent insulinotropic polypeptide (GIP) and mitogen-activated protein kinase (MAPK)-induced Nox1 activation [ 21 ].…”