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Regulation of rat interstitial collagenase gene expression in growth cartilage and chondrocytes by vitamin D 3 , interleukin‐1β, and okadaic acid
Author(s) -
Grumbles R.M.,
Shao L.,
Jeffrey J.J.,
Howell D.S.
Publication year - 1996
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/(sici)1097-4644(19961215)63:4<395::aid-jcb2>3.0.co;2-o
Subject(s) - collagenase , chondrocyte , endocrinology , interstitial collagenase , medicine , microbiology and biotechnology , protein kinase c , staurosporine , chemistry , messenger rna , cartilage , gene expression , biology , kinase , biochemistry , enzyme , gene , anatomy
The interstitial collagenase produced by the rat growth plate chondrocytes is the homologue of the human collagenase‐3, or matrix metalloproteinase‐13. This enzyme is responsible for the loss of collagen during hypertrophy of chondrocytes and for the degradation of transverse septa in long bone growth. Rachitic rats (42 days, male Sprague‐Dawley) had an 8‐fold higher level of collagenase mRNA in the hypertrophic versus proliferative zone of growth plate cartilage. Intramuscular injection of 1,25‐dihydroxyvitamin D 3 (1,25‐(OH) 2 D 3 ; 1.0 μg/kg body weight) in rachitic rats increased collagenase mRNA another 1.5‐fold in the hypertrophic zone. The regulation of collagenase gene by 1,25‐(OH) 2 D 3 and interleukin (IL)‐1β in cultured proliferative chondrocytes was studied by means of steady‐state mRNA and half‐life determination of mRNA using the transcriptional inhibitor actinomycin D, and nuclear run‐on transcription analyses. Treatment of cells with 1,25‐(OH) 2 D 3 (10 ‐6 M) and IL‐1β (2 ng/ml) increased collagenase mRNA 8‐ and 13‐fold, respectively. Additionally, the collagenase mRNA half‐life was increased by 1,25‐(OH) 2 D 3 and IL‐1β. In the presence of a protein kinase C inhibitor, staurosporine, 1,25‐(OH) 2 D 3 induction of collagenase mRNA was blocked. Here the addition of phorbol 12‐myrisate 13‐acetate (PMA) to activate protein kinase C increased collagenase mRNA 10‐fold. However, in the presence of staurosporine (50 nM), PMA induction was blocked, whereas IL‐1β was not. IL‐1β is known to activate several phosphorylation pathways. Okadaic acid (500 nM), a protein phosphatase inhibitor, increased the relative collagenase mRNA abundance 10‐fold. The rate of the rat collagenase gene transcription in nuclei was increased with 1,25‐(OH) 2 D 3 , IL‐1β and okadaic acid. In separate experiments, the collagenase promoter was ligated to a reporter plasmid and the plasmid was transfected into chondrocytes. The results showed that 1,25‐(OH) 2 D 3 , IL‐1β, and PMA increased reporter activity 2.5‐, 2.8‐, and 3.27‐fold, respectively. Thus, there are multiple nuclear and cytoplasmic mechanisms by which cartilage modulators regulate rat interstitial collagenase gene expression. © 1996 Wiley‐Liss, Inc.