2型糖尿病血清Afamin蛋白组学研究

Proteomics Research of Serum Afamin in Type 2 Diabetes

Afamin是与2型糖尿病(T2DM)密切相关的糖蛋白,本研究主要利用免疫印迹(Western Blot)和酶联免疫(ELISA)的方法研究T2DM血清Afamin的变化趋势,并进一步利用同位素标记相对和绝对定量(iTRAQ)方法以及免疫印迹的方法探讨并验证T2DM病人血清中Afamin糖基化修饰状态的变化。结果显示,T2DM病人Afamin条带灰度较健康对照显著增强(p=0.0101),血清Afamin在T2DM组和健康对照组浓度分别6.03±0.19μg/mL和5.07±2.12μg/mL,且两组之间具有显著性差异(p=0.0019);在Afamin高甘露糖型糖基化修饰异质性分析中,原血清以及Con A洗脱蛋白质中均检测到Afamin,而流穿蛋白质中未检测到Afamin,即所有血清Afamin都存在高甘露糖型糖链;iTRAQ标记定量数据显示血清高甘露糖型Afamin在T2DM中发生显著性上调,Western Blot结果验证了这一变化趋势(p=0.0462)。

Afamin is a sort of glycoprotein closely related to type 2 diabetes(T2DM).Western Blot and ELISA(enzyme linked immunosorbent assay)were used to analyze the changing trend of serum Afamin from T2DM,iTRAQ(isobaric tags for relative and absolute quantification)was applied to explore the glycosylation modified state of serum Afamin from T2DM,and Western Blot was adopted to validate this data.The results showed that Afamin appeared up-regulated significantly in T2DM(p=0.0101).The average content of serum Afamin was 6.03±0.19μg/mL in T2DM,which was 5.07±2.12μg/mL in the health control(p=0.0019).In high mannose heterogeneity analysis,we could see Afamin developing bands in crude serum and serum proteins eluted from Con A.However,we could not see Afamin band in the flow through.The results indicated that all of serum Afamin had high mannose glycan.The results of iTRAQ showed high mannose type Afamin was up-regulated in T2DM and Western Blot validated this data(p=0.0462).