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Results for “Matrix Metalloproteinases, Secreted”

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PubMed1992

Rapidly destructive arthropathy of the hip. Studies on bone resorptive factors in joint fluid with a theory of pathogenesis.

The mechanism of joint destruction in rapidly destructive coxopathy was studied by analyzing bone resorptive factors in the joint fluid. Prostaglandins were found to play a partial role in joint destruction. Some cases of rapidly destructive coxopathy revealed elevated levels of interleukin-1 beta (IL-1 beta) in the joint fluid. Electrophoretic analysis of proteolytic enzymes in polyacrylamide gel containing sodium dodecyl sulfate and copolymerized gelatin demonstrated that the resorptively active peptides have relative molecular weights (M(r)) of approximately 92,000, 72,000, and lower than 60,000. Cultured cells from synovia obtained perioperatively secreted matrix metalloproteinase 2 (MMP-2) with an M(r) of 72,000 and matrix metalloproteinase 3(MMP-3) with an M(r) of 57,000. Synovial cells from the patients with coxarthrosis secreted fewer proteolytic enzymes. Prostaglandins, IL-1 beta, MMP-2, and MMP-3 could act synergetically as promotors in the rapid destruction of the hip joint.

Bone Matrix↗

PubMed2006

Membrane type-matrix metalloproteinases in idiopathic pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is characterized by fibroblast expansion and extracellular matrix accumulation. Some secreted matrix metalloproteinases (MMPs) as MMP2 are highly upregulated in IPF lungs. Membrane-type (MT)-MMPs participate in the activation of pro-MMP2. However, they have not been examined in IPF. METHODS: Type I transmembrane MT-MMPs, MT1, MT2, MT3, and MT5-MMP were analyzed by real-time PCR and immunohistochemistry in IPF and normal lungs. MMP-2 was also immunolocalized and evaluated by gelatin zymography in BAL fluids. Additionally, the MT-MMPs were examined by real time PCR in lung fibroblasts stimulated with TGF-beta1 and IFN-gamma. RESULTS: MT1-MMP, was the most highly expressed followed by MT2- and MT5-MMP, and by a moderate expression of MT3-MMP. Regarding their localization, MT1- and MT2-MMPs were found in alveolar epithelial cells, MT3-MMP in fibroblasts from fibroblastic foci and alveolar epithelial cells and MT5-MMP in basal bronchiolar epithelial cells and in areas of squamous metaplasia. MMP2 was localized in alveolar and basal bronchiolar epithelial cells and fibroblasts, and increased active enzyme was observed in BAL fluids. In lung fibroblasts, TGF-beta1 induced a strong upregulation of MT3-MMP, both at the gene and protein level. This effect was blocked by genistein, a protein tyrosin kinase inhibitor and partially repressed by SB203580 a p38 MAP kinase inhibitor. IFN-gamma had no effect. CONCLUSIONS: MT-MMPs are expressed in IPF, in the same cell types as MMP2. Mostly by different types of epithelial cells a pivotal component in the aberrant remodeling of the lung microenvironment. Interestingly MT3-MMP that was found in fibroblastic foci was upregulated in vitro by TGF-beta1 a potent profibrotic mediator.

Bronchoalveolar Lavage Fluid↗

PubMed2004

Increased intercellular communication through gap junctions may contribute to progression of osteoarthritis.

Our aim was to support the hypothesis of a specific association between gap junctions in synovial tissue and the presence of osteoarthritis, as evidenced by differences between osteoarthritis and non-osteoarthritis synovia in the number of gap junctions, the amount of gap-junction protein, and the amount of enzymatic activity produced through a pathway mediated by gap-junction intercellular communication. An average of 4.41 gap junctions were found per 100 cells counted in the osteoarthritis synovia, compared with 1.00 in the controls. The amount of the gap-junction protein connexin 43 in synovial lining cells was approximately 50% greater in patients with osteoarthritis. Synovial lining cells from patients with osteoarthritis produced matrix metalloproteinases constitutively and, at higher levels, in response to stimulation by interleukin-1 beta. In both cases, intercellular communication through gap junctions was shown to be critical to the ability of the cells to secrete matrix metalloproteinases. Overall, the results indicated that gap junctions between synovial lining cells were altered significantly in patients with osteoarthritis, as a consequence of the disease process or as part of the causal chain. In either case, gap junctions seem to be a rational therapeutic target.

