Latent Collagenase Research Articles

Regulation of matrix metalloproteinases is at the heart of myocardial remodeling

remodeling of the myocardium is a recognized risk factor contributing to the pathogenesis of cardiovascular disease, with the development of ventricular dilatation being a fundamental component of this structural remodeling (16). A recognized prerequisite in order for dilatation to occur is the degradation of the myocardial extracellular matrix (ECM) (2–4, 7, 9, 11, 15). The presence of an abundant latent collagenase system closely associated with the interstitial collagen matrix in the heart was first recognized by Montfort and Perez-Tamayo in 1975 (10), and since that time over 20 known endogenous matrix metalloproteinases (MMPs) have been identified, each having specificity for one or more ECM components (15). Although numerous studies have established that MMP activation mediates myocardial remodeling in cardiac pathologies, the endogenous control systems that regulate the induction of MMP expression and activation are still relatively poorly understood. Notwithstanding, remarkable progress has been made with respect to elucidating MMP biology and how it relates to myocardial matrix remodeling. In this vein, Zavadzkas et al. (19) reports that persistent myocardial extracellular MMP inducer (EMMPRIN) expression caused adverse myocardial remodeling consisting of left ventricular (LV) dilatation, hypertrophy, and fibrosis associated with increased levels of membrane type-1 (MT1)-MMP and active MMP-2 in aging mice. Thus this study is the first to establish mechanistic evidence that increased EMMPRIN levels can cause LV remodeling, resulting in heart failure. Accordingly, this study should inspire focused efforts seeking to elucidate the signaling pathways responsible for the induction of EMMPRIN in cardiovascular disease, as well as the mechanisms by which EMMPRIN induces MMP expression. Functional Role of EMMPRIN in the Heart EMMPRIN (also known as Basigen, collagenase stimulatory factor, and CD147) is a novel 58-kDa transmembrane glycoprotein of the Ig superfamily (1). EMMPRIN has been most extensively investigated with respect to tumor invasion and metastasis, having been implicated in those studies as a factor contributing to the induction of MMP expression (1, 6, 17). However, significantly increased myocardial EMMPRIN expression has also previously been reported in patients with end-stage cardiomyopathy (8, 14). Some studies have identified cardiac myocytes as the cellular source of this increase in EMMPRIN (13, 14). In the vasculature, EMMPRIN was colocalized with macrophage infiltrates in atherosclerotic intima and vascular smooth muscle cells from human carotid endarterectomy specimens (18). This study found that stimulation with the proinflammatory cytokines interleukin-1β and tumor necrosis factor-α increased the expression of both EMMPRIN and MMPs in bone marrow-derived human monocyte cultures (18). Thus the cytokine-mediated regulation of EMMPRIN expression may prove to be the underlying mechanism responsible for the induction of MMPs and ECM remodeling in the heart (5, 8). Further evidence that EMMPRIN is induced in myocardial inflammatory cells is derived from the association of EMMPRIN upregulation with the increased expression of MT1-MMP on monocytes isolated from patients following an acute myocardial infarction (12). Therefore, in the context of pathological myocardial remodeling, EMMPRIN upregulation in multiple cell types appears to be critical to the transcriptional regulation of MMPs. Selective Induction of Specific MMPs by EMMPRIN Based upon the findings reported by Zavadzkas et al. (19), together with these past observations, an important aspect of EMMPRIN biology that requires further clarification is the supposition that EMMPRIN induces a limited portfolio of specific MMPs within the myocardium. If this should prove to be true, it would allow the implementation of targeted drug interventions aimed at either the specific MMPs mediating adverse myocardial remodeling or the upstream signaling pathways responsible for their induction. This would represent an important advance beyond the currently available broad spectrum MMP inhibitory compounds that have thus far been studied. Hence several important issues deserving of future research emphasis remain with respect to the regulation of MMPs in myocardial remodeling. These include the targeted modulation of the induction of MMP expression by EMMPRIN and other comparable pathways, delineation of the individual role of specific MMP isoforms in mediating myocardial remodeling, and identification of endogenous mechanisms regulating the activation of MMPs. In summary, Zavadzkas et al. (19) found that the selective induction of specific MMPs concomitant with the chronic myocardial overexpression of EMMPRIN was sufficient to produce adverse LV remodeling and heart failure. Accordingly, identifying the signal transduction pathways mediating EMMPRIN expression in cardiovascular disease, as well as novel approaches by which to target EMMPRIN-induced MMP expression, constitute important avenues of further research.

