Tissue Inhibitor Of Metalloproteinase-1 Production Research Articles

Cyclic adenosine 3‘,5‘-monophosphate suppresses interleukin 1-induced synthesis of matrix metalloproteinases but not of tissue inhibitor of metalloproteinases in human uterine cervical fibroblasts.

Interleukin 1 (IL-1) mediates many cellular functions, but the signal transduction mechanisms of its actions are not clearly understood. Here, we have examined the exact participation of cAMP in the IL-1-induced production of the precursors of matrix metalloproteinase (MMPs) and their specific inhibitor, tissue inhibitor of metalloproteinases (TIMP) in human uterine cervical fibroblasts. IL-1 significantly augmented the production of proMMP-1 (vertebrate procollagenase), proMMP-3 (prostromelysin), and TIMP without detectable changes in the intracellular level of cAMP. Dibutyryl cAMP (Bt2cAMP) and the cAMP elevating agent (forskolin) did not replace IL-1 as MMP inducers. On the contrary, the IL-1-mediated induction of proMMP-1 and proMMP-3 was significantly suppressed by treatment of the cells with Bt2cAMP, forskolin, or theophylline. The suppressive effect of Bt2cAMP on the IL-1-induced production of proMMP-1 and -3 was not due to the inhibition of zymogen secretion, but resulted from the decrease in the steady-state levels of proMMP-1 and proMMP-3 mRNAs. In contrast, Bt2cAMP slightly enhanced the IL-1-induced production of TIMP. The synthesis of proMMP-2 (72-kDa progelatinase/type IV procollagenase) was not altered by IL-1 and/or Bt2cAMP. These results suggest, first, that induction of proMMP-1 and -3 synthesis may share similar transduction pathways but they are distinct from those for proMMP-2 and TIMP synthesis and, second, that cAMP does not function as a second messenger in the MMPs' induction upon IL-1 stimulation in human uterine cervical fibroblasts. Thus, it is further suggested that the system that increases the intracellular cAMP level may be involved in negative regulation of proMMP-1 and -3 production.

Calmodulin differentially modulates the interleukin 1-induced biosynthesis of tissue inhibitor of metalloproteinases and matrix metalloproteinases in human uterine cervical fibroblasts

The effects of a specific calmodulin inhibitor, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7) on the synthesis of tissue inhibitor of metalloproteinases (TIMP) and precursor of matrix metalloproteinase 1/tissue collagenase (proMMP-1) and matrix metalloproteinase 3/stromelysin (proMMP-3), were examined using human uterine cervical fibroblasts in culture. When the cells were treated with human recombinant interleukin 1 alpha, the synthesis of TIMP, proMMP-1, and proMMP-3 was greatly enhanced along with the increase in the steady-state levels of mRNAs for respective proteins. The treatment of the cells with human recombinant interleukin 1 alpha and W-7 further augmented the production of proMMPs-1 and -3 and the accumulation of their mRNAs. In contrast, TIMP production and its steady-state mRNA level were reduced considerably under these conditions. Similar observations were made with another calmodulin inhibitor, trifluoperazine, but not with N-(6-aminohexyl)-1-naphthalenesulfonamide, the weakest inhibitor for calmodulin. These results indicate that calmodulin is required for the interleukin 1-enhanced synthesis of TIMP but it is a suppressor for the synthesis of proMMPs-1 and -3.

Fibroblast heterogeneity in collagenolytic response to cyclosporine.

To investigate the mechanism of cyclosporine (CS)-induced fibrotic gingival enlargement, the effect of CS on the collagenolytic activities of 14 different human gingival fibroblast strains derived from healthy individuals with non-inflammed gingiva was examined in vitro. There was marked heterogeneity among individuals in basal levels of collagenase activity, and there was also variation among the subpopulations derived from one strain. Fibroblasts from different individuals also varied markedly in their collagenolytic response to CS (0.1 to 0.75 micrograms/ml). In most strains, CS decreased collagenase activity, but in some, the drug caused no change or significantly increased activities. In most of the subpopulations CS significantly decreased collagenolytic activity. Two of the fibroblasts strains and the subpopulations described above were examined for the production of immunoreactive collagenase and tissue inhibitor of metalloproteinase (TIMP). The two strains made similar amounts of collagenase, but differed markedly in TIMP levels; CS affected their collagenase production differently but had similar effects on TIMP. Among the subpopulations there was variation in the production of collagenase, although none made detectable levels of TIMP; they also varied in the production of both proteins in response to CS. In two of the subpopulations and in both strains at some concentrations, the effect of CS on the relative levels of collagenase and TIMP could account for the decreased collagenase activity; i.e., the level of collagenase was unchanged or decreased, and TIMP production was unchanged or increased. This study demonstrates the variation among individuals as well as intrastrain heterogeneity of human gingival fibroblasts with regard to collagenase activity and the production of collagenase and TIMP. The heterogeneity of the collagenolytic response of different gingival fibroblast strains and their subpopulations to CS treatment may partly explain the susceptibility of only some individuals to CS-induced gingival enlargement.

