Expression and Significance of Plasma miR-1181 in Patients with Type 2 Diabetes Mellitus
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摘要:
目的 miRNA可作为预测糖尿病发生和发展的生物标志物,探索miR-1181在2型糖尿病(Type 2 Diabetes Mellitus,T2DM)患者的表达情况。 方法 运用Agilent miRNA芯片对T2DM患者(n = 5)和正常对照组(n = 5)血浆样本进行差异筛选,荧光定量PCR技术(quantitative Real Time PCR,qRT-PCR)验证出差异表达的miRNA;通过生物信息学方法预测其靶基因并绘制miRNA-靶基因-代谢通路关系网络图,扩大样本检测血浆中差异表达的miRNA:miR-1181和靶基因MAP2K2、MAPK12表达水平。 结果 miRNA芯片筛选和qRT-PCR验证出与T2DM相关的差异表达miRNA:miR-1181,生物信息学分析得出miR-1181靶基因有CCND1、PI3KR2、MAP2K2和MAPK12;T2DM患者血浆中 miR-1181的表达水平明显下降(P < 0.001),而靶基因MAPK12 mRNA的表达水平明显升高(P < 0.01)。 结论 2型糖尿病患者血浆miR-1181水平降低,可能与其抑制靶基因MAPK12对MAPK通路影响有关。 Abstract:Objective To verify that miRNA can be used as biomarkers to predict the occurrence and development of diabetes mellitus (DM), and to explore the expression of miR-1181 in patients with type 2 diabetes mellitus (Type 2 Diabetes Mellitus). Methods Agilent miRNA chip was used to screen plasma samples from patients with T2DM (n = 5) and normal control group (n = 5). The differentially expressed miRNA were verified by fluorescence quantitative PCR (qRT-PCR). The target genes were predicted by bioinformatics methods, and the relationship network diagram of miRNA - target gene - metabolic pathway was drawn. The differentially expressed miRNA: miR-1181, MAP2K2 and MAPK12 in plasma were detected by expanding samples. Results miRNA chip screening and qRT-PCR verified the differentially expressed miRNA associated with T2DM: miR-1181.Bioinformatics analysis showed that the target genes of miR-1181 were CCND1, PI3KR2, MAP2K2 and MAPK12. The expression of miR-1181 in the plasma of T2DM was significantly decreased (P < 0.001). While expression of MAPK12 mRNA was increased (P < 0.01). Conclusion The decrease of plasma miR-1181 in T2DM patients may be related to inhibiting effect of target gene MAPK12 on MAPK pathway. -
Key words:
- miR-1181 /
- Type 2 Diabetes /
- Target Genes /
- MAPK12
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表 1 qRT-PCR 引物及退火温度列表
Table 1. qRT-PCR Primer and annealing temperature list
引物名称 序列(5′to3′) 退火温度Tm(℃) U6-F CTCGCTTCGGCAGCACATATACT 58 U6 -R ACGCTTCACGAATTTGCGTGTC cel-miR-39-5p-RT GTCGTATCCAGTGCAGGGTCCGAGGTATTCGCACTGGATACGACTATTAC cel-miR-39-5p-F TGGGAGCTGATTTCGTCTTG 59.7 hsa-mir-1181-RT GTCGTATCCAGTGCAGGGTCCGAGGTATTCGCACTGGATACGACCGGCTC hsa-mir-1181-F AATAACCGTCGCCGCCACCCG 72.5 GAPDH-F: GGTTGTCTCCTGCGACTTCA 58.2 GAPDH-R TGGTCCAGGGTTTCTTACTCC 58.2 MAP2K2-F: CACTATCAACCCTACCATCGC 57.3 MAP2K2-R: CGCTTCCTTTGCTGCTCAT 58.8 MAPK12-F TCTGTCCTGACCAACGCAA 57.6 MAPK12-R AAGGGACTCAAAGTATGGATGG 57.6 表 2 患者和对照组临床资料比较[(
$\bar x \pm s$ ),M(P25,P75)]Table 2. Clinical data of patients between two groups
项目 对照组(n = 65) T2DM组(n = 65) 性别(男/女) 34/31 36/29 年龄(岁) 33.5 ± 11.69 56.27 ± 12.28▲ BMI(kg/m2) 22.09 ± 3.18 23.80 ± 3.58▲ FPG(mmol/L) 5.0(4.72,5.44) 7.05(5.45,9.7) ▲ TC(mmol/L) 4.47 ± 0.52 4.21 ± 1.0▲ TG(mmol/L) 1.1(0.79,1.64) 1.61 (1.3,2.4) ▲ LDL(mmol/L) 2.48 ± 0.44 2.88 (2.41,3.11) ▲ HDL(mmol/L) 1.37(1.21,1.68) 0.98 (0.83,1.12) ▲ UA(μmol/L) 344.01(300.25,382.5) 320.3(284.4,354.15) Cr(μmol/L) 76.0(62.5,85.25) 62.75 (52.03,70.7) eGFR(ml/min*1.73m2) 106.76 ± 17.04 108.97 ± 28.15 UACR(mg/g) - 26.8(12.34,50.86) 注:正态性分布资料以(${{\bar x} } \pm s$)表示,非正态性资料以M(P25,P75)表示,与对照组比较,▲P < 0.05。BMI:体重指数;FPG:空腹血糖;TC:总胆固醇;TG:甘油三酯;LDL:低密度脂蛋白;HDL:高密度脂蛋白;UA:尿酸;Cr:肌酐;eGFR:估计肾小球滤过率;UACR:尿白蛋白肌酐比。 表 3 miR-1181靶基因预测结果
Table 3. Prediction Results of miR-1181 Target Genes
miRNA Transcript Total score Total energy Max score Max energy Strand miRNA
lengthTranscript
lengthTranscript
PostionmiR-1181 ZBTB4 150 −18.25 150 −18.25 3 21 5961 5584 miR-1181 MAP2K2 157 −25.76 157 −25.76 1 21 1759 1645 miR-1181 MAPK12 156 −15.92 143 −15.92 2 21 1778 1644 -
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