Interaction of beta 1-anticollagenase from human plasma with collagenases from various tissues and competition with alpha 2-macroglobulin.

β1-Anticollagenase from human plasma forms an inactive enzyme-inhibitor complex with active human polymorphonuclear leukocyte collagenase [Macartney, H. W. and Tschesche, H. (1983) Eur. J. Biochem. 130, 85–92]. The inhibitor was shown to contain a reactive sulfhydryl group necessary for inhibition of the collagenase. Inhibition of the enzyme by β1-anticollagenase was shown to take place via a thiol/disulfide interchange reaction similar to the mechanism reported for the reaction between polymorphonuclear leukocyte collagenase and the polymorphonuclear collagenase inhibitor [Macartney, H. W. and Tschesche, H. (1980) FEBS Lett. 119, 327–332]. The observations made in this paper have shown that the collagenases obtained from other human tissues, such as rheumatoid synovial fluid, embryonic skin fibroblasts, diabetic skin fibroblasts and normal skin fibroblasts, all react with β1-anticollagenase via a thiol/disulfide interchange mechanism, and are inhibited in a 1:1 stoichiometric reaction. Reaction of the inhibitor with the collagenases can be blocked by alkylation of the inhibitor's free sulfhydryl group with iodoacetamide, or by reaction of the inhibitor with disulfide-containing compounds such as cystine, oxidised glutathione, insulin, relaxin or trypsinogen. The β1-anticollagenase–collagenase complexes (latent enzyme) can all be reactivated with disulfide-containing compounds, such as those mentioned above. An activation process has been reported for human latent polymorphonuclear leukocyte collagenase via a thiol/disulfide interchange as catalysed by the glutathione cycle in a peroxidase-coupled reaction to glucose metabolism [Tschesche, H. and Macartney, H. W. (1981) Eur. J. Biochem. 120, 183–190]. The same physiological activation process is capable of reactivating β1-anticollagenase–collagenase complexes in a reversible manner. It was further shown in competition experiments with β1-anticollagenase (and with human leukocyte collagenase inhibitor) and α2-macroglobulin that vertebrate collagenases were preferentially inhibited by the low-molecular-weight inhibitors. It could also be shown that there was no transfer of the enzyme to α2-macroglobulin from either β1-anticollagenase–collagenase, or leukocyte inhibitor–collagenase complexes indicating a true inhibitory function for the β1-anticollagenase and the leukocyte collagenase inhibitor.

The collagenase inhibitor from human polymorphonuclear leukocytes. Isolation, purification and characterisation.

A new collagenase inhibitor of latent human polymorphonuclear collagenase, initially reported by Macartney and Tschesche [Hoppe-Seyler's Z. Physiol. Chem. 361, 298–299 (1980) and FEBS Lett. 119, 327–332 (1980)], has been purified to apparent homogeneity. We described several methods of purification of this inhibitor, but the fastest and most reproducible method is by passage of purified latent human polymorphonuclear leukocyte collagenase on activated thiol-Sepharose 4-B (preceding paper in this journal). Active collagenase is eluted from the column and the collagenase inhibitor is retarded. Elution of the inhibitor is achieved by eluting the column with thiol compounds in the equilibration buffer. Further purification of the inhibitor is obtained by gel filtration on Sephacryl S-300 and ion-exchange chromatography on DEAE-Sephacel. The inhibitor exhibits a strict specificity for mammalian collagenases. It inhibits collagenases isolated from human synovial fluid, human normal and diabetic skin fibroblasts and human embryonic skin fibroblasts. The inactive complexes are all subject to activation by several disulfide compounds, such as oxidised glutathione by the thiol/disulfide interchange reaction already described. The purified inhibitor was shown by sodium dodecyl sulfate/polyacrylamide gel electrophoresis (10% gel) to be homogeneous and displayed an apparent molecular weight of 24500. Titration of the inhibitor with Ellmans reagent revealed that it contains one free sulfhydryl group, a prerequisite for its inhibitory activity. The inhibitor inhibits human polymorphonuclear leukocyte collagenase in a strict 1:1 stoichiometric reaction. The amino acid composition of the complex formed from the inhibitor and active enzyme reproduces that of the latent collagenase thus confirming that the latent enzyme is a 1:1 complex. The inhibitor is an acidic protein having an isoelectric point at pH 5.5, which would be expected from the amino acid analysis. The composition reveals a glycoprotein of about 240 residues containing a carbohydrate moiety composed of N-acetylglucosamine, mannose, galactose and glucose. The amino acid composition of the inhibitor is not identical to that of β1-anticollagenase from human plasma.

