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Characterization of interstitial collagenases in jaw cyst wall
Abstract
Neutral salt extracts of 14 specimens of jaw cysts were prepared. Histopathological analysis showed that the specimens consisted of 6 radicular cysts, 6 dentigerous cysts, 1 residual cyst, and 1 odontogenic keratocyst. One periapical granuloma, 1 dental follicle and a sample of clinically healthy oral mucosa were similarly processed and used as controls. Measurement of collagenase activity by monitoring the formation of specific degradation products of type I and II collagen in solution by SDS-PAGE demonstrated that all the cyst extracts contained collagenase, some of which was endogenously activated. Cyst wall collagenase preferably degraded type I over type II collagen, which suggests that the degradation was due to MMP-1 (matrix metalloproteinase-1) rather than the MMP-8 type. This was further supported by the doxycycline-inhibition profile of cyst collagenase, which was similar to that of MMP-1. Part of the cyst wall collagenase was in latent proenzyme form and probably derived, at least in part, from the newly synthesized intracellular collagenase pool. Latent cyst collagenase was efficiently activated with phenylmercuric chloride and to a lesser extent by gold (I) thioglucose and NaOCl. Western-blotting, using specific antibodies against collagenase from human polymorphonuclear neutrophilic leukocytes (MMP-8) and from fibroblasts (MMP-1), revealed a typical 55/45 kDa doublet; also MMP-8 in the latent 80 kDa form and fragmented to 65 kDa active species were found. These results suggest the presence of MMP-1 and, to a lesser extent, MMP-8 type collagenase in the cyst wall.(ABSTRACT TRUNCATED AT 250 WORDS)
... Recent studies involving the components of the connective tissue in KCOT revealed some resemblance with the stromal components of aggressive tumors such as ameloblastoma: high frequency of stromal myofibroblasts 38 , differences in the collagen fibers of the extracellular matrix 40 , high enzymatic activity with increased matrix metalloproteinases (MMPs) 41,42 and mast cell tryptase 43 , and increased expression of receptor activator of nuclear factor-kB (RANK), RANK ligand (RANKL), and osteoprotegerin (OPG). 44 Overall, a series of genetic and molecular mechanisms, not yet fully understood, appears to promote the development and progression of tumors via a multiple step mechanism. ...
... Enucleation (14) Marsupialization (2) 29 0 Forsell et al. 49 75 Enucleation (one piece) (28) Enucleation (several pieces) (41) Marsupialization (5) 18 56 60 Brondum et al. 50 44 Enucleation + osteotomy (32) Marsupialization + enucleation (12) 25 0 Eufinger et al. 51 49 ...
... Marsupialization (3) Enucleation (78) One piece (23) Curetage (31) Cystectomy (11) Carnoy (5) 100 39 13 29 27 20 Dammer et al. 41 38 Enucleation (32) Enucleation + Carnoy (2) Resection (0) 4,6 50 0 Chow 44 70 Enucleation (68) Enucleation + Carnoy (23) Resection (1) 8,8 4,3 0 Bataineh et al. 52 31 Resection (31) 0 Browne 53 Marsupialization (12) Enucleation (72) 25 23,6 Schmidt 54 26 Enucleation + liquid nitrogen 11,5 Stoelinga 45 80 ...
Keratocystic odontogenic tumor (KCOT), formerly referred to as odontogenic keratocyst, is a benign neoplasm of odontogenic origin which may present an aggressive and infiltrative behavior leading to high recurrence rates. A review of the various treatment modalities, ranging from simple enucleation to radical surgery is portrayed in relation to clinical, radiological, histopathological and molecular features. Although prognostic factors based on clinico-pathologic and immunohistochemical findings for determining the potential for recurrence of KCOT still remains unclear, its use for determining the potential for recurrence of KCOT after surgical treatment may become important to successfully manage this neoplasm's aggressive behavior. The key element for future management of KCOTs will probably be based on thorough knowledge of the biological basis of this tumor, thereby enabling a more tailored treatment approach.
... IL-1α, as noted above is thought to play a crucial role in the expansion of OKCs. After marsupialization and decompression, IL-1α and cytokeratin [10] disappear from the cyst lining. B-cell lymphoma-2 protein (Bcl-2) which protects cells from early death through apoptosis is found in cyst linings (as well as neoplasms) thus contributing to loss of control of cell proliferation. ...
... Recent studies have also high-lighted the importance of the epithelial -mesenchymal interaction in the expansion of the odontogenic cysts [9,10]. Collagen, responsible for maintaining the functional integrity of tissues including the odontogenic apparatus, is thought to play a role in pathogenesis and expansion of odontogenic cyst. ...
Large cystic lesions of the jaws are often marsupialized in order to reduce their size prior to further surgical treatment by enucleation with or without adjunctive procedures. Experience with this procedure is an initial period of rapid reduction in size followed by stabilization of the healing process when little or no decrease in cavity size occurs. At this stage enucleation is usually contemplated. Research in expansion of cysts has highlighted the role of myofibroblasts and cytokines expressed from the epithelial and stromal cells. The role of interstitial fluid flow and its effect on osteogenesis is well known. Marsupialization probably reduces the cystic cavity by multiple mechanisms such as contractility of myofibroblasts, decrease in bone-resorbing cytokines and increased osteoblastic activity. Application of negative pressure, physiological loading of bone, LIPUS and electromagnetic stimulation may positively influence various physiological processes which enhance healing.
... The various types of MMPs were studied by Western blotting with specific poly-and monoclonal antibodies. [28][29][30][31] Purified MMPs from human peripheral blood neutrophils as well as human gingival fibroblasts and keratinocytes (MMP-1, -2, -8, and -9) were used as described in earlier papers. [28][29][30][31][32][33] Recombinant MMPs (MMP-1, -3, -7, -12, -13, -14, and -20), kindly provided by Drs. ...
... [28][29][30][31] Purified MMPs from human peripheral blood neutrophils as well as human gingival fibroblasts and keratinocytes (MMP-1, -2, -8, and -9) were used as described in earlier papers. [28][29][30][31][32][33] Recombinant MMPs (MMP-1, -3, -7, -12, -13, -14, and -20), kindly provided by Drs. G. Murphy, V. Knäuper, and C. López-Otín were also used in the experiments. ...
Bisphosphonates are a group of drugs capable of inhibiting bone resorption, and are thus used for the treatment of bone diseases, such as Paget's disease, osteoporosis, and for bone metastases of malignant tumors. Their primary cellular target is considered to be the osteoclast. The molecular mechanisms responsible for the downregulation of bone resorption by bisphosphonates have remain unclear. We have discovered that various matrix metalloproteinases (MMPs) are inhibited in vitro by several bisphosphonates. This novel finding may, in part, explain the efficacy of bisphosphonates in their current indications in humans. In enzyme activity tests using purified and recombinant enzymes, we have observed the inhibition of MMP-1, -2, -3, -7, -8, -9, -12, -13, and -14 by clondronate, alendronate, pamidronate, zolendronate, nedrinate, and clodrinate. The IC50s range from 50 to 150 μM. We have also shown that clodronate can downregulate the expression of MT1-MMP protein and mRNA in several cell lines. Additionally, several bisphosphonates decrease the degree of invasion of malignant melanoma (C8161) and fibrosarcoma (HT1080) cells through artificial basement membrane (Matrigel) in cell cultures at IC50s of 50-150 μM and below. Having low toxicity and proven to be well tolerated after several years in human use, bisphosphonates have the potential to become one of the main MMP-inhibitors for MMP-related human soft and hard tissue-destructive diseases in the near future.
... [5,[7][8][9] Recent studies have focused on the importance of the epithelial-mesenchymal interactions in odontogenic cysts and have demonstrated that the expansion involves degradation of bone matrix and cell attachment to the ECM components. [15][16][17] Vedtofte et al, demonstrated that transplanted keratocyst epithelium in nude mice retained its typical histological appearance only when supported by its own stroma. Thus, it can be suggested that the biological behavior of odontogenic keratocyst is dependent not only on the epithelium but also on the underlying stroma. ...
... Thus, inflammatory cells could affect the arrangement of collagen. [15][16][17] In the present study, the predominant color of collagen fibers around the tumor islands in odontogenic tumors was orangish red. When the comparison of color of collagen fibers of odontogenic tumors with that of odontogenic keratocysts and dentigerous cysts was done, there was no significant difference statistically (P > 0.05). ...
