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Results for “Matrix Metalloproteinases, Secreted”

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PubMed1996

Increased matrix metalloproteinase-9 secretion in short-term tissue cultures of prostatic tumor cells.

We have examined the expression of 2 tumor-associated metalloproteinases, MMP-2 and MMP-9, in 48 primary cultures of prostatic carcinoma (PRCA) and 33 cultures of benign prostatic hyperplasia (BPH). PRCA cultures secrete significantly more MMP-9 than their benign counterparts. Secreted MMP-2 did not differ significantly in cultures but was lower in PRCA cultures. Two cultures of benign origin exhibited high MMP-9 secretion and growth patterns consistent with a malignancy. Both cases were followed and successively re-evaluated histologically and rediagnosed as organ-confined PRCA. MMP expression in culture may be of predictive value in the identification of incidental PRCA. MMP-9 secretion and its ratio with MMP-2 were highest in epithelial cultures from invasive, metastatic tumors when compared both to disease confined to prostate gland and to locally extensive disease. MMP-9 secretion was greatest also in cultures derived from tissues of high Gleason histological grade. Active MMP-9 species were detected in 15 cultures (31%) of PRCA. Active MMP-2 species were observed in cultures of both BPH and PRCA origin in almost the same amounts. Although average levels were not significantly different, as a ratio to proform species, a significant elevation was observed in cultures of PRCA origin. We propose, therefore, that an elevated expression of MMP-9 and a high ratio of MMP-9 to MMP-2 in short-term prostate epithelial cultures is of potential diagnostic and prognostic significance.

Diagnosis, Differential↗

PubMed2000

Differential regulation of the secretions of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinases-1 from human keratinocytes in culture.

To investigate the pathophysiological roles of matrix metalloproteinase (MMP)-9 and tissue inhibitor of metalloproteinases (TIMP)-1 in the differentiation of keratinocytes, we used human keratinocytes in culture to examine the gelatinolytic activity of MMP-2 and MMP-9 by zymographic analysis, MMP-9 mRNA by reverse transcriptase-polymerase chain reaction, and TIMP-1 mRNA by Northern blot analysis. Adding of calcium (1 mM) to the culture medium increased the activity of MMP-9 and MMP-9 mRNA without affecting MMP-2 activity or the expression of TIMP-1 mRNA. These results suggest that keratinocytes regulate the expressions of MMP-9 and TIMP-1 differently during the process of differentiation.

Base Sequence↗

PubMed2003

Latanoprost stimulates secretion of matrix metalloproteinases in tenon fibroblasts both in vitro and in vivo.

PURPOSE: To investigate the presence and the possible role of different matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in Tenon capsule fibroblasts. These enzymes are essential for the control of tissue remodeling in the context of wound repair. This aspect is important to further the understanding of and possibly to influence the scarring process of filtering blebs after glaucoma surgery. METHODS: Untreated and latanoprost-treated human Tenon fibroblasts were examined for the presence of MMPs and TIMPs on the mRNA and protein levels. Assays performed included RT-PCR, real-time RT-PCR, immunocytochemistry, Western blot analysis, flow cytometry, and zymography. To investigate the changes in vivo, conjunctival specimens of rabbits treated with latanoprost eye drops were examined by immunohistochemistry. RESULTS: In all assays, both MMP-3 and TIMP-2 were detected. With the real-time RT-PCR technique, MMP-1, -2, -3, -7, -9, and -14 and TIMP-1 and -2 were detected. An upregulation of MMP-3 and TIMP-2 after latanoprost treatment of the fibroblasts was shown and found to occur on the mRNA and the protein levels. The upregulation of MMP-3 and TIMP-2 was confirmed in vivo. CONCLUSIONS: Tenon fibroblasts contain the ability on the mRNA level to synthesize all enzymes of the MMP and TIMP family that are related to remodeling of the extracellular matrix. The levels of MMP-3 and TIMP-2 increase after treatment with latanoprost. Tenon fibroblasts may be the target cells for attempts to influence the tissue levels of MMPs and TIMPs in the context of conjunctival wound healing after glaucoma surgery.