Characterisation of beta 1-anticollagenase from human plasma and its reaction with polymorphonuclear leukocyte collagenase by disulfide/thiol interchange.

A beta 1-serum component, beta 1-anticollagenase, capable of inhibiting various mammalian tissue collagenases, was isolated from human plasma by gel filtration, affinity chromatography and ion-exchange chromatography. The inhibitor contains 1-2 free sulfhydryl groups, which are a prerequiste for inhibitory activity and for binding to the thiol-Sepharose affinity support. Alkylation of beta 1-anticollagenase by iodoacetamide blocks inhibitory activity. The inhibitor was purified to apparent homogeneity and exhibited a Mr = 30500 determined by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The amino acid and carbohydrate composition was determined. According to its composition and the isoelectric focussing beta 1-anticollagenase is an acidic protein with an isoelectric point of 5.6. Inhibition of human leukocyte collagenase proceeds in a strong 1 : 1 stoichiometric reaction. The mechanism of this association takes place by a disulfide/thiol interchange reaction as has been previously indicated for human leukocyte collagenases in forming the latent enzyme [Macartney, H. W. and Tschesche, H. (1980) FEBS Lett. 119, 327-332]. The beta 1-anticollagenase--leukocyte-collagenase complex (latent enzyme) is activatable by disulfide-containing compounds such as cystine, oxidised glutathione, insulin, relaxin, trypsinogen and others, but not by 179,203-di(S-carboxymethyl)trypsinogen, or its trypsin derivative. Compounds containing inaccessible disulfide bonds, e.g. chymotrypsin, or sulfhydryl groups, e.g. D-penicillamine, do not activate the complex. Activation is, however, easily obtained with the oxidised-glutathione-generating system myeloperoxidase/H2O2/glutathione as was previously demonstrated for the human leukocyte latent collagenase activatable in a phagocytosis-simulated respiratory burst [Tschesche, H. and Macartney, H. W. (1981) Eur. J. Biochem. 120, 183-190].

Proteolytic Mechanisms in the Ovulatory Folliculo-Luteal Transformation

The connective tissue matrix of the wall of ovarian follicles is degraded and remodeled during ovulatory rupture and formation of the corpus luteum. Ovarian surface epithelial cells in close contact with the apical wall of preovulatory ovine follicles secrete a urokinase-type plasminogen activator in response to surge levels of gonadotropins. Urokinase activates latent collagenases and stimulates release of tumor necrosis factor &#102 from thecal endothelium. Tumor necrosis factor &#102 progressively induces matrix metalloproteinase gene expression, apoptosis, and inflammatory necrosis. Collagenolysis and cellular death are a prelude to stigma formation and ovarian rupture. Ovulation is blocked by intrafollicular injection of TNF &#102 or MMP-2 antisera. Unruptured follicles luteinize but are deficient in collagenous/vascularized trabeculae, and produce less progesterone than their control luteal counterparts. It appears that TNF &#102 , via MMP-2 induction, contributes to the reorganization of an ovulatory follicle into a fully competent corpus luteum.

Proteolytic Mechanisms in the Ovulatory Folliculo-Luteal Transformation

The connective tissue matrix of the wall of ovarian follicles is degraded and remodeled during ovulatory rupture and formation of the corpus luteum. Ovarian surface epithelial cells in close contact with the apical wall of preovulatory ovine follicles secrete a urokinase-type plasminogen activator in response to surge levels of gonadotropins. Urokinase activates latent collagenases and stimulates release of tumor necrosis factor f from thecal endothelium. Tumor necrosis factor f progressively induces matrix metalloproteinase gene expression, apoptosis, and inflammatory necrosis. Collagenolysis and cellular death are a prelude to stigma formation and ovarian rupture. Ovulation is blocked by intrafollicular injection of TNF f or MMP-2 antisera. Unruptured follicles luteinize but are deficient in collagenous/vascularized trabeculae, and produce less progesterone than their control luteal counterparts. It appears that TNF f , via MMP-2 induction, contributes to the reorganization of an ovulatory follicle into a fully competent corpus luteum.

