Correction of glutathione deficiency for treatment of diabetic nephropathy

描述(由申请人提供)： 氧化应激和谷胱甘肽(GSH)失衡是糖尿病肾病发病机制的主要因素。循环单核细胞参与这一过程，因为这些细胞携带高氧化负担，并在糖尿病肾组织中发现。目前治疗糖尿病肾病氧化应激的选择有限，而且只有部分有效，因此人们对开发新策略的兴趣很高。N-乙酰半胱氨酸(NAC)和水飞蓟(MTH)植物黄酮木脂素是具有互补抗氧化特性的营养补充剂。这两种补充剂都能直接中和有毒的自由基，但更重要的是，NAC是细胞内产生GSH的底物，而MTH黄酮木脂素是许多参与GSH代谢的细胞酶的诱导者，包括谷胱甘肽还原酶(GSH-R)、谷胱甘肽过氧化物酶(GSH-Px)、谷胱甘肽S转移酶(GST)和超氧化物歧化酶(SODase)。我们认为，联合口服补充NAC和MTH黄酮木脂素可以减少蛋白尿以及氧化应激和炎症的尿液和全身表现，这是T2 DM和相关肾病患者的特征。我们预计这些效果将以最小的副作用或没有副作用，以及良好的患者耐受性实现。为了验证这一假设，我们提出了一项双盲随机、安慰剂对照、五臂先导研究，其中包括T2 DM和已确诊肾病患者的剂量范围成分。干预措施包括单独和联合口服一种水平的NAC和两种水平的MTH黄酮木素或安慰剂，为期三个月。主要的结果将是尿白蛋白排泄，这是肾小球损伤的标志。次要结果指标将是肾小管损伤的标志物α-1微球蛋白，以及尿中炎性细胞因子和C-C趋化因子的排泄量，即肾脏炎症的标志物。在同一患者的血浆和外周血单核细胞中，我们将分析GSH含量和GSH代谢酶的活性。此外，我们将分析血浆和尿糖蛋白质组，重点是那些作为炎症细胞信使和激素的糖蛋白。这些变量将根据治疗分配和积极治疗的普遍血液和尿液水平进行监测。在整个试验过程中，我们将监测这种联合治疗的安全性和耐受性。这一假说的验证将为NAC和MTH黄酮木素联合治疗的有效性、安全性和耐受性提供初步证据，并将为未来设计更明确的临床研究和T2 DM和糖尿病肾病动物模型的机制研究奠定必要的第一步。 公共卫生相关性： 氧化应激(OS)是指有毒的促氧化剂化学物质的大量产生和低释放，在2型糖尿病(T2 DM)和糖尿病肾病患者中非常常见。退伍军人有很高的T2 DM、糖尿病肾病和相关OS的患病率。OS会导致糖尿病肾病的进展，导致蛋白在尿中溢出，最终需要透析或肾移植。在大多数情况下，可用的治疗方法无法阻止糖尿病肾病的进展和尿液中蛋白质的溢出。我们怀疑这是因为没有传统的治疗方法可以完全纠正糖尿病肾病的OS。我们建议测试两种有效的抗氧化营养补充剂N-乙酰半胱氨酸和水飞蓟提取物水飞蓟素的组合是否能够纠正患有T2 DM和糖尿病肾病的退伍军人的OS和尿蛋白溢出。

DESCRIPTION (provided by applicant): Oxidative stress and glutathione (GSH) imbalance are major contributors to the pathogenesis of diabetic nephropathy. Circulating monocytes participate to this process since these cells carry a high oxidative burden and are found in diabetic renal tissue. Current options for the treatment of oxidative stress in diabetic nephropathy are limited and only partially effective, thus interest in the development of new strategies is high. N-acetylcysteine (NAC) and the milk thistle (MTh) plant flavonolignans are nutritional supplements with complementary antioxidant properties. Both supplements are capable of neutralizing directly toxic free radicals but, more importantly, NAC is substrate for the intracellular generation of GSH and the MTh flavonolignans are inducers of many cellular enzymes participating in GSH metabolism, including GSH-reductase (GSH-R), GSH-peroxidase (GSH-Px), GSH-S-transferases (GST) and superoxide dismutase (SOD). We propose that combined oral supplementation of NAC and MTh flavonolignans will reduce proteinuria and urinary and systemic manifestations of oxidative stress and inflammation, which are characteristically observed in patients with T2DM and related nephropathy. We expect these effects to be achieved with minimal or no side effects, and with good patient tolerance. To test this hypothesis, we propose a double-blind randomized, placebo-controlled, five-arm pilot study which includes a dose ranging component in patients with T2DM and established nephropathy. Intervention will consist of the individual and combined oral administration of one level of NAC and two levels of MTh flavonolignans or placebo for three months. The primary outcome measure will be urinary excretion of albumin, a marker of glomerular injury. Secondary outcome measures will be alpha-1 microglobulin, a marker of tubular injury, and urinary excretion of inflammatory cytokines and C-C chemokines, i.e. markers of renal inflammation. In plasma and in peripheral blood monocytes from the same patients, we will analyze GSH content and activity of GSH metabolizing enzymes. In addition, we will analyze the plasma and urine glycoproteome, with focus on those glycoproteins serving as inflammatory cell messengers and hormones. These variables will be monitored in relation to both treatment allocation and prevalent blood and urine levels of the active treatment. Throughout the trial, we will monitor the safety and tolerability of this combination treatment. Validation of the proposed hypothesis will provide preliminary evidence of efficacy, safety and tolerability of combination treatment with NAC and MTh flavonolignans and it will be the necessary first step for the future design of more definitive clinical investigation and of mechanistic research in animal models of T2DM and diabetic nephropathy. PUBLIC HEALTH RELEVANCE: Oxidative stress (OS) consists of high production and low disposal of toxic pro-oxidant chemicals, and it is very common in patients with Type 2 Diabetes Mellitus (T2DM) and diabetic kidney disease. Veterans have high prevalence of T2DM, diabetic kidney disease, and associated OS. OS causes progression of diabetic kidney disease, with spilling of protein in the urine and, eventually, with the need for dialysis or kidney transplantation. In most cases, the available treatments cannot stop the progression of diabetic kidney disease and the spilling of protein in the urine. We suspect this is because no conventional treatments can completely correct OS in diabetic kidney disease. We propose to test if the combination of two potent antioxidant nutritional supplements, N-acetylcysteine and the milk thistle extract silymarin, is capable of correcting the OS and the spilling of protein in the urine in Veterans with T2DM and diabetic kidney disease.