The present study was undertaken to detect and compare the pattern of collagen fibers in odontogenic cysts and also to find out if this methodology could be used to predict the aggressive nature of odontogenic cysts by comparing with the odontogenic tumors.
The collagen in the wall of 11 odontogenic keratocysts, 14 dentigerous cysts and 14 radicular cysts was studied histochemically by staining sections with picrosirius red and examining under polarizing microscope. This was compared to 10 cases of odontogenic tumors using Z test of proportion at 1% and 5%.
In dentigerous cysts, odontogenic keratocysts and odontogenic tumors, the predominant color of collagen fibers birefringence was found to be orangish red, whereas in radicular cysts the collagen fiber was of green color.
Similar birefringence pattern of collagen fibers between dentigerous cysts, odontogenic keratocysts and odontogenic tumors may indicate that these lesions have a common histogenesis with a broad spectrum of biological behavior and belong to the same group, i.e., are developmental in origin. Different patterns of radicular cysts suggest different biological behavior and a positive role of inflammation on polarization color of collagen fibers.
... A participação da MMP-9 na angiogênese é de extrema importância, uma vez que esta protease tem como substrato o colágeno tipo IV e a laminina, componentes da membrana basal de vasos sangüíneos. Assim, promove a degradação necessária ao brotamento vascular induzido pelos fatores angiogêncios (ASAHI et al., 2001;KENAGY et al., 1997;LEE et al., 2007;VALABLE et al., 2005;KEISER, 1998 , 2001;IURLARO et al., 1999;KENAGY et al., 1997;KOIVUNEN et al., 1999;LEE et al., 2007;VU et al., 1998;ZHENG et al., 2006 LORETO, 2005;LIN et al., 1997;LIN et al., 2002;SHIN et al., 2002;SILVEIRA et al., 2007;SOARES et al., 2007;TERONEN et al., 1995a, TERONEN et al., 1995bWAHLGREN et al., 2001;WAHLGREN et al., 2002;WAHLGREN et al., 2003). ...
... Estudos recentes sobre a biologia das lesões periapicais também destacam a atuação das MMPs, na patogênese das lesões periapicais, atuando ativamente na regulação da proliferação epitelial e migração do tecido epitelial, na degradação tecidual e remodelação da cápsula fibrosa e da parede cística, bem como nos mecanismos de expansão inerentes destas lesões, solubilizando a matriz óssea e regulando o início da reabsorção ( KUBOTA et al., 2000;CALTABIANO;LORETO, 2005;LIN et al., 1997;LIN et al., 2002;SHIN et al., 2002;SILVEIRA et al., 2007;SOARES et al., 2007;TERONEN et al., 1995a, TERONEN et al., 1995bWAHLGREN et al., 2001;WAHLGREN et al., 2002;WAHLGREN et al., 2003). ...
Tese (doutorado)—Universidade de Brasília, Faculdade de Ciências da Saúde, 2008. A angiogênese tem sido implicada na patogênese das lesões periapicais e para que ela ocorra é necessária a atuação de potentes fatores pró-angiogênicos e a participação das MMPs na degradação da matriz extracelular. O objetivo deste estudo foi investigar a imunoexpressão da MMP-9, do VEGF e do FvW em granulomas periapicais (GP), cistos periapicais (CP) e cistos periapicais residuais (CPR) no intuito de verificar a participação da angiogênese no desenvolvimento destas lesões. A análise quantitativa do VEGF nas células do tecido conjuntivo revelou número de médio de células imunorreativas semelhante nos GP e CP (564,9 e 565,0, respectivamente) e maior que nos CPR (443,9), não havendo diferença estatisticamente significativa. Lesões com intenso infiltrado inflamatório apresentaram maior número de células imunorreativas ao VEGF em comparação com lesões que apresentavam leve infiltrado inflamatório (p<0.01). No revestimento epitelial, o VEGF revelou forte expressão tanto em CP (55%) quanto em CPR (70%), padrão observado também nos casos de epitélio atrófico (66,7%) e hiperplásico (55,6%), não havendo diferença estatisticamente significativa entre o dois tipos de cistos (p>0.05) e entre os cistos de revestimento epitelial atrófico e hiperplásico (p>0.05). O número médio de vasos imunorreativos ao FvW foi maior nos CP (250,8) que nos GP (210,4) e nos CPR (217,0), sem haver diferença estatisticamente significativa entre os grupos (p>0.05). Lesões que apresentavam intenso infiltrado inflamatório exibiram maior número de vasos imunomarcados que aquelas com leve ou moderado infiltrado inflamatório (p<0.05). Não houve correlação entre a quantidade de células imunorreativas para o VEGF e o número de vasos imunomarcados com o anticorpo anti-FvW (r=0.224; p=0,118). A MMP-9 revelou marcada tendência à forte expressão na maioria dos casos de GP (70%) e CP (70%) e fraca expressão nos casos de CPR (60%). Houve associação entre a forte expressão da MMP-9 nas células endoteliais e um intenso infiltrado inflamatório (p=0,055). Além disso, foi observado maior número de células imunopositivas para o VEGF (p<0.01) e maior número de vasos imunorreativos para o FvW (p=0,051) em lesões com forte expressão de MMP-9. Os resultados do presente estudo sugerem que o VEGF pode atuar como importante estímulo à expressão de MMP-9 em células endoteliais de vasos sangüíneos, influenciando o processo de angiogênese em granulomas periapicais, cistos periapicais e cistos periapicais residuais. Adicionalmente, a intensidade do infiltrado inflamatório desempenha importante influência no número de vasos sangüíneos, identificados através da expressão do FvW, na expressão de MMP-9 em células endoteliais e na expressão de VEGF em células do tecido conjuntivo, nas lesões do presente estudo. Por fim, a forte expressão do VEGF no revestimento epitelial dos cistos periapicais residuais sugere a existência de um potencial de crescimento nestas lesões, provavelmente em virtude da manutenção de um status de permeabilidade vascular aumentada. ______________________________________________________________________________________ ABSTRACT Angiogenesis has been related to the pathogenesis of periapical lesions. So that it occurs is necessary the action of powerful angiogênic factors and the participation of MMPs in the degradation of the extracellular matrix. The purpose of this study was to investigate the immunoexpression of MMP-9, VEGF and vWF in periapical granulomas (PG), periapical cysts (PC) and residual periapical cysts (RPC) to ensure the participation of angiogenesis in the development of these injuries. The quantitative analysis of VEGF in the connective tissue cells revealed similar average number of cells expressing this factor in PG and PC (564,9 e 565,0, respectively) and higher than in RPC (443,9), without statistically significant difference. There was a positive association between the intensity of the inflammatory infiltrate and the average number of cells expressing VEGF (p<0.01). In the epithelial lining, VEGF was strongly expressed by both PC (55%) as RPC (70%), pattern observed also in cases of atrophic (66,7%) and hyperplastic epithelium (55,6%), without difference statistically significant between the two types of cysts (p>0.05) and between atrophic and hyperplastic epithelial lining cysts (p>0.05). The average number of vessels expressing vWF was higher in PC (250,8) than in PG (210,4) and the RPC (217,0), without statistically significant difference between groups (p>0.05). Injuries showing intense inflammatory infiltrate exhibited greater number of vessels immunoreactive than those with mild or moderate inflammatory infiltrate (p<0.05). There was no correlation between the number of cells immunoreactive to VEGF and the number of vessels immunoreactive to the anti-vWF (r=0.224, p=0.118). The MMP-9 showed marked tendency to strong expression in most cases of GP (70%) and PCs (70%) and low expression in cases of CPR (60%). There was strong association between the expression of MMP-9 on endothelial cells and an intense inflammatory infiltrate (p=0055). Moreover, it was observed greater number of cells immunexpressing VEGF (p<0.01) and highest number of vessels immunoreactive to vWF (p=0051) in lesions with strong expression of MMP-9. The results of this study suggest that VEGF can act as an important stimulus for expression of MMP-9 in endothelial cells of blood vessels, influencing the angiogenic process in PG, PC and PRC. Additionally, the intensity of the inflammatory infiltrate plays important influence on the number of blood vessels, identified by the expressions of vWF, in the expression of MMP-9 in endothelial cells and the expression of VEGF in connective tissue cells in the lesions of this study. Finally, the strong expression of VEGF in the epithelial lining of the RPC suggests that there is a potential for growth in these injuries, probably because of the maintenance of a increased vascular permeability status.
