Enzymatic Regulation of the Podocyte

描述（由申请人提供）：肾小球性肾病呈上升趋势，目前尚无明确的蛋白尿性肾病治疗方法。在过去的几年里，我们已经表明，足细胞足突（FP）消失，蛋白尿和肾脏疾病的进展，可以特异性地诱导一种新的细胞溶质组织蛋白酶L变体（cCatL）的行动。 cCatL 因此作为调节性蛋白酶，其在肾小球损伤期间被诱导切割GTP酶发动蛋白、突触足蛋白以及CD 2AP。 这些裂解事件导致了一个新的概念，其特征在于FP消失足细胞酶疾病。在这次竞争性更新中，我们寻求澄清中和cCatL的创新方法，以更有效地治疗肾小球疾病。我们将检验抑制足细胞中cCatL活性将改善肾小球疾病的假设。 为了验证这一假设，我们将进行计算机模拟和细胞培养研究的组合，并利用我们的新型小鼠模型，该模型可诱导cCatL的特异性足细胞表达。 在诱导足细胞cCatL后，该小鼠发生足细胞FP消失和蛋白尿。我们将利用这种小鼠并研究cCatL被一组称为丝氨酸蛋白酶抑制剂的酶抑制。 我们最初的重点将是SERPINB 13（hurpin），它是经典的位于细胞溶质的皮肤和已知的组织蛋白酶L的特异性抑制剂。我们在足细胞中发现了可诱导的表达，并为SERPINB 13创建了足细胞特异性敲除动物，还拥有SERPINB 13转基因小鼠，以研究这些酶在面对高cCatL时将如何表现。 我们还将分析足细胞碱化对cCatL活性的影响，因为正常足细胞胞内pH接近7.3，但在疾病中更接近p7.0时cCatL活性的最佳pH。我们有初步数据显示，在LPS小鼠模型中，使用例如L-谷氨酰胺补充剂的碱化足细胞被保护免于蛋白尿。 总之，生物化学，细胞培养和遗传动物模型将用于一种新的多方面的方法来解决蛋白尿性肾病。

DESCRIPTION (provided by applicant): Glomerular kidney diseases are on the rise with no definitive treatment for proteinuric renal disorders available yet. Over the past years, we have shown that podocyte foot process (FP) effacement, proteinuria and kidney disease progression can be specifically induced by the action of a novel cytosolic cathepsin L variant (cCatL). cCatL hereby acts as regulatory protease that is induced during glomerular injury cleaving th GTPase dynamin, synaptopodin as well as CD2AP. These cleavage events have led to a novel concept that characterizes FP effacement as podocyte enzymatic disease. In this competitive renewal, we seek to clarify innovative ways to neutralize cCatL in an effort to treat glomerular disease more effectively. We will test the hypothesis that inhibition of cCatL activity in podocytes will improve glomerular disease. To test this hypothesis, we will perform a combination of in silico and cell culture studies and make use of our novel mouse model that is inducible for specific podocyte-expression of cCatL. Upon induction of podocyte cCatL, this mouse develops podocyte FP effacement and proteinuria. We will utilize this mouse and study inhibition of cCatL by a group of enzymes called serpins. Our initial focus will be on SERPINB13 (hurpin) which is classically located in the cytosol of skin cels and known to be a specific inhibitor for cathepsin L. We found inducible expression in podocytes and have created podocyte-specific knockout animals for SERPINB13 and also possess SERPINB13 transgenic mice to study how these enzymes will perform in the face of high cCatL. We will also analyze the effects of podocyte alkalinization on cCatL activity as normal podocyte intracellular pH is close to 7.3 but in disease is closer to the pH optimum for cCatL activity at p 7.0. We have preliminary data showing that alkalinized podocytes using for example L-glutamine supplementation are protected from proteinuria in the LPS mouse model. In sum, biochemical, cell culture and genetic animal models will be utilized in a novel multifaceted approach to tackle proteinuric renal diseases.