Animals↗

PubMed1993

Secretion of matrix metalloproteinases by human endometrial cells in vitro.

At each menstrual cycle, the uterine endometrium undergoes intense remodelling. Of the many factors implicated in tissue remodelling, the matrix metalloproteinases (MMPs) play a central role owing to their capacity to degrade the extracellular matrix. The aim of this study was to examine the nature and cellular origin of endometrial proteases as evaluated in culture systems with clearly characterized cell types. Endometrial cells from hysterectomy specimens were prepared using collagenase digestion. Bone-marrow-derived cells (a known source of proteases) were removed by immunopurification. Cells were cultured on different substrates (matrigel, agarose, glass or plastic). Purity of cell preparations was examined by immunocytochemistry, and proteases were characterized by zymography on SDS-PAGE containing gelatin. The cell phenotype in culture was largely influenced by the type of substrate. Gelatin-degrading enzymes detected in culture supernatants of stromal and epithelial cells had molecular masses ranging from 42 to 248 kDa, and were identified as metalloproteinases. We conclude that human endometrial stromal and epithelial cells express several matrix metalloproteinases, the expression of which clearly depended on the purity of cell preparation, on cell adhesion and on the nature of the substrate on which the cells grew. These enzymes might be involved in endometrium remodelling, blastocyst implantation and trophoblast invasion.

Cells, Cultured↗

PubMed1992

Secretion of matrix metalloproteinases in cultured stromal cells isolated from a giant cell tumor of bone.

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Bone Neoplasms↗

PubMed2005

[Effect of hypoxia on production and secretion of matrix metalloproteinases in tumor cells].

BACKGROUND & OBJECTIVE: The malignant tumor tissues usually suffer from hypoxia due to its fast growth, which increases its invasive ability. This study was to explore changes of matrix metalloprotei-nases (MMPs) in tumor cells under hypoxia in vitro. METHODS: Zymography was used to determine the changes of activities of MMP-2 and MMP-9 in pancreatic cancer cell line SW1990 cultured under hypoxia. Western blot, and reverse transcription-polymerase chain reaction (RT-PCR) were used to detect protein and mRNA levels of MMP-2 and MMP-9. RESULTS: Under normal circumstances, activities of MMPs in SW1990 cells were relatively low. When SW1990 cells were cultured under hypoxia, the activity of MMP-9 increased by about 5.5 times that in normal circumstances, while the activity of MMP-2 didn't change. The same changes in protein level of MMP-9 had been confirmed by Western blot, while protein level of MMP-2 didn't change. mRNA level of MMP-9 was higher in cells cultured under hypoxia for 12 h than in control cells and CoCl(2)-cultured tumor cells by 46.1% and 56.4%, but no change of MMP-9 was seen when hypoxia inducible factor-1alpha (HIF-1alpha) was produced in SW1990 cells stimulated by CoCl(2). CONCLUSIONS: Production of MMP-9 in SW1990 cells is enhanced significantly under hypoxia, and its activity altered correspondingly; but those of MMP-2 had no change. This alteration has no relation with enhancement of HIF-1alpha under hypoxia.

Cell Hypoxia↗

PubMed2001

A conserved sequence within the propeptide domain of membrane type 1 matrix metalloproteinase is critical for function as an intramolecular chaperone.

