Biomedical subjects

T Sorsa

Publications and source records attributed to T Sorsa.

At least 127 records · Page 7Linked to original sources

PubMed1998

Inhibition of MMP synthesis by doxycycline and chemically modified tetracyclines (CMTs) in human endothelial cells.

Doxycycline is a commonly used broad-spectrum antibiotic. Recently, it has been shown that it also inhibits the activity of mammalian collagenases and gelatinases, an activity unrelated to its antimicrobial efficacy. In this study, we show that doxycycline not only inhibits MMP-8 and MMP-9 (gelatinase B) activity, but also the synthesis of MMPs in human endothelial cells. Doxycycline (50 microM) completely inhibited the phorbol-12-myristate-13-acetate (PMA)-mediated induction of MMP-8 and MMP-9, as measured by Western blotting and gelatin zymography, respectively. The inhibition was also observed at the mRNA level. No effect was observed on the expression of MMP-2 and of the MMP inhibitors TIMP-1 and TIMP-2. Chemically modified tetracyclines (CMTs) showed an inhibition similar to that of doxycycline, albeit less efficient. These observations demonstrate that endothelial cells display a specific regulation of MMPs, which may have implications for the pharmaceutical interaction in angiogenesis and angiogenesis-related diseases.

Anti-Bacterial Agents↗

PubMed1998

Tetracyclines inhibit connective tissue breakdown by multiple non-antimicrobial mechanisms.

A seminal experiment involving a germ-free rat model of connective tissue breakdown (followed soon thereafter by a series of in vitro studies) identified an unexpected non-antimicrobial property of tetracyclines (TCs). This ability of TCs to inhibit matrix metalloproteinases (MMPs) such as collagenase was found to reflect multiple direct and indirect mechanisms of action, and to be therapeutically useful in a variety of dental (e.g., adult periodontitis) and medical (e.g., arthritis, osteoporosis, cancer) diseases. The site on the TC molecule responsible for its MMP-inhibitory activity was identified which led to the development of a series of chemically modified non-antimicrobial analogs, called CMTs, which also have therapeutic potential but do not appear to induce antibiotic side-effects. Longitudinal double-blind studies on humans with adult periodontitis have demonstrated that a sub-antimicrobial dose of doxycycline (previously reported to suppress collagenase activity in the periodontal pocket) is safe and effective and has recently been approved by the FDA as an adjunct to scaling and root planing.

Animals↗

PubMed1998

The effects of chemically modified tetracyclines (CMTs) on human keratinocyte proliferation and migration.

Chemically modified non-antimicrobial tetracycline derivatives and other low-molecular-weight synthetic matrix metalloproteinase (MMP) inhibitors inhibited keratinocyte migration. Since 72-kDa gelatinase-A (MMP-2) was the major gelatinase in our culture conditions, the results suggest that this MMP may be important in the regulation of keratinocyte mobility. On the other hand, we measured only gelatinase activities (MMP-2 and -9) present in culture medium, and therefore the results do not reveal how the inhibitors affect other MMPs as well as MMP levels close to the cell membranes. Overall, CMTs were found to be efficient in the inhibition of keratinocyte migration.

Cell Division↗

PubMed1998

High levels of expression of collagenase-3 (MMP-13) in pathological conditions associated with a foreign-body reaction.

A foreign-body-type host response can contribute to the induction and release of collagenolytic tissue-destructive enzymes of pathogenetic significance. Our aim was to analyse collagenase-3 in two conditions with putative involvement of foreign-body reactions. Synovial membrane-like tissue samples were obtained from cases of aseptic loosening of a total hip replacement (THR) and osteoarthritis (OA). The reverse transcription polymerase chain reaction (RT-PCR) disclosed that all the samples from patients contained collagenase-3 mRNA compared with only three out of ten control samples. The identity of the RT-PCR amplification product was confirmed by nucleotide sequencing. Immunohistochemical staining showed that collagenase-3 was present in endothelial cells, macrophages and fibroblasts, including those found in the synovial lining. This finding was confirmed by avidin-biotin-peroxidase complex-alkaline phosphatase-anti-alkaline phosphatase double staining and the specificity of the staining by antigen preabsorption using recombinant human collagenase-3. Collagenase-3 was released into the extracellular space and thus found in the synovial fluid in all patient samples as shown by Western blotting. The similar extent of collagenase-3 expression in aseptic loosening and OA compared with the low expression in control synovial membrane suggests involvement of a similar, foreign-body-based pathogenetic component in both. Comparative analysis of collagenase-3 and of foreign particles indicates that paracrine factors rather than phagocytosis per se are responsible for the induction of collagenase-3. We suggest that due to its localisation and substrate specificity, collagenase-3 may play a significant pathogenetic role in accelerating tissue destruction in OA and in aseptic loosening of a THR.