... The higher inflammatory activity index in the periapical area indicated delayed repair in an MMP positively-stained area for this group. Although MMP expression had been immunolocalized in human apical periodontitis previously (24)(25)(26)(27)(36)(37)(38), this is the first study to compare MMP expression in teeth with apical periodontitis submitted to different clinical protocols of root canal therapy. ...
... MMPs play an important role in dental pulp tissue destruction (25,26) and it has been proposed that MMP-1, MMP-8, MMP-9, and MMP-13 together are involved in jaw cyst expansion (27,(36)(37)(38). In teeth submitted to root canal treatment using calcium hydroxide as the root canal dressing, a lower inflammatory index was observed, accompanied by an increased percentage of fibroblasts. ...
The objective of this study was to investigate the expression of matrix metalloproteinases (MMPs) in apical periodontitis and during the periapical healing phase after root canal treatment.
Apical periodontitis was induced in dog teeth, and root canal treatment was performed in a single visit or by using an additional calcium hydroxide root canal dressing. One hundred eighty days after treatment the presence of inflammation was examined, and tissues were stained to detect bacteria. Bacterial status was correlated to the degree of tissue organization, and to further investigate molecules involved in this process, tissues were stained for MMP-1, MMP-2, MMP-8, and MMP-9. Data were analyzed by using one-way analysis of variance followed by Tukey test or Kruskal-Wallis followed by Dunn test.
Teeth with apical periodontitis that had root canal therapy performed in a single visit presented an intense inflammatory cell infiltrate. Periapical tissue was extremely disorganized, and this was correlated with the presence of bacteria. Higher MMP expression was evident, similar to teeth with untreated apical periodontitis. In contrast, teeth with apical periodontitis submitted to root canal treatment with calcium hydroxide presented a lower inflammatory cell infiltrate. This group had moderately organized connective tissue, lower prevalence of bacteria, and lower number of MMP-positive cells, similar to healthy teeth submitted to treatment.
Teeth treated with calcium hydroxide root canal dressing exhibited a lower percentage of bacterial contamination, a lower MMP expression, and a more organized extracellular matrix, unlike those treated in a single visit. This suggests that calcium hydroxide might be beneficial in tissue repair processes.
... MMPs are a family of metaldependent endopeptidases, which are secreted as inactive proenzymes (zymogens) and activated in tissues by cleavage of the propeptide (19)(20)(21)(22)(23). Although MMPs have been reported in periapical lesions (11,(24)(25)(26)(27)(28)(29)(30), a direct comparison of MMPs in cysts and granulomas has not been undertaken. Since MMPs must be activated to exert their function, it is also important to localize areas of proteinase activity within lesions. ...
... Using in situ zymography, we demonstrated the presence and increased levels of active MMPs in the inflammatory cell zone in periapical granulomas compared to cysts. Although MMP expression has been immunolocalized in human periapical lesions (11,(24)(25)(26)(27)(28)34), this is the first study to identify areas of MMP enzymatic activity. ...
The inability to distinguish periapical cysts from granulomas before performing root canal treatment leads to uncertainty in treatment outcomes because cysts have lower healing rates. Searching for differential expression of molecules within cysts or granulomas could provide information with regard to the identity of the lesion or suggest mechanistic differences that may form the basis for future therapeutic intervention. Thus, we investigated whether granulomas and cysts exhibit differential expression of extracellular matrix (ECM) molecules.
Human periapical granulomas, periapical cysts, and healthy periodontal ligament tissues were used to investigate the differential expression of ECM molecules by microarray analysis. Because matrix metalloproteinases (MMP) showed the highest differential expression in the microarray analysis, MMPs were further examined by in situ zymography and immunohistochemistry. Data were analyzed by using one-way analysis of variance followed by the Tukey test.
We observed that cysts and granulomas differentially expressed several ECM molecules, especially those from the MMP family. Compared with cysts, granulomas exhibited higher MMP enzymatic activity in areas stained for MMP-9. These areas were composed of polymorphonuclear cells (PMNs) in contrast to cysts. Similarly, MMP-13 was expressed by a greater number of cells in granulomas compared with cysts.
Our findings indicate that high enzymatic MMP activity in PMNs together with MMP-9 and MMP-13 stained cells could be a molecular signature of granulomas unlike periapical cysts.
... ProMMP-1 was purified by conventional and affinity column chromatographies from culture medium of human synovial fibroblasts and activated by 1 mM (APMA) or a brief exposure to trypsin [9,10]. Interstitial collagenase was also extracted from jaw cyst wall tissue shown to contain MMP-1 by Western-blot [11,12] (Fig. 1C); the jaw cyst MMP-1 existed in an endogenously active form [11,13,14]. Both MMP-1 species were pretreated with buffer and several indicated concentrations of clodronate for 60 minutes, subsequently the samples were incubated together with type I collagen for 8 hours at 22°C. ...
... ProMMP-1 was purified by conventional and affinity column chromatographies from culture medium of human synovial fibroblasts and activated by 1 mM (APMA) or a brief exposure to trypsin [9,10]. Interstitial collagenase was also extracted from jaw cyst wall tissue shown to contain MMP-1 by Western-blot [11,12] (Fig. 1C); the jaw cyst MMP-1 existed in an endogenously active form [11,13,14]. Both MMP-1 species were pretreated with buffer and several indicated concentrations of clodronate for 60 minutes, subsequently the samples were incubated together with type I collagen for 8 hours at 22°C. ...
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.
... Periapical lesions are most commonly of endodontic origin and are related to pulp infection (1,2). Bacteria and their by-products can exit the root canal system through the apical foramen and cause an inflammatory response in the periapical tissues (3,4) and resorption of the alveolar bone surrounding the root (5). Most endodontic lesions can be classified as periapical granuloma or periapical cyst (6,7,8). ...
The most common lesions of endodontic origin are granuloma and periapical cyst. They are often asymptomatic and are discovered after a routine radiographic examination. The aim of this study was to review the most common and important periapical lesions (root canal and granuloma) and to present simple scientific guidelines to recognize their difference. Attempts were made not only to locate or describe the size of the lesion, but also to diagnose and classify them radiographically as granuloma or periapical cyst. From the presentation of our case reports we can conclude that the current accuracy of clinical and radiological examinations to distinguish between periapical cyst and granuloma is high, but the gold standard for accurately diagnosing and identifying periapical lesions is his-tological examination.
... Elas podem agir tanto em processos fisiológicos quanto em patológicos, dependendo de diversos fatores locais que atuam como co-fatores na reorganização da matriz orgânica tecidual. O interesse em desenvolver inibidores sintéticos das MMPs que possam ser usados em terapias médicas e odontológicas é o que movimenta muitas pesquisas nessa área.Dentário 6,4,5,7,18,20,21,22,23,27,28,32,62,68,69,118,119,125,128,130, 140, 157, 159, 167, 200, 201, 202, 203, 206, 207, 209, 213, 250 Estética 8,1,32,50,54,56,72,73,74,75,78,86,87,88,98,99,101,115,117,118,119,120,121,122,124,125,126,129,130,131,132, 136, 207, 209, 211, 212, 220 Idosos 9,133,134,135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 148, 149, 150, ...
... Dentição Permanente 61,200,211,212 Doença Periodontal 20,79,81,82,88,128,130,135,136,145,149,160,240,241 E Epidemiologia 2,134,263 Estética 8,1,32,50,54,56,72,73,74,75,78,86,87,88,98,99,101,115,117,118,119,120,121,122,124,125,126,129,130,131,132,136,207,209,211,212,220 Estética dentária 117, 119 ...
... Our findings of significantly increased levels of serum MDA, IL-6, TNF-alpha, and MMP-9 in patients when compared to controls align with a previous study that suggest RC formation may be due to the degradation of collagen via MMPs (i.e., the main stop in the loss of periodontal supporting tissue) [17]. However, another study suggests the primary factor in RC formation is due to collagenases [18]. The separation of epithelium from the connective tissues results in lesion progression and recurrence [19][20]. ...
Background
Matrix metalloproteinase-9 (MMP-9) and antioxidants are associated with the pathogenesis of cysts and may initiate and sustain the formation of new capillaries.
Objective
The objective of this study was to determine the association of oxidative stress and the production of inflammatory mediators MMP-9 and interleukin 6 (IL-6) in systemic events in radicular cyst growth.
