基于基因表达谱研究淫羊藿总黄酮干预老年大鼠海马炎性衰老的效果与机制

Intervention outcome and mechanism of epimedium total flavonoids on aged rat hippocampus during inflamm-aging progress based on gene expression profile

目的探讨炎性衰老的炎性细胞因子网络机制,及淫羊藿总黄酮(epimedium total flavonoids,EF)干预的效果与机制。方法清洁级健康雄性SD大鼠分为3组:4月龄(4 m)组、24月龄(24 m)组和24月龄加EF干预(24 m+EF)组,每组10只。自满21月龄当天开始,24 m+EF组予EF灌胃,24 m组给予等量蒸馏水灌胃,均连续90 d。取各组大鼠海马组织,应用炎症细胞因子与受体基因芯片对海马组织分别进行基因表达检测,绘制基因表达谱,筛选有表达差异的基因。海马组织匀浆,离心取上清液,运用酶联免疫吸附法检测上清液中炎性细胞因子肿瘤坏死因子-α(TNF-α)、白介素-6(IL-6)和IL-10蛋白表达。结果在海马组织中,与4 m组相比,24 m组表达上调的促炎性反应细胞因子与受体基因有9个,表达下调的抗炎性细胞因子与受体基因有6个。与24 m组比较,24 m+EF组中表达上调的抗炎性细胞因子与受体基因有8个,表达下调的促炎性细胞因子与受体基因有11个。24 m组海马组织中TNF-α和IL-6水平都显著高于4 m组(P<0.01);与24 m组比较,24 m+EF组中TNF-α和IL-6水平都明显降低,IL-10水平显著升高(P<0.01)。与4 m组相比,24 m组海马组织上清液中TNF-α/IL-10和IL-6/IL-10比值均显著升高(P<0.01);24 m+EF组TNF-α/IL-10和IL-6/IL-10比值均比24 m组显著降低(P<0.01)。结论老年大鼠海马存在促炎性细胞因子表达上调,导致促-抗炎性细胞因子网络和促-抗炎反应体系平衡失调,这可能是炎性衰老中出现高促炎性反应状态的原因和炎性衰老发生的新机制;EF可能通过下调促炎性细胞因子表达和上调抗炎性细胞因子表达,重塑促-抗炎性细胞因子网络和促-抗炎性反应体系新的平衡,达到干预炎性衰老的目的。

Objective To investigate the mechanisms of inflammatory cytokines network in the development of inflamm-aging and EF intervention outcome and its mechanism.Methods Healthy male Sprague Dawley rats were divided into 3 groups： 4 m young group with 4 months rats,24 m aged group with 24 months rats treated with distilled water intragastric administration and 24 m＋EF group with 24 months rats treated with EF by intragastric administration for 3 months from 21 months,10 each group.The gene expression profiles of rat hippocampal tissues from rats in every group were analyzed by using inflammatory cytokines and receptors cDNA microarray,respectively.The differential expressed genes were screened.At the same time,the levels of inflamm-aging-related inflammatory cytokines proteins expressions of TNF-α,IL-6 and IL-10 in rat hippocampal tissues were evaluated by enzyme linked immunosorbent assay. Results In rat hippocampal tissues,compared with those in 4 m group,there were 9 differentially up-regulated expressed pro-inflammarory genes and 6 down-regulated expressed anti-inflammatory genes in 24 m group.Compared with those in 24 m group,there were 8 up-regulated expressed anti-inflammatory genes and 11 down-regulated expressed pro-inflammatory genes in 24 m＋EF group.Compared with those in 4 m group,the levels of TNF-α and IL-6 in the supernatant from rat hippocampal tissues in 24 m group increased significantly（P〈0.01）.As compared with those in 24 m group,the levels of TNF-α and IL-6 in the supernatant from rat hippocampal tissues in 24 m＋EF group decreased significantly（P〈0.01）.But the level of IL-10 in EF＋24 m group was significantly higher than that in 24 m group（P〈0.01）.The ratio of TNF-α to IL-10 and IL-6 to IL-10 in 24 m group increased significantly as compared with those in 4 m group（P〈0.01）,and decreased significantly in 24 m＋EF group as compared with those in 24 m group（P〈0.01）. Conclusions There is upregulation of mRNA and protein expressions of pro-inflammatory cytokines and receptors in the aged rat hippocampus,which may eventually contribute to disequilibrium of the network of pro-and anti-inflammatory cytokines and receptors and disequilibrium of response of pro-and anti-inflammatory system and high inflammatory state,which provides new mechanistic insight into inflamm-aging.EF may interfere in inflamm-aging via rebuilding the new equilibrium of the network of pro-and anti-inflammatory cytokines and receptors and equilibrium of response of pro-and anti-inflammatory system,which might be caused by downregulation of expressions of pro-inflammatory cytokines and receptors and upregulation of expressions of anti-inflammatory cytokines and receptors.