Cloning and characterization of a cDNA encoding the baboon tissue inhibitor of matrix metalloproteinase-1 (TIMP-1)

Abstract

A baboon aortic smooth muscle cell (SMC) cDNA library was screened for the presence of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) by polymerase chain reaction (PCR); oligodeoxyribonucleotide primers corresponding to the coding frame of the known human TIMP-I gene were used as primers. Sequencing of the PCR-amplified baboon cDNA demonstrated only eight single-nucleotide (nt) mismatches, when compared with the coding frame of human TIMP-1. The authenticity of the PCR-amplified TIMP-I cDNA was further confirmed by clonal screening of the library with the PCR probe and sequencing of positive clones. On Northern blots from cultured baboon SMC, the baboon cDNA hybridized to a TIMP-l-specific mRNA of 800 bp. Phorbol ester (PMA) treatment of cultured baboon SMC produced a 2.5-fold increase in TIMP-1 transcript. TIMP-1 transcripts were also demonstrated in cultures of endothelial cells and fibroblasts obtained from baboon arteries. Immunohistochemical analysis demonstrated that TIMP-1 protein is localized to the adventitial layer of baboon artery. We conclude that TIMP-1 is a conserved molecule across species and localized to the tunica adventitia of baboon vessels.