Materials and methods
Fifty patients (34 men, 16 women) with periapical granulomas and radicular cysts were included in this cross-sectional study. Twenty subjects (12 men, eight women) with no signs of periodontal diseases were recruited as controls. Blood serum levels of MMP-9, IL-6, superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione peroxidase (GPx) were recorded. We also recorded body mass index (BMI) and tumor necrosis factor-alpha (TNF-alpha) levels.
Results
The mean age of the test group patients and control patients was 45.9 and 48.8 years, respectively. The BMI of test group patients (23.77± 3.88 kg/m²) was higher than that of the controls (27.98 ± 3.88 kg/m²; p ≤ 0.000). Levels of serum MDA (p ≤ 0.033), IL-6 (p ≤ 0.041), TNF-alpha (p ≤ 0.004), and MMP-9 (p ≤ 0.033) were significantly increased in patients as compared with control values. SOD (p ≤ 0.003) and GPx (p ≤ 0.033) levels were significantly reduced in patients as compared with controls.
Conclusion
Oxidative imbalance and the increased production of inflammatory mediators may be associated with systemic events in radicular cysts. Bone-resorbing mediators and proinflammatory cytokines that were evaluated in the study (MMP-9, IL-6, C-reactive protein, TNF-alpha) were also elevated in the serum of the ailing group, thus documenting a well-established role for these circulating biochemical variables in the course of the progression and pathogenesis of radicular cyst development.
... Thus, inflammatory cells could affect the arrangement of collagen. [20][21][22] But in the developmental cysts because of their longstanding duration may cause degeneration of the stroma in the form of hyalinization and may probably explain such an exhibition by the collagen fibers. These regions also exhibited either a change in phenotype or thickness of overlying epithelium. ...
Introduction:
Odontogenic lesions represent a range of conditions, the features of which probably depend on the stage of induction towards tooth formation reached prior to neoplastic or hamartomatous proliferation. It has been also suggested that inductive changes may allow progression from one type of odontogenic tumor to another. The epithelium also plays an important role in the pathogenesis of these lesions; even stroma is likely to play an equally important role in the pathogenesis and biological behavior. So, this study was performed to investigate, compare, and correlate different types of collagen fibers in odontogenic cysts and odontogenic tumors.
Materials and methods:
Thirty each pre-diagnosed odontogenic cysts and tumors were histochemically analyzed using a special stain (Picrosirius red stain) and polarizing microscopy.
Results:
Seven cases (99%) of inflammatory cysts exhibited predominantly greenish-yellow birefringence indicating procollagen, intermediate, or pathologic collagen fibers suggestive of loosely packed collagen fibers. Predominant yellowish-orange birefringence exhibited by 21 cases (99%) of developmental cysts was comparable to the yellowish-orange and orangish-red to red birefringence exhibited by odontogenic tumors suggesting tightly packed fibers.
Conclusions:
The Picrosirius red stain in conjunction with polarizing microscopy serves as a specific and sensitive tool in characterizing collagen fibers in odontogenic cysts and odontogenic tumor.
... Thus, inflammatory cells could affect the arrangement of collagen. [6,14] Recently, it has been proposed that inflammation is necessary to trigger the initiation of fibrosis; chronic inflammation leads to tissue destruction, ongoing wound healing responses, and eventually fibrosis. This could explain the presence of orange-red fibers which were also present in significant amount in radicular cysts. ...
Background: Lesions of odontogenic origin comprise the heterogeneous group ranging from hamartomatous proliferations, cysts to benign and malignant tumors. Interplay between the epithelium and connective tissue can be assumed to play a significant role in the pathogenesis of odontogenic cysts. Aims and Objectives: A study was taken up to show the role of picrosirius red (PSR) stain to demonstrate the fibers and also to assess the difference in the nature of the fibers (different color patterns) and to find out the role of it, if any in the pathogenesis and biological behavior of the commonly occurring odontogenic cysts. Materials and Methods: Collagen fibers of 30 cases of odontogenic cysts (10 radicular cysts, 10 odontogenic keratocysts (OKC's), and 10 dentigerous cysts) were studied by staining the sections with PSR stain and examining them under bright field and polarizing microscope. Results: Sixty-seven percentage of the thin collagen fibers and 55% of the thick fibers in radicular cyst showed green-yellow birefringence. Fifty-seven percentage of the thin collagen fibers and 15% of the thick fibers in OKC showed green-yellow birefringence. Eighty-two percentage of the thin collagen fibers and 66% of the thick fibers in dentigerous cysts showed green-yellow birefringence. Rest of the fibers showed orange-red birefringence. Statistical analysis with one-way ANOVA was significant with a P < 0.01 only for thick fibers. Moreover, comparison of polarization colors of thick fibers of odontogenic cysts with duration of the lesion gave statistically significant results. Conclusion: The observations in the present study with respect to color profiles of the collagen fibers in the three commonly occurring odontogenic cysts possibly explain the biological behavior of the lesions. The predominant orange-red birefringence in OKC's in comparison to radicular and dentigerous cysts suggests that OKC's exhibit well organized and tightly packed fibers. This may possibly explain the reason for the poorer prognosis of OKC's. It is suggested that though the epithelium plays an important role in the pathogenesis of these lesions, even stroma is likely to play an equally important role in the pathogenesis and biological behavior. © 2015 Journal of Orofacial Sciences | Published by Wolters Kluwer - Medknow.
... Matrix metalloproteinase (MMP) are a large family of enzymes which take part in many proteolysis associated pathological conditions [3]. Role of MMPs and their tissue inhibitors has also been associated to several different cysts types found in maxillofacial region, including follicular cysts [4][5][6]. However, all the mechanisms how the cyst develops and eventually destructs the bone have not been completely clarified. ...
Objective:
Mechanisms of the dentigerous cyst formation from the normal eruption follicle is unknown but disturbances in the proteolytic activity have been suspected, since the growth of these cysts is accompanied by local bone destruction. The aim of the present study was to evaluate the expression of matrix metalloproteinases (MMP) in human dental dentigerous cysts and healthy dental follicles.
Materials and methods:
We studied 10 patients with dentigerous cysts and 10 healthy dental follicles from the lower jaw in respect to their immunoexpression of MMPs -8, -9, -25, and -26 and tissue inhibitor of metalloproteinases -1 (TIMP-1).
Results:
MMP-8 was expressed slightly more in cyst epithelium than in odontogenic epithelium of healthy controls dental follicle but the difference lacked statistical difference. Other MMPs and TIMP-1 did not differ regarding the studied specimens.
Conclusion:
Differences in MMP expression cannot solely explain the cyst expansion suggesting the potential involvement of other osteolytic mechanisms.
... 8 The macrophages, T-lymphocytes, and matrix metalloproteinases (MMP-1 and -2) in the cyst wall may provide a continuous source of boneresorptive metabolites. [9][10][11][12] In addition, cytokines can cause lesions to expand, regardless of the root canal's microbial status. Earlier, it was believed that all of the cyst lining should be removed to prevent recurrence of the lesion. ...
Objective:
To evaluate the efficacy of surgical fenestration (partial enucleation) in resolution of large periapical inflammatory cyst-like lesions.
Method: Seven cases (5M: 2F, average age: 29.4 yrs.) of large periapical cyst-like lesions, associated with non-vital permanent teeth were recruited in the study. Informed consent was taken. Endodontic treatment was completed as per the standard protocol. Under local anesthesia, flap was reflected, the thinned out bone was removed and a small piece of cystic lining was cut. No attempt was made to completely enucleate the cystic lining. The exposed cavity was then gently curetted, and copious irrigation with saline was done. The flap was sutured back with 3/0 silk. Pre and post treatment clinical and radiographic record was maintained. The cases were followed up at 3, 6,12 and 18 months.
Result:
Post-operative healing was satisfactory, both clinically and on radiographic evaluation. The periapical lesion had either healed completely or showed decrease in the size of the lesion, indicating healing. Six of the seven cases were diagnosed as periapical cyst on histology while in one case the tissue was crushed and could not be commented upon.
Conclusion:
Based on the present case-series, it is concluded that surgical fenestration is a simple, minimally invasive and least traumatic method of treatment of large periapical inflammatory cyst-like lesion. It is not necessary to completely enucleate the lining, if the cyst is of inflammatory origin, secondary to endodontic infection. This technique also provides tissue for biopsy for differential diagnosis to rule out other lesions of grevious prognosis.
