Thrombospondin1 antagonists and diabetic nephropathy

描述（由申请方提供）：尽管增加了血糖控制并使用了肾素-血管紧张素系统抑制剂，但糖尿病肾病仍然是终末期肾病的主要原因。转化生长因子-<$（transforming growth factor-<$，TGF-<$）是糖尿病肾病发病机制中的关键分子因子。TGF-β的表达及其活性在糖尿病中增加。TGF-β是一种生物学上的潜在分子，必须转化为其活性形式，以诱导纤维化效应。该活化步骤代表了TGF-β生物活性调节的主要点。我们确定了血小板反应蛋白1（TSP 1）作为糖尿病中TGF-β激活的分子调节剂。TSP 1蛋白表达增加的糖尿病肾病介质，如葡萄糖和血管紧张素II。体外研究和STZ诱导的糖尿病伴高血压大鼠模型显示，四氨基酸肽（LSKL）对TSP 1依赖性TGF-β激活的拮抗作用阻断了葡萄糖和血管紧张素II对TGF-β活性的刺激，细胞外基质的产生，并预防和逆转了心肌纤维化。当通过腹膜内注射给药时，LSKL肽在该模型和其他非糖尿病肾纤维化和肝纤维化模型中是TGF-β依赖性纤维化的有效拮抗剂。LSKL肽代表了治疗糖尿病肾病的有效治疗策略：该肽具有选择性地仅抑制由于TSP 1介导的活化引起的TGF-β活性的致病性增加的独特优势。这将LSKL与其他抑制TGF-β作用的策略区分开来，这些策略不区分TGF-β活性的稳态水平和病理性过度。在这些研究中，我们将在新开发的1型糖尿病遗传小鼠模型（129/SvEv Ins 2秋田）中检验LSKL肽的给药通过阻断TGF-β的活化来改善肾功能和减轻肾纤维化的假设，该模型具有显著的蛋白尿和系膜硬化。在特定目标1中，小鼠将在15周的时间内每周三次接受LSKL或对照（LSAL）肽的腹膜内注射，并将评价肾功能、形态学和TGF-β信号传导。具体目标2将确定靶向血管紧张素II 1型受体和TSP 1的联合治疗方法是否具有增加的益处。重要的是，该提案将解决TGF-β的稳态功能是否受到TSP 1依赖性激活的拮抗作用的影响。具体目标3将解决阻断TSP 1激活的TGF-β是否对TGF-β的稳态功能具有有害影响，这些功能涉及全身组织病理学、肿瘤发生率、免疫细胞特征和皮肤伤口愈合。这些研究将有助于建立这种TSP 1依赖性TGF-β激活的肽拮抗剂作为糖尿病肾病新治疗药物的效用。

DESCRIPTION (provided by applicant): Despite increased glycemic control and use of renin-angiotensin system inhibitors, diabetic nephropathy remains a leading cause of end stage renal disease. The fibrogenic cytokine, transforming growth factor-¿ (TGF-¿), is a key molecular factor in the pathogenesis of diabetic nephropathy. The expression of TGF-¿ and its activity are increased in diabetes. TGF-¿ is expressed as a biologically latent molecule that must be converted to its active form in order to induce fibrogenic effects. This activation step represents a major point of regulation of TGF-¿ bioactivity. We identified thrombospondin 1 (TSP1) as the molecular regulator of TGF-¿ activation in diabetes. TSP1 protein expression is increased by mediators of diabetic nephropathy such as glucose and angiotensin II. In vitro studies and a rat model of STZ-induced diabetes with hypertension showed that antagonism of TSP1-dependent TGF-¿ activation by a four amino acid peptide (LSKL) blocked glucose and angiotensin II stimulation of TGF-¿ activity, extracellular matrix production, and prevented and reversed myocardial fibrosis. The LSKL peptide when administered by intraperitoneal injection is an effective antagonist of TGF-¿ -dependent fibrosis in this model and in other non-diabetic models of renal and hepatic fibrosis. The LSKL peptide represents an effective therapeutic strategy for treatment of diabetic nephropathy: this peptide has the unique advantage of selectively inhibiting only the pathogenic increases in TGF-¿ activity due to TSP1-mediated activation. This distinguishes LSKL from other strategies for inhibiting TGF-¿ action, which do not discriminate between homeostatic levels and pathologic excesses of TGF-¿ activity. In these studies, we will test the hypothesis that administration of the LSKL peptide improves renal function and attenuates renal fibrosis by blocking activation of TGF- ¿ in a newly developed genetic mouse model (129/SvEv Ins2 Akita) of type 1 diabetes, which has significant proteinuria and mesangial sclerosis. In Specific Aim 1, mice will receive thrice weekly i.p. injections of LSKL or control (LSAL) peptide over a 15 week period and renal function, morphology, and TGF-¿ signaling will be evaluated. Specific Aim 2 will determine whether a combined therapeutic approach that targets both the angiotensin II type 1 receptor and TSP1 has increased benefit. Importantly, this proposal will address whether homeostatic functions of TGF-¿ are compromised by antagonism of TSP1-dependent activation. Specific Aim 3 will address whether blockade of TSP1-activated TGF-¿ has deleterious effects on homeostatic functions of TGF-¿ with respect to systemic histopathology, tumor incidence, immune cell profile, and dermal wound healing. These studies will help establish the utility of this peptide antagonist of TSP1-dependent TGF-¿ activation as a novel therapeutic for diabetic nephropathy.