Aged↗

PubMed2003

Calcium-induced activation and truncation of promatrix metalloproteinase-9 linked to the core protein of chondroitin sulfate proteoglycans.

In the leukemic macrophage cell-line THP-1, a fraction of the secreted matrix metalloproteinase 9 (MMP-9) is linked to the core protein of chondroitin sulfate proteoglycans (CSPG). Unlike the monomeric and homodimeric forms of MMP-9, the addition of exogenous CaCl2 to the proMMP-9/CSPG complex resulted in an active gelatinase due to the induction of an autocatalytic removal of the N-terminal prodomain. In addition, the MMP-9 was released from the CSPG through a process that appeared to be a stepwise truncation of both the CSPG core protein and a part of the C-terminal domain of the gelatinase. The calcium-induced activation and truncation of the MMP-9/CSPG complex was independent of the concentration of the complex, inhibited by the MMP inhibitors EDTA, 1,10-phenanthroline and TIMP-1, but not by general inhibitors of serine, thiol and acid proteinases. This indicated that the activation and truncation process was not due to a bimolecular reaction, but more likely an intramolecular reaction. The negatively charged chondroitin sulfate chains in the proteoglycan were not involved in this process. Other metal-containing compounds like amino-phenylmercuric acetate (APMA), NaCl, ZnCl2 and MgCl2 were not able to induce activation and truncation of the proMMP-9 in this heterodimer. On the contrary, APMA inhibited the calcium-induced process, whereas high concentrations of either MgCl2 or NaCl had no effect. Our results indicate that the interaction between the MMP-9 and the core protein of the CSPG was the causal factor in the calcium-induced activation and truncation of the gelatinase, and that this process was not due to a general electrostatic effect.

Animals↗

PubMed2004

Clinical and biochemical results of the metalloproteinase inhibition with subantimicrobial doses of doxycycline to prevent acute coronary syndromes (MIDAS) pilot trial.

BACKGROUND: Vulnerable plaque demonstrates intense inflammation in which macrophages secrete matrix metalloproteinases (MMPs) that degrade the fibrous cap, ultimately leading to rupture, in situ thrombosis, and an associated clinical event. Thus, inhibition of MMP activity or more general suppression of vascular inflammation are attractive targets for interventions intended to reduce plaque rupture. We hypothesized that subantimicrobial doses of doxycycline (SDD) (20 mg twice daily) would benefit patients with coronary artery disease by reducing inflammation and MMP activity and thus possibly prevent coronary plaque rupture events. METHODS AND RESULTS: We conducted a prospective, randomized, double-blind, placebo-controlled pilot study of 6 months of SDD or placebo treatment to reduce inflammation and prevent plaque rupture events. A total of 50 patients were enrolled, of whom 24 were randomized to placebo and 26 to SDD. At 6 months, there was no difference in the composite endpoint of sudden death, fatal myocardial infarction (MI), non-fatal MI, or troponin-positive unstable angina in SDD compared with placebo-treated patients (8.4% versus 0%, P=0.491). Biochemical markers of inflammation were assessed in plasma at study entry and after 6 months of therapy in 30 patients. In SDD-treated patients, high-sensitivity C-reactive protein (CRP) was reduced by 46% from 4.8+/-0.6 microg/mL to 2.6+/-0.4 microg/mL (P=0.007), whereas CRP was not significantly reduced in placebo patients. Interleukin (IL)-6 decreased from 22.1+/-3.7 pg/mL at baseline to 14.7+/-1.8 pg/mL at 6 months in SDD-treated patients (P=0.025) but did not decrease significantly in placebo-treated patients. On zymography, pro-MMP-9 activity was reduced 50% by SDD therapy (P=0.011), whereas it was unchanged by placebo treatment. CONCLUSIONS: SDD appears to exert potentially beneficial effects on inflammation that could promote plaque stability. These findings should be investigated in a larger study.

Aged↗

PubMed2005

Posttranslational modifications of decidual IGFBP-1 by steroid hormones in vitro.