... The role of the extracellular matrix (ECM) in the pathogenesis and behavior of epithelial odontogenic lesions has been demonstrated in some publications [8,9,17,18], suggesting that these lesions could be dependent on epithelial-mesenchymal interactions [9,17,[19][20][21][22][23][24]. The ECM is a dynamic, intricate network of macromolecules that plays an important role in the maintenance of a correct microenvironment for basic cell functions, such as celladhesion, proliferation, and differentiation [25][26][27][28]. ...
Dentigerous cyst (DC) and keratocystic odontogenic tumor (KOT) are odontogenic lesions arising from epithelial elements, such as those observed in dental follicles (DF), that have been part of the tooth forming apparatus. These lesions show different clinical and histological characteristics, as well as distinct biological behavior. This study aimed to qualify and quantify collagen and elastic fibers by means of histochemical techniques with light and confocal laser microscopic methods in three odontogenic entities. Eleven DF, 13 DC (n=10 with inflammation, n=3 without inflammation) and 13 KOT were processed to the following techniques: Hematoxylin and Eosin, Masson's Trichrome, Picrosirius, Direct Blue, and Orcein. DF and DC without inflammation exhibited collagen with similar characteristics: no parallel pattern of fiber orientation, thick fibers with dense arrangement, and absence of distinct layers. A comparison between DC with inflammation and KOT revealed similar collagen organization, showing distinct layers: thin collagen fibers with loose arrangement near the epithelium and thick fibers with dense arrangement in distant areas. The only difference found was that KOT exhibited a parallel collagen orientation in relation to the odontogenic epithelia. It may be suggested that the connective tissue of DC is a reactive tissue, inducing an expansive growth associated with fluid accumulation and inflammatory process, which in turn may be present as part of the lesion itself. In KOT, loosely arranged collagen may be associated with the behavior of the neoplastic epithelium.
... 8 The macrophages, T-lymphocytes, and matrix metalloproteinases (MMP-1 and -2) in the cyst wall may provide a continuous source of boneresorptive metabolites. [9][10][11][12] In addition, cytokines can cause lesions to expand, regardless of the root canal's microbial status. Earlier, it was believed that all of the cyst lining should be removed to prevent recurrence of the lesion. ...
Various conservative approaches have been utilized to manage large periapical lesions. This article presents a relatively new, very conservative technique known as surgical fenestration which is both diagnostic and curative. The technique involves partially excising the cystic lining, gently curetting the cystic cavity, performing copious irrigation, and closing the surgical site. This technique allows for decompression and allows the clinician the freedom to take a biopsy of the lesion, as well as perform other procedures such as root resection and retrograde sealing, if required. As the procedure does not perform a complete excision of the cystic lining, it is both minimally invasive and cost-effective. The technique and the concepts involved are reviewed in 4 cases treated with this novel surgical approach.
... It is constitutively expressed at low levels under normal physiological conditions; however, its expression may increase markedly in pathological conditions. MMP-1 is present in jaw cyst wall extracts and cyst fluids (Teronen et al., 1995). MMP-8 was first cloned from messenger RNA (mRNA) extracted from the peripheral leucocytes of a patient with chronic granulocytic leukemia (Hasty et al., 1990). ...
Matrix metalloproteinases (MMPs) are a major group of enzymes that regulate cell–matrix composition. The MMPs are zinc-dependent endopeptidases known for their ability to cleave one or several extracellular matrix (ECM) constituents, as well as nonmatrix proteins. This review focuses on structural and functional elements of MMPs, and their roles in physiological and pathological processes, in which it is believed the MMPs play an important, or even indispensable role. According to their structural and functional characteristics, MMP family members have been classified into six different but closely related subgroups with fairly characteristic but often overlapping substrate specificities. MMP synthesis and functions are regulated by transcriptional activation, post-transcriptional processing (release of pro-domain, cell surface shedding), and control of activity by a family of endogenous inhibitors collectively known as tissue inhibitors of metalloproteinases (TIMPs). The balance of MMPs to TIMPs therefore determines matrix turnover, where either an excess of MMPs or a deficit of TIMPs may result in excess ECM degradation. Moreover, the recent development of selective and non-selective inhibitors of MMPs would also provide new insights in the knowledge of the relationship between activation of inflammatory cells and tissue remodeling and propose new therapeutic possibilities to the treatment of inflammatory disease.
... The role of the extracellular matrix (ECM) in the pathogenesis and behavior of epithelial odontogenic lesions has been demonstrated in some publications [8,9,17,18], suggesting that these lesions could be dependent on epithelial-mesenchymal interactions [9,17,[19][20][21][22][23][24]. The ECM is a dynamic, intricate network of macromolecules that plays an important role in the maintenance of a correct microenvironment for basic cell functions, such as celladhesion, proliferation, and differentiation [25][26][27][28]. ...
Dentigerous cyst (DC) and keratocystic odontogenic tumor (KOT) are odontogenic lesions arising from epithelial elements, such as those observed in dental follicles (DF), that have been part of the tooth forming apparatus. These lesions show different clinical and histological characteristics, as well as distinct biological behavior. This study aimed to qualify and quantify collagen and elastic fibers by means of histochemical techniques with light and confocal laser microscopic methods in three odontogenic entities. Eleven DF, 13 DC (n=10 with inflammation, n=3 without inflammation) and 13 KOT were processed to the following techniques: Hematoxylin and Eosin, Masson's Trichrome, Picrosirius, Direct Blue, and Orcein. DF and DC without inflammation exhibited collagen with similar characteristics: no parallel pattern of fiber orientation, thick fibers with dense arrangement, and absence of distinct layers. A comparison between DC with inflammation and KOT revealed similar collagen organization, showing distinct layers: thin collagen fibers with loose arrangement near the epithelium and thick fibers with dense arrangement in distant areas. The only difference found was that KOT exhibited a parallel collagen orientation in relation to the odontogenic epithelia. It may be suggested that the connective tissue of DC is a reactive tissue, inducing an expansive growth associated with fluid accumulation and inflammatory process, which in turn may be present as part of the lesion itself. In KOT, loosely arranged collagen may be associated with the behavior of the neoplastic epithelium.
... Several other studies have also substantiated the effect of MMPs along with tissue inhibitor of metalloproteinases and collagenases in cyst walls 6,7 . These cell products have been found to be stimulated by mast cell derivatives 7-9 . ...
Aim: Mast cells have been hypothesized to play a significant role in pathogenesis of odontogenic cysts. The aim of this study was to evaluate mast cell distribution in cystic lining and the capsule to formulate a mechanism of cystic expansion. Methods: Ten formalin-fixed paraffin embedded tissue blocks each of OKC, dentigerous and radicular cysts were selected. Toluidine blue staining (1% in 1% NaCl solution) was done in 5μm thick sections and counting performed in 10 areas using an ocular grid. Areas counted were divided into 4 zones: intraepithelial, subepithelial, intermediate and deep zones (Group I, II, III and IV respectively). Statistical analysis: Mean ±S.D. was calculated in each group followed by paired ‘T’ test. Results: Mast cells had greatest concentration in subepithelial zone. 'T' test showed no significant differences between group I and II zones in OKC but a highly significant difference between groups I and II in dentigerous cyst. Radicular cysts showed a significant difference between groups II and III. Conclusion: Mast cell degranulation releases numerous hydrolytic enzymes that facilitate breakdown of capsular matrix increasing the hydrostatic pressure due to raised osmolality. Influx of tissue fluids results in their enlargement coupled with resorption at the bone-cyst interface.
... Matrix metalloproteinases (MMPs) are genetically distinct but structurally related proteinases consisting of five subgroups: collagenases, gelatinases, stromelysins, membrane-type MMPs, and other MMPs [5]. MMPs are Zn 2+ -and Ca 2+ -requiring enzymes capable of degrading almost all extracellular matrix (ECM) and basement membrane (BM) components in normal tissue remodelling, and especially in tissuedestructive diseases, including odontogenic tumours and cysts [6][7][8]. ...
