Research for the medical treatment of rheumatoid arthritis by endogenous inhibitor of aggrecanase

Aggrecan是关节软骨承重不良和减震的主要原因。在退行性关节疾病中，如类风湿性关节炎(RA)和骨关节炎(OA)，细胞外基质的丢失会导致关节软骨的破坏。聚集素蛋白降解是导致细胞外基质分解的早期事件之一。据报道，基质金属蛋白酶是软骨退变的关键酶。最近的研究表明，Aggrecanase-1和Aggrecanase-2是ADAMTS(一种带有血栓反应蛋白基序的去整合素和金属蛋白酶)家族的成员，这些酶被认为在类风湿关节炎或骨性关节炎软骨聚集素的磨损中起着关键作用。基质金属蛋白酶组织抑制因子-3(TIMP-3)是一种有效的聚集聚糖酶-1和-2的抑制因子，但TIMP-3是否是唯一的聚集聚糖酶抑制因子尚不清楚。因此，在考虑降解性关节疾病的新药时，确定一种特定的内源性侵袭酶抑制剂至关重要。本研究的目的是鉴定Aggrecanase-1的内源抑制物，并研究Aggrecanase-1与其内源抑制物相互作用的机制。以人Aggrecanase-1的催化结构域为诱饵，进行酵母双杂交筛选，并将含有该诱饵的EGY48酵母菌株转化到人肝cDNA文库中。我们鉴定了血浆丝氨酸蛋白酶抑制剂中的α-1-抗胰凝乳素和α-1-抗胰凝乳素为猎物。

Aggrecan is greatly responsible for bad-bearing and shock-absorbing junction of the joint cartilage. In degradative articular diseases such as rheumatoid arthritis(RA) and osteoarthritis(OA), loss of extracellular matrices occurs resulting in the destruction of joint cartilage. Aggrecan proteolysis is one of the early events leading to the breakdown of the extracellular matrix. Matrix metalloproteinases have been reported to be the key enzymes in cartilage degeneration. Recent studies show that aggrecanase-1 and aggrecanase-2 are members of the ADAMTS(a disintegrin and metalloproteinase with thrombospondin motifs) family, and these enzymes are considered to play a pivotal role in the abrasion of cartilage aggrecan in RA or OA. Tissue inhibitor of matrix metalloproteinase-3(TIMP-3) is a potent inhibitor of aggrecanase-1 and -2, but whether TIMP-3 is the only inhibitor of aggrecanases remains unknown. The identification of a specific endogenous aggrecanase inhibitor therefore would be of critical importance in considering new medication for degradative joint diseases. The aims of the current study were to identify the endogenous inhibitor of aggrecanase-1 and to characterize the mechanism of interaction between aggrecanase-1 and its endogenous inhibitor We performed the yeast two-hybrid screen using the catalytic domain of human aggrecanase-1 as a bait, and an EGY48 yeast strain containing the bait plasmid was transformed with a human liver cDNA library plasmid. We identified alpha-1-antitrypsin and alpha-1-antichymotrypsin, members of plasma serine proteinase inhibitors, as preys.