miRNAs调控RPS14影响MDS细胞生物学行为的研究

As a key factor in regulation of gene post-transcriptional, microRNAs have been identified to play an important role in carcinogenesis in various tumors. Myelodysplastic syndrome(MDS) is a clonal myeloid disease with refractory quantitative and qualitative abnormalities of hemocytes and its pathogenesis is poorly understood. Some researches have showed that abnormal expressions of some microRNAs have close relationship with the pathogenesis of MDS. However, little research has been conducted in roles of microRNAs in pathogenesis of MDS and the relationship between miRNAs and target proteins. Recently, low RPS14 expression is found common in all kinds of myelodysplastic syndromes , but its mechanism is unclear. Based on the fact, we plan to do our research from the following three aspects:①Screening and identification of miRNAs targeting RPS14 predicted by bioinformatics; ②Detection of expressions of RPS14 and miRNAs targeting RPS14 in MDS patients;③Analysis of influence of miRNAs on RPS14 expression and proliferation, differentiation and apoptosis of hematopoietic stem cells and evaluation of the role of miRNAs acting as a potential therapeutic target in MDS. This research will help reveal the pathogenesis of MDS from a new angle and provide new ideas for the diagnosis, treatment and prognosis evaluation of MDS..

miRNAs作为基因转录后水平调控的关键因素之一，已被证实在多种肿瘤的发生中发挥重要作用。骨髓增生异常综合征（MDS）是一种难治性血细胞质、量异常的髓系肿瘤，发病机制不明。已有研究表明某些miRNAs的异常表达与MDS的发病关系密切,但对于miRNAs参与MDS发病的作用机理以及MDS中miRNAs与靶蛋白之间的关系鲜有报道。最近发现核糖体蛋白小亚基14（RPS14）在各种类型的MDS中普遍表达低下，但机制尚不清楚。基于此，本课题拟从以下三个方面展开研究：①筛选和验证生物信息学预测的靶向调控RPS14的miRNAs；②检测调控RPS14的miRNAs及RPS14在MDS中的表达；③建立相对稳定的调控RPS14的miRNAs突变小鼠模型，探讨miRNAs对RPS14的表达及造血干/祖细胞分化、增殖和凋亡的影响。有望从新的角度揭示MDS的发病机制，为MDS的诊断、治疗及预后判断提供新的思路。

miRNAs作为基因转录后水平调控的关键因素之一，已被证实在多种肿瘤的发生中发挥重要作用。骨髓增生异常综合征（MDS）是一种难治性血细胞质、量异常的髓系肿瘤，发病机制不明。已有研究表明某些miRNAs的异常表达与MDS的发病关系密切,但对于miRNAs参与MDS发病的作用机理以及MDS中miRNAs与靶蛋白之间的关系鲜有报道。有研究发现核糖体蛋白小亚基14（RPS14）在各种类型的MDS中普遍表达低下，但机制尚不清楚。本课题在此基础上展开研究，通过生物信息学软件及荧光素酶报告系统筛选及鉴定出在MDS中靶向调控RPS14的miR-223,然后由免疫荧光结果筛选出高表达RPS14的细胞株（K562细胞株-慢性粒细胞白血病细胞株）。应用慢病毒感染的方法，通过分别构建过表达和沉默miR-223的载体，转染MDS细胞株后应用流式方法测定感染效率，并进一步应用PCR在RNA水平验证感染率，从而成功获得外源性过表达或抑制调控RPS14的miR-223的目标实验细胞。应用MTT实验及流式对感染慢病毒miR-223后的SKM-1及K562细胞株进行增殖和凋亡检测后提示：过表达慢病毒miR-223后促进细胞增殖，抑制凋亡；沉默后可促进细胞凋亡，抑制增殖。Western blot证实miR-223的过表达可以促进RPS14基因沉默。细胞分化实验发现miR-223可抑制红系分化。研究初步明确了miR-223通过靶向调控RPS14参与MDS干细胞发病的作用机制。