The propeptide domain of secreted matrix metalloproteinases (MMPs) is responsible for maintaining the latency of these proteinases. Recently, the propeptide domain of the prototype membrane type matrix metalloproteinase (MT1-MMP) was demonstrated to act as an intramolecular chaperone (Cao, J., Hymowitz, M., Conner, C., Bahou, W. F., and Zucker, S. (2000) J. Biol. Chem. 275, 29648-29653). In the current study, the role of an unique four-amino acid sequence in the propeptide domain of MT1-MMP was examined. The sequence (42)YGYL(45) is conserved in the propeptide domain of all six members of the MT-MMP subfamily, but not in secreted MMPs. Mutant MT1-MMP cDNAs coding for alanine substitutions (single and double amino acid sequences) in this conserved propeptide region were transfected into COS-1 cells deficient in endogenous MT1-MMP. As demonstrated by immunofluorescence, mutant MT1-MMP protein was synthesized and displayed on the plasma membrane of transfected cells. Alanine substitutions within the (42)YGYL(45) sequence proved to be detrimental for enzyme function in terms of activation of proMMP-2 and binding TIMP-2 to the cell surface (MT1-MMP serves as a cell surface receptor for TIMP-2). In contrast to wild-type MT1-MMP-transfected cells, mutant MT1-MMP-transfected cells were incapable of degrading and migrating on a fibronectin substrate. These data indicate that the conserved (42)YGYL(45) sequence within the propeptide domain of MT-MMPs is required for intramolecular chaperone function of these intrinsic membrane proteinases.

Alanine↗

PubMed2000

Wild-type p53 gene transfer inhibits invasion and reduces matrix metalloproteinase-2 levels in p53-mutated human melanoma cells.

The tumor suppressor gene p53 has inhibitory effects on cell growth and angiogenesis and induces apoptosis when overexpressed in melanoma and in a variety of tumor cells by adenovirus-mediated gene transfer. The invasive ability of tumor cells, facilitating local infiltration and metastasis, is related to matrix metalloproteinase levels. In melanoma, matrix metalloproteinase-2 and matrix metalloproteinase-9 have a prominent role in this process. The aim of this study was to evaluate whether wild-type p53 overexpression, obtained by a recombinant adenovirus vector (AdCMV.p53), affects cell invasiveness through modulation of matrix metalloproteinase-2 and matrix metalloproteinase-9. Two human melanoma cell lines were used in this study: the SK-MEL-110, carrying a mutated p53 gene, and the SK-MEL-147, carrying the wild-type p53 gene. SK-MEL-110 cells infected with AdCMV.p53 exhibited decreased invasion capability from day 1 after infection, compared with cells not infected or infected with the control vector AdCMV.Null. This reduced invasiveness was associated with decreased matrix metalloproteinase-2 levels in conditioned media whereas no changes were detected in matrix metalloproteinase-9 secreted levels. No modulation in matrix metalloproteinase-2 mRNA levels was detectable, however, after wild-type p53 gene transfer. Furthermore, protein expression of secreted tissue inhibitor of metalloproteinase-2 was not altered by AdCMV.p53 treatment. In contrast, in SK-MEL-147 cells, AdCMV.p53 did not affect cell invasiveness and levels of secreted matrix metalloproteinase-2. Gene transfer of wild-type p53 inhibited proliferation of both cell lines, showing that also SK-MEL-147 cells respond to wild-type p53 overexpression. This novel mechanism of action of wild-type p53 gene transfer may contribute to its antitumor effect by downregulating cell invasion and matrix metalloproteinase-2 secreted levels in mutated p53 human melanoma cell lines.

Adenoviridae↗

PubMedDate not supplied

Developmental differences in the expression and modulation of extracellular matrix proteases and inhibitors in mouse skin fibroblasts.