Matrix metalloproteinases (MMPs) collectively degrade extracellular matrix and basement membrane proteins in chronic inflammation and bone-destructive lesions. This study examined the ability of immunoglobulin-producing plasma cells, typically present in sites of chronic inflammation, to express collagenases (MMP-8 and -13) in vivo and in vitro. Phorbol-12-myristate-13-acetate, interleukin-6, and tumour necrosis factor-alpha and heparin with the tumour promoter or cytokines potently enhanced (up to nine-fold) MMP-8 and -13 expression by the RPMI 8226 myeloma cell line, as evidenced by western blotting and semi-quantitative reverse transcriptase-polymerase chain reaction. Immunohistochemical analysis and in situ hybridization revealed that plasma cells expressed MMP-8 and -13 focally in periapical granulomas, odontogenic cysts, and malignant plasmacytomas. MMP-8 and MMP-13 from plasma cells can participate in bone organic matrix destruction at sites of chronic inflammation and neoplastic growth. Since MMP-13 was more frequently expressed than MMP-8 in plasma cells of strongly recurring keratocysts and malignant plasmacytomas, it is concluded that plasma cell MMP-13 has a particularly important role in benign and malignant bone-destructive lesions.
Objective
The aim of this study is to evaluate the osteogenesis effect following jaw cyst surgery by comparing three scenarios: no use of filling materials, the use of a medical collagen sponge alone, and the combined application of Bio-Oss bone powder with a medical collagen sponge. This comparison aims to provide evidence for selecting appropriate filling materials for post-surgical treatment of jaw cysts.
Methods
This research involved 52 patients with jaw cysts at the Zhongshan City Stomatological Hospital from October 2022 to September 2023. Patients were randomly assigned to three groups: Group A (control group with no filling material), Group B (medical collagen sponge), and Group C (Bio-Oss bone powder combined with a medical collagen sponge). CBCT scans were taken before surgery and at 3 and 6 months post-operation. The Mimics 21.0 software was utilized to analyze changes in the volume of the cyst cavity and the bone density values.
Results
Significant statistical differences in changes to the volume of the cyst cavity and bone density were observed among the three groups (P < 0.01). Group C demonstrated the most effective osteogenesis, with a reduction in cyst cavity volume by 51.74% and an increase in bone density by 321.24% at 3 months post-operation. By 6 months, the cyst cavity volume had decreased by 80.54%, and the bone density had increased by 422.72%. Group B had the second-best outcomes, while Group A had the least effective bone regeneration.
Conclusion
The study concluded that the combination of Bio-Oss bone powder and medical collagen sponge leads to the most effective osteogenesis in patients undergoing surgery for jaw cysts. The use of a medical collagen sponge alone was the second most effective, while no filling material resulted in the least effective postoperative bone regeneration.
The odontogenic keratocyst (OKC) is classified by The World Health
Organization as an epithelial developmental cyst. It represents about 5-10% of all
odontogenic cysts.
The OKC may be found in patients ranging from infancy to old age with a
peak incidence in the second and third decades. A second minor peak is referred by some
authors in the fifth decade or later. Males are more frequently affected than females with
a male-to-female ratio of about 1.5:1.
These cysts are diagnosed by histological examination. Most of the times
there are no associated signs and symptoms. Significantly more OKC are found in the
mandible, with the majority being found in the molar, angle, and ramus area. It is
commonly seen associated with impacted teeth.
Radiographically, OKC appears as a unilocular, lobulated or multilocular
radiolucency. It may be seen on any location resembling other pathologies such as:
radicular cyst, residual cyst, gengival cyst of the adult, periodontal lateral cyst,
dentigerous cyst and nasopalatin cyst. The multilocular type resembles the amelolastoma.
The clinical and radiographic features of the OKC are not pathognomonic.
In this study, we have confirmed the increased difficulty in diagnosing an
OKC based on clinical and radiographic information. The pre-operative suspicious of an
OKC is low even among surgeons. It also shows that even without clinical and
radiographic pathognomonic characteristics, the pre-operative diagnose of an OKC is
based on the radiographic type of image and the localization of the cyst. It is more
frequently suspected among multilocular radiolucencies situated on the molar, angle and
ramus area.
Based on its symptomless aggressive clinical behaviour, its diverse location
and on the fact that it is diagnosed by its histological features, it is adviseable that this
pathology is considered on the differential diagnosis of other pathologies such as
radicular, residual and dentigerous cyst.
This study also reinforces that it is mandatory to analyse all lesions surgically
removed; this decision should not be left to the surgeon's opinion
Clinical differentiation between cystic lesions of endodontic and non-endodontic origin is of importance because correct diagnosis may affect treatment decision making. Most radicular cysts are treated with conservative approaches and, therefore, are not surgically removed. The objective of this study was to determine the accuracy of clinical diagnosis of periapical lesions as compared to the histological findings, and to evaluate various associated factors. All biopsy specimens submitted for histological evaluation from 2002 to 2009 were assessed. Only cases of periapical lesions with complete patient data and clinical diagnosis were included. Sensitivity, specificity and accuracy of the clinical diagnosis were calculated and various patient-related factors were evaluated. Of the 4,908 cases, 183 met inclusion criteria. Histologically, there were 171 lesions of radicular cysts and 12 cases of non-endodontic cysts, including OKC and Incisive Canal Cyst. The diagnostic accuracy for clinical diagnosis for radicular cysts was 91.84% and 91.84% for non-endodontic cysts. There was a high accuracy of clinical differentiation between cystic lesions of endodontic and non-endodontic origin. However, some non-endodontic lesions may be incorrectly diagnosed clinically as lesions of endodontic origin. Histological evaluation may be necessary for the correct diagnosis. Further clinical studies are needed to evaluate clinical examination and histological diagnosis of periapical lesions.
Interleukin-1α (IL-1α) is strongly expressed in odontogenic keratocysts. In this study, we investigated the effects of IL-1α on the activation of matrix metalloproteinase-2 (MMP-2) in the fibroblasts isolated from odontogenic keratocysts. Odontogenic keratocyst fibroblasts secreted a latent form of MMP-2 (proMMP-2) spontaneously. Type I collagen induced the activation of the proMMP-2, and recombinant human IL-1α (rhIL-1α) further enhanced the type I collagen-induced activation of proMMP-2 in a dose-dependent manner. The rhIL-1α-induced activation of proMMP-2 was inhibited by anti-human IL-1α antibody. A reverse-transcription/polymerase chain-reaction (RT-PCR) and Western immunoblotting demonstrated that the expression of membrane-type 1 matrix metalloproteinase (MT1-MMP) mRNA and protein was increased in the fibroblasts when the cells were cultured on type I collagen, and the expression was further enhanced by rhIL-1α. Thus, IL-1α may up-regulate proMMP-2 activation by increasing the expression of MT1-MMP in the fibroblasts isolated from odontogenic keratocysts synergistically with type I collagen.
Cysts of the Oral and Maxillofacial Regions is a seminal text for those working in oral pathology, oral medicine, oral & maxillofacial surgery and radiology. This fourth edition reflects advances in immunohistochemistry, molecular biology and human genetics, which have contributed to the understanding of the etiology, pathogenesis, pathology and treatment of these lesions. This book is a comprehensive treatise on cysts occurring in the oral and maxillofacial regions, covering clinical features, epidemiology, radiology, pathogenesis and pathology.
The aim of the present study was to evaluate the expression and distribution of different classes of matrix metalloproteinases (MMPs) in radicular cysts and periapical granulomas.
Twenty consecutive specimens of radicular cysts and 20 of periapical granulomas were selected. Expression of MMP-2, -9, -8, -13, -3 was immunohistochemically evaluated. The intensity of expression of the MMPs was evaluated using a semi-quantitative analysis: low = +; intermediate = ++; high = +++.
Positive expression of MMPs was present with different distribution. MMP-9 expressed differently in the lesions. Indeed, in periapical granulomas low expression was found in endothelial cells and fibroblasts, whilst high intensities were only detected in inflammatory cells. On the contrary, in radicular cysts the high intensities were mainly present in keratinocytes and fibroblasts. MMP-8 was mainly expressed in inflammatory cells of periapical granulomas. MMP-2 and -3 presented a low intensity of expression in both groups. MMP-13 showed a variable pattern of distribution in the different cell types of the two different lesions.
The present investigation supports the role of MMPs in the inflammatory process leading to the development of radicular cysts and periapical granulomas. The results of the present study suggested that the increased enlargement of radicular cysts, compared to periapical granulomas, might be related to a higher expression of MMP-9. On the other hands, the higher intensity of expression of MMP-8 in periapical granulomas could be related to an active inflammatory process. MMP-8 could play an important role in the inflammation processes during the development of periapical lesions.
