Method for Analysis of Matrix Degradation by CCN2 Through the MMP/TIMP System
Many studies have shown effects of members of the CCN family on matrix synthesis and accumulation but few have examined degradative pathways. This scarcity of information is in part due to the lack of suitable model systems. Here we describe methods for making rhCCN2 and also for the preparation of a biosynthetically labeled matrix substrate that can be used to measure the effect of CCN on cellular or secreted degradative pathways.
Key wordsCCN2 Connective tissue growth factor Matrix metalloproteinase Tissue inhibitor of matrix metalloproteinase Matrix degradation pathways
The work described in this chapter was supported by NH&MRC and Rebecca L Cooper Medical Research Foundation grants, to Professor Stephen Twigg and Associate Professor Susan McLennan.
- 4.Kim HS, Nagalla SR, Oh Y et al (1997) Identification of a family of low-affinity insulin-like growth factor binding proteins (IGFBPs): characterization of connective tissue growth factor as a member of the IGFBP superfamily. Proc Natl Acad Sci U S A 94:12981–12986CrossRefPubMedPubMedCentralGoogle Scholar
- 9.Yang DH, Kim HS, Wilson EM et al (1998) Identification of glycosylated 38-kDa connective tissue growth factor (IGFBP-related protein 2) and proteolytic fragments in human biological fluids, and up-regulation of IGFBP-rP2 expression by TGF-β in Hs578T human breast cancer cells. J Clin Endocrinol Metab 83:2593–2596PubMedGoogle Scholar
- 12.Min D, Lyons JG, Jia J et al (2006) 2-Methoxy-2,4-diphenyl-3(2H)-furanone-labeled gelatin zymography and reverse zymography: a rapid real-time method for quantification of matrix metalloproteinases-2 and -9 and tissue inhibitors of metalloproteinases. Electrophoresis 27(2):357–364CrossRefPubMedGoogle Scholar