Mechanisms of Neuropeptides Action in Diabetes

神经肽在糖尿病中的作用机制

• 批准号: 8811708
• 负责人: ARISTIDIS VEVES
• 金额: $ 13.05万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-30 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8811708
• 关键词: Ablation; Affinity; Animal Model; Asthma; Blood Vessels; Bronchodilator Agents; Calcium; Calcium Channel; Calcium Channel Blockers; Calcium-Sensing Receptors; Cell Degranulation; Cell membrane; Cells; Chemicals; Comparative Study; Complex; Cromolyn Sodium; Dermal; Development; Diabetes Mellitus; Diabetic Foot Ulcer; Diabetic wound; Drug Formulations; Enhancers; Future; Genetic; IL6 gene; IgE Receptors; Inflammation Mediators; Inflammatory; Interleukin-6; Investigation; Ligation; Mast Cell Stabilizer; Mediating; Medical; Membrane Potentials; Modeling; Mus; Neuropeptides; Outcome; Parents; Patients; Pharmaceutical Preparations; Phenotype; Play; Process; Property; Proteins; Research; Role; STIM1 gene; Skin; Smooth Muscle Myocytes; Streptozocin; T-Cell Activation; T-Lymphocyte; Testing; Topical application; Tumor Necrosis Factor-alpha; Wound Healing; channel blockers; comparative efficacy; cytokine; diabetic; diabetic wound healing; drug discovery; efficacy testing; extracellular; improved; in vivo; inflammatory marker; interest; macrophage; mast cell; mouse model; parent grant; pre-clinical; prophylactic; prototype; public health relevance; reagent standard; receptor; response; small molecule; standard of care; wound

 DESCRIPTION (provided by applicant): On-going research in our lab, which is supported by the parent grant, has begun to clarify the critical role played by activated mast cells (MC) in the development of diabetic foot ulcers (DFU). More specifically, we found that in the skin from patients with diabetes (DM) the number of activated MC is significantly higher than healthy controls and correlates with dermal inflammatory cells and dermal inflammatory cytokines, as well as with circulating levels of inflammatory markers such as IL6 and TNFα. Furthermore, we were able to demonstrate in a murine model of diabetic wound healing that the number of degranulated MC was significantly higher in the skin of streptozotocin-induced diabetic (STZ-DM) mice. Treatment with the MC stabilizer disodium cromoglycate (DSCG) significantly accelerated wound healing, accompanied with significantly reduced amount of degranulated MC. In addition, DSCG considerably reduced the M1 macrophage phenotype, both pre- and post-wounding in this animal model. Thus, our preclinical proof-of-concept results strongly support that the inhibition of MC degranulation is a viable approach for treating DFU. MC degranulation is controlled by an elevated level of cytosolic calcium that is mediated by the stored operated calcium (SOC), and to a lesser extent, by the receptor potential canonical (TRPC) channels. The best characterized SOC channel is the calcium selective orai, also known as calcium-release activated calcium (CRAC) channel that is expressed not only by MC but also by T cells as well as vascular smooth muscle cells. Activation of MC stimulates the opening of the orai channels for calcium influx. Small molecule orai/CRAC channel blockers have been shown to potently inhibit MC degranulation and T-cell activation. In this revised proposal, we plan to evaluate the effects of calcium channel blockers in our mouse model of diabetic wound healing. Our main hypothesis is that calcium channel blockers are more efficacious in improving diabetic wound healing. In the first specific aim, we will focus on the synthesis of the prototype calcium channel blockers and formulation development for topical drug administration. The synthetic process for each compound will require three chemical steps from available reagents and standard purification technics such as column separation and recrystallization. Because of the intrinsic hydrophobic properties of the prototype compounds, we will also employ solubilizing enhancers to formulate the drugs for topical delivery in in vivo efficacy studies. In the second specific aim, we will test the efficacy of developed prototypes in animal models of diabetic wound healing. For this, we will employ the streptozotocin-induced diabetic (STZ-DM) wild-type (WT) C57BL/6J mouse model that was used in the parent application to evaluate its ability to improve wound healing either used prophylactically before the wound creation or applied topically post wounding. We will also compare the efficacy of this treatment to the only commercially available MC stabilizer, DSCG.

 描述（由适用提供）：由父母赠款支持的我们实验室中正在进行的研究已开始阐明激活的肥大细胞（MC）在 糖尿病足溃疡（DFU）的发展。更具体地说，我们发现，在糖尿病患者（DM）的皮肤中，活化的MC的数量显着高于健康对照，并且与皮肤炎性细胞和皮肤炎性细胞因子以及与IL6和TNFα等炎性标记物的循环水平相关。此外，我们能够在糖尿病伤口愈合的鼠模型中证明，在链球菌诱导的糖尿病（STZ-DM）小鼠的皮肤中，脱粒的MC的数量明显更高。用MC稳定剂二钠二钠（DSCG）处理可显着加速伤口愈合，并伴有明显降低的MC量。此外，在该动物模型中，DSCG认为降低了M1巨噬细胞表型，无论是在该动物模型中的前后。这是，我们的临床前概念证明结果强烈支持MC脱粒是一种可行的DFU方法。 MC脱粒受到升高的胞质钙水平的控制，该钙是由储存的工作钙（SOC）介导的，并且在较小程度上是由接收器电位规范（TRPC）通道介导的。最佳特征的SOC通道是钙选择性ORAI，也称为钙释钙活化钙（CRAC）通道，不仅由MC，而且由T细胞以及血管平滑肌细胞表达。 MC的激活刺激了Orai通道的打开钙影响。小分子ORAI/CRAC通道阻滞剂已被证明可能抑制MC脱粒和T细胞激活。在这项修订的建议中，我们计划评估钙通道阻滞剂在我们的糖尿病伤口愈合模型中的影响。我们的主要假设是钙通道阻滞剂在改善糖尿病伤口愈合方面更有效。在第一个特定目的中，我们将重点关注局部药物给药的原型钙通道阻滞剂和配方开发的合成。每种化合物的合成过程将需要从可用试剂和标准纯化技术（例如柱分离和重结晶）中进行三个化学步骤。由于原型化合物的固有疏水性能，我们还将采用硫化增强剂来制定药物以在体内效率研究中局部递送。在第二个特定目标中，我们将测试开发原型在糖尿病伤口愈合模型中的有效性。为此，我们将采用链霉菌素诱导的糖尿病（STZ-DM）野生型（WT）C57BL/6J小鼠模型，该模型在母体应用中用于评估其改善伤口愈合的能力。我们还将将此处理的功效与唯一可商购的MC稳定器DSCG进行比较。