Affordable Access

Publisher Website

Direct extraction and estimation of collagenase(s) activity by zymography in microquantities of rat myocardium and uterus

Authors
Publisher
Elsevier Inc.
Publication Date
Volume
26
Issue
3
Identifiers
DOI: 10.1016/0009-9120(93)90025-2
Keywords
  • Collagenase
  • Zymography
  • Rat Heart
  • Rat Uterus
  • Gelatinase
  • Metallproteinase
Disciplines
  • Biology
  • Chemistry

Abstract

Abstract Measurement of collagenolytic activity is of interest to a wide variety of investigators using mammalian tissue. In order to develop a method that would quantitative active collagenase from microquantities of human tissue, we employed zymography to the heart and uterus of neontal, adult, and postpartum rats. Collagenase rapidly cleaves native collagen into two fragments, which at 37°C form gelatin. Gelatin can also by hydrolyzed by collagenase, but at slower rate, and therefore we used gelatin to quantitate the amount of collagenase present in heart and uterine tissue and developed a method for the direct extraction of collagenase from small quantities of rat myocardium. Our method was found to be comparable with the chemical method reported by Masue et al. (Anal Biochem 1977; 17:215-21). The enzyme, which was not detected in normal adult rat cardiac tissue, was found to exist entirely in latent form and demonstrated typical properties of a mammalian collagenase/ gelatinase after activation by trypsin and plasma. We observed a 60–80% increase in collagenase activity after activation by these proteases and estimated that there is approximately 5 ± 2 pg of procollagenase per μg of normal adult rat left ventricle. Collagenolytic activity in the postpartum rat heart was found to be slightly (∼2–5%) reduced when compared to the adult heart but it was increased in the neonatal heart and postpartum uterus. This method allows for the rapid quantitative and qualitative measurement of collagenase activity in a variety of tissues containing collagenase/gelatinase activity. Our results indicate the most collagenase in the myocardium exists in latent form.

There are no comments yet on this publication. Be the first to share your thoughts.