Ganoderma spore lipid inhibits N-methyl-N-nitrosourea-induced retinal photoreceptor apoptosis in vivo

Abstract

The purpose of the present study was to investigate the effect of Ganoderma spores lipid (GSL) on Bax, Bcl-xl and Caspase-3 expression in damaged retina and to address the effect of GSL on photoreceptor cell lesions induced by N-methyl-N-nitrosourea (MNU). Thirty 50-day-old female Sprague–Dawley rats were divided randomly into five groups to detect the dose–response effect of GSL by electroretinogram (ERG) analysis. Four groups received different daily GSL doses (0.5, 1, 2 and 4 g/kg, respectively) and one control group received no treatment. Sixty rats were divided randomly into an untreated MNU model control group and the GSL treatment group. Retina tissue samples were obtained sequentially 0 h before and 1, 3, 7 and 10 d after MNU injection. Expressions of Bax, Bcl-xl and Caspase-3 were detected by RT-PCR and immunofluorescence assays, then photoreceptor cell apoptosis was confirmed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling (TUNEL) signals. GSL had a dose–response effect on retina ERG and reversed retinal photoreceptor damage induced by MNU. RT-PCR analysis demonstrated that transcription levels of Bax, Bcl-xl and Caspase-3 in MNU control group and GSL treatment group were all upregulated on 1 d ( p < 0.01) and peaked on 3 d ( p < 0.01) after MNU injection compared to before MNU injection. GSL treatment significantly decreased mRNA levels of Bax on 1, 3, 7 and 10 d vs. MNU group (all p < 0.01) and mRNA levels of Caspase-3 on 1, 3, 7 d ( p < 0.01) and 10 d ( p < 0.05) vs. MNU group. Bcl-xl was clearly upregulation on 1, 3, 7 and 10 d vs. MNU group (all p < 0.01). Expression ratios of Bax/Bcl-xl were raised after MNU injection on 1 and 3 d vs. 0 h before MNU (both p < 0.01), peaked on 3 d, then dropped after GSL treatment on 1, 3, 7 and 10 d vs. MNU group (all p < 0.01). Immunofluorescence assays showed GSL decreased Bax and Caspase-3 positive staining levels in retina and increased Bcl-xl level. TUNEL-positive cells were evoked only in the outer nuclear layer and peaked on 3 d in rats receiving MNU ( p < 0.01 vs. 0 h before MNU). GSL administration decreased apoptosis levels significantly, and the apoptotic indexes (AIs) of the GSL group were lower than those of MNU group on 1 and 3 d (both p < 0.01). Taken together, these data suggest that GSL may regulate the expressions of Bax, Bcl-xl and Caspases-3, inhibiting MNU-induced rat photoreceptor cell apoptosis and protecting retinal function.