Roles of the ovarian surface epithelium in ovulation and carcinogenesis.

Although ovarian mechanisms of ovulation have been a subject of investigation for more than a century, essential regulatory pathways remain uncertain. A role for the ovarian surface epithelium in ovulation has recently been demonstrated. Ovarian surface epithelial cells in close contact with the apical wall of preovulatory ovine follicles secrete a urokinase-type plasminogen activator in response to surge concentrations of (locally delivered) gonadotrophins. Urokinase activates latent collagenases and stimulates release of tumour necrosis factor alpha from thecal endothelium. Tumour necrosis factor alpha progressively induces matrix metalloproteinase gene expression, apoptosis and inflammatory necrosis. Collagenolysis and cellular death are a prelude to stigma formation and ovarian rupture. Epithelium exfoliated from the dome of ovulatory follicles is replenished by generative stem cell replication and migration from the wound edges. Common epithelial ovarian cancer has been related to successive bouts of ovulation and mitosis. The integrity of the DNA of surface cells circumjacent to the ovarian rupture site is compromised during the ovulatory process. Clonal expansion of an epithelial cell with damaged (unrepaired) DNA is a putative factor in carcinogenesis. Ovarian cancer is a deadly insidious disease because typically it is asymptomatic until the malignancy has reached beyond the ovaries.

Roles of the ovarian surface epithelium in ovulation and carcinogenesis

Although ovarian mechanisms of ovulation have been a subject of investigation for more than a century, essential regulatory pathways remain uncertain. A role for the ovarian surface epithelium in ovulation has recently been demonstrated. Ovarian surface epithelial cells in close contact with the apical wall of preovulatory ovine follicles secrete a urokinase-type plasminogen activator in response to surge concentrations of (locally delivered) gonadotrophins. Urokinase activates latent collagenases and stimulates release of tumour necrosis factor alpha from thecal endothelium. Tumour necrosis factor alpha progressively induces matrix metalloproteinase gene expression, apoptosis and inflammatory necrosis. Collagenolysis and cellular death are a prelude to stigma formation and ovarian rupture. Epithelium exfoliated from the dome of ovulatory follicles is replenished by generative stem cell replication and migration from the wound edges. Common epithelial ovarian cancer has been related to successive bouts of ovulation and mitosis. The integrity of the DNA of surface cells circumjacent to the ovarian rupture site is compromised during the ovulatory process. Clonal expansion of an epithelial cell with damaged (unrepaired) DNA is a putative factor in carcinogenesis. Ovarian cancer is a deadly insidious disease because typically it is asymptomatic until the malignancy has reached beyond the ovaries.

A fibronectin fragment induces type II collagen degradation by collagenase through an interleukin-1-mediated pathway.

To examine the effects of a fibronectin (FN) fragment containing the COOH-terminal heparin-binding domain (HBFN-f) on chondrocyte-mediated type II collagen (CII) cleavage by collagenase and proteoglycan (PG) degradation in articular cartilage in explant culture. Intact FN or HBFN-f was added to explant cultures of mature bovine articular cartilage. We investigated collagenase-mediated cleavage of CII caused by HBFN-f in explant cultures using a new immunoassay for detection and measurement of the primary collagenase cleavage site of CII. CII denaturation in cartilage was also measured using a specific enzyme-linked immunosorbent assay. Degradation of PG (principally aggrecan) was analyzed by a dye-binding assay. APMA and/or a matrix metalloproteinase 13 (MMP-13) preferential inhibitor or interleukin-1 receptor antagonist (IL-1Ra) were added to some cultures to examine the presence of latent procollagenase or the involvement of MMP-13 or IL-1, respectively, in cartilage breakdown induced by HBFN-f. Secretion of MMP-3 and MMP-13 into media was detected by immunoblotting. In contrast to intact FN, HBFN-f was shown to stimulate CII cleavage by collagenase in a dose-dependent manner following PG degradation, similar to cartilage breakdown induced by IL-1. Treatment with HBFN-f also resulted in elevated denaturation of CII. Immunoblotting demonstrated that HBFN-f enhanced pro-matrix metalloproteinase 13 (proMMP-13) production as well as that of proMMP-3. APMA, which activates latent proMMPs, enhanced the HBFN-f-mediated cleavage of CII by collagenase. An MMP-13 preferential inhibitor or IL-1Ra suppressed HBFN-f-induced collagen cleavage to control levels. Our data demonstrate that HBFN-f can induce early PG degradation and subsequent CII cleavage. The latter is probably mediated by early proMMP-13 induction involving an IL-1-dependent pathway. Activation of latent collagenase is delayed. This new information, together with existing data on other FN fragments, reveals that increased levels of these fragments, found in diseased joints such as in osteoarthritis and rheumatoid arthritis, may stimulate cartilage breakdown by mechanisms of the kind demonstrated in the present study.

