miR-98及调节相关因子在Ⅱ型糖尿病大动脉早期病变中的作用

The large artery complication could cause the main death and disable to diabetic individual. However, the mechanisms involved in the pathogenesis of vascular complications in Non-insulin-dependent diabetes mellitus (NIDDM, type-2 diabetes).NIDDM are poorly understood. Many studies showed that the important roles of miRNA in pathogenesis of diabetes..In precious study, we found that the expression of miR-98 was decreased obviously in the adventitia and the endomembrane in the diabetic GK rats compared with control groups. The regulating factors of miR-98 may be TRB2 and CyclinD2 by using microRNA TargetScan software analysis. After the reporter plasmid (pcDNA-GFP-UTR vector) was constructed, miR-98 was co-transfected with the reporter plasmid into rat RAOEC cells. The intensities of ?uorescence from the miR-98 groups were all decreased in comparison to the control group, which indicated that miR-98 could regulate TRB2-3'-UTR expression. TRB2 expression was found to be highly upregulated in the aorta wall of GK rat compared with the Wistar rat, which support that miR-98 and TRB2 attend the artery pathogenesis of diabetes. Based on precious study，we futher explore the roles of miR-98 in regulating the TRB2/Cyclin D2 by molecular biology or immune methods, and analyze roles of miR-98 and its relative factors in the early pathogenesis of vascular complications in NIDDM. This study will be crucial to diabetic prevetion and provide new target or data for diabetic therapy.

大血管病变是糖尿病个体致死、致残的主要原因之一，具体的病变机制尚不清楚。研究表明miRNA在糖尿病中起重要作用。.前期研究，我们发现与对照组相比，miR-98在糖尿病GK大鼠大动脉内膜、外膜表达都明显降低。通过microRNA软件分析发现，miR-98调节的因子可能有Cyclin D2、TRB2等。进而构建了pcDNA-GFP-UTR载体，与miR-98共转染大鼠内皮细胞株，结果表明miR-98对TRB2-3'-UTR起调控作用，而TRB2在Ⅱ型糖尿病大鼠模型大血管内膜和外膜表达量都明显增高，说明二者参与了糖尿病大血管病变过程。在上述基础上，本课题将利用分子生物学、免疫学等技术，从细胞和个体水平，深入研究miR-98对TRB2、Cyclin D2及相关因子的调节作用，进而分析miR-98及相关因子在Ⅱ型糖尿病大动脉早期病变中的作用，为Ⅱ型糖尿病血管病变的预防和治疗提供新的靶点和科学资料。

大血管病变是糖尿病个体致死、致残的主要原因之一，具体的病变机制尚不清楚。研究表明miRNA在糖尿病中起重要作用。本课题利用分子生物学、免疫学等技术，从细胞和个体水平，深入研究miR-98对TRB2、Cyclin D2及相关因子的调节作用，及其在糖尿病发病中的作用。. (1) miR-98与TRB2的在2型糖尿病血管病变中的作用：取模型鼠大动脉组织，应用免疫组织化学、ELISA、逆转录-聚合酶链反应和实时定量PCR检测相关指标。流式细胞仪和荧光显微镜检测RAOEC细胞GFP表达。TRB2在糖尿病大鼠血管内皮细胞和外膜中表达明显高于对照组。RAOEC细胞表明miR-98可抑制TRB2的表达。进一步发现mR-98在模型鼠动脉外膜和内膜表达明显减少。然后，我们检测了TRB2底物Akt的表达，我们发现内膜细胞Akt 308位苏氨酸和外膜473位丝氨酸的磷酸化水平比对照组明显减少。miR-98可调节TRB2的表达，二者在2型糖尿病鼠大动脉发病中发挥重要作用。TRB2和miR-98可作为糖尿病鼠大动脉早期治疗的靶点。. (2) miR-98与Cyclin D2在2型糖尿病血管病变中的作用：研究中我们发现，与低浓度相比，高浓度的葡萄糖（4.5g/L）能明显促进RAOEC细胞增殖，明显增加Cyclin D2表达和p-RB1水平。Cyclin D2-3’UTR有miR-98的调控位点，Western blot证实了miR-98能调控Cyclin D2和p-RB1的表达。进一步结果表明miR-98能诱导RAOEC细胞凋亡，具体机理与调节Bcl-2、Bax、Caspase 9等表达有关。高浓度的葡萄糖（4.5g/L）处理后，可降低导RAOEC细胞miR-98表达。在STZ-诱发的糖尿病模型鼠中发现，miR-98在大动脉表达明显减少，而Cyclin D2和p-RB1的表达明显增高。结果表明高浓度葡萄糖能诱导Cyclin D2高表达，降低miR-98的表达；通过调节Cyclin D2的水平，miR-98能抑制血管内皮细胞增殖，为2型糖尿病血管病变的预防和治疗提供新的靶点和科学资料。