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目的研究模拟失重环境对大鼠肾上腺髓质激素分泌及miRNA-375表达的影响。方法健康成年雄性Wistar大鼠56只,随机分为0h(正常对照)、6h、12h、1d、3d、5d、7d共7个组(n=8)。采用Morey-Holton尾悬吊法建立模拟失重动物模型。各组大鼠实验结束后眼球取血,取双侧肾上腺。采用放射免疫分析法检测血浆肾上腺髓质激素[多巴胺(DA)、去甲肾上腺素(NE)、肾上腺素(E)]水平,RT-PCR法检测肾上腺组织酪氨酸羟化酶(TH)mRNA和miRNA-375的表达,Western blotting和免疫组化技术检测肾上腺髓质TH蛋白表达。结果尾悬吊模拟失重条件下大鼠血浆肾上腺髓质激素波动明显,6~12h出现一过性下降,随后急剧升高,3d时形成峰值。RT-PCR结果显示,各组大鼠肾上腺组织TH mRNA表达波动明显,6h时呈现一过性下降,随即上升,1d时达峰值,之后呈下降趋势,5d时形成谷值。大鼠肾上腺髓质TH蛋白表达以尾悬吊3d时染色最强。肾上腺髓质中miRNA-375表达表现为尾悬吊早期(6h)呈一过性显著升高,随后快速下降,12h、1d和3d时表达受到明显抑制,表达量显著下降,5d时表达量增加,7d时显著升高(P<0.05)。结论失重环境可影响大鼠肾上腺髓质激素分泌,其机制可能与miRNA-375通过复杂的信号通道调控TH表达有关。
Objective To investigate the effects of simulated weightlessness on adrenomedullin secretion and miRNA-375 expression in rats. Methods 56 healthy male Wistar rats were randomly divided into 7 groups (n = 8): 0h (normal control), 6h, 12h, 1d, 3d, 5d, 7d. Animal model of simulated weightlessness was established by Morey-Holton tail suspension method. Eyeballs were taken from rats in each group after the experiment, bilateral adrenal glands were taken. The levels of plasma adrenomedullin (DA, NE, adrenaline (E)] were measured by radioimmunoassay. The expression of tyrosine hydroxylase (TH) mRNA in adrenal tissue was detected by RT- And miRNA-375 expression, Western blotting and immunohistochemical detection of adrenal medulla TH protein expression. Results The rat plasma adrenomedullin showed significant fluctuations after simulated weightlessness in the tail suspension, transiently decreased from 6 to 12 hours, then increased sharply and peaked at 3 days. RT-PCR results showed that the expression of TH mRNA in adrenal tissues of rats in each group fluctuated significantly, and then decreased transiently at 6h, then increased and then reached a peak at 1d, then decreased, and formed a valley at 5d. The expression of TH protein in adrenal medulla of rats was the strongest when it was dyed with tail suspension. Expression of miRNA-375 in adrenal medulla showed a transient increase in early post-suspension (6h), followed by a rapid decrease. Expression of miRNA-375 was significantly inhibited at 12h, 1d and 3d, and increased at 5d , Significantly increased at 7d (P <0.05). CONCLUSION: Weightlessness can affect the secretion of adrenomedullin in rats, which may be related to the regulation of TH expression by miRNA-375 through complex signaling pathways.