摘要
目的 通过转录组学分析评价短链脂肪酸(SCFAs)降低脓毒症小鼠认知功能障碍的机制.方法 SPF级健康成年雄性C57BL/6小鼠60只,8～12周龄,体质量20～25 g,采用随机数字表法分为3组(n=20):假手术组(Sham组)、脓毒症组(SEP组)和短链脂肪酸+脓毒症组(SCFAs+SEP组).采用盲肠结扎穿孔(CLP)的方法制备小鼠脓毒症模型.SCFAs+SEP组于CLP前14 d,在饮用水中添加67.5 mmol/L乙酸盐、40 mmol/L 丁酸盐、25.9 mmol/L丙酸盐,并持续至CLP术后14 d.分别于术后第4、7和14天行Y迷宫实验,Y迷宫实验小鼠数量为小鼠当日存活数量.术后第14天取SEP组和SCFAs+SEP组小鼠(n=4)麻醉后处死取脑组织行转录组测序,利用GO数据库、KEGG数据库进行富集分析.最后每组随机处死5只小鼠取脑组织采用Western blot法验证测序结果.结果 与Sham组相比,SEP组小鼠术后第4、7和14天进入新异臂次数减少,新异臂停留时间缩短(P＜0.05);与SEP组相比,SCFAs+SEP组小鼠术后第4、7和14天进入新异臂次数增加,新异臂停留时间延长(P＜0.05).转录组学分析结果:与SEP组相比,SCFAs+SEP组共筛选出438个显著的差异表达基因,其中175个基因表达上调,263个基因表达下调.与Sham组相比,SEP组小鼠脑组织γ-氨基丁酸A型受体亚基rho2(GABRR2)表达上调(P＜0.05);与SEP组相比,SCFAs+SEP组小鼠脑组织GABRR2表达下调(P＜0.05);Sham组和SCFAs+SEP组小鼠脑组织GABRR2表达差异无统计学意义(P＞0.05).结论 SCFAs可能通过下调GABRR2表达降低脓毒症小鼠认知功能障碍.
Abstract
Objective To evaluate the mechanism of short-chain fatty acids(SCFAs)reducing cognitive dysfunction in septic mice through transcriptomic analysis.Methods Sixty SPF healthy adult male C57BL/6 mice,aged 8-12 weeks,weighing 20-25 g,were divided into 3 groups(n=20 each)using a random number table method:sham operation group(Sham group),sepsis group(SEP group),and SCFAs+sepsis group(SCFAs+SEP group).Cecal ligation and puncture(CLP)was used to simulate a sep-sis model in anesthetized animals.In SCFAs+SEP group,67.5 mmol/L acetate,40 mmol/L butyrate,and 25.9 mmol/L propionate were added to drinking water starting from 14 days before CLP until 14 days after CLP.Y maze tests were conducted on days 4,7 and 14 after surgery,with the number of mice in the Y maze test being the number of surviving mice on that day.On day 14 after surgery,4 mice were selected in SEP group and SCFAs+SEP group and sacrificed after anesthesia,and brain tissues were obtained to per-form transcriptome sequencing,and enrichment analysis was performed using GO database and KEGG data-base.Finally,5 mice were randomly sacrificed in each group,and brain tissues were collected and Western blot analysis was performed to verify the sequencing results.Results Compared with Sham group,the num-ber of times they entered the novel arm on days 4,7 and 14 after surgery were significantly reduced,and the time spent in the novel arm was shortened in SEP group(P＜0.05).Compared with SEP group,the number of times they entered the novel arm on days 4,7 and 14 after surgery was significantly increased,and the time spent in the novel arm was prolonged in SCFAs+SEP group(P＜0.05).The results of transcriptomic a-nalysis showed that 438 significantly differentially expressed genes were identified in SCFAs+SEP group,of which the expression of 175 genes was up-regulated and the expression of 263 genes was down-regulated compared with SEP group.The expression of gamma-aminobutyric acid receptor rho-2(GABRR2)in the brain tissues was significantly up-regulated in SEP group as compared with Sham group(P＜0.05).Com-pared with SEP group,the expression of GABRR2 in the brain tissue was significantly down-regulated in SCFAs+SEP group(P＜0.05).There was no significant difference in the expression of GABRR2 in brain tis-sues between Sham group and SCFAs+SEP group(P＞0.05).Conclusions SCFAs may reduce cognitive dysfunction by down-regulating the expression of GABRR2 in septic mice.
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