灵芝孢子油对N-甲基-N-亚硝酸脲诱导的大鼠视网膜损伤中Caspase-3表达的影响

Effect of ganoderma spore oil on aspase-3 expression in rat retina with photoreceptor damage induced by N-methyl-N-nitrosourea

目的观察灵芝孢子油(ganoderma spore oil,GSO)对N-甲基-N亚硝酸脲(N-methyl-N-nitrosourea,MNU)诱导的视网膜外核层细胞损伤的作用及对Caspase-3时空表达的影响。方法50d♀SD大鼠随机分为正常对照(NC)组、MNU造模(MNU)组、二十二碳六烯酸处理(DHA)组和灵芝孢子油处理(GSO)组。各组分别于MNU造模前0h及造模后1d、3d、7d、10d处死大鼠取材,采用RT-PCR、免疫荧光技术检测视网膜中Caspase-3表达和分布的变化,TUNEL法检测视网膜细胞凋亡情况。结果①RT-PCR检测:与NC组比较,MNU组1d、3d、7d视网膜Caspase-3表达增强(P<0.01);GSO组和DHA组1d、3d低于MNU组(P<0.01),1dGSO组表达较DHA组低(P<0.01)。②免疫荧光检测:在3d,GSO组和DHA组Caspase-3表达水平低于MNU组,GSO组较DHA组稍弱。③TUNEL法检测:NC组在视网膜各层均未发现凋亡细胞,MNU造模后可见外核层细胞凋亡。GSO组和DHA组在1d、3d外核层细胞凋亡百分率少于对应的MNU组(P<0.01),3d GSO组低于DHA组(P<0.01)。结论灵芝孢子油可能通过下调视网膜Caspase-3的表达,阻抑MNU诱导的视网膜外核层光感受器细胞凋亡的作用。

Aim To investigate the effect of ganoderma spore oil （GSO） on Caspase-3 expression in damaged retina and discuss its role in photoreceptor cell lesion induced by N-methyl-N-nitrosourea （ MNU ）. Methods 120 female Sprague-Dawley rats （50-day-old） were divided into 4 groups randomly, including normal control group （NC） , untreated MNU model control group （ MNU ） , doeosahexaenoie acid treatment group （DHA） and ganoderma spore oil treatment group （GSO）. Retina tissue samples were obtained sequentially 0 hour before MNU injection and 1, 3, 7, 10 days after MNU injection. Expressions of 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. Results （1） Caspase-3 mRNA expression in MNU group up-regulated on 1 d and peaked on 3 d compared with NC （P 〈0. 01 ）. The expression levels of DHA and GSO group decreased obviously on 1 d,3 d （P 〈0. 01 vs MNU group）, furthermore, that of GSO group on 1 d was lower than that of DHA group （P 〈 0. 01 ）. （2） Immunofluorescence assays showed that Caspase-3 expression of DHA and GSO group in retina decreased clearly on 3 d compared with MNU group. Moreover, GSO down-regulated Caspase-3 expression in retina a little bit more than DHA did on 3 d. （3） Photoreceptor cell apoptosis was only evoked in the outer nuclear layer and peaked on 3 d in MNU administered rats. The apoptotic indexes （AI） of GSO group and DHA group were lower than those of MNU group on 1 d,3 d, respectively （P 〈0. 01）. Compared with that of DHA group, AI of GSO group was reduced significantly only on 3 d （ P 〈 0.01 ）. Conclusion GSO may inhibit MNU-induced rat photoreceptor cell apoptosis via down-regulation of Caspases-3 expression.