Odontoblast function seen as the response of dentinal tissue to dental caries.

Microbes are responsible for the initiation and maintaining of carious processes. They have an efficient machinery for dissolving crystalline hydroxyapatite. When initiating carious processes, microbial acid formation determines the rate of the process in enamel. When the process reaches dentin, the micro-environment changes. Dential fluid in dentin tubules is the liquid where dissolving products of apatites are destroyed. Inorganic composition of dentinal fluid, however, is not altered much during the carious process, indicating that a functional secretory domain is working to pump the dissolved calcium and phosphate ions out of the fluid. Activation of odontoblast alkaline phosphatase and dentin latent collagenases is the known cellular event during the carious process in dentin. Because the caries lesion is by definition undermining, this suggests that, in this degradation process, the extracellular compartment, crystalline hydroxyapatite is dissolved by microbial acids, and a mixture of proteinases degrades the organic matrix. The degradation products of collagen and other matrix components in dentinal fluid must be transported either through the caries lesion in the enamel to saliva or through the odontoblast to the pulp (active transport). This facilitates further processing of the degradation products intracellularly during the passage through the cell.

Proteolytic and Cellular Death Mechanisms in Ovulatory Ovarian Rupture

Collagen breakdown and cellular death (apoptosis and inflammatory necrosis) within the apex of preovulatory ovine follicles are hallmarks of impending ovarian rupture. An integrative mechanism is presented whereby gonadotropic stimulation of urokinase-type plasminogen activator secretion by ovarian surface epithelial cells bordering the preovulatory follicle elicits a localized increase in tissue plasmin, which activates latent collagenases and secretion of tumor necrosis factor-α (TNF-α) from thecal endothelium. TNF-α potentiates collagenolysis (via matrix metalloproteinase gene expression) and (at elevated concentrations) mediates epithelial/vascular dissolution. Incidental damage to DNA of ovarian surface epithelial cells circumjacent to the ruptured follicle is a putative etiological factor in ovarian cancer.

Human mast cells produce matrix metalloproteinase 9.

Extracellular matrix-destructive enzymes, like matrix metalloproteinases (MMP), have been recognized in the process of inflammation and tissue remodeling and repair. The affected tissues often contain markedly increased numbers of mast cells. Although mast cells are capable of activating latent collagenase and proMMP, it has so far been unknown whether human mast cells themselves produce and secrete MMP9. In this study, MMP9 production by cord blood-derived cultured human mast cells and HMC-1 human mast cells was examined by reverse-transcriptase PCR, gelatin zymography and Western blot analysis using an antibody against MMP9. Cultured mast cells and HMC-1 cells treated with phorbol 12-myristate 13-acetate were shown to express MMP9 mRNA, and the cultured conditioned media from these cells showed gelatinolytic activity, identical with MMP9. Immunohistochemical examination was performed to detect MMP9 in tissue mast cells; mast cells localized in the skin, lung and synovial tissue showed strongly positive reactions for MMP9. Thus, these findings indicate that human mast cells can produce MMP9, which might contribute to extracellular matrix degradation and absorption in the process of allergic and nonallergic responses.

Interstitial collagenase (MMP-1) activity in human ovarian tissue

The collagen content and collagenase activity were estimated in human ovarian interstitial tissue devoid of all visible follicles in menstruating, fertile as well as climacteric women. The mean total collagenase activity in ovarian specimens taken during both follicular (n = 10, 3.97 ± 0.58 U/g wet weight, ww) and luteal phase (n = 10, 3.39 ± 1.24 U/g ww) of the normal menstrual cycle along with total collagen concentration (184.8 ± 41.0 vs. 194.4 ± 30.5 μg/mg ww, respectively) did not differ. Total collagenase activity of climacteric gonads (n = 5, 1.55 ± 0.71 U/g ww) was lower than in specimens collected during both follicular and luteal phase (p = 0.0002 and p = 0.017, respectively). About 23% of the total collagenase activity in follicular phase ovarian extracts and only about 1% in luteal phase ovarian preparations was found in the latent form. The percentage of latent collagenase in ovarian tissue during the follicular phase was negatively correlated with the day of the menstrual cycle (r = -0.93, p = 0.007). Extracellular matrix remodelling in the human ovary can be correlated with the functional status of the follicular unit.