To investigate developmental differences in the wound repair process between fetal and adult skin fibroblasts, we studied the expression of plasminogen activator, plasminogen activator inhibitor, matrix metalloproteinase, and tissue inhibitor of metalloproteinase in E-15, E-17, newborn and adult mouse skin fibroblasts cultured within three dimensional matrices of either collagen or fibrin. Fibrin overlay and reverse overlay analyses revealed that mouse skin fibroblasts secreted tissue plasminogen activator and type1 plasminogen activator inhibitor. However, only E-15 and E-17 fibroblasts secreted the active form of tissue plasminogen activator, while in newborn and adult fibroblasts tissue plasminogen activator was conjugated to type1 plasminogen activator inhibitor. Only adult fibroblasts expressed a high level of active type1 plasminogen activator inhibitor. Gelatin zymography revealed that the predominant matrix metalloproteinase secreted by all the mouse fibroblasts was gelatinase A (matrix metalloproteinase -2). Matrix metalloproteinase -2 was partially activated in the adult fibroblasts cultured within a collagen matrix. The tissue inhibitor of metalloproteinase-2 was expressed by all fibroblasts, but levels were highest in the newborn and adult fibroblasts. When E-15 fibroblasts were cultured within a fibrin matrix, tissue plasminogen activator was downregulated. Transforming growth factor-betadownregulated tissue plasminogen activator while upregulating type1 plasminogen activator inhibitor, and platelet-derived growth factor enhanced tissue plasminogen activator expression in E-15 fibroblasts. Therefore, plasminogen activator and its inhibitor, and matrix metalloproteinase and its associated tissue inhibitor are differentially expressed in fetal and adult fibroblasts, and their expression is controlled by extracellular matrix components and growth factors present in wounds.

Animals↗

PubMed2005

Regulation of pituitary hormones and cell proliferation by components of the extracellular matrix.

The extracellular matrix is a three-dimensional network of proteins, glycosaminoglycans and other macromolecules. It has a structural support function as well as a role in cell adhesion, migration, proliferation, differentiation, and survival. The extracellular matrix conveys signals through membrane receptors called integrins and plays an important role in pituitary physiology and tumorigenesis. There is a differential expression of extracellular matrix components and integrins during the pituitary development in the embryo and during tumorigenesis in the adult. Different extracellular matrix components regulate adrenocorticotropin at the level of the proopiomelanocortin gene transcription. The extracellular matrix also controls the proliferation of adrenocorticotropin-secreting tumor cells. On the other hand, laminin regulates the production of prolactin. Laminin has a dynamic pattern of expression during prolactinoma development with lower levels in the early pituitary hyperplasia and a strong reduction in fully grown prolactinomas. Therefore, the expression of extracellular matrix components plays a role in pituitary tumorigenesis. On the other hand, the remodeling of the extracellular matrix affects pituitary cell proliferation. Matrix metalloproteinase activity is very high in all types of human pituitary adenomas. Matrix metalloproteinase secreted by pituitary cells can release growth factors from the extracellular matrix that, in turn, control pituitary cell proliferation and hormone secretion. In summary, the differential expression of extracellular matrix components, integrins and matrix metalloproteinase contributes to the control of pituitary hormone production and cell proliferation during tumorigenesis.

Adenoma↗

PubMed1999

Phase I study of intrapleural batimastat (BB-94), a matrix metalloproteinase inhibitor, in the treatment of malignant pleural effusions.

Tumor cells and associated stromal cells secrete matrix metalloproteinases (MMPs), contributing to invasion, angiogenesis, and metastasis. Batimastat (BB-94) is a broad-spectrum MMP inhibitor that causes resolution of ascites and/or tumor growth delay in animal models of breast, ovarian, and colorectal cancer. We recruited 18 patients with cytologically positive malignant pleural effusions into a Phase I study of intrapleural BB-94. Three patients received single doses of BB-94 at each dose level: 15, 30, 60, 105, 135, and 300 mg/m2. Two patients were retreated with a second course at 60 and 105 mg/m2. BB-94 was detectable in plasma 1 h after intrapleural administration, and peak levels of 20-200 ng/ml occurred after 4 h to 1 week. BB-94 persisted in the plasma for up to 12 weeks, at levels exceeding the IC50s for target MMPs. Peak values were higher, and persistence in the plasma was longer after higher doses of BB-94. The treatment was well tolerated. Toxic effects included low-grade fever for 24-48 h (6 of 18 patients, 33%) and reversible asymptomatic elevation of liver enzymes (8 patients, 44%). Toxicity seemed unrelated to BB-94 dose or plasma levels. Sixteen patients evaluable for response required significantly fewer pleural aspirations in the 3 months after BB-94 compared with the 3 months before. Seven patients (44%) required no further pleural aspiration until death/last follow-up. After 1 month, patients treated with 60-300 mg/m2 BB-94 had significantly better dyspnea scores, indicating improved exercise tolerance, compared with baseline scores the day after BB-94. The maximum tolerated intrapleural dose remains to be defined, but it is clear that intrapleural BB-94 is well tolerated, with evidence of local activity.

