Nitric Oxide and gastric motility in female diabetics

女性糖尿病患者的一氧化氮和胃动力

• 批准号: 7758382
• 负责人: PANDU R GANGULA
• 金额: $ 18.13万
• 依托单位: MEHARRY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7758382
• 关键词: Abdominal Pain; Accounting; Affect; American; Anabolism; Attenuated; Biological; Biopterin; Body Weight decreased; Chronic; Clinic; Clinical; Complications of Diabetes Mellitus; Data; Diabetes Mellitus; Dimerization; Disease; Dyspepsia; Enzymes; Event; Experimental Models; Female; Functional disorder; Gastric Emptying; Gastric Tissue; Gastrointestinal Motility; Gastroparesis; Guanosine Triphosphate; Health; Hyperglycemia; Impairment; In Vitro; Insulin; Lead; Mechanics; Mediating; Messenger RNA; Molecular; Muscle Contraction; Nausea and Vomiting; Nervous system structure; Neurons; Neurotransmitters; Nitric Oxide; Nitric Oxide Synthase Type I; Non-Insulin-Dependent Diabetes Mellitus; Pathogenesis; Pathway interactions; Patients; Production; Progressive Disease; Rattus; Recurrence; Regulation; Relaxation; Reporting; Retina; Role; Sex Bias; Sex Characteristics; Solid; Stomach; Streptozocin; Structure; Supplementation; Symptoms; Syndrome; Testing; Tissues; Type I Insulin; Vomiting; Woman; Work; base; cell motility; cofactor; design; diabetic; diabetic gastroparesis; diabetic rat; dimer; enzyme activity; gastrointestinal; in vivo; inhibitor/antagonist; kidney vascular structure; motility disorder; neuron loss; new therapeutic target; protein expression; research study; tetrahydrobiopterin

DESCRIPTION (provided by applicant): Gastroparesis is a disabilitating disease affecting predominantly young women. The biological basis of this disorder and its associated gender bias remains poorly understood. Recent studies have implicated a role for dysregulation of neuronal nitric oxide synthase (nNOS) in myenteric neurons and we surmised that gender differences in nitrergic control of gastric motility may possibly account for observed vulnerability of females to diabetic gastroparesis. Our preliminary data suggests that a significant impairment of nitrergic relaxation and delayed gastric emptying for solids was demonstrated selectively in females after diabetes induction with streptozotocin (STZ). Most importantly, we have also shown that changes in nitrergic relaxation in both healthy and diabetic rats correlates well with the state of dimerization of nNOS1 but not with the expression of total nNOS (1, 2 and 3). Dimerization of nNOS is essential for activity of this enzyme but has not been previously studied in gastrointestinal tissue. In this proposal, we have focused on the role of tetrahydrobiopterin (BH4), an essential cofactor for nNOS activity that is required to maintain the homodimeric structure of nNOS. BH4 levels in tissue in turn depend on the activity of GTP cyclohydrolase1 (GTPCH1), the rate limiting enzyme in the de novo synthesis of BH4. In preliminary experiments we have demonstrated that GTPCH1 mRNA and protein expression are down regulated in the diabetic female gastric tissues along with significantly reduced BH4 content, suggesting reduced synthesis of biopterins leading to decreased nitrergic relaxation. In support of this, the GTPCH1 inhibitor, 2, 4 diamino-6-hydroxypyrimidine (DAHP), reduced nNOS1 dimerization, NO release and nitrergic relaxation of gastric tissue in normal female rats in vitro. Conversely, the exogenous addition of BH4 to female gastric tissues in vitro reverses a hyperglycemia-induced decrease in nitrergic relaxation. In addition, in vivo BH4 treatment attenuated reduced nNOS activity in diabetic gastric tissue. We therefore hypothesize that impaired biosynthesis of gastric BH4 accounts for the decrease in nNOS activity and nitrergic relaxation in female diabetic gastroparesis. To test this hypothesis we propose the following specific aims: Specific Aim 1: To determine whether diabetes results in impairment of the BH4 biosynthetic pathway, nNOS dimerization, nNOS activity and nitrergic relaxation in female rat gastric tissues. Specific Aim 2: To investigate whether BH4 supplementation restores delayed gastric emptying, impaired gastric nNOS dimerization, nNOS activity, NO synthesis and nitrergic relaxation in female diabetic rats. The data from these studies will provide important information as to the mechanisms of regulation of nNOS function in normal and diabetic female rat gastric tissues and thereby enhance our understanding of the pathophysiology of gastroparesis. Further, it may lead to the discovery of novel therapeutic targets for the management of this challenging clinical syndrome.Diabetes mellitus is a chronic progressive disease that affects 16 million Americans. Diabetes causes several complications that affect retina, kidney, vascular, gastrointestinal, and nervous system. The mechanism through which diabetic complications develop is unclear. Gastric dysmotility or gastropathy is one of the vigorous complications of diabetic mellitus in clinics and can cause disabling symptoms including nausea, vomiting and weight loss and often leads to delayed or accelerated gastric emptying. Reports indicate that 80% of patients are women suffering with this problem. Although the exact pathogenesis remains unknown, there is fairly convincing evidence in experimental models that diabetes results in the malfunctioning of specific neurons that produce the neurotransmitter nitric oxide (NO). This condition is due to loss of expression of the enzyme responsible for NO production (neuronal nitric oxide synthase: nNOS) and can be reversed with insulin replacement. The molecular events leading to loss of nNOS expression and altered (delay or accelerate) gastric emptying (mechanical due to loss of neuronal control of stomach muscle contractions and relaxations) in diabetic stomachs remain unknown. The data from these studies will provide important information as to the mechanisms of regulation of NO mediated gastric emptying and stomach contractions in diabetic female rat. Further, it may lead to the discovery of novel therapeutic targets for the management of this challenging clinical syndrome.

