Immunohistochemical studies on cell proliferation and neuroblast differentiation induced by sodium butyrate and pyridoxine in the dentate gyrus of a mouse model of D-galactose induced aging

Abstract

The hippocampus is an important region in memory. Newborn neurons are continuously produced in certain brain regions; The subgranular zone of the hippocampal dentate gyrus is one of these regions. Neurogenesis declines critically as being aged, and cognition is also related closely to age. Some studies have reported that D-galactose (D-gal) generates reactive oxygen species (ROS) during D-gal metabolism, and chronic treatment of D-gal induces age-related phenotype in the mouse brain. Many factors influence neurogenesis. Pyridoxine, which was used in this study, is one of factors that influences neurogenesis. We confimed that pyridoxine showed effects on cell proliferation and neuroblast differentiation by upregulating GABAergic system in the mouse dentate gyrus. Sodium butyrate is a member of histone deacetylase (HDAC) inhibitor, and some studies have reported that these acetylation and deacetylation play a key role in regulating gene expression during cell proliferation and differentiation. The goal of this study was to investigate effects of the treatment of pyridoxine and sodium butyrate on neurogenesis in the dentate gyrus of a mouse model of D-gal induced aging using immunohistochemistry and western blot analysis. In the first experiment, we confirmed the neurogenic effects of pyridoxine in the mouse hippocampus. The treatment of pyridine (350 mg/kg, intraperitoneally) increased glial fibrillary acidic protein (GFAP)-, double cortin (DCX)-, and Ki67- positive cells without inducing neuronal death and microglia activation. It also increased the protein level of glutamic acid decarboxylase (GAD) 67, which is an enzyme for GABA synthesis. In the second experiment, we examined the synergisteic effects of sodium butyrate on neurogenesis induced by pyridoxine. Sodium butyrate (300 mg/kg, subcutaneously) alone did not show significant increase in cell proliferation, but sodium butyrate in combination with pyridoxine robustly enhanced cell proliferation and neurogenesis in the mouse dentate gyrus. In the last experiment, we investigated the effects of treatment with sodium butyrate combined with pyridoxine on cell proliferation and neuroblast differentiation in the dentate gyrus of a mouse model of D-galactose induced aging. In the morris water maze test, D-gal-treated mice had longer escape latency than control mice, but the treatment of sodium butyrate combined with pyridoxine shortened the time of escape, which was almost similar to the control mice. In addition, D-gal-treated mice showed a significant reduction in cell proliferation and neuroblast differentiation compared to control mice, but the treatment of sodium butyrate combined with pyridoxine markedly increased the numbers of Ki67-positive cells and DCX-positive neuroblasts. In short, pyridoxine enhanced cell proliferation and neuroblasts differentiation by upregulation of GABAergic system. Furthermore, combinational treatment of pyridoxine with sodium butyrate showed synergistic effects on neurogenesis. In a D-gal induced aging model, pyridoxine in combination with sodium butyrate restored the age-related reduction of memory, cell proliferation, and neuroblast differentiation.