Insulin-like growth factor binding protein-1 (IGFBP-1) appears to regulate insulin-like growth factors (IGFs; IGF-I and IGF-II) biological activity within the local environment of human placenta by modulating IGFs interaction with their receptors. Considering that posttranslational modifications of IGFBP-1 such as phosphorylation and proteolysis affect its affinity for IGFs, this study was undertaken to identify the role of estrogen and progesterone in this regard. The conditioned media of steroid hormone-treated decidual cells were evaluated using different approaches using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and non-denaturing PAGE following immunoblotting as well as zymographys that contained gelatin and IGFBP-1 as substrates. Our results demonstrated that medroxy progesterone acetate (MPA) treatment increased both phosphorylated and non-phosphorylated decidual-secreted IGFBP-1, whereas 17beta-estradiol (E2) treatment attenuated its phosphorylated forms. Furthermore, the results of zymography revealed that steroid hormones regulated the activity of decidual-secreted matrix metalloproteinases (MMP)-2 and -9, in which E2 treatment up-regulated the MMP-9 activity. Finally, it was demonstrated in our study that decidual-secreted MMP-9 was capable of degrading human amniotic fluid-derived IGFBP-1. In conclusion, our data implicate steroid hormones in the control of IGF system activities at the embryo-maternal interface, at least in part, through their effects on the post-translation changes of decidual-secreted IGFBP-1 such as its phosphorylation and/or proteolysis.

Adult↗

PubMed2003

Pro-inflammatory cytokine-induced matrix metalloproteinase-1 (MMP-1) secretion in human pancreatic periacinar myofibroblasts.

Matrix metalloproteinases (MMPs) are the proteases involved in the degradation of the extracellular matrix. MMP-1 is thought to be one of the key enzymes in fibrolysis, a process closely related to tissue remodeling. In the present study, we investigated MMP-1 secretion from human pancreatic periacinar myofibroblasts in response to pro-inflammatory cytokines IL-1beta and TNF-alpha. We also attempted to clarify the intracellular signaling pathways mediating the cytokine-induced MMP-1 secretion. MMP-1 secretion was measured by an enzyme-linked immunosorbent assay. MMP-1 molecules were analyzed by Western blotting. MMP-1 mRNA expression was evaluated by Northern blotting. IL-1l and TNF-alpha stimulated the MMP-1 secretion in a dose- and time-dependent manner. Ninety percent of MMP-1 was secreted as inactive form (pro-MMP-1). The effects of IL-1beta and TNF-alpha were significantly inhibited by PD98059 MEK/ERK inhibitor). In contrast, SB203580 (p38 MAPK inhibitor), GF109203X (PKC inhibitor), and PDTC (NF-kappaB inhibitor) did not alter the MMP-1 secretion induced by IL-1beta and TNF-alpha. These effects were also observed at them RNA level. In conclusion, in human pancreatic periacinar myofibroblasts, MMP-1 secretion was regulated by the pro-inflammatory cytokines via the MEK/ERK cascade. Thus, human pancreatic periacinar myofibroblasts may play an important role in the remodeling of damaged pancreatic tissue in chronic pancreatitis via MMP-1 secretion.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2004

Transinactivation of the epidermal growth factor receptor tyrosine kinase and focal adhesion kinase phosphorylation by dietary flavonoids: effect on invasive potential of human carcinoma cells.

Focal adhesion kinase (FAK), a member of a growing family of structurally distinct protein tyrosine kinases (PTK), has been linked to specific phosphorylation events, and the elevation of FAK activity in human carcinoma cells correlated with increased invasive potential. Transactivation of the epidermal growth factor receptor (EGFR) tyrosine kinase activity is proposed to stimulate cell migration and the subsequent activation of downstream signaling pathways. Quercetin (Qu) and luteolin (Lu), are potent PTK inhibitors as well as putative chemopreventive agents. The present work, we demonstrate that Qu and Lu at concentration of 20 microM transinactivated EGFR tyrosine kinase activity with marked reduction in phosphotyrosyl level of 170, 125, 65, 60 and 42 kDa cellular proteins, and induced apoptosis in MiaPaCa-2 cells. The 125 kDa protein was further identified as a FAK by immunoprecipitation and immunoblotting analyses. Tumor cells treated with Lu or Qu dampened the phosphorylation of FAK. In addition, our data clearly demonstrated that tumor cells responded to Qu and Lu by parallel reductions in the levels of phosphorylated FAK and the secreted matrix metalloproteinase (MMP) that may lead to the suppression of invasive potential and cell migration in vitro. While the molecular mechanism of FAK regulation of MMP secretion in tumor cells remains unclear, our results suggested that blockade of the EGFR-signaling pathway may contributed to the net effect. As suggested in the current study, targeting EGFR and FAK with the objective of modulating their regulatory pathways could offer prospects for the treatment of EGFR-responsive cancers in the future.