Evidence suggests that metalloprotease expression may affect the biological behavior of odontogenic lesions. This study was conducted to review the literature about the role of metalloproteases in the development of odontogenic lesions. A search was carried out using one database, MEDLINE, via PubMed. Only articles written in English were included. Abstracts of all articles retrieved in the electronic search were evaluated for their relevance. Three articles met
inclusion criteria. They analyzed the role of MMP-2, MMP-8 and MMP-13 in radicular cysts, dentigerous cysts and keratocystic odontogenic tumors, and of MMP-1, MMP-7 and MMP-27 in keratocystic odontogenic tumors. The immunostaining technique used for all studies was similar, differing only in type of staining used. Different immunoreactivity results were found in the studies. The pattern of metalloprotease expression in odontogenic lesions was different from the pattern found in other lesions. In the studies analyzed, there was a signifi cant positive immunoreactivity for metalloproteases in odontogenic lesions, particularly in keratocystic odontogenic tumors, a fi nding that may explain KCOT aggressiveness.
IN MICROSCOPY, AS IN NATURE, ONE RECOGNIZES ONLY WHAT ONE ALREADY KNOWS.
Routine histopathological diagnostic reports and publications based on retrospective reviewing of such, perpetuate the notion that nearly half of all apical periodontitis lesions are cysts. Studies based on meticulous serial sectioning of apical lesions retrieved in toto show that the actual prevalence of cysts is only about 15% of all apical periodontitis lesions. Periapical cysts exist in two structurally distinct classes, namely those containing cavities completely enclosed in epithelial lining (periapical true cysts) and those containing epithelium-lined cavities that are open to the root canals (periapical pocket cysts). From a clinical point of view, small cysts, particularly the pocket cysts, can heal after non-surgical root canal therapy whereas large cysts, mainly true cysts, are less likely to be resolved without surgical intervention.
Mast cells are granule-containing cells in mucosal and connective tissues that are known to play a central role in allergic and inflammatory responses owing to pro-inflammatory mediators. Cysts in jaws are among the most common expansive, benign and destructive bone lesions; at some stage they are associated with chronic inflammation. Earlier studies have identified mast cells in odontogenic cysts (OC). We investigated the presence and distribution of mast cells and compared their number in different types of radicular cysts (RC), dentigerous cysts (DC) and odontogenic keratocysts (OKC). Ten cases each of RC, DC and OKC diagnosed clinically and histopathologically were selected and stained with 1% toluidine blue. The greatest number of mast cells/mm(2) was found in RC. The fewest mast cells/mm(2) were found in OKC. The subepithelial zones of all cysts contained more mast cells than the deeper zones.
Heparanase is an endo-β-D: -glucuronidase enzyme which degrades heparan sulfate glycosaminoglycan side chains of proteoglycans in the extracellular matrix and in basement membranes. The aim of this study was to evaluate the expression of heparanase in periapical granulomas (PGs) and radicular cysts (RCs). Immunohistochemistry was used to assess heparanase expression in PGs and RCs. Parameters including stain intensity, location and cell type were used to characterize heparanase expression in the periapical lesions. Ordered categories (from weak to strong) were used to compare the level of heparanase staining in the PG and RC groups. Both epithelial cells and inflammatory cells were positive for heparanase. The relative staining of the epithelial cells was strong, whereas the relative staining of the inflammatory cells was weak. Significant differences in immunohistochemical staining of epithelial cells were observed between RCs and PGs (p = 0.002). The relative expression of heparanase in epithelial cells in RCs was strong. In PGs, lesions with few or no epithelial cells, heparanase was predominantly expressed weakly by inflammatory cells. PGs and RCs have the same infectious origin. Therefore, the different cellular sources of heparanase in these periapical lesions may imply that this enzyme has specific pathogenetic functions in RCs and PGs.
The aim of this study was to investigate levels of matrix metalloproteinase-8 (MMP-8) and substance P (SP) in gingival crevicular fluid (GCF) during root canal treatment (RCT) of nonvital teeth.
Patients scheduled for nonsurgical RCT were prospectively selected; all patients provided informed consent. GCF samples were collected from teeth scheduled for RCT and their contralateral teeth across 3 different time periods. MMP-8 and SP levels were measured using enzyme-linked immunosorbent assay (ELISA). Data were analyzed using a mixed model analysis and the Pearson correlation analysis.
Patients' subjective pain levels were significantly related to both MMP-8 and SP levels. MMP-8 and SP levels in GCF were decreased during RCT, and they showed a positive correlation with each other (P < .05).
This study demonstrated that periradicular inflammation of endodontic origin can elevate SP and MMP-8 levels in GCF.
The aim of this study was to evaluate the biological profile of odontogenic epithelium by immunolabeling of epidermal growth factor receptor (EGFR), Ki-67 and survivin in keratocystic odontogenic tumors (KOT), dentigerous cysts (DC), and pericoronal follicles (PF). Immunohistochemical analysis was performed in 13 KOTs, 14 DCs and 9 PFs. Immunolabeling was analyzed in the basal and suprabasal layers of KOTs and DCs, and in the islands of odontogenic epithelium and/or reduced enamel epithelium of PFs. KOTs showed the highest proliferation rate among the three groups, mainly in suprabasal layers. EGFR immunolabeling was observed mainly in the cytoplasm in basal and suprabasal layers of KOTs and in the suprabasal layer of DCs. Immunolabeling in both membrane and cytoplasm was greater in PFs. In PFs, membrane-only staining was observed. Survivin immunolabeling showed a greater percentage of positive cells (scoring +++) in the suprabasal layer of KOTs. In DCs, both layers showed similar percentages of cells scoring +++; PFs showed the highest percentage of these cells. In KOTs, epithelial cells showed stimulus-independent neoplastic proliferative characteristics, suggesting the presence of a suprabasal proliferative compartment, maintained by inhibition of apoptosis. In DCs, the basal layer seemed to proliferate in response to stimulus. Although PFs showed low proliferative activity, the expression of EGFR indicates that some cells have a high capacity to respond to stimuli, which could probably explain the origin of odontogenic lesions.
It is a general belief that large cyst-like periapical lesions and apical true cysts caused by root canal infection are less likely to heal after nonsurgical root canal therapy. Nevertheless, there is no direct evidence to support this assumption. A large cyst-like periapical lesion or an apical true cyst is formed within an area of apical periodontitis and cannot form by itself. Therefore, both large cyst-like periapical lesions and apical true cysts are of inflammatory and not of neoplastic origin. Apical periodontitis lesions, regardless of whether they are granulomas, abscesses, or cysts, fail to heal after nonsurgical root canal therapy for the same reason, intraradicular and/or extraradicular infection. If the microbial etiology of large cyst-like periapical lesions and inflammatory apical true cysts in the root canal is removed by nonsurgical root canal therapy, the lesions might regress by the mechanism of apoptosis in a manner similar to the resolution of inflammatory apical pocket cysts. To achieve satisfactory periapical wound healing, surgical removal of an apical true cyst must include elimination of root canal infection.
The objective of this study was to determine the expression of matrix metalloproteinase-9 (MMP-9) in apical periodontitis lesions.
Nineteen epithelialized and 18 nonepithelialized apical periodontitis lesions were collected after periapical surgery. After histological processing, serial sectioning, H&E staining, and microscopic analysis, 10 epithelialized and 10 nonepithelialized lesions were selected for immunohistochemical analysis for MMP-9 and CD 68. At least one third of each specimen collected was frozen at -70 degrees C for further mRNA isolation and reverse transcription into cDNA for real-time-PCR procedures. Geometric averaging of multiple housekeeping genes normalized MMP-9 mRNA expression level.
Polymorphonuclear neutrophils, macrophages and lymphocytes presented MMP-9 positive immunostaining in both types of lesions. When present, epithelial cells were also stained. The number and the ratio of MMP-9(+)/total cells were greater in nonepithelialized than epithelialized lesions (P = .0001) presenting a positive correlation to CD68(+)/total cells (P = .045). Both types of lesions presented increased MMP-9 expression (P < .0001) when compared to healthy periapical ligaments. However, no significant differences were observed for MMP-9 mRNA expression between ephithelized and nonephithelized lesions.