Adult↗

PubMed1996

Identification of matrix metalloproteinases and their inhibitor in the articular disc of the craniomandibular joint of the rabbit.

Connective tissue cells synthesize and secrete matrix metalloproteinases (MMPs), a family of matrix-degrading enzymes (comprising collagenases, gelatinases and stromelysins), which are capable of degrading all the constituent molecules of connective tissues at physiological pH. This investigation documents the synthesis and distribution of MMPs and their inhibitor TIMP-1 (tissue inhibitor of metalloproteinases-1) in the developing articular disc of the craniomandibular joint of the rabbit using indirect immunofluorescence microscopy. Cells of the disc synthesised all three classes of MMPs as well as TIMP-1 in all regions of the disc at all stages examined. MMPs and TIMP-1 were detected as bright intracellular accumulations probably within Golgi vesicles and as occasional diffuse, matrix-bound deposits. These results suggest that MMP-mediated matrix remodelling is a prominent feature of growth in craniomandibular joint disc.

Animals↗

PubMed2006

Modulation of matrix metalloproteinase-9 (MMP-9) secretion in B lymphopoiesis.

The matrix metalloproteinases (MMPs) are proteolytic enzymes that degrade the extracellular matrix, thus involved in cellular migration. The extent and role of MMPs secretion in primary non-transformed B cells, and specifically during early stages of development in the bone marrow (BM), has been barely unveiled. Herein, we investigated the secretion of MMP-9 during B lymphopoiesis and its modulation in response to different mitogens and cytokines. To do so, we used our BM culture system and well-studied mutated mouse models to isolate the different B cell populations. Our results show that MMP-9 is spontaneously secreted throughout B lymphopoiesis, and that the level of secreted MMP-9 is developmentally regulated. Using reverse transcription-PCR, we found that IFNbetaR is expressed throughout B cell development, while tumor necrosis factor (TNF)-alphaR-p55 and IFNgammaR expressions are initiated only at the pre-B stage. We found that TNFalpha stimulates MMP-9 secretion in transitional cells, whereas IFNs suppress MMP-9 secretion in immature cells. LPS and phorbol 12-myristate 13-acetate suppressed MMP-9 secretion in transitional cells, whereas LPS and concanavalin A stimulated MMP-9 secretion in mature B cells. We conclude that B lymphocyte development is accompanied with MMP-9 secretion and the developing cells are competent to modify this secretion upon different immune stimuli.

Animals↗

PubMed1991

Inhibition of tumor invasion of smooth muscle cell layers by recombinant human metalloproteinase inhibitor.

Matrix metalloproteinases secreted by tumor cells play an important role in the proteolytic degradation of the extracellular matrix during invasion. In a previous study, we showed that the degradation of extra-cellular matrices by human HT 1080 fibrosarcoma cells is suppressed by endothelial cells. The identification of inhibitors of metalloproteinases secreted by endothelial cells led us to postulate that these inhibitors were responsible for the suppressive effect (Cancer Res., 46: 3580-3586, 1986). In the present study, we have investigated the inhibitory activity of one of these inhibitors designated metalloproteinase inhibitor (MI)/tissue inhibitor of metalloproteinases (TIMP)-2 on the degradation and invasion of rat smooth muscle cell matrices by two invasive tumor cell lines, the c-Ha-ras-1 transfected rat embryo cell line 4R and the HT 1080 human fibrosarcoma cell line. The inhibitor was obtained in recombinant form from the culture medium of Chinese hamster ovary cells transfected with a human MI complementary DNA. Recombinant MI/TIMP-2 inhibited several matrix metalloproteinases identified in the culture medium of the tumor cell lines including interstitial collagenase. Mr 72,000 gelatinase (type IV collagenase), and Mr 92,000 gelatinase. Approximately 70% inhibition of the degradation of smooth muscle cell matrices was observed when the recombinant inhibitor was present along with cultured cells at a concentration of 10 micrograms/ml. Similarly, inhibition of the penetration of a multilayer of growing smooth muscle cells and their surrounding matrix was demonstrated. The inhibitor had no effect on cell growth or attachment. Thus, recombinant MI/TIMP-2, like TIMP, is a potent inhibitor of tumor invasion. Since both inhibitors are secreted by endothelial cells (J. Biol. Chem., 264: 17445-17453, 1989), they may play an important role in protecting large blood vessels from invasion.

