摘要
目的探讨铁蓄积小鼠骨量下降与微小RNA（miRNA，miR）差异性表达的关系。方法12周龄雄性ICR小鼠（n=12）随机分为两组：铁蓄积组（FAC组）及对照组（Con组），分别腹腔注射枸橼酸铁胺（FAC）及等量生理盐水，干预8周后检测血清铁蛋白，微计算机断层扫描技术（Micro-CT）检测小鼠股骨骨量，并分析相关骨结构参数。小鼠全血分离自细胞提取RNA，进行Taqman miRNA芯片检测表达谱差异，实时荧光定量聚合酶链反应（FQ-PCR）检测验证差异miRNA。利用Targetscan及miRDB数据库预测差异miRNA的下游靶基因，并对预测的下游靶基因进行生物信息学分析，包括细胞成分、生物过程、分子功能和京都基因和基因组百科全书（KEGG）通路分析。结果FAC组干预8周后，血清铁蛋白[（218．2±29．5）μg/L]相较于对照组[（30．6±9．9）μg/L]明显升高（P=0．000）。股骨三维重建示铁蓄积小鼠骨质结构破坏，骨量、骨小梁厚度（Tb．Th）及骨体积分数（BV／TV）显著下降（P=0．001、0．005、0．021），而骨小梁间隙（Th．Sp）升高（P=0．000）。miRNA芯片结果：初步筛选出20个miRNA表达上调：mmu-miR-29b-3p、mmu-miR-324-3p、mmu-miR-133a-5p、mmu-miR-214-5p、mmu-miR-22-3p、mmu-miR-34a-5p、mmu-miR-31-5p，mmu-miR-143-5p，mmu-miR-423-3p、mmu-miR--223、mmu-miR-155、mmu-miR-106a、mmu-miR-2861、mmu-miR-148a、mmu-miR-96、mmu-miR-449a-5p，mmu-miR-423-5p、mmu-miR-204-5p、mmu-miR-211、mmu-miR-23b及7个miRNA表达下调：mmu-miR-18a．3p、mmu-miR-223-3p、mmu-miR-199a-5p、mmu-miR-196a-5p、mmu-miR-30c-5p、mmu-miR-15b、mmu-miR-130b。其中mmu-miR-423-5p表达差异最为显著，q-PCR验证结果显示miR-423-5p的表达趋势与测序结果一致。Targetsean和miRDB预测并筛选miR-423-5p下游靶基因共91个。生物信息学分析表明miR-423-5p的靶基因在磷脂酰肌醇3激酶（P13K）-蛋白激酶B（Akt）、Rap1、Ras及丝裂原活化蛋白激
Objective To explore the correlation between bone loss and the differential expression of microRNA （ miRNA, miR） in mice with iron accumulation. Methods 12 - week - old male ICR mice （ n = 12） were randomly divided into two groups ： FAC group and Control group. Intraperitoneal injections of iron citrate amine （FAC） and equivalent saline were conducted in these two groups. Serum fen＇itin level was detected after 8 weeks of intervention. Micro - CT was used to detect bone volume and bone structure parameters in mice. The whole blood of mice were collected and RNA from white blood cells were extracted. Differential expression profiles of miRNA chips was conducted, and the resuh was verified by real - time fluorescence quantitative polymerase chain reaction（ FQ- PCR） . Targetscan and miRDB were used to predict the downstream target genes of miRNA. Bioinformatics analysis of target gene was conducted, including celluar component, biologic process, molecular function and kyoto encyclopedia of genes and genomes （KEGG） pathway analysis. Results Serum ferritin （218.2 ± 29. 5） μg/L was significantly higher than that in the control group （30. 6 ±9. 9） μg/L after 8 weeks. Micro - CT analysis showed a destruction of bone structure in iron accumulation mice. Bone mass, trabecular thickness （Tb. Th） and bone volume fraction （BV/TV） decreased significantly （P = 0. 001, 0. 005, 0. 021 ） in iron accumulation mice, while trabecular separation/spacing （Tb. Sp ） increased, miRNA chip results： 20 miRNAs up regulated： mmu-miR-29b-3p, mmu - miR - 324 - 3p, mmu - miR - 133a - 5p, mmu - miR - 214 - 5p, mmu-miR-22-3p, mmu - miR - 34a - 5p, mmu - miR - 31 - 5p, mmu - miR - 143 - 5p, mmn- miR-423-3p, mmu- miR-223, mmu- miR- 155, mmu- miR- 106a, mmu- miR-2861, mmu - miR - 148a, mmu-miR-96, mmu - miR - 449a - 5p, mmu - miR - 423 - 5p, mmu-miR-204-5p, mmu- miR- 211, mmu- miR- 23b and 7 miRNAs down regulated： mmu-miR-18a-3p, mmu-miR-223 -3p, mmu-miR-199a-5p, mmu-miR-196a-Sp, mmu-miR-30c-5
作者
朱和平
周建刚
蒋建农
张雷炎
王强
徐又佳
Zhu Heping;Zhou Jiangang;Jiang Jiannong;Zhang Leiyan;Wang Qiang;Xu Youjia(Department of Orthopedics, the Second Affiliated Hospital of Soochow University, Suzhou 215004, China （Zhu HP, Xu Y.];Department of Orthopedics, the AJfiliated Yixing Hospital of Jiangsu University, Yixing 214200, China （ Zhu HP, Zhou JG, Jiang JN, Zhang LY, Wang Q)
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2018年第7期1308-1311,共4页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金（81572179）
江苏省临床专项（BL2014044）
苏州市临床医学中心（SZZXJ201504）
苏州市民生科技项目（SS201634）
苏州大学附属第二医院优势学科群项目（XKQ2015001）
无锡市卫生计生科研青年项目（Q201732）
