Tat-Endophilin A1 Fusion Protein Protects Neurons from Ischemic Damage in the Gerbil Hippocampus: A Possible Mechanism of Lipid Peroxidation and Neuroinflammation Mitigation as Well as Synaptic Plasticity

Abstract

The present study explored the effects of endophilin A1 (SH3GL2) against oxidative damage brought about by H2O2 in HT22 cells and ischemic damage induced upon transient forebrain ischemia in gerbils. Tat-SH3GL2 and its control protein (Control-SH3GL2) were synthesized to deliver it to the cells by penetrating the cell membrane and blood–brain barrier. Tat-SH3GL2, but not Control-SH3GL2, could be delivered into HT22 cells in a concentration- and time-dependent manner and the hippocampus 8 h after treatment in gerbils. Tat-SH3GL2 was stably present in HT22 cells and degraded with time, by 36 h post treatment. Pre-incubation with Tat-SH3GL2, but not Control-SH3GL2, significantly ameliorated H2O2-induced cell death, DNA fragmentation, and reactive oxygen species formation. SH3GL2 immunoreactivity was decreased in the gerbil hippocampal CA1 region with time after ischemia, but it was maintained in the other regions after ischemia. Tat-SH3GL2 treatment in gerbils appreciably improved ischemia-induced hyperactivity 1 day after ischemia and the percentage of NeuN-immunoreactive surviving cells increased 4 days after ischemia. In addition, Tat-SH3GL2 treatment in gerbils alleviated the increase in lipid peroxidation as assessed by the levels of malondialdehyde and 8-iso-prostaglandin F2α and in pro-inflammatory cytokines such as tumor necrosis factor-α, interleukin-1β, and interleukin-6; while the reduction of protein levels in markers for synaptic plasticity, such as postsynaptic density 95, synaptophysin, and synaptosome associated protein 25 after transient forebrain ischemia was also observed. These results suggest that Tat-SH3GL2 protects neurons from oxidative and ischemic damage by reducing lipid peroxidation and inflammation and improving synaptic plasticity after ischemia.

Tat-malate dehydrogenase fusion protein protects neurons from oxidative and ischemic damage by reduction of reactive oxygen species and modulation of glutathione redox system

“…In addition, we confirmed the delivery of 1 mg/kg Tat-MDH1 into the gerbil hippocampus 6 h after ischemia. These results are consistent with previous studies showing that Tat-cargo were efficiently and effectively translocated into HT22 cells and the gerbil hippocampus 24,25,35,36 . In this study, we observed that treatment with H 2 O 2 significantly decreased cell viability, and simultaneous treatment with H 2 O 2 and Tat-MDH1 ameliorated the reduction in cell viability in a concentration-dependent manner, whereas surviving cells were significantly increased after treatment with 4 μM Tat-MDH1 based on 5-CFDA-AM staining.…”

Neuroprotective Effects of Purpurin Against Ischemic Damage via MAPKs, Bax, and Oxidative Stress Cascades in the Gerbil Hippocampus

“…To elucidate the possible role of 6 mg/kg purpurin against ischemia, we observed the morphology of astrocytes and microglia as well as pro-inflammatory cytokines in the hippocampus because a recent study showed the anti-inflammatory roles of purpurin in RAW 264.7 murine macrophage cells [ 14 ]. The animals were sacrificed 6 h after ischemia to measure IL-1β, IL-6, and TNF-α levels in the hippocampus because these levels are significantly increased in the early period of ischemia [ 29 , 30 , 43 ]. In addition, the IL-1 receptor antagonist showed neuroprotective effects against ischemic damage in rats [ 44 ].…”

“…To elucidate the mechanisms of purpurin’s effects against ischemic damage, animals ( n = 5 in each group) were euthanized with 75 mg/kg alfaxalone and 10 mg/kg xylazine 6 h and 4 days after ischemia/reperfusion, when pro-inflammatory cytokine levels were significantly increased and returned to control levels, respectively [ 29 , 30 ]. In brief, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α levels were measured in the hippocampus based on comparisons with linear calibration curves generated using IL-1β, IL-6, and TNF-α standard solutions by using their respective enzyme immunoassay kits (BioSource International Inc., Camarillo, CA, USA).…”

Neuroprotective Effects of Purpurin Against Ischemic Damage via Anti-inflammatory and MAPK Pathway

“…To elucidate the possible role of 6 mg/kg purpurin against ischemia, we observed the morphology of microglia and pro-in ammatory cytokines in the hippocampus because a recent study showed the antiin ammatory roles of purpurin in RAW 264.7 murine macrophage cells [14]. The animals were sacri ced 6 h after ischemia to measure IL-1β, IL-6, and TNF-α levels in the hippocampus because these levels are signi cantly increased in the early period of ischemia [26,27,39]. In addition, the IL-1 receptor antagonist showed neuroprotective effects against ischemic damage in rats [40].…”

“…To elucidate the mechanisms of purpurin's effects against ischemic damage, animals (n = 5 in each group) were euthanized with 75 mg/kg alfaxalone and 10 mg/kg xylazine 6 h after ischemia/reperfusion, when pro-in ammatory cytokine levels were signi cantly increased [26,27]. In brief, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α levels were measured based on comparisons with linear calibration curves generated using IL-1β, IL-6, and TNF-α standard solutions.…”