사람 관절 연골의 세포외기질 발현 측정을 위한 cDNA chip 제작

초록

The extracellular matrix (ECM) is the substance which provides mechanical supprot and physical strenght to tissue, organs and the organism as a whole. Recently, ECM is known to exert profound influences on the behavior of cells also. Articular cartilage is composed of a hydrated extensive ECM which is synthesized, assembled and regulated by chondrocytes. The balance between ECM formation and catabolism is thought to paly an important role in the maintenance of normal articular cartilage. Osteoarthritis, which is characterized by destruction of artiuclar cartilage, has been viewed as a failure of the normal balance of ECM metabolism. However, most studies have focused on the aggrecan and major collagen, and little is known about other ECMs. In this study, we manufactured an ECM-related cDNA array to analyze multiple ECMs and related enzumes simultaneously, and analyzed several biochemical and biomechanical effects of the ECMs metabolism of artiuclar chondoryctes. Total RNA was isolate from PCR primers, cloned in pGEM T-Easy Vector. The PCR primer desined wiht primer selectionbased on mRNA sequence and were pruchased from Bioneer. Cloned cDNAs were spotted on CMT-CAPS Coated slides. Total RNA was prepared from human articular chodnrocytes at various treatments using Trizol reagent. Two color fluorescence (Cy-3, Cy-5) labeled probes were prepared by RT-PCR, were hybridized to the manufactured cDNA array overnight. The hybridized cDNA array was scanned and analyzed. The signals are normalized againest the house keeping gene, GAPDH. There are difference expression of ECM between control and treatment. From theses results, we can confirm that our manufactured ECM=related cDNA array provides an excellent method for studying patterns of ECM gene expression in human articular cartilage.