Cell Adhesion↗

PubMed2003

Secretion of macrophage urokinase plasminogen activator is dependent on proteoglycans.

The importance of proteoglycans for secretion of proteolytic enzymes was studied in the murine macrophage cell line J774. Untreated or 4beta-phorbol 12-myristate 13-acetate (PMA)-stimulated macrophages were treated with hexyl-beta-d-thioxyloside to interfere with the attachment of glycosaminoglycan chains to their respective protein cores. Activation of the J774 macrophages with PMA resulted in increased secretion of trypsin-like serine proteinase activity. This activity was completely inhibited by plasminogen activator inhibitor 1 and by amiloride, identifying the activity as urokinase plasminogen activator (uPA). Treatment of both the unstimulated or PMA-stimulated macrophages with xyloside resulted in decreased uPA activity and Western blotting analysis revealed an almost complete absence of secreted uPA protein after xyloside treatment of either control- or PMA-treated cells. Zymography analyses with gels containing both gelatin and plasminogen confirmed these findings. The xyloside treatment did not reduce the mRNA levels for uPA, indicating that the effect was at the post-translational level. Treatment of the macrophages with xylosides did also reduce the levels of secreted matrix metalloproteinase 9. Taken together, these findings indicate a role for proteoglycans in the secretion of uPA and MMP-9.

Amiloride↗

PubMed2001

Identification of a matrix-degrading phenotype in human tuberculosis in vitro and in vivo.

Tuberculous meningitis is characterized by cerebral tissue destruction. Monocytes, pivotal in immune responses to Mycobacterium tuberculosis, secrete matrix metalloproteinase-9 (MMP-9), which facilitates leukocyte migration across the blood-brain barrier, but may cause cerebral injury. In vitro, human monocytic (THP-1) cells infected by live, virulent M. tuberculosis secreted MMP-9 in a dose-dependent manner. At 24 h, MMP-9 concentrations increased 10-fold to 239 +/- 75 ng/ml (p = 0.001 vs controls). MMP-9 mRNA became detectable at 24--48 h. In contrast, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) gene expression and secretion were similar to constitutive levels from controls at 24 h and increased just 5-fold by 48 h. In vivo investigation revealed MMP-9 concentration per leukocyte in cerebrospinal fluid (CSF) from tuberculous meningitis patients (n = 23; median (range), 3.19 (0.19--31.00) ng/ml/cell) to be higher than that in bacterial (n = 12; 0.23 (0.01--18.37) ng/ml/cell) or viral meningitis (n = 20; 0.20 (0.04--31.00) ng/ml/cell; p < 0.01). TIMP-1, which was constitutively secreted into CSF, was not elevated in tuberculous compared with bacterial meningitis or controls. Thus, a phenotype in which MMP-9 activity is relatively unrestricted by TIMP-1 developed both in vitro and in vivo. This is functionally significant, since MMP-9 concentrations per CSF leukocyte (but not TIMP-1 concentrations) were elevated in fatal tuberculous meningitis and in patients with signs of cerebral tissue damage (unconsciousness, confusion, or neurological deficit; p < 0.05). However, MMP-9 activity was unrelated to the severity of systemic illness. In summary, M. tuberculosis-infected monocytic cells develop a matrix-degrading phenotype, which was observed in vivo and relates to clinical signs reflecting cerebral injury in tuberculous meningitis.