Carcinoma↗

PubMed2004

Bone marrow endothelial cells increase the invasiveness of human multiple myeloma cells through upregulation of MMP-9: evidence for a role of hepatocyte growth factor.

The migration of multiple myeloma (MM) cells from the circulation into the bone marrow (BM) implicates that they must have the capacity to cross the BM endothelium including the subendothelial basement membrane. In this study, human CD138+ MM cells were immunomagnetically isolated from BM samples of MM patients and their invasion through Matrigel, that is, a reconstituted basement membrane, was determined. We demonstrated that primary MM cells have the capacity to transmigrate through basement membrane and that this invasiveness was considerably increased when assessed on Matrigel filters coated with BM endothelial cells (EC) (4LHBMEC line) (transendothelial invasion). The isolated MM cells were shown by zymography to secrete matrix metalloproteinase (MMP)-9 and anti-MMP-9 antibodies inhibited transendothelial invasion, indicating that MMP-9 is involved in this process. BM EC were found to increase the MMP-9 secretion in MM cells, indicating that EC enhance MM cell invasion through stimulation of MMP-9 secretion. BM EC were found to produce hepatocyte growth factor (HGF), and this cytokine also stimulated MMP-9 secretion in MM cells, while anti-HGF antibodies significantly inhibited EC-stimulated MM cell invasion. In summary, our findings provide evidence that MM cell-BM EC interactions enhance the invasion of human MM cells through stimulation of MMP-9 secretion.

Bone Marrow Cells↗

PubMed2003

Inhibitory effect of methanol extract of Euonymus alatus on matrix metalloproteinase-9.

Various extracts prepared from stems of Euonymus alatus were tested for cytotoxic activity on human hepatocellular carcinoma cell line, Hep3B cells using the XTT assay method. Also, the extracts were investigated the inhibitory effects on matrix metalloproteinase (MMP)-9 activity using gelatin zymography. The methanol extract, hexane and ethyl acetate fraction exhibited weak cytotoxic activity (IC(50) of >100 microg/ml). However, butanol (IC(50)=65 microg/ml) and chloroform (IC(50)=85 microg/ml) fraction exhibited strongly cytotoxic activity. Gelatin zymography showed that the Hep3B cells secreted matrix metalloproteinase (MMP), probably including MMP-9, which may be involved in tumor cell invasion and metastasis. The butanol fraction showed stronger inhibitory effect of proteolytic activity than other fractions. Also, the butanol fraction was able to decrease the proteolytic activity of MMP-9 in a concentration-dependent manner on zymography. These results suggest that the butanol fraction from E. alatus has highly inhibitory effect on MMP-9 in comparatively low cytotoxicity.

Antineoplastic Agents↗

PubMed1998

Protein tyrosine phosphorylation in signalling pathways leading to the activation of gelatinase A: activation of gelatinase A by treatment with the protein tyrosine phosphatase inhibitor sodium orthovanadate.

Fibroblasts in monolayer culture secrete gelatinase A (MMP2; 72 kDa type IV collagenase) only in its proenzyme form. Unlike other secreted matrix metalloproteinases, progelatinase A is refractory to activation by serine proteinases. Disparate agents, including monensin, cytochalasin D, and concanavalin A, have been found to mediate the activation of gelatinase A zymogen secreted by fibroblast monolayers. Our finding that monensin-mediated activation can be reversed by the protein tyrosine kinase inhibitor genistein (Li et al., Experimental Cell Research 232 (1997) 332) prompted us to investigate the effect of the specific inhibitor of protein tyrosine phosphatases, sodium orthovanadate, on progelatinase A activation. Treatment of fibroblast monolayers with orthovanadate also results in the secretion of activated gelatinase A. This activation is dose- and time-dependent, requires protein synthesis, and is associated with cell membranes. Vanadate-mediated activation does not occur in the presence of herbimycin A, a protein tyrosine kinase inhibitor. As with progelatinase activation mediated by monensin, concanavalin A, and cytochalasin D, orthovanadate treatment results in increased synthesis of the membrane proteinase MT1-MMP, that can catalyze the activation of progelatinase A. Protein tyrosine kinase inhibitors are able to prevent the increase of MT1-MMP mRNA, as shown by Northern blot and RT-PCR. In addition, orthovanadate potentiates the effects of monensin and concanavalin A. While treatment with monensin or concanavalin A result only in an increase of the putative activator MT1-MMP, orthovanadate also reduces the production of the specific inhibitor TIMP-2. These experiments implicate protein tyrosine phosphorylation in the signal transduction pathways which lead to the activation of progelatinase A.

