Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Atrophoderma is a rare dermal disorder characterized by a patchy distribution of areas apparently devoid of elastic fibers. Skin fibroblast cultures were established from the normal and affected dermis of a patient with this disorder. Human tropoelastin was identified in culture medium by use of electroblotting and anti-elastin antisera. An enzyme-linked immunosorbent assay was used to establish that significantly less elastin accumulated in the media of cultured cells from lesional fibroblasts over a 3-d period. Since elastin biosynthesis in most tissues is under pretranslational control, molecular hybridization to a nick-translated genomic elastin probe was performed; however, elastin messenger RNA levels were equivalent in both cell strains. Both strains produced less elastin than did normal skin fibroblasts. Extracellular proteolysis of elastin was evaluated as a possible mechanism. Elastase activity was increased and porcine tropoelastin was degraded four times faster, on a per-cell basis, in lesional fibroblast cultures than in cells derived from an unaffected site. The two cell strains exhibited no significant differences in collagen production or collagenase activity. These results are the first demonstration of elastin production by cultured human skin fibroblasts, and they suggest that the primary defect in atrophoderma may be a result of enhanced degradation of newly synthesized elastin precursors.
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PMID:Demonstration of elastin gene expression in human skin fibroblast cultures and reduced tropoelastin production by cells from a patient with atrophoderma. 397 23

1. Insoluble elastin has been prepared by several different methods from adult bovine and calf ligamentum nuchae. Highly purified tropoelastin has been prepared from copper-deficient porcine aorta. 2. Amino acid analyses indicated that all preparations, except that obtained from calf ligamentum nuchae by using an EDTA extraction followed by collagenase digestion (preparation E6), were typical of pure elastin having high concentrations of hydrophobic and low concentrations of hydrophilic amino acids. Preparation E6 was found to contain approx. 40% collagen. 3. The determination and composition of the carbohydrates associated with these preparations is reported. With the exception of preparation E6, the insoluble elastins contained only trace amounts of neutral sugars (0.13-0.35%, w/w) and amino sugars (0.01-0.06%, w/w). The porcine tropoelastin contained virtually no carbohydrate. 4. The results suggest that carbohydrate analyses can yield valuable information about the purity of elastin preparations.
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PMID:Carbohydrate content of insoluble elastins prepared from adult bovine and calf ligamentum nuchae and tropoelastin isolated from copper-deficient porcine aorta. 500 Jun 44

We have investigated potential mechanisms for blister formation by assaying proteolytic enzymes in the blister fluids of patients with various bullous diseases. Blister fluids were obtained from patients with dermatitis herpetiformis (DH), bullous pemphigoid (BP), chronic bullous disease of childhood (CBDC), and pemphigus vulgaris (PV). The cells were recovered by centrifugation, and the supernatants as well as the cell pellets were assayed first for collagenase activity using [3H]proline-labeled type I collagen as substrate. Collagenase activity could be detected in most cases with DH, BP, and CBDC, while no activity was found in 2 cases of PV or in 5 control blister fluids obtained from suction blisters induced in healthy control subjects. Elastase activity was assayed in the same blister fluids by using a synthetic substrate succinyl-(L-alanyl)3-paranitroanilide or soluble [14C]valine-labeled tropoelastin. High levels of elastase activity were present in all DH patients, while lower, but clearly detectable, levels were found in BP, CBDC, and PV. The enzyme activity in BP was inhibited by Na2EDTA, but not by phenylmethylsulfonyl fluoride (PMSF), and Ca2+ stimulated the activity, suggesting that the enzyme in BP was a metalloproteinase. In cell-free supernatants of the DH blister fluids, the elastase activity was markedly decreased by PMSF, indicating that most of the enzyme activity was due to a serine protease. The cells recovered from DH blister fluids also contained high levels of elastase activity which could be inhibited by PMSF but not by Na2EDTA. Thus, in DH, the elastase activity is probably derived from polymorphonuclear leukocytes abundantly present in the lesions. The results indicate that active proteases are present in the blister fluids of skin diseases, and they may play a mechanistic role in the blister formation by degrading connective tissue components of the dermis and the dermal-epidermal junction.
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PMID:Demonstration of collagenase and elastase activities in the blister fluids from bullous skin diseases. Comparison between dermatitis herpetiformis and bullous pemphigoid. 630 88

We have established that treatment of cultured human skin fibroblasts with tropoelastin or with heterogenic peptides, obtained after organo-alkaline or leukocyte elastase hydrolysis of insoluble elastin, induces a high expression of pro-collagenase-1 (pro-matrix metalloproteinase-1 (pro-MMP-1)). The identical effect was achieved after stimulation with a VGVAPG synthetic peptide, reflecting the elastin-derived domain known to bind to the 67-kDa elastin-binding protein. This clearly indicated involvement of this receptor in the described phenomenon. This notion was further reinforced by the fact that elastin peptides-dependent MMP-1 up-regulation has not been demonstrated in cultures preincubated with 1 mm lactose, which causes shedding of the elastin-binding protein and with pertussis toxin, which blocks the elastin-binding protein-dependent signaling pathway involving G protein, phospholipase C, and protein kinase C. Moreover, we demonstrated that diverse peptides maintaining GXXPG sequences can also induce similar cellular effects as a "principal" VGVAPG ligand of the elastin receptor. Results of our biophysical studies suggest that this peculiar consensus sequence stabilizes a type VIII beta-turn in several similar, but not identical, peptides that maintain a sufficient conformation to be recognized by the elastin receptor. We have also established that GXXPG elastin-derived peptides, in addition to pro-MMP-1, cause up-regulation of pro-matrix metalloproteinase-3 (pro-stromelysin 1). Furthermore, we found that the presence of plasmin in the culture medium activated these MMP proenzymes, leading to a consequent degradation of collagen substrate. Our results may be, therefore, relevant to pathobiology of inflammation, in which elastin-derived peptides bearing the GXXPG conformation (created after leukocyte-dependent proteolysis) bind to the elastin receptor of local fibroblasts and trigger signals leading to expression and activation of MMP-1 and MMP-3, which in turn exacerbate local connective tissue damage.
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PMID:Conformational dependence of collagenase (matrix metalloproteinase-1) up-regulation by elastin peptides in cultured fibroblasts. 1108 20

In this review are presented the last new results of our research group dealing with the molecular structures (atomic level) of tropoelastin, elastin and elastin derived peptides studied by using essentially methods of bioinformatics (theoretical predictions and molecular modelling) linked to experimental circular dichroism spectroscopic studies. We already had characterized both the local secondary structure and some parts of the tertiary structure of the tropoelastin and elastin molecules (human, bovine...), by using either theoretical predictions (local secondary structure, linear epitopes...) and/or experimental data (optical spectroscopic methods: Raman scattering, infrared absorption, circular dichroism). Except the cross-linking regions which are in helical conformations, the whole tropoelastin structure displays a lot of beta-reverse turns which usually belong to irregular structures in proteins. These turns play a key role in other regularly structures orientation (alpha-helix, beta-strand), thus they are very important in the native protein 3D architecture. It is particularly true for human tropoelastin, because its sequence is rich in glycines and prolines, and these residues are frequently met in beta-turns (a beta-turn is made of four consecutive residues which are stabilized by an hydrogen bond). Several types of beta-turns can be defined with the dihedral angles values phi and psi of the two central residues. Thus, by using a very recent updated set of propensities for the amino acid residues to belong to given types of reverse beta-turns (extracted from a reference set of known 3-D structures of globular proteins), we have determined, (by using our home made software COUDES), for all possible tetrapeptides of the human tropoelastin sequence, the distribution and the characterization of the possible type of turns. Thus, it is shown that the locations and/or the types of these reverse beta-turns reveal a regularity and are not all random. This confirms our hypothesis that intra-molecular elasticity of tropoelastin could be explained by the possibility of transitions between conformations involving short beta-strands and beta-turns. This result is of great interest in the construction (by using molecular biology) of elastic biomaterials derived from the elastin sequence (particularly, the elastin derived peptides corresponding to the sequence exon 21--(exon 24--exon 24...). Our study permit also to predict the conformations of specific elastin derived peptides which could have interesting biological activity. Peptides resulting from the degradation of elastin, the insoluble polymer of tropoelastin and responsible for the elasticity of vertebrate tissues, can induce biological effects and notably the regulation of matrix metalloproteinases (MMP-s) activity. Recently, it was proposed that some elastin derived hexapeptides resulting from circular permutations of VGVAPG (a three fold repetition sequence in exon 24 of human tropoelastin) possess MMP-1 production and activation regulation properties. This effect depends on the presence of the tropoelastin specific membraneous receptor 67 KDa EBP (Elastin Binding Protein). Our results obtained by using both circular dichroism spectroscopy and linear predictions confirmed the hypothesis of a structure dependent mechanism with a possibly occurring type VIII beta-turn on the first four residues of the GXXPG sequence consensus which is only present among all active peptides. Thus, we have performed extensive molecular dynamics studies, in both implicit and explicit solvent, on these active and inactive elastin derived hexapeptides. Using our own analysis method of pattern recognition of the types of the beta-reverse-turns followed during the molecular dynamics trajectory, we found that active and inactive peptides effectively form two well distinct conformational groups in which active peptides preferentially adopt conformation close to type VIII GXXP (beta-reverse-turn. The structural role of the C terminal G residue could also be explained. Additional molecular simulations on (VGVAPG)2 and (VGVAPG)3 show the formation of two or three GXXP tetrapeptides adopting a structure close to type VIII beta-reverse-turn, suggesting a local conformational preference for this motif. This observation of a specific structural single and/or repeated motif is in agreement with the circular dichroism spectra of the involved (VGVAPG)1, (VGVAPG)2 and (VGVAPG)3 peptides and then it can be proposed that their biological activities have to be linear. The final aim of this type of work is to understand more about the sequence/structure/function/activity relationships of those structured peptides in order to propose specific sequences (corresponding to specific structures) for best biological activity results.
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PMID:[A turning point in the knowledge of the structure-function-activity relations of elastin]. 1172 5

HT-1080 fibrosarcoma cells express at their plasma membrane the elastin-binding protein (EBP). Occupancy of EBP by elastin fragments, tropoelastin or XGVAPG peptides was found to trigger procollagenase-1 (proMMP-1) overproduction by HT-1080 cells at the protein and enzyme levels. RT-PCR analysis indicated that elastin peptides did not modify the MMP-1 mRNA steady state levels, suggesting the involvement of a post-transcriptional mechanism. We previously reported that binding of elastin peptides to EBP induced other matrix metalloproteinases (MMP-2 and MT1-MMP) expression. Since those peptides were here found to also accelerate the secretion of urokinase from HT-1080 cells, culture medium was supplemented with plasminogen together with elastin peptides at aims to induce or potentiate MMPs activation cascades. In such conditions, plasmin activity was generated and exacerbate proMMP-1 and proMMP-2 activation. As a consequence, elastin peptides and plasminogen-treated HT-1080 cells displayed a significant type I collagen matrix invasive capacity.
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PMID:Cumulative influence of elastin peptides and plasminogen on matrix metalloproteinase activation and type I collagen invasion by HT-1080 fibrosarcoma cells. 1196 74

Synthesis of extracellular matrix (ECM) proteins and their degradation by matrix metalloproteinases (MMP) are part of the dermal remodeling resulting from chronic exposure of skin to ultraviolet radiation (UVR). We have compared two alternative mechanisms for these responses, namely, a direct mechanism in which UV-B or UV-A is absorbed by fibroblasts and an indirect mechanism in which cytokines, produced in skin in response to UVR, stimulate production of the ECM proteins and MMP. These studies were carried out on human dermal fibroblasts grown in contracted, free-floating 9 day old collagen gels as a dermal equivalent. Synthesis of tropoelastin, collagen, fibrillin, MMP-1, -2, -3 and -9 and tissue inhibitors of metalloproteinases (TIMP)-1 and -2 were measured. Tropoelastin, collagen and fibrillin levels were stable between days 4 and 10, and MMP and TIMP decreased by day 10. Neither UV-B (2.5-50 mJ/cm2) nor UV-A (2-12 J/cm2) altered synthesis of ECM proteins, but UV-A increased MMP-1 and -3 production. Tropoelastin synthesis increased in response to transforming growth factor-beta1 (5 ng/mL) treatment. Both interleukin-1beta and tumor necrosis factor-alpha (10 ng/mL) decreased fibrillin messenger RNA levels but increased MMP-1, -3 and -9 synthesis markedly. Collagen synthesis was not modulated by UV-B, UV-A or cytokine treatment. These results indicate that certain cytokines may have greater effects on production of ECM proteins and MMP than absorption of UV-B and UV-A by fibroblasts grown in dermal equivalents and suggest that the former pathway may play a role in the dermal remodeling in photoaged skin.
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PMID:Effects of UVR and UVR-induced cytokines on production of extracellular matrix proteins and proteases by dermal fibroblasts cultured in collagen gels%. 1497 20

To investigate the effects of topically applied 17beta-estradiol on the expression of extracellular matrix proteins in aged human skin, 17beta-estradiol (0.01%) and its vehicle (70% propylene glycol, 30% ethanol) were applied to aged (68-82 y, eight females and five males) human buttock skin under occlusion for 2 wk (three times per week). Topical 17beta-estradiol was found to increase the expression of type 1 procollagen mRNA and protein significantly in human aged skin in vivo. In addition, metalloproteinase (MMP-1 protein levels were reduced by topical 17beta-estradiol. The expressions of TGF-beta1, TGF-beta type II receptor, and Sma and Mad related (Smad)3 were increased by topical 17 beta-estradiol in aged human skin, and TGF-beta1 neutralizing antibody inhibited 17beta-estradiol-induced procollagen synthesis in cultured fibroblasts. We also found that the expressions of tropoelastin and fibrillin-1 mRNA and protein, and elastic fibers in aged skin were also increased by topical 17beta-estradiol. Topical 17beta-estradiol also increased keratinocyte proliferation and the epidermal thickness in aged human skin. We also observed the same effects of topical 17beta-estradiol in young skin. In conclusion, our results suggest that topical 17beta-estradiol treatment may improve the cutaneous function of aged human skin by improving the connective tissue and increasing epidermal thickness.
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PMID:Topical application of 17beta-estradiol increases extracellular matrix protein synthesis by stimulating tgf-Beta signaling in aged human skin in vivo. 1595 89

Skin aging can be attributed to photoaging (extrinsic) and chronological (intrinsic) aging. Photoaging and intrinsic aging are induced by damage to human skin attributable to repeated exposure to ultraviolet (UV) irradiation and to the passage of time, respectively. In our previous report, eicosapentaenoic acid (EPA) was found to inhibit UV-induced matrix metalloproteinase-1 (MMP-1) expression in human dermal fibroblasts. Therefore, we investigated the effects of EPA on UV-induced skin damage and intrinsic aging by applying EPA topically to young and aged human skin, respectively. By immunohistochemical analysis and Western blotting, we found that topical application of EPA reduced UV-induced epidermal thickening and inhibited collagen decrease induced by UV light. It was also found that EPA attenuated UV-induced MMP-1 and MMP-9 expression by inhibiting UV-induced c-Jun phosphorylation, which is closely related to UV-induced activator protein-1 activation, and by inhibiting JNK and p38 activation. EPA also inhibited UV-induced cyclooxygenase-2 (COX-2) expression without altering COX-1 expression. Moreover, it was found that EPA increased collagen and elastic fibers (tropoelastin and fibrillin-1) expression by increasing transformin growth factor-beta expression in aged human skin. Together, these results demonstrate that topical EPA has potential as an anti-skin-aging agent.
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PMID:Photoprotective and anti-skin-aging effects of eicosapentaenoic acid in human skin in vivo. 1646 81

Prematurely born infants who require oxygen therapy often develop bronchopulmonary dysplasia (BPD), a debilitating disorder characterized by pronounced alveolar hypoplasia. Hyperoxic injury is believed to disrupt critical signaling pathways that direct lung development, causing BPD. We investigated the effects of normobaric hyperoxia on transforming growth factor (TGF)-beta and bone morphogenetic protein (BMP) signaling in neonatal C57BL/6J mice exposed to 21% or 85% O(2) between postnatal days P1 and P28. Growth and respiratory compliance were significantly impaired in pups exposed to 85% O(2), and these pups also exhibited a pronounced arrest of alveolarization, accompanied by dysregulated expression and localization of both receptor (ALK-1, ALK-3, ALK-6, and the TGF-beta type II receptor) and Smad (Smads 1, 3, and 4) proteins. TGF-beta signaling was potentiated, whereas BMP signaling was impaired both in the lungs of pups exposed to 85% O(2) as well as in MLE-12 mouse lung epithelial cells and NIH/3T3 and primary lung fibroblasts cultured in 85% O(2). After exposure to 85% O(2), primary alveolar type II cells were more susceptible to TGF-beta-induced apoptosis, whereas primary pulmonary artery smooth muscle cells were unaffected. Exposure of primary lung fibroblasts to 85% O(2) significantly enhanced the TGF-beta-stimulated production of the alpha(1) subunit of type I collagen (Ialpha(1)), tissue inhibitor of metalloproteinase-1, tropoelastin, and tenascin-C. These data demonstrated that hyperoxia significantly affects TGF-beta/BMP signaling in the lung, including processes central to septation and, hence, alveolarization. The amenability of these pathways to genetic and pharmacological manipulation may provide alternative avenues for the management of BPD.
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PMID:Hyperoxia modulates TGF-beta/BMP signaling in a mouse model of bronchopulmonary dysplasia. 1707 23


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