Metabolic Markers of Carbonyl, Oxidative and Nitrosative Stress in the DCCT/EDIC

描述（由申请人提供）：葡萄糖水平升高已被确定为1型糖尿病微血管和大血管并发症进展的罪魁祸首和/或风险因素。我们对DCCT结束时测定的晚期糖基化终产物（AGEs）的生化性质和临床意义的研究表明，皮肤胶原AGEs是DCCT结束后16年微血管和亚临床心血管并发症的长期预测因子。这种预测作用与HbA 1c、年龄、糖尿病病程和其他风险因素无关。然而，在实验室水平上测定皮肤中的特异性AGEs是不可行的，除了内源性荧光。因此，如何将这些令人兴奋的发现转化为更可行的替代方法成为一项紧迫的任务。 利用PAR公告，我们提出了一个实用的和临床相关的假设，即测定血清和尿液中羰基（AGEs）、氧化和亚硝化应激的蛋白标志物（“代谢应激标志物”），单独或一组，在基线或纵向，将使我们能够预测DCCT/EDIC试验中哪些1型糖尿病患者发生微血管和大血管并发症的风险长期增加（主要终点）。为了实现这一目标，一个由凯斯西储大学DCCT/EDIC研究小组的生物化学家和临床糖尿病学家调查员以及乔治华盛顿大学生物统计中心DCCT数据协调中心的统计学家组成的团队已经成立。 将使用具有同位素稀释的强大液相色谱质谱法生成数据，该方法可准确测定9种标志物，即2种糖化产物（果糖-赖氨酸和葡萄糖窗格），2种糖氧化产物（羧甲基-赖氨酸（CML）和羧乙基-赖氨酸（CEL），2种氧代醛产物（乙二醛和甲基乙二醛氢咪唑酮），2种金属催化氧化产物（烯丙基赖氨酸和2-氨基己二酸）和一种氧化产物（蛋氨酸亚砜）。将通过荧光HPLC法测定戊糖苷和亚硝基化产物3-硝基酪氨酸。将对统计模型进行测试，以确定这些血清和尿液标记物与皮肤AGEs的关系，对并发症的长期风险的贡献，以及对已知风险因素的调整是否会削弱这些标记物的预测能力。这项研究的成功结果可能会导致实施“代谢应激面板”作为辅助HbA 1c在临床环境中评估糖尿病并发症的代谢危险因素。

DESCRIPTION (provided by applicant): Elevated glucose levels have been identified as culprits and/or risk factors for the progression of microvascular and macrovascular complications in Type 1 diabetes. Our studies on the biochemical nature and clinical significance of advanced glycation endproducts (AGEs) determined near DCCT closeout demonstrated that skin collagen AGEs are robust long-term predictors of microvascular and subclinical cardiovascular complications 16 years after the end of DCCT. This predicting effect is, independent of HbA1c, age, diabetes duration, and other risk factors. However, the determination of specific AGEs in skin is not feasible at the bench level, except for intrinsic ski fluorescence. Therefore, how to translate these exciting findings to a more feasible alternative approach becomes a pressing task. Taking advantage of the PAR announcement, we propose to investigate a pragmatic and clinically relevant hypothesis that determination of serum and urine protein markers of carbonyl (AGEs), oxidative and nitrosative stress ("metabolic stress markers"), individually or a set, at baseline or longitudinally, will allow us to predict which individuals with Type 1 diabetes in the DCCT/EDIC trial are at long-term increased risk of developing micro- and macrovascular vascular complications (primary endpoints). To achieve this goal, a team consisting of biochemists and a clinical diabetologist investigator from the DCCT/EDIC Research Group at Case Western Reserve University and statisticians at theDCCT Data Coordinating at the Biostatistics Center of the George Washington University has been assembled. Data will be generated using a powerful liquid chromatography mass spectrometry method with isotope dilution that accurately determines 9 markers, i.e. 2 glycation products (fructose-lysine and glucose pane), 2 glycoxidation products (carboxymethyl-lyinse (CML) and carboxyethyl-lysine (CEL), 2 oxoaldehyde products (glyoxal and methylglyoxal hydroimidazolones), 2 products of metal catalyzed oxidation (allysine and 2-amino adipic acid) and one oxidation product (methionine sulfoxide). Pentosidine and the nitrosylation product 3-nitrotyrosine will be determined by fluorescence HPLC. Statistical models will be tested to determine how these serum and urine markers relate to skin AGEs, contribute to long-term risk of complications, and whether adjustment for known risk factors attenuates the predicting power of these markers. A successful outcome of this research may lead to the implementation of a "metabolic stress panel" as adjunct to HbA1c for the evaluation of metabolic risk factors for diabetic complications in the clinical setting.