Adult↗

PubMed2007

Induction of matrix metalloproteinase gene expression in an endothelial cell line by direct interaction with malignant cells.

Mouse endothelial TKD2 cells in monolayers were cocultured with various human cell lines for 24 h, and the expression of several secreted matrix metalloproteinases (MMP) and cell adhesion molecules was examined by real-time reverse transcription-polymerase chain reaction using mouse-specific primers. Coculture with normal fibroblasts did not elicit the expression of these molecules, but coculture with cancer cells induced the expression of MMP-3, MMP-9 and MMP-10 mRNA in endothelial cells, and in normal mouse embryonic fibroblasts. The induction of MMP mRNA was dependent on direct cell adhesion, as separate culture of A549 cells in Boyden chambers did not induce MMP mRNA, and neutralizing antibody against VLA-4 abolished the induction. An inhibitor of phosphatidylinositol-3-phosphate kinase strongly suppressed the induction of MMP-3, MMP-9 and MMP-10 mRNA, and expression of the dominant-negative mutant of phosphatidylinositol-3-phosphate kinase also decreased the induction. It was suggested that intracellular reactive oxygen species (ROS) levels were increased in TKD2 cells following adhesion to cancer cells. ROS scavengers decreased the levels of MMP induction, and roterone, an inhibitor of mitochondrial complex I, strongly suppressed the induction of MMP-3, MMP-9 and MMP-10. The depletion of mitochondria in TKD2 cells decreased the induction of MMP-9, but the induction of MMP-3 and MMP-10 was not affected. These results indicate that the adhesion of cancer cells to endothelial cells activates several distinct signaling pathways to induce MMP gene expression, and the pathways for MMP-3, MMP-9 and MMP-10 are partly different. For the induction of MMP-9, mitochondria participate in induction, possibly through the production of ROS.

Animals↗

PubMed2003

Thrombin stimulates production of fibronectin by human proximal tubular epithelial cells via a transforming growth factor-beta-dependent mechanism.

BACKGROUND: Tubulointerstitial fibrosis contributes to the progression of many forms of glomerular disease and to end-stage renal failure. Inflammatory mediators generated during glomerular injury may induce tubulointerstitial lesions by stimulating tubular cells. Thrombin has multiple biological functions in addition to its role in haemostasis and has been detected in the urine of patients with glomerular diseases. The present study investigated whether thrombin can modulate the production of fibronectin (FN) in cultured human proximal tubular epithelial cells (PTEC). METHODS: Cultured PTEC were incubated with or without thrombin to examine the effect of thrombin on FN production in PTEC. FN and transforming growth factor-beta (TGF-beta) levels were measured in culture supernatants by enzyme-linked immunosorbent assay (ELISA). Expression of FN mRNA was analysed by reverse transcriptase-polymerase chain reaction. Effects of thrombin on matrix metabolism were examined by enzyme immunoassay for the detection of secreted matrix metalloproteinase (MMP) and its inhibitors (TIMPs) as well as by zymography. RESULTS: Thrombin stimulated FN secretion in PTEC. Thrombin also stimulated TGF-beta secretion in PTEC in a dose-dependent manner. Expression of FN mRNA by PTEC was augmented by thrombin. The stimulatory effect of thrombin on FN secretion was inhibited by neutralizing antibodies against TGF-beta but not by an irrelevant antibody. Thrombin-induced FN secretin was also inhibited by thrombin inhibitors, such as antithrombin III, hirudin and argatroban. Although thrombin stimulated TIMP-1 and -2 secretion by PTEC, the stimulatory effect of thrombin on MMP-2 was not statistically significant. Thrombin did not affect the expression of MMP-2 in zymography studies. CONCLUSIONS: We found that thrombin stimulates FN production in PTEC without causing matrix degradation, an effect that may contribute to the formation of tubulointerstitial fibrosis associated with glomerular disease. The stimulatory effect of thrombin on FN production in PTEC is, at least in part, mediated by TGF-beta.

Actins↗