酸枣仁提取物靶向调控促进骨骼增长的miRNA筛选  

miRNA screening for targeted regulation of bone growth by semen ziziphi spinosae extract

作　　者：罗石任 谢艳 张丽 殷娜 Luo Shiren;Xie Yan;Zhang Li;Yin Na(Henan Provincial Luoyang Orthopedic-Traumatological Hospital(Henan Provincial Orthopedic Hospital),Luoyang 471002,Henan Province,China)

机构地区：[1]河南省洛阳正骨医院(河南省骨科医院),河南省洛阳市471002

出　　处：《中国组织工程研究》2022年第35期5658-5664,共7页Chinese Journal of Tissue Engineering Research

基　　金：河南省中医药科学研究专项课题(2018ZY1023),项目参与人:谢艳,罗石任。

摘　　要：背景:生长激素大部分在慢波睡眠期间分泌,而慢波睡眠启动和延长的一个决定因素是5-羟色胺。作者前期研究发现,酸枣仁提取物可以通过升高5-羟色胺1A受体的表达,使其与5-羟色胺结合,延长慢波睡眠,从而促进生长激素的分泌,促进骨骼增长。同时发现,酸枣仁提取物可以下调脑组织5-羟色胺2A受体的表达,会使5-羟色胺1A受体失去竞争抑制与5-羟色胺更好地结合,从而促进慢波睡眠的延长,促进生长激素的分泌,引发骨骼增长。目的:证明酸枣仁提取物可以通过降低5-羟色胺2A受体促进小鼠骨骼增长,并筛选由酸枣仁提取物调控靶向5-羟色胺2A受体促进骨骼增长的miRNA。方法:(1)昆明种小鼠分为空白对照组、酸枣仁提取物高、低剂量组、阳性对照组和5-羟色胺2A受体选择性抑制剂组(以下简称抑制剂组),每组10只。空白对照组小鼠每日灌胃去离子水,酸枣仁提取物高、低剂量组分别灌胃酸枣仁提取物混悬液0.16,0.32 mg/g,阳性对照组灌胃酸枣仁皂甙A标准品溶液,抑制剂组灌胃酸枣仁提取物水溶液0.32 mg/g,同时灌胃最后3 d每天侧脑室注射10μg M100907。灌胃后第25天观察药物对小鼠体长的影响,ELISA法观察药物对血清生长激素、脑组织5-羟色胺2A受体表达的影响。(2)SD大鼠分为空白对照组、酸枣仁提取物组、阳性对照组和5-羟色胺2A受体选择性抑制剂组(以下简称抑制剂组),于灌胃第3天开始,观察药物对慢波睡眠的影响。(3)然后用芯片法检测药物引起骨骼增长的小鼠与普通小鼠脑组织差异表达的miRNAs,并优选以5-羟色胺2A受体为靶基因的miRNA,同时进行qRT-PCR验证。结果与结论:(1)用药25 d后,高剂量组小鼠体长明显长于空白对照组(P<0.01);小鼠血清生长激素水平明显高于空白对照组和抑制剂组(P<0.01,P<0.01);酸枣仁提取物组脑组织5-羟色胺2A受体表达水平明显降低,低于抑制剂组(P<0.01);(2)酸BACKGROUND:G rowth hormone is secreted during slow-wave sleep in large proportion,and the initiation and prolongation of slow-wave sleep is closely linked with serotonin.Previous studies have shown that semen ziziphi spinosae extra ct can increase the expression of serotonin 1 A recepto r.The combination of serotonin 1 A receptor and serotonin can prolong slow-wave sleep,thereby promoting the secretion of growth hormone and bone growth.In addition,semen ziziphi spinosae extra ct can down-regulate the expression of serotonin 2 A receptor in brain tissue,which can promote the combination of serotonin 1 A receptor and serotonin,thereby promoting the prolongation of slow-wave slee p,the secretion of growth hormone,and bone growth.OBJECTIVE:To prove that semen ziziphi spinosae extract can promote bone growth in mice by down-regulating the expression of serotonin 2 A receptor and to screen miRNAs to rgeting serotonin 2 A receptor and promoting bone growth.METHODS:(1)Kunming mice were divided into blank control group,high-and low-dose groups of semen ziziphi spinosae extract(hereinafter referred to as high-and low-dose groups),positive control group,and serotonin 2 A receptor selective inhibitor group(hereinafter refe rred to as inhibitor group),with 10 mice in each group.Mice in the blank control group were gavaged with deionized water every day,the high-and low-dose groups were gavaged with 0.16 and0.32 mg/g semen ziziphi spinosae extra ct suspension respectively,the positive control group was gavaged with semen ziziphi spinosae saponin A standard solution,and the inhibitor group was gavaged with semen ziziphi spinosae extract aqueous solution.Gavage was administered to each group at the same pe riod,and 10μg of M100907 was injected into the lateral ventricle of each mouse once a day for the last 3 days.Changes in the body length of mice were observed,and changes in the expression of serum growth hormone and brain tissue serotonin 2 A receptor were observed by ELISA.(2)Sprague-Dawley rats were divided into blank cont

关 键 词：酸枣仁提取物 5-羟色胺2A受体 小鼠 骨骼增长 MIRNA 大鼠 慢波睡眠 

分 类 号：R459.9[医药卫生—治疗学] R336[医药卫生—临床医学]