描述（由申请人提供）：胃轻瘫是一种致残性疾病，主要影响年轻女性。这种疾病的生物学基础及其相关的性别偏见仍然知之甚少。最近的研究表明，神经型一氧化氮合酶（nNOS）在肌间神经元的失调的作用，我们推测，性别差异的氮能控制胃动力可能是观察到的女性糖尿病胃轻瘫的脆弱性。我们的初步数据表明，一个显着的损害氮能松弛和延迟固体胃排空表现出选择性的女性糖尿病诱导后，链脲佐菌素（STZ）。最重要的是，我们还表明，健康和糖尿病大鼠的氮能松弛的变化与nNOS 1的二聚化状态密切相关，但与总nNOS的表达无关（1，2和3）。nNOS的二聚化对于这种酶的活性是必不可少的，但以前没有在胃肠道组织中进行过研究。在这个建议中，我们集中在四氢生物蝶呤（BH 4）的作用，一个必要的辅因子的nNOS活性，需要保持nNOS的同源二聚体结构。组织中的BH 4水平反过来取决于GTP环水解酶1（GTPCH 1）的活性，GTP环水解酶1是BH 4从头合成的限速酶。在初步的实验中，我们已经证明，GTPCH 1 mRNA和蛋白质表达下调，在糖尿病女性胃组织沿着显着减少BH 4含量，表明减少合成的生物蝶呤，导致减少氮能松弛。GTPCH 1抑制剂2，4-diamino-6-hydroxypyrimidine（DAHP）在体外可减少正常雌性大鼠胃组织nNOS 1二聚化、NO释放和氮能舒张。相反，外源性添加BH 4的女性胃组织在体外逆转高血糖症诱导的氮能松弛的减少。此外，体内BH 4治疗减弱了糖尿病胃组织中nNOS活性的降低。因此，我们推测，胃BH 4的生物合成受损占nNOS活性和氮能松弛女性糖尿病胃轻瘫的下降。为了验证这一假设，我们提出了以下具体目标：具体目标1：确定糖尿病是否会导致BH 4生物合成途径，nNOS二聚化，nNOS活性和雌性大鼠胃组织中的氮能松弛的损害。具体目标二：研究补充BH 4是否能恢复雌性糖尿病大鼠胃排空延迟、胃nNOS二聚化受损、nNOS活性、NO合成和氮能舒张。这些研究的数据将提供重要的信息，正常和糖尿病雌性大鼠胃组织中的nNOS功能的调节机制，从而提高我们对胃轻瘫的病理生理学的理解。糖尿病是一种慢性进行性疾病，影响着1600万美国人。糖尿病引起影响视网膜、肾脏、血管、胃肠道和神经系统的几种并发症。糖尿病并发症发生的机制尚不清楚。胃动力障碍或胃病是糖尿病临床上严重的并发症之一，可引起包括恶心、呕吐和体重减轻在内的致残性症状，并常导致胃排空延迟或加速。报告显示，80%的患者是患有这一问题的妇女。虽然确切的发病机制尚不清楚，但在实验模型中有相当令人信服的证据表明，糖尿病导致产生神经递质一氧化氮（NO）的特定神经元发生故障。这种情况是由于负责NO产生的酶（神经元型一氧化氮合酶：nNOS）的表达丧失，并且可以通过胰岛素替代来逆转。在糖尿病胃中导致nNOS表达丧失和胃排空改变（延迟或加速）（由于神经元对胃肌肉收缩和舒张的控制丧失而引起的机械性）的分子事件仍然未知。这些结果为进一步研究NO调节糖尿病雌性大鼠胃排空和胃收缩的机制提供了重要信息。此外，它可能会导致发现新的治疗靶点，用于管理这种具有挑战性的临床综合征。