Tissue inhibitor of metalloproteinases-1 (TIMP-1) RNA is expressed at elevated levels in malignant non-Hodgkin's lymphomas

Secreted metalloproteinases (MPs) and their specific inhibitors (TIMPs, tissue inhibitors of MPs) are important mediators of extracellular matrix metabolism. Previous studies have linked either excessive MP release or reduced TIMP-1 production to the invasive and metastatic phenotypes of cancer cells. In the present study we investigated the relationship between the expression of TIMP-1 and the clinical behavior of 28 non-Hodgkin's lymphomas. Northern blot analysis showed that levels of TIMP-1 mRNAs correlated directly with clinical aggressiveness: tumors in the high-grade category contained the highest levels of TIMP-1 transcripts approaching those found in maximally growth factor-stimulated fibroblasts in vitro. In situ hybridization localized the TIMP-1 expression to stromal cells of endothelial and fibroblastic origin. In contrast, transcripts hybridizing with metalloproteinase gene probes (interstitial collagenase and 72-Kd type IV collagenase) were expressed at very low levels in malignant lymphomas and their expression was not coordinately regulated with that of TIMP-1. The majority of tumors expressed either interstitial collagenase or 72-Kd type IV collagenase, and only a small number expressed both. Interstitial collagenase transcripts were only detected in high-grade tumors. The relative levels of TIMP-1 expression did not correlate with the degree of fibrosis of the tumors. Our data suggest the importance of tumor-stromal interactions in non-Hodgkin's lymphomas, and moreover, our results indicate a possible relationship between high-level, localized expression of TIMP-1 and the malignant phenotype of high-grade advanced-stage lymphomas.

Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) RNA Is Expressed at Elevated Levels in Malignant Non-Hodgkin's Lymphomas

Secreted metalloproteinases (MPs) and their specific inhibitors (TIMPs, tissue inhibitors of MPs) are important mediators of extracellular matrix metabolism. Previous studies have linked either excessive MP release or reduced TIMP-1 production to the invasive and metastatic phenotypes of cancer cells. In the present study we investigated the relationship between the expression of TIMP-1 and the clinical behavior of 28 non-Hodgkin's lymphomas. Northern blot analysis showed that levels of TIMP-1 mRNAs correlated directly with clinical aggressiveness: tumors in the high-grade category contained the highest levels of TIMP-1 transcripts approaching those found in maximally growth factor-stimulated fibroblasts in vitro. In situ hybridization localized the TIMP-1 expression to stromal cells of endothelial and fibroblastic origin. In contrast, transcripts hybridizing with metalloproteinase gene probes (interstitial collagenase and 72-Kd type IV collagenase) were expressed at very low levels in malignant lymphomas and their expression was not coordinately regulated with that of TIMP-1. The majority of tumors expressed either interstitial collagenase or 72-Kd type IV collagenase, and only a small number expressed both. Interstitial collagenase transcripts were only detected in high-grade tumors. The relative levels of TIMP-1 expression did not correlate with the degree of fibrosis of the tumors. Our data suggest the importance of tumor-stromal interactions in non-Hodgkin's lymphomas, and moreover, our results indicate a possible relationship between high-level, localized expression of TIMP-1 and the malignant phenotype of high-grade advanced-stage lymphomas.

Hormonal regulation of collagenolysis in uterine cervical fibroblasts. Modulation of synthesis of procollagenase, prostromelysin and tissue inhibitor of metalloproteinases (TIMP) by progesterone and oestradiol-17 beta.

Rabbit uterine cervical fibroblasts produced a large amount of matrix metalloproteinases (MMPs) such as collagenase (MMP-1) and stromelysin (MMP-3) and a small relatively amount of tissue inhibitor of metalloproteinases (TIMP). When cells were treated with progesterone or oestradiol-17 beta, both steroids concurrently decreased the level of procollagenase and prostromelysin in the culture media and the steady-state levels of the respective mRNAs. On the other hand, the level of TIMP in the culture media and the steady-state level of its mRNA were simultaneously increased by these steroids. Similarly, the suppression of production of MMPs and the augmentation of TIMP production by both steroids were observed with interleukin 1 (IL-1)-treated cells, but the action of progesterone was more effective than that of oestradiol-17 beta in the IL-1-untreated and -treated cells. These results suggest that collagenolysis in uterine cervical fibroblasts is negatively regulated by steroid hormones via the acceleration of TIMP production and the suppression of synthesis of MMPs at the pretranslational level.