Mast cell collagenase correlates with regression of pulmonary vascular remodeling in the rat.

Pulmonary vascular remodeling, produced by cell hypertrophy and extracellular matrix protein synthesis in response to hemodynamic stress, regresses after reduction of blood pressure, possibly by proteolysis of structural proteins. To test this postulate, we assessed the breakdown of extracellular matrix proteins and expression of collagenase and elastase in pulmonary arteries of rats exposed to hypoxia (10% O2 for 10 d) followed by normoxia. During hypoxia, contents of collagen and elastin increased in pulmonary arteries and latent rat interstitial collagenase was expressed without increased collagenolytic activity or mRNA levels. At 3 days after normoxia, collagen and elastin contents decreased coincident with the new appearance of activated collagenase and transient increases in collagenolytic and elastolytic activities. The amount of immunoreactive collagenase, localized predominately in connective tissue-type mast cells, was increased in the adventitia and media of hypertensive vessels. We conclude that mast cells containing latent collagenase are recruited into the outer walls of pulmonary arteries during remodeling. It is possible that mast cell-derived collagenase contributes to collagen breakdown in pulmonary arteries during early recovery from hypoxia and plays a role in restoration of vascular architecture.

Human Collagenase-3 Is Expressed in Malignant Squamous Epithelium of the Skin

Co-expression of several members of the matrix metalloproteinase (MMP) family is a characteristic of human carcinomas. To investigate the role of the recently cloned collagenase-3 (MMP-13) in epidermal tumors, we studied samples representing malignant (basal and squamous cell carcinoma, Paget's disease), pre-malignant (Bowen's disease, solar keratosis), and benign (keratoacanthoma, seborrheic keratosis, linear epidermal nevus) tumors. Basal cell carcinomas expressed collagenase-3 mRNA in focal areas of keratinized cells, the squamous differentiation of which was confirmed by positive immunostaining for involucrin. Apoptosis was observed in central parts of these foci. In squamous cell carcinomas, collagenase-3 expression was detected at the epithelial tumor front and less frequently in the surrounding stromal cells. Collagenase-3 mRNA co-localized with immunostaining for laminin-5, an adhesion molecule suggested to participate in the migration of tumor cells. The pre-malignant and benign tumors were mostly negative for collagenase-3. Stromelysin-1, a potential activator of latent collagenases, was frequently expressed by stromal cells surrounding the malignant tumors, and the two MMPs occasionally co-localized in keratotic foci. Our results demonstrate that in basal cell carcinomas, expression of collagenase-3 is associated with terminal differentiation of epithelial cells. Furthermore, the gene is activated during skin carcinogenesis, and we suggest a role for collagenase-3 in degradation of the extracellular matrix associated with malignant epithelial growth.

Modulation by hypergravity of extracellular matrix macromolecules in in vitro human dermal fibroblasts

In vitro human dermal fibroblasts were submitted to normal gravity (1 g) or to chronic hypergravity (20 g) over a period of 8 days. Changes in organization of extracellular matrix molecules were seen by indirect immunofluorescence. In the fibronectin layer, bundles of fibrils were gathered together leading to a disorganisation of the normal parallel pattern of fibers seen in control cultures. Type I collagen fibrils appeared with wooly outlines in controls whereas thick fibers were closely packed in 20-g cultures. A moderate increase of type III collagen fibril density was observed. No elastic fibers were seen in control or in 20-g cultures. In the culture medium, the release of soluble elastin (ELISA) and type I and III collagens (RIA) was undisturbed. Assays of enzymes involved in the remodeling of extracellular matrix showed an increase of cellular elastase activity (10%) and a decrease of the spontaneously active collagenase. Nevertheless, the total collagenase activity, (activated by trypsin), was increased by up to 30%. These data show a significant rise of the latent collagenase activity and suggest that release of the tissue inhibitor of metalloproteinase (TIMP1) was enhanced by hypergravity.

Characterization of interstitial collagenases in jaw cyst wall.