The present data suggest the participation of several inflammatory cells, mainly CD68(+) cells, in the MMP-9 expression in apical periodontitis lesions. MMP-9 could be actively enrolled in the extracellular matrix degradation in apical periodontitis lesions.
Inflammatory and developmental cysts of the jaws are relatively common bone destructive lesions in the human maxillofacial skeleton but their pathogenesis is still poorly understood. In this study the role of mast cells (MC), and mast cell tryptase in particular, was evaluated in the pathophysiology of bone resorption and jaw cyst formation in different types of cysts. The distribution of MC and the amount of tryptase in histological tissue sections were determined by immunohistochemistry using monoclonal antihuman tryptase antibodies and the results were quantitated by using an image analyzing system. The amount of tryptase was further studied by Western-blotting and measurement of trypsin-like activity from the neutral salt extracts obtained from different types of jaw cysts. In contrast to control tissue, high trypsin-like activities and abundant immunoreactive tryptase were observed in the extracts of all types of cysts studied (radicular, dentigerous and keratocyst). In tissue sections the highest amount of tryptase-positive staining was observed in radicular cysts (mean 6.2% of reference area) and the lowest amount in keratocysts (mean 2.1% of reference area, P < 0.01). MC were found to be located in inflammatory cell-rich tissue areas and just beneath the cyst epithelium. Importantly, MC located at the border of bone were observed to be degranulated, indicating high activity of MC and release of tryptase at the regions of early bone destruction. Based on previous findings addressing the role of mast cell tryptase in proteolytic cascades, and the known association of MC with osteoporosis, we suggest that mast cells and mast cell tryptase may contribute significantly to jaw cyst tissue remodelling during growth of a cyst, and to the destruction of the surrounding bone, resulting in jaw cyst expansion.
The exact molecular mechanisms of the loosening of a dental implant are not well-known. The characteristics of implant sulci are similar to those of periodontal sulci regarding gingival crevicular fluid (GCF) and peri-implant sulcular fluid (PISF). Proteolytic enzymes, matrix metalloproteinases (MMPs), participate in peri-implant tissue remodeling. Clodronate is a well-tolerated bisphosphonate-group drug currently used in bone-resorption-related diseases in humans. The mechanisms of bisphosphonate action are not clarified. Collagenase activity in diseased PISF was significantly higher than in the clinically healthy group. Immunoblotting disclosed that diseased PISF contained increased immunoreactives MMP-8 compared with the healthy PISF. The residual latent collagenase activity in the diseased PISF was activated by gold thioglucose and inhibited completely by 100 microM of doxycycline closely resembling pure neutrophil collagenase (MMP-8). The presence of MMP-8 in diseased but not in clinically healthy PISF may prove to be a useful biochemical indicator to monitor peri-implant health and disease. Pure human neutrophil collagenase (MMP-8) and the MMP-8 present in PISF and in the GCF of both loosening implants and periodontitis-affected teeth were efficiently inhibited in vitro by clodronate (50% inhibition [IC50] was achieved by 150 microM of clodronate), an osteoactive, antiresorptive bisphosphonate. Furthermore, the new finding suggests an extended and hitherto-undescribed potential for clodronate in preventing the loosening of both implants and teeth, based on a dual beneficial effect: prevention of both bone resorption/osteolysis and of soft tissue/dental ligament destruction. Potential new therapeutic indications based on the collagenase-inhibiting effect of clodronate provide potential new therapeutic indications for a variety of diseased involving connective tissue breakdown, such as periodontal disease, arthritides, and tumor invasion.
To investigate the mechanisms involved in expansion of radicular cysts, monoclonal antibodies against interstitial collagenase (MMP-1) and tissue inhibitor of metalloproteinases-1 (TIMP-1) were used to localize the sites of MMP-1 and TIMP-1 expression in 30 radicular cysts. Positive MMP-1 staining was detected in the lining epithelium and subepithelial fibroblasts, macrophages, endothelial cells and osteoblasts/osteocytes in all specimens. Positive TIMP-1 staining was identified in osteoblasts/osteocytes and endothelial cells of all specimens, and in the lining epithelium and subepithelial fibrous connective tissue wall of five radicular cysts with an intense inflammatory cell infiltrate. The number and distribution of positive cells for MMP-1 or TIMP-1 varied widely among individual specimens, but strong immunostaining was constantly detected at sites with prominent subepithelial inflammation. Results here support the hypothesis that MMP-1 may play an important role in the expansion of radicular cysts. The absence of TIMP-1 expression in lining epithelium and subepithelial fibroblasts and macrophages in most cases studied indicated that an imbalance between MMP-1 and TIMP-1 production may lead to radicular cyst expansion.
During the past few decades several authors have perpetuated the notion that nearly half of all periapical lesions are radicular cysts. A few studies, based on meticulous serial sectioning of periapical lesions retrieved in toto, have shown that the actual incidence of radicular cyst is only about 15% of all periapical lesions. Equally significant was the discovery in 1980 and recent confirmation that radicular cysts exist in two structurally distinct classes namely, those containing cavities completely enclosed in epithelial lining (periapical true cysts) and those containing epithelium-lined cavities that are open to the root canals (periapical pocket cysts). From a clinical point of view a periapical pocket cyst may heal after conventional root canal therapy whereas an apical true cyst is less likely to be resolved without surgical intervention.
Bisphosphonates are a group of drugs capable of inhibiting bone resorption, and are thus used for the treatment of bone diseases, such as Paget's disease, osteoporosis, and for bone metastases of malignant tumors. Their primary cellular target is considered to be the osteoclast. The molecular mechanisms responsible for the downregulation of bone resorption by bisphosphonates have remain unclear. We have discovered that various matrix metalloproteinases (MMPs) are inhibited in vitro by several bisphosphonates. This novel finding may, in part, explain the efficacy of bisphosphonates in their current indications in humans. In enzyme activity tests using purified and recombinant enzymes, we have observed the inhibition of MMP-1, -2, -3, -7, -8, -9, -12, -13, and -14 by clondronate, alendronate, pamidronate, zolendronate, nedrinate, and clodrinate. The IC50s range from 50 to 150 microM. We have also shown that clodronate can downregulate the expression of MT1-MMP protein and mRNA in several cell lines. Additionally, several bisphosphonates decrease the degree of invasion of malignant melanoma (C8161) and fibrosarcoma (HT1080) cells through artificial basement membrane (Matrigel) in cell cultures at IC50s of 50-150 microM and below. Having low toxicity and proven to be well tolerated after several years in human use, bisphosphonates have the potential to become one of the main MMP-inhibitors for MMP-related human soft and hard tissue-destructive diseases in the near future.
The collagen in the walls of 15 keratocysts was studied histochemically by staining sections with picrosirius red and examining them with polarizing microscopy. This was compared to 15 cases of dentigerous cyst and 15 cases of radicular cyst. Polarization colours of the collagen fibres were recorded according to their width. No differences were found between the polarization colours of thin fibres (<0.8 microm) in all three lesions; the polarization colours of thick fibres (1.6-2.4 microm) in keratocysts were significantly more greenish-yellow when compared with those of dentigerous cysts and radicular cysts. The staining of the collagen fibres in the keratocysts is similar to that reported in odontogenic neoplasms, which suggests that the stroma of keratocysts could be regarded not just as a structural support of the cyst wall, but as playing a part in the neoplastic behaviour of the cyst.
Interleukin-1alpha (IL-1alpha) and matrix metalloproteinase-9 (MMP-9) are thought to be involved in odontogenic cyst expansion. In this study, we investigated the effects of IL-1alpha on the secretion and activation of MMP-9 in odontogenic jaw cysts. An active form of MMP-9 was present in odontogenic keratocyst (6 of 8 cases) fluids more frequently than dentigerous cyst (3 of 10 cases) and radicular cyst (3 of 10 cases) fluids, although proMMP-9 was present in all cyst fluids. Odontogenic keratocyst fragments in explant culture secreted a larger amount of IL-1alpha than dentigerous cyst and radicular cyst fragments in explant culture, and spontaneously secreted both proMMP-9 and an active form of MMP-9. The fragments of dentigerous cysts and radicular cysts secreted a small amount of proMMP-9, but no active form of MMP-9. Exogenously added recombinant human IL-1alpha (rhlL-1alpha) increased the secretion and activation of proMMP-9 in the fragments of dentigerous cysts and radicular cysts. The epithelial cells isolated from odontogenic keratocysts secreted IL-1alpha and proMMP-9