Base Sequence↗

PubMed1997

Modulation of matrix metalloproteinase-7 (matrilysin) secretion in coculture of human colon carcinoma cells with fibroblasts from orthotopic and ectopic organs.

Matrix metalloproteinase-7 (MMP-7) is a member of the family of matrix-degrading metalloproteinases that are believed to contribute to the complex process of cancer invasion and metastasis. The secretion level of MMP-7 as assayed by immunoblot analysis was low but distinct in the culture medium of a human colon carcinoma cell line, WIDr, whereas none of the fibroblasts secreted the detectable level of MMP-7. The coculture of WiDr with various human fibroblasts from orthotopic (colon) and ectopic (thyroid, brain, lung, and skin) organs significantly stimulated the secretion of MMP-7 compared with the cultures of individual cells. Reverse transcriptase-polymerase chain reaction analysis and RNA blot analysis suggested that this enhancement occurred at a pretranslational level. The extent of the stimulation was widely varied by the fibroblasts used and was dependent on the cellular ratios and density in the coculture. There may exist a tendency that fibroblasts of orthotopic origin stimulate more extensively than do those of ectopic origin. Moreover, in the coculture of high cell density, normal fibroblasts from the ectopic organs reduced the MMP-7 secretion. The stimulation of MMP-7 secretion may be partially mediated through soluble factor(s); however, direct cell-cell interactions would be required for maximum stimulation. The enhanced MMP-7 secretion was also observed in coculture of colon fibroblasts with other colorectal carcinoma cell lines such as RCM-1 and SW837, which secreted hardly detectable levels of MMP-7 in the individual culture. These results suggest that MMP-7 secretion by colon carcinoma cells is influenced by specific interactions between the carcinoma cells and host fibroblasts.

Cell Communication↗

PubMed1997

Intracellular activation of gelatinase A (72-kDa type IV collagenase) by normal fibroblasts.

Normal fibroblasts cultured as monolayers secrete matrix metalloproteinases (MMP), including gelatinase A (72-kDa type IV collagenase) as inactive zymogens. Previously we found that normal fibroblasts cultured in a type I collagen lattice (dermal equivalent) secrete active gelatinase A. Here we show that the activation of progelatinase A occurs within the cell and that the activator copurifies with Golgi membranes. Cell extracts of fibroblasts cultured in collagen lattices contain active 62-kDa gelatinase A at least 4-6 h before active enzyme is detected in the culture medium. Pulse-chase experiments confirm these results. The activator is membrane-bound and localizes to the Golgi-enriched fraction. Highly purified plasma membranes from lattice cultures are unable to convert gelatinase A from the zymogen to its active form. The activator may be a metalloproteinase because EDTA prevents activation of exogenous proenzyme by membrane fractions. Membrane-type MMP1, the enzyme thought to be responsible for activation of gelatinase A on the plasma membrane of tumor cells, shows no significant change in either mRNA or protein levels during lattice culture. Intracellular levels of gelatinase A mRNA and protein increase during the culture period, and tissue inhibitor of metalloproteinases concentration does not change. Because of the greater availability of tissue inhibitor of metalloproteinases-free proenzyme as a substrate for the activator, it is possible that membrane-type MMP1 is the activating enzyme. In that case, malignant transformation may involve a change in the localization of the activator to the plasma membrane.

Biomarkers↗

PubMed2002

Macrophage metalloproteinases degrade high-density-lipoprotein-associated apolipoprotein A-I at both the N- and C-termini.