Neutral salt extracts of 14 specimens of jaw cysts were prepared. Histopathological analysis showed that the specimens consisted of 6 radicular cysts, 6 dentigerous cysts, 1 residual cyst, and 1 odontogenic keratocyst. One periapical granuloma, 1 dental follicle and a sample of clinically healthy oral mucosa were similarly processed and used as controls. Measurement of collagenase activity by monitoring the formation of specific degradation products of type I and II collagen in solution by SDS-PAGE demonstrated that all the cyst extracts contained collagenase, some of which was endogenously activated. Cyst wall collagenase preferably degraded type I over type II collagen, which suggests that the degradation was due to MMP-1 (matrix metalloproteinase-1) rather than the MMP-8 type. This was further supported by the doxycycline-inhibition profile of cyst collagenase, which was similar to that of MMP-1. Part of the cyst wall collagenase was in latent proenzyme form and probably derived, at least in part, from the newly synthesized intracellular collagenase pool. Latent cyst collagenase was efficiently activated with phenylmercuric chloride and to a lesser extent by gold (I) thioglucose and NaOCl. Western-blotting, using specific antibodies against collagenase from human polymorphonuclear neutrophilic leukocytes (MMP-8) and from fibroblasts (MMP-1), revealed a typical 55/45 kDa doublet; also MMP-8 in the latent 80 kDa form and fragmented to 65 kDa active species were found. These results suggest the presence of MMP-1 and, to a lesser extent, MMP-8 type collagenase in the cyst wall.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of collagen degradation in the rat myocardium after infarction

Fibrillar collagens, essential for maintaining the structural integrity of the myocardium, are degraded by matrix metalloproteinase (MMP-1). In other tissues collagenolysis is an important component of wound healing. Here we examined collagen degradation in the myocardium after infarction. Collagenase activity, measured by zymography, and expression of matrix metalloproteinase (MMP-1) and tissue inhibitor of metalloproteinase (TIMP) mRNA, detected by Northern blotting and in situ hybridization, in the rat heart 6 h to 28 days after left coronary artery ligation were studied. Sham-operated rats served as controls. Infarcted left ventricle was compared to non-infarcted right ventricle and interventricular septum and to sham-operated tissues. We found a transient increase in collagenase activity in the infarcted left ventricle, which began at day 2 (4.5-fold increase compared to controls), peaked at day seven (6.5-fold increase) and declined thereafter, together with a concomitant increase and contribution in collagenolytic activity of gelatinases (MMP-2 and MMP-9). An increase in collagenase mRNA was not seen until day 7 and only in the infarcted ventricle, while changes in MMP-1 activity or mRNA expression were not observed at remote sites or in sham-operated controls. Transcription of TIMP mRNA was observed at 6 h (two-fold increase) in the infarcted ventricle, peaked on day two after MI (eight-fold increase) and slowly decreased thereafter. No change in TIMP mRNA expression was observed at remote sites or in sham-operated controls. Cells responsible for transcription of MMP-1 and TIMP mRNA were fibroblast-like cells, not inflammatory or endothelial cells. At the site of infarction post-translational activation of latent collagenase (MMP-1) plays a greater role in the wound healing response than transcription of collagenase mRNA. Collagenase mRNA is synthesized when the latent extracellular pool of MMP-1 is reduced through the activation of latent collagenases and gelatinases. TIMP mRNA synthesis is regulated by the activation of MMPs with the balance between collagenase activation and TIMP inhibition determining the amount of collagenolysis in infarcted tissue.

High Free and Latent Collagenase Activity in Psoriatic Arthritis Synorial Fluid

Journal Article High Free and Latent Collagenase Activity in Psoriatic Arthritis Synorial Fluid Get access G. PARTSCH, G. PARTSCH Ludwig Boltzmann Institute of RheumatologyVienna Oberlaa, Austria Search for other works by this author on: Oxford Academic PubMed Google Scholar J.S. SMOLEN J.S. SMOLEN Ludwig Boltzmann Institute of RheumatologyVienna Oberlaa, Austria Search for other works by this author on: Oxford Academic PubMed Google Scholar Rheumatology, Volume 34, Issue 1, January 1995, Pages 83–84, https://doi.org/10.1093/rheumatology/34.1.83-a Published: 01 January 1995 Article history Accepted: 06 October 1994 Published: 01 January 1995

High Free and Latent Collagenase Activity in Psoriatic Arthritis Synorial Fluid

High Free and Latent Collagenase Activity in Psoriatic Arthritis Synorial Fluid