Adult↗

PubMed1998

Mast cells in bronchiectasis.

Bronchiectasis (BE) is a chronic severe inflammatory lung disease characterized by frequent bacterial infections and polymorphonuclear neutrophil-dominated inflammatory reaction. We have attempted to elucidate the role of mast cells (MCs) in BE lung inflammation by measuring in the bronchoalveolar lavage fluid (BALF) the MC-derived tryptase levels by radioimmunoassay and immunoblotting and also by measuring the tryptase-like activities in 36 BE patients and in 14 healthy controls. The amount of MC in the lung tissue was assessed by immunohistochemical staining of resected lung tissue samples. Based on the clinical and radiological parameters the patients were divided into subgroups according to the severity of the disease. The MC tryptase concentrations (microg/L; median (range)) in BALF of BE patients were higher compared to healthy controls (4.7(1.4-20.1) and 2.0 (0.1-3.5), respectively, P < 0.01). Tryptase concentrations in the groups of mild, moderate and severe BE were 3.8 (0.9-10.8), 4.3 (3.0-12.6) and 9.6 (1.2-20.1), respectively. All the values differed significantly from those observed in the healthy controls. The tryptase-like activities (nmol/sec/L) in BE patients were also markedly increased (174 (31-2874)) compared to healthy controls (28 (9-45) P < 0.0001). The tryptase-like activities in the patient subgroups were 45 (36-598) in mild, 91 (31-1437) in moderate and 1336 (37-2874) in severe BE. Again, all values differed significantly from those observed in the healthy controls. Moreover, immunoblot experiments disclosed the most intensive immunoreactivity of the 27.5 kD tryptase monomer in BALF of patients with severe BE followed by weaker immunoreactivity in groups of moderate and mild BE and in healthy controls. No significant difference could be observed in the amount of tryptase-positive cells between BE patients and controls. However, the presence of degranulated MCs was more evident in BE lung tissue. Significant correlation could also be observed between the degree of activation of latent procollagenase and tryptase concentration (r = 0.8, P = 0.0004) in BALF of individual BE patients. The observed strong correlation between tryptase levels and disease severity suggests that MCs may be involved in the inflammatory reaction in the BE lung. Importantly, the high levels of tryptase, observed also in patients with mild BE, suggests that activation of and proteinase release from MCs may be one of the reasons for the perpetuation of tissue injury even during the clinically quiescent periods in BE.

Adult↗

PubMed1998

Functional sites of chemically modified tetracyclines: inhibition of the oxidative activation of human neutrophil and chicken osteoclast pro-matrix metalloproteinases.