Atheromatous plaques contain various cell types, including macrophages, endothelial cells and smooth-muscle cells. To investigate the possible interactions between secreted matrix metalloproteinases and high-density lipoprotein (HDL) components, we tested the above cell types by culturing them for 24 h. HDL(3) (HDL subfractions with average sizes of between 8.44 nm for HDL(3A) and 7.62 nm for HDL(3C)) were then incubated in their cell-free conditioned media. Proteolytic degradation of apolipoprotein A-I was observed with macrophages, but not with endothelial-cell- or muscle-cell-conditioned supernatant. Absence of calcium or addition of EDTA to incubation media prevented all proteolytic processes. The identified apolipoprotein A-I fragments had sizes of 26, 22, 14 and 9 kDa. Two-dimensional electrophoresis and MS resolved the 26 and the 22 kDa components and identified peptides resulting from both N- and C-terminal cleavage of apolipoprotein A-I. The higher abundance of C- than N-terminally cleaved peptides agrees with data in the literature for a fully structured alpha-helix around Tyr(18) compared with an unstructured region around Gly(185) and Gly(186). The flexibility in the latter region of apolipoprotein A-I may explain its susceptibility to proteolysis. In our experimental set-up, HDL(3C) was more extensively degraded than the other HDL(3) subclasses (HDL(3A) and HDL(3B)). Proteolytic fragments produced by metalloproteinase action were shown by gel filtration and electrophoresis to be neither associated with lipids nor self-associated.

Animals↗

PubMed2002

[Effects of blastocyst surface oligosaccharide LeY on secretion and expression of matrix metalloproteinase in vivo].

The effects of oligosaccharide LeY on expression and secretion of matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase 1 (TIMP1) of embryos were investigated in cultured mouse blastocysts by zymography, immuno-dot blots and RT-PCR after blocking the oligosaccharide on embryo surface with specific monoclonal antibody AH6. The cultured blastocysts were treated with antibody AH6 for 3 h, then the antibody was removed and culture was continued, and the expression and secretion of MMP and TIMP1 were analyzed at 1.5 h, 3 h and 6 h. It was shown that, only 1.5 h after blocking, the expression of MMP2 gene and MMP9 gene in embryo was significantly declined, and expression of TIMP1 was slight increased, then secretions of MMP2, MMP9 were also declined at 3 h, but secretion of TIMP1 had no significant change during the period of observation. The results indicate that LeY may play a role of regulating the expression and secretion of MMP of mouse embryo at pre-implantation stage, by controlling the expression of MMP2 gene and MMP9 gene.

Animals↗

PubMed2004

Mechanisms of abruption-induced premature rupture of the fetal membranes: Thrombin enhanced decidual matrix metalloproteinase-3 (stromelysin-1) expression.

OBJECTIVE: The aim of the study was to evaluate thrombin and progestin effects on matrix metalloproteinase-3 expression in term decidual cells as a mechanism of abruption-related preterm delivery. STUDY DESIGN: Decidual cells were isolated by standard techniques, purified to homogeneity, grown to confluence, and passaged. Cultures were primed with 10 (-8) M estradiol or estradiol plus 10 (-7) progestin and then incubated in a defined medium with corresponding steroid(s) plus or minus thrombin or the protease-activated thrombin receptor-1 agonist for 24 hours. Secreted matrix metalloproteinase-3 levels were assessed by enzyme-linked immunosorbent assay, and immunoblotting and messenger RNA levels were measured by Northern blotting and quantitative reverse transcription-polymerase chain reaction. RESULTS: Immunoreactive matrix metalloproteinase-3 levels were inhibited 66% by estradiol plus progestin versus estradiol ( P < .05). Thrombin elicited a dose-dependent reversal in this progestin inhibition, producing a 2.5-fold increase at 2.5 U/mL ( P < .05) that attained 33% of matrix metalloproteinase-3 levels in parallel incubations with estradiol plus thrombin. Protease-activated thrombin receptor-1 agonist mimicked 60% of thrombin-enhanced matrix metalloproteinase-3 output. Immunoblotting validated the enzyme-linked immunosorbent assay results. Northern blotting and quantitative reverse transcription-polymerase chain reaction demonstrated corresponding effects on steady-state messenger RNA levels. CONCLUSION: Abruption-generated thrombin promotes preterm delivery by mediating fetal membrane extracellular matrix degradation via enhanced decidual cell matrix metalloproteinase-3 expression, whereas progesterone blunts this thrombin-induced effect.

