Untersuchungen zur biologischen Rolle von neuen Kazal-typ Inhibitoren LEKTI-2 und LEKTI-3

通过对健康皮肤角质层抗菌肽的分析，发现了两种新的“淋巴上皮型”蛋白酶抑制剂(lekti-2和lekti-3)。虽然重组lekti-2具有抗菌特性，是激肽释放酶5(KLK-5)的选择性抑制剂，但重组lekti-3缺乏抗菌活性，但对胰蛋白酶KLK5和胰凝乳酶KLK7都有很强的抑制活性。这两种酶在表皮脱屑中起着至关重要的作用，这种脱屑在Netherton综合征中受到干扰，Netherton综合征是一种炎症性皮肤病，由于lekti-1基因缺陷导致lekti-1缺乏症，导致皮肤屏障破坏。为了研究这两种新的LEKTI基因在脱皮中是否具有相似的重要性，将产生lekti-2基因敲除小鼠和随后的lekti-3基因敲除小鼠以及过度表达这些LEKTI基因的转基因小鼠。在我实验室的这个合作项目中，将克隆小鼠lekti-2和-3的同源基因，分析其在小鼠中的组织表达，然后重组表达，用于分析抗菌和蛋白水解酶抑制特性，特别是对脱屑相关激肽释放酶的抑制。我们将进一步产生用于免疫组织化学的mLEKTI-2和-3抗体以及用于lekti-2和3-的ELISA，以分析其在各种小鼠模型脱屑中的作用。

Analyses of healthy skin stratum corneum for antimicrobial peptides led to the discovery of two novel „lympho-epithelial Kazal-Type” protease inhibitors (LEKTI-2 and LEKTI-3). Whereas recombinant LEKTI-2 has antimicrobial properties and is a selective inhibitor of Kallikrein 5 (KLK-5), recombinant LEKTI-3 lacks antimicrobial activity, but exhibits potent inhibitory activities against both, tryptic KLK5 and chymotryptic KLK7. These two proteases have an essential role in the epidermal desquamation, which is disturbed in Netherton-Syndrome, an inflammatory skin disease with disrupted skin barrier due to a LEKTI-1 gene defect that causes LEKTI-1 deficiency. This was proven in LEKTI-1 k.o.- mice, which show a phenotype resembling Netherton syndrome patients.To study whether both novel LEKTIs are of similar importance in desquamation, LEKTI-2- und later also LEKTI-3 knock-out mice as well as transgenic mice overexpressing these LEKTIs will be generated. In this cooperation project in my laboratory murine LEKTI-2- and -3 orthologs will be cloned, its tissue expression in mice analysed and then recombinantly expressed for analyses of antimicrobial and protease-inhibiting properties, in particular for inhibition of desquamation-relevant kallikreins. We will further generate mLEKTI-2 and -3-antibodies for immunohistochemistry and ELISAs for LEKTI-2- and 3- to analyse its role in desquamation of various mouse models.