OBJECTIVE: We studied the relative ability of 6 different chemically modified non-antimicrobial analogs of tetracycline (CMT) to inhibit human and chicken matrix metalloproteinases (MMP) in vitro. The ability of tetracycline and its analogs to inhibit MMP appears to depend on the Ca++/Zn++ binding site at C11 (carbonyl oxygen) and C12 (OH group) of the molecule, which is lacking in CMT-5, the pyrazole derivative of tetracycline. This significant property of CMT-5 was used to differentiate between the effects of CMT on already active MMP versus the oxidative activation of latent MMP (pro-MMP). METHODS: Cultured chicken osteoclast conditioned medium and purified human neutrophil progelatinase (MMP-9) and pro-collagenase (MMP-8) were assayed for proteinase activities using gelatin and collagen, respectively. The pro-MMP were activated either by preincubation with 1 mM aminophenylmercuric acetate (APMA) or 100 microM sodium hypochlorite (NaOCI). CMT were added either to the preincubation mixtures together with NaOCl or after activation of pro-MMP with NaOCl. RESULTS: All CMT tested, except CMT-5, inhibited APMA or NaOCl activated pro-MMP. However, CMT-5 (like the other CMT), inhibited the oxidative activation of pro-MMP by NaOCl when added together by scavenging the reactive oxygen species. The degradation of type-I collagen by chicken osteoclast conditioned medium was probably due to MMP-2 and/or MMP-13. CONCLUSION: Oxidative activation of pro-MMP may be crucial during soft tissue/bone destruction in the inflammatory diseases, including the arthritides. Our results indicate that the Ca++/Zn++ binding site of CMT is not essential for inhibition of the oxidative activation of pro-MMP.

Animals↗

PubMed1998

Activation of periprosthetic connective tissue in aseptic loosening of total hip replacements.

In aseptic loosening of initially well inserted total hip prostheses, implant wear debris and cyclic mechanical loading lead to a foreign body type of chronic inflammatory reaction, then to osteolysis, and finally to loosening of the implant. In the present work the reactive and adaptive changes of the periprosthetic tissues and pseudojoint were characterized by analysis of the local cell proliferation. Immunohistochemical demonstration of proliferating cells was performed by application of affinity purified rabbit antihuman Ki-67 antibodies to periprosthetic tissues obtained from revision operations for loose total hip prostheses. The fibrous areas and, in particular, the cell rich, vascular areas of the interface tissue (between implant and bone) and the pseudocapsule around aseptically loosened implants contained higher numbers of proliferating cells than the tissues around well fixed implants. In addition, the pseudosynovial lining occasionally contained some Ki-67 positive proliferating cells. Somewhat surprisingly, proliferating vascular endothelial cells were relatively rare. These findings suggest that reactive (interface tissues) and adaptive (pseudojoint and capsule formed around the artificial joint) tissue changes in loosening total hip prostheses comprise proliferation of local fibroblastlike cells. It is concluded that periprosthetic tissues of the loosened total hip prosthesis represent activated mesenchymal tissue.

Adult↗

PubMed1998

Expression of stem cell factor (SCF) and SCF receptor (c-kit) in synovial membrane in arthritis: correlation with synovial mast cell hyperplasia and inflammation.

OBJECTIVE: Stem cell factor (SCF), the ligand for the SCF receptor (c-kit) expressed on precursors and mature mast cells (MC), is a major agonist for human MC (e.g., SCF induces MC development, chemotaxis, activation, proliferation of MC precursors, mediates MC adhesion, and changes MC releasability). We investigated expression of SCF and c-kit in synovial membrane with particular reference to the mechanism of local MC hyperplasia and inflammation in arthritis. METHODS: We conducted single and double labeling immunohistochemistry (ABC, APAAP, indirect immunofluorescence techniques) with antibodies to SCF, c-kit, MC tryptase, Ki-67 antigen (marker for proliferating cells), and CD68 (monocyte/macrophage marker). Synovial specimens analyzed were from 31 patients: traumatic arthritis (TrA, n=9), osteoarthritis (OA, n=12), and rheumatoid arthritis (RA, n=10). Control experiments were performed on human lung, skin, and buccal mucosa tissues, on the HMC-1 mast cell line, and isolated lung MC. Morphometry was performed by computerized image analysis. RESULTS: Synovial c-kit expression was found to be restricted to MC, whereas SCF is detected in synovial lining cells, stromal fibroblasts, monocyte/macrophages, endothelial cells, and in vascular basement membranes. SCF staining was localized to MC as well, but it was not possible to specify whether this represents SCF produced by or bound (via c-kit) to MC. In inflamed synovial membranes/areas, SCF was found to be redistributed into the extracellular matrix. Redistribution of SCF was accompanied by degranulation and/or accumulation of c-kit+ MC, the hyperplasia of which correlated positively with histologic inflammation/inflammatory cell densities, but did not appear to involve MC proliferation in situ. These findings appeared to be common for all the conditions (TrA, OA, RA) studied. CONCLUSION: In addition to the demonstration/characterization of SCF and c-kit protein expression in human synovium, results of this study suggest the hypothesis that, in arthritis, local mobilization of SCF may play a role in the development of synovial MC hyperplasia without inducing in situ proliferation of MC, and that the synovial SCF/MC c-kit system may contribute to the local nonspecific inflammatory response/arthritic flares in TrA, OA, and RA.