Dexamethasone selectively modulates basal and lipopolysaccharide-induced metalloproteinase and tissue inhibitor of metalloproteinase production by human alveolar macrophages.

To define the capacity of glucocorticoids to regulate tissue damage associated with inflammation more clearly, we have studied the effects of dexamethasone on human alveolar macrophage secretion of both a variety of metalloproteinases and also the counter-regulatory tissue inhibitor of metalloproteinases (TIMP). We found that dexamethasone selectively and coordinately inhibited expression of the following human metalloproteinases: interstitial collagenase, stromelysin, and the 92-kDa type IV collagenase, as well as TIMP. Both basal and LPS-stimulated cells exhibited similar degrees of inhibition, with greater than 50% decrease in secretion of all enzymes and TIMP observed at dexamethasone concentrations of greater than or equal to 10(-8) M in serum-containing medium. The effects of dexamethasone were mediated at a pretranslational level. In summary, our results indicate that glucocorticoids suppress the matrix-degrading phenotype that is characteristic of mature human mononuclear phagocytes, and block the effects of the most potent known signal for upregulation of metalloproteinase secretion. Similar actions in vivo would serve to limit tissue damage associated with the inflammatory response.

Bone-derived growth factors modulate collagenase and TIMP (tissue inhibitor of metalloproteinases) activity and type I collagen degradation by mouse calvarial osteoblasts

The finding that osteoblasts synthesize collagenase has led to the hypothesis that bone cells play a major role in bone resorption by degrading the surface osteoid layer, thereby preparing the underlying mineralized bone for osteoclastic action. To further understand the mechanisms regulating osteoid removal, mouse calvarial osteoblasts were cultured on 14C-labelled type I collagen films and the abilities of (i) bovine bone matrix extracts and (ii) purified or recombinant human growth factors, to modify their collagenolytic behaviour were investigated. EDTA/Tris-HCl extracts of bone matrix containing growth factor activity, exerted a dose-dependent inhibition of type I collagenolysis by osteoblasts stimulated with 1,25-dihydroxyvitamin D 3 (1,25(OH) 2D 3, 10 ng/ml). Inhibition was accompanied by a reduction in collagenase activity and an increase in free TIMP (tissue inhibitor of metalloproteinases) in the culture medium. Transforming growth factor-β, epidermal growth factor, platelet-derived growth factor and the acidic and basic fibroblast growth factors all mimicked these effects. In contrast, insulin-like growth factors-I and-II did not inhibit type I collagenolysis, only partially inhibited collagenase activity, and did not stimulate TIMP production by either 1,25(OH) 2D 3-treated or untreated cells. These findings provide additional evidence for the tight control exerted on the proteolytic activity of osteoblasts and the importance of TIMP in its regulation. They suggest strongly that the conversion (coupling) of the initial resorptive phase of the bone remodelling cycle to one of deposition, may be mediated by polypeptide growth factors either produced rocally by osteoblasts, or released by proteolysis from the bone matrix.

Tumor necrosis factor bifunctionally regulates matrix metalloproteinases and tissue inhibitor of metalloproteinases (TIMP) production by human fibroblasts

The production of tissue inhibitor of metalloproteinases (TIMP) in human uterine cervical fibroblasts was increased by human recombinant tumor necrosis factor α (hrTNF) at a low concentration (0.005 ng ml ) but the elevated synthesis was suppressed in a dose-dependent manner at higher concentrations (up to 50 ng ml ). In contrast, the production of collagenase (EC 3.4.24.7) and stromelysin was stimulated at all the corresponding concentrations. In contrast, human recombinant interleukin-1α (hrIL-1, 10 ng ml ) coordinately induced these enzymes and TIMP production. The reduction of the elevated TIMP production by TNF was not due to the inhibition of TIMP secretion. These results suggest that TNF modulates the extracellular matrix degradation in human fibroblasts bifunctionally by the suppression of TIMP production in addition to the acceleration of matrix metalloproteinases production. Furthermore, the fact that TNF and IL-1 differently controlled the production of TIMP suggests that the signal pathway of TNF for TIMP production is different from that of IL-1.