Base Sequence↗

PubMed2001

Insulin-like growth factor (IGF)-II inhibition of endometrial stromal cell tissue inhibitor of metalloproteinase-3 and IGF-binding protein-1 suggests paracrine interactions at the decidua:trophoblast interface during human implantation.

In human pregnancy, insulin-like growth factor (IGF)-II messenger RNA (mRNA) is expressed at the maternal-fetal interface exclusively by the placental trophoblast. Highest levels are expressed by the invading extravillous trophoblasts, which also secrete matrix metalloproteinases as they degrade the decidual extracellular matrix. In contrast, the maternal decidua expresses high levels of IGF-binding protein (IGFBP)-1 and tissue inhibitors of matrix metalloproteinase (TIMPs), both of which inhibit trophoblast invasiveness in vitro. The present study investigated the hypothesis that IGF-II may serve as a paracrine modulator of maternal restraints on invasion, by examining its effects on TIMP-3 and IGFBP-1 expression by decidualized endometrial stromal cells. Human endometrial stromal cells were decidualized in vitro with progesterone (P), after which 0-130 nM IGF-II and IGF analogs were added. IGFBP-1 in conditioned medium was assayed by immunoradiometric assay. In addition, Northern analyses were conducted using a PCR-generated 421-bp complementary DNA (cDNA) fragment corresponding to nucleotides 132-553 of the human TIMP-3 cDNA, and a 934-bp EcoRI fragment of the human IGFBP-1 cDNA. TIMP-3 mRNA transcripts of 2.2, 2.5, and 4.4 kilobases were detected in decidualized stromal cells not treated with IGF-II, but not detected in nondecidualized stromal cells, consistent with its known induction upon decidualization and in response to P. In decidualized stromal cells, IGF-II and Des(1-6) IGF-II, an analog with reduced affinity for IGFBPs, caused a dose-dependent inhibition of TIMP-3 mRNA expression. Long R(3) IGF-I, an IGF analog with minimal affinity for IGFBPs, also significantly inhibited (79 +/- 0.3%) TIMP-3 mRNA expression in these cells at 6 nM. Decidualized stromal cells secreted IGFBP-1 and expressed a 1.5-kilobase IGFBP-1 transcript, which was not detected in nondecidualized cells, consistent with its known induction upon decidualization and in response to P. IGF-II caused a dose-dependent inhibition of IGFBP-1 mRNA expression and protein secretion in decidualized stromal cells when added in molar excess of endogenous IGFBP-1 levels, with virtually complete inhibition at higher concentrations of IGF-II (65 and 130 nM). By comparison, Long R(3) IGF-I inhibited IGFBP-1 expression with a 50% effective dose of 0.2-0.4 nM. These data suggest that the invading trophoblast has the capacity, via IGF-II, to inhibit maternal restraints on trophoblast invasiveness by regulating decidual TIMP-3 and IGFBP-1.

Cells, Cultured↗

PubMed2006

Macrophage matrix metalloproteinase-2/-9 gene and protein expression following adhesion to ECM-derived multifunctional matrices via integrin complexation.

Macrophages are commonly observed at the biomaterial-tissue interface and interact with the extracellular matrix (ECM) mainly by integrin receptors to play a critical role in ECM turnover by secreting matrix metalloproteinases (MMPs). To investigate beta1 and beta3 containing integrin-mediated adhesion and subsequent MMP-2/-9 protein and gene expression in human blood-derived monocytes, biofunctional peptides immobilized onto flexible polyethylene glycol (PEG) arms were grafted onto a gelatin-based interpenetrating network (IPN). Adherent monocyte density was dramatically greater in the presence of RGD immobilized onto flexible PEG arms of the gelatin-based IPN. Pretreatment of monocytes with either anti-integrin beta1 or beta3 led to a significant decrease in adherent cell density on RGD-PEG-grafted IPNs. MMP-2 and MMP-9 protein and MMP-9 mRNA expression increased in the presence of IPNs initially, independent of ligand identity. Anti-integrin beta1 or beta3 antibody pretreatment of monocytes led to a general decrease in MMP-2/-9 protein expression. These results demonstrate the importance of beta1 and beta3 containing integrins in mediating monocyte adhesion onto RGD immobilized onto flexible PEG arms of the IPN. The results also reveal that MMP-2/-9 protein and gene expression is influenced by the presence of gelatin and not the ligands immobilized on the PEG arms of the IPN.

Cell Adhesion↗