Adult↗

PubMed1997

Matrix metalloproteinase-8 is expressed in rheumatoid synovial fibroblasts and endothelial cells. Regulation by tumor necrosis factor-alpha and doxycycline.

Neutrophil collagenase (matrix metalloproteinase-8 or MMP-8) is regarded as being synthesized exclusively by polymorphonuclear neutrophils (PMN). However, in vivo MMP-8 expression was observed in mononuclear fibroblast-like cells in the rheumatoid synovial membrane. In addition, we detected MMP-8 mRNA expression in cultured rheumatoid synovial fibroblasts and human endothelial cells. Up-regulation of MMP-8 was observed after treatment of the cells with either tumor necrosis factor-alpha (10 ng/ml) or phorbol 12-myristate 13-acetate (10 nM). Western analysis showed a similar regulation at the protein level. The size of secreted MMP-8 was 50 kDa, which is about 30 kDa smaller than MMP-8 from PMN. Conditioned media from rheumatoid synovial fibroblasts contained both type I and II collagen degrading activity. However, degradation of type II collagen, but not that of type I collagen, was completely inhibited by 50 microM doxycycline, suggesting specific MMP-8 activity. In addition, doxycycline down-regulated MMP-8 induction, at both the mRNA and protein levels. Thus MMP-8 exerts markedly wider expression in human cells than had been thought previously, implying that PMN are not the only source of cartilage degrading activity at arthritic sites. The inhibition of both MMP-8 activity and synthesis by doxycycline provides an incentive for further studies on the clinical effects of doxycycline in the treatment of rheumatoid arthritis.

Arthritis, Rheumatoid↗

PubMed1997

Identification and structural and functional characterization of human enamelysin (MMP-20).

A cDNA encoding a new human matrix metalloproteinase (MMP) has been cloned from RNA prepared from odontoblastic cells. The open reading frame of the cloned cDNA codes for a polypeptide of 483 amino acids and is extensively similar to the sequence of recently described porcine enamelysin, suggesting that the isolated cDNA codes for the human homologue of this enzyme. Human enamelysin (MMP-20) has a domain organization similar to other MMPs, including a signal peptide, a prodomain with the conserved motif PRCGVPD involved in maintaining enzyme latency, a catalytic domain with a Zn-binding site, and a COOH-terminal fragment similar to the sequence of hemopexin. The calculated molecular mass of human enamelysin is about 54 kDa, which is similar to that of collagenases or stromelysins. However, this human MMP lacks a series of structural features distinctive of these subfamilies of MMPs. The full-length human enamelysin cDNA has been expressed in Escherichia coli, and the purified and refolded recombinant protein is able to degrade synthetic peptides used as substrates of MMPs, confirming that human enamelysin belongs to this family of proteases. Furthermore, the recombinant human enamelysin is able to degrade amelogenin, the major protein component of the enamel matrix. On the basis of its degrading activity on amelogenin, and its highly restricted expression to dental tissues, we suggest that human enamelysin plays a central role in the process of tooth enamel formation. Finally, we have found that the human enamelysin gene (MMP-20) maps to chromosome 11q22, clustered to at least seven other members of the MMP gene family.

Amino Acid Sequence↗

PubMed1997

Activation of type IV procollagenases by human tumor-associated trypsin-2.

Increased production of proteinases, such as matrix metalloproteinases (MMPs), is a characteristic feature of malignant tumors. Some human cancers and cell lines derived from them also express trypsinogen, but the function of the extrapancreatic trypsin has remained unclear. In this study we cloned and sequenced trypsinogen-2 cDNA from human COLO 205 colon carcinoma cells and characterized the ability of the enzyme to activate latent human type IV procollagenases (proMMP-2 and proMMP-9). As shown by cloning and N-terminal amino acid sequencing, the amino acid sequence of tumor-associated trypsin-2 is identical to that of pancreatic trypsin-2. We found that both pancreatic trypsin-2 and tumor cell-derived trypsin-2 are efficient activators of proMMP-9 and are capable of activating proMMP-9 at a molar ratio of 1:1000, the lowest reported so far. Human trypsin-2 was a more efficient activator than widely used bovine trypsin and converted the 92-kDa proMMP-9 to a single 77-kDa product that was not fragmented further. The single peptide bond cleaved by trypsin-2 in proMMP-9 was Arg87-Phe88. The generation of the 77-kDa species coincided with the increase in specific activity of MMP-9. In contrast, trypsin-2 only partially activated proMMP-2. Trypsin-2 cleaved the Arg99-Lys100 peptide bond of proMMP-2 generating 62-65-kDa MMP-2 species. Trypsin-2-induced proMMP-2 and -9 conversions were inhibited by tumor-associated trypsin inhibitor added either prior to or during activation indicating that proMMPs were not activated autocatalytically. Trypsin-2 also activated proMMPs associated with tissue inhibitor of matrix metalloproteinases, the complexes of which are thought to be the major MMP forms in vivo. The ability of human tumor cell-derived trypsin-2 to activate latent MMPs suggests a role for trypsin-2 in initiating the proteinase cascade that mediates tumor invasion and metastasis formation.

Amino Acid Sequence↗

PubMed1997

Proteolytic processing regulates receptor specificity and activity of VEGF-C.

The recently identified vascular endothelial growth factor C (VEGF-C) belongs to the platelet-derived growth factor (PDGF)/VEGF family of growth factors and is a ligand for the endothelial-specific receptor tyrosine kinases VEGFR-3 and VEGFR-2. The VEGF homology domain spans only about one-third of the cysteine-rich VEGF-C precursor. Here we have analysed the role of post-translational processing in VEGF-C secretion and function, as well as the structure of the mature VEGF-C. The stepwise proteolytic processing of VEGF-C generated several VEGF-C forms with increased activity towards VEGFR-3, but only the fully processed VEGF-C could activate VEGFR-2. Recombinant 'mature' VEGF-C made in yeast bound VEGFR-3 (K[D] = 135 pM) and VEGFR-2 (K[D] = 410 pM) and activated these receptors. Like VEGF, mature VEGF-C increased vascular permeability, as well as the migration and proliferation of endothelial cells. Unlike other members of the PDGF/VEGF family, mature VEGF-C formed mostly non-covalent homodimers. These data implicate proteolytic processing as a regulator of VEGF-C activity, and reveal novel structure-function relationships in the PDGF/VEGF family.

Amino Acid Sequence↗

PubMed1997

Salivary mucous glycoprotein MG1 in Sjögren's syndrome.

The aim of the study was to develop and apply a rapid method for the simultaneous analysis of the concentration and molecular weight of the human high-molecular weight mucin MG1 in small volumes of unprocessed saliva from healthy controls and from patients with Sjögren's syndrome (SS). In high performance liquid chromatography (HPLC) with a TSK 5000 PW size exclusion column, MG1 eluted with a retention time 10.6 min corresponding to a M(r) of 2 to 2.5 x 10(6). Molecular weight changes under various experimental conditions are compatible with the suggestion that the MG1 complex is composed of four 660 x 10(3) glycosylated subunits connected by disulphide bridges and associated with a 25-35 x 10(3) Da link protein. In SS the molecular weight of MG1 was normal and its concentration was high in resting (190 vs. 70 micrograms/ml, P = 0.001) but not in stimulated (46 vs. 48 micrograms/ml, P > 0.05) saliva; MG1 concentration in resting SS saliva did not vary in parallel with protein and the interindividual differences were considerable. Size exclusion HPLC is a rapid and reproducible method suitable for isolation and analysis of salivary MG1 from small volumes of unprocessed samples. The molecular weight or subunit structure of MG1 were not altered in SS. The high concentration of MG1 in resting saliva in SS, may be explained by the concentration effect, or alternatively by the low water retaining capacity, which may play an important pathogenic role in xerostomia of SS.

Chromatography, High Pressure Liquid↗

PubMed1997

Role of nitric oxide in Sjögren's syndrome.

OBJECTIVE: To measure levels of salivary nitrite (NO2-) and to localize nitric oxide synthases (NOS) in the labial salivary glands (LSGs) of patients with Sjögren's syndrome (SS). METHODS: NO2- was measured by the Griess reaction. LSGs were analyzed using NADPH-diaphorase histochemical and immunohistochemical studies to determine the constitutive NOS (neuronal [ncNOS] and endothelial [ecNOS]) and inducible NOS (iNOS) isoforms. RESULTS: The NO2- concentration (mean +/- SEM 307 +/- 51 microM versus 97 +/- 16 microM; P < 0.05) and output (166 +/- 46 nmoles/minute versus 37 +/- 7 nmoles/minute) were increased in SS patients compared with healthy control subjects. NADPH-diaphorase was found in some nerve fibers and endothelial cells, and, in SS, was found in myoepithelial, acinar, and ductal epithelial cells, but in only a few inflammatory cells. In SS, ncNOS-immunoreactive nerve fibers were sparse and ecNOS was found in a minority of the CD31-positive vascular endothelial cells and acinar cells, whereas iNOS was localized in myoepithelial, acinar, and ductal epithelial cells, often together with tumor necrosis factor alpha. CONCLUSION: Nitrite was found in normal human saliva. NO produced by ncNOS probably acts as a nonadrenergic, noncholinergic neurotransmitter, whereas that produced by ecNOS exerts a vasodilatory effect. SS patients had increased NO2- concentrations, with most of the superfluous salivary NO being produced not by the immigrant inflammatory cells, but rather, by the resident salivary gland cells. NO may contribute to inflammatory damage and acinar cell atrophy in SS.

Adult↗

PubMed1997

Matrix metalloproteinase 13 (collagenase 3) in human rheumatoid synovium.

OBJECTIVE: To show the eventual presence and extent of production of matrix metalloproteinase 13 (MMP-13, or collagenase 3) in rheumatoid synovial tissue samples and extracts, and to assess the inhibition characteristics of recombinant MMP-13. METHODS: Immunohistochemical avidin-biotin-peroxidase complex staining/morphometry was used to analyze MMP-13-positive cells in situ. Neutral salt extraction of synovial tissue, electrophoresis of the extract in different buffer systems, and Western blotting were also used. The inhibitory properties of doxycycline, clodronate, pamidronate, and D-penicillamine for recombinant enzyme were determined with a soluble type II collagen assay. RESULTS: MMP-13 was detected in fibroblast- and macrophage-like mononuclear cells in the synovial lining and stroma and in vascular endothelial cells. The overall expression of MMP-13 in these cells in the synovial stroma was high in rheumatoid arthritis (86 +/- 12%) compared with osteoarthritis (17 +/- 5%) patient samples (P = 0.0027). In a high-pH native electrophoresis gel, immunoreactivity to anti-MMP-1 and anti-MMP-13 were clearly separated, with anti-MMP-13-immunoreactive material migrating faster than anti-MMP-1-immunoreactive material. Finally, in contrast to MMP-1 and MMP-8, MMP-13 was found to be relatively resistant to the inhibitory effects of doxycycline and clodronate in vitro. CONCLUSION: Due to its localization in synovial tissue, its substrate profile, increased expression, and relative resistance to known MMP inhibitors, MMP-13 is suggested to play a major role in the pathogenesis of tissue destruction in rheumatoid arthritis.

Adult↗

PubMed1997

Macrophage-colony stimulating factor (M-CSF) is increased in the synovial-like membrane of the periprosthetic tissues in the aseptic loosening of total hip replacement (THR).

The aim of the study was to assess the eventual presence, cellular localization and extent of expression of the osteoclast activating cytokine M-CSF (CSF-1) in the periprosthetic tissues around loose total hip replacement (THR). Synovial-like membrane was obtained from the implant-to-bone interface and pseudocapsule from ten total hip revisions performed for aseptic loosening and compared to ten hip synovial tissue samples obtained from ten patients who had primary THR for osteoarthritis. Avidin-biotinperoxidase complex (ABC) and alkaline phosphatase-anti-alkaline phosphatase (APAAP) methods were used for staining and VIDAS image analysis for quantification. M-CSF was mainly produced by macrophages, which often contained wear particles, but also by some fibroblasts and vascular endothelial cells. The number of cells containing (per one mm2 tissue) clearly increased in the interface (1585 +/- 212; p < 0.01) and pseudocapsular (1456 +/- 248; p < 0.01) tissue compared to synovial tissue (543 +/- 118). The present findings suggest, that inflammatory foreign-body type of response enhances expression of M-CSF in cases of aseptic loosening of THR. M-CSF produced in the synovial-like membrane in the implant-bone interface may contribute to activation of osteoclasts in periprosthetic bone and thus to loosening.

Adult↗

PubMed1997

A matrix metalloproteinase inhibitor reduces bone-type collagen degradation fragments and specific collagenases in gingival crevicular fluid during adult periodontitis.

OBJECTIVE AND DESIGN: To determine whether an inhibitor of matrix metalloproteinases (MMPs), administered to human subjects in a dental school research clinic, can reduce bone-type collagen degradation fragments in oral inflammatory exudates containing excessive levels of collagenase. MATERIALS AND SUBJECTS: Gingival crevicular fluid (GCF) was collected from 18 subjects with adult periodontitis whose clinical findings (gingival inflammation, pocket depth, and bone loss on radiographs) predicted excessive MMP activity in their periodontal pockets. TREATMENT: One month before the baseline appointment, plaque and calculus were removed from the teeth by supra- and subgingival scaling. After collection of GCF from 8-12 pocket sites per subject and recording of clinical indices, 12 of the 18 subjects were treated with doxycycline at a low dosage (20 mg b.i.d.) known via an extensive literature to suppress mammalian MMP activity by a non-antimicrobial mechanism. The remaining 6 subjects were followed without drug treatment. METHODS: At the baseline, 1 and 2-month appointments, GCF samples were analyzed for ICTP. (carboxyterminal peptide, a pyridinoline-containing fragment of Type I collagen) and osteocalcin by radioimmunoassay, as well as collagenolytic enzyme activity and MMP species (Western blot). Statistical analyses were determined by ANOVA. RESULTS: GCF ICTP and functional collagenase activity (but not osteocalcin levels) were significantly reduced (p < 0.05) in the doxycycline-treated subjects at both 1 and 2 month evaluations: there was no such change in the non-treated subjects. Western blots revealed that neutrophil-type collagenase (MMP-8) was the predominant MMP; MMP-13, which has been associated with pathologic collagenolysis including bone resorption, was detected in human GCF for the first time and was more substantially reduced than MMP-8. CONCLUSION: This is the first demonstration in human subjects of the simultaneous reduction of excessive MMP activity with concomitant reduction in levels of collagen degradation fragments. The findings are potentially applicable to a wide variety of human diseases characterized by excessive collagenase activity.

Adult↗

PubMed1997

Inhibition of matrix metalloproteinase-1 by dichloromethylene bisphosphonate (clodronate).

Interstitial collagenase present in human jaw cyst extract and purified human fibroblast-type collagenase (MMP-1) were both efficiently inhibited in vitro by clodronate, an osteoactive, antiresorptive bisphosphonate. The IC50 of clodronate to inhibit MMP-1 is 150 microM. These findings suggest an extended and hitherto undescribed properties for clodronate/biphosphonates in prevention and treatment of tissue degradation in both bone and soft tissue destructive diseases.

Bone Diseases↗