Interleukin 6 enhances the production of tissue inhibitor of metalloproteinases (TIMP) but not that of matrix metalloproteinases by human fibroblats

We have investigated the effect of interleukin 6 (IL-6) on the synthesis of tissue inhibitor of metalloproteinases (TIMP) and matrix metalloproteinases (MMPs), collagenase (MMP-1) and stromelysin (MMP-3) using human skin and uterine cervical fibroblasts. IL-6 did not modulate the expression of MMPs by these fibroblasts, but the production of TIMP was enhanced by IL-6 in a dose dependent manner, whereas IL-1 stimulated the production of both MMPs and TIMP. The combination of IL-6 and IL-1 further augmented IL-1-induced MMPs and TIMP production. The results provide the first evidence that IL-6 participates in the catabolism of the extracellular matrix components by modulating the effects of IL-1 on MMPs and TIMP synthesis as well as its direct effects on the synthesis of TIMP by connective tissue cells.

Production of collagens, collagenase and collagenase inhibitor during the dedifferentiation of articular chondrocytes by serial subcultures

Rabbit articular chondrocytes were cultured in monolayer and the progressive loss of their differentiated phenotype was monitored from passage to passage. The cell densities achieved in confluent cultures decreased abruptly between the primoculture and the second or third subculture, and more slowly thereafter, reflecting parallel morphological changes. The synthesis of collagen (but not that of other proteins) decreased sharply, and a smaller proportion of collagen was incorporated into the matrix. Cells in primoculture synthesized mainly the cartilage-specific collagens, types II and XI, which were mostly deposited in the matrix, but no type I nor III collagen. With increasing passages, the synthesis of type II collagen decreased progressively while that of types I and III collagens increased, the latter being almost completely released in the culture medium. Simultaneously, the production of type XI collagen was apparently switched to that of type V. Fully differentiated confluent chondrocytes in primoculture produced the collagenase inhibitor TIMP (tissue inhibitor of metalloproteinases) but no detectable procollagenase; their production of procollagenase was, however, induced by interleukin 1. The production of TIMP increased from passage to passage. A spontaneous production of procollagenase was only occasionally observed in confluent cultures of dedifferentiated chondrocytes. However, interleukin 1 induced an always higher production of procollagenase from dedifferentiated chondrocytes than from cells in primoculture.

Sequence of human tissue inhibitor of metalloproteinases and its identity to erythroid-potentiating activity

Collagen fibres form the stable architecture of connective tissues and their breakdown is a key irreversible step in many pathological conditions. The destruction of collagen is usually initiated by proteinases, the best known of which is the metalloproteinase collagenase (EC 3.4.24). Collagenase and related metalloproteinases are regulated at the level of their synthesis and secretion, through the action of specific stimuli such as hormones and cytokines, and also at the level of their extracellular activity through the action of a specific inhibitor, TIMP (tissue inhibitor of metalloproteinases), which irreversibly forms inactive complexes with metalloproteinases. Although the mechanisms governing the production of TIMP are unknown, immunologically identical forms of this glycoprotein have been detected in a wide variety of human body fluids and cell and tissue culture media. We therefore suggested that under physiological conditions this ubiquitous inhibitor predominates over active metalloproteinases and that tissue destruction may arise when any perturbation of this controlling excess arises. However, further progress towards testing this theory has been hindered by a lack of knowledge about the structure of TIMP and insufficient material for studying it in model systems. Here we describe the structure of TIMP predicted from its complementary DNA, its synthesis in Escherichia coli and transfected animal cells, and the finding that it is identical to a protein recently reported to have erythroid-potentiating activity (EPA).

The effects of razoxane (ICRF 159) on the production of collagenase and inhibitor (TIMP) by stimulated rabbit articular chondrocytes

Monolayer cultures of rabbit chondrocytes were stimulated to produce collagenase with conditioned medium from human peripheral blood mononuclear cells (MCM), and the ability of Razoxane to modulate the production of collagenase and specific tissue inhibitor of metalloproteinases (TIMP) was studied. Collagenase production was inhibited and TIMP increased by Razoxane, in a dose-dependent manner, when cells were treated daily for 3 days. Over this period the effect of Razoxane was progressive; 50 μg/ml or less had no effect at day 1 but 50 μg/ml was effective by day 3. The effectiveness of Razoxane was inversely related to the degree of MCM stimulation and the confluency of the culture. On removal of the drug, chondrocytes stimulated with MCM recovered their ability to produce collagenase, and TIMP production returned to near normal. The results suggest that the ability of Razoxane to reduce collagenase and increase TIMP production may correlate with its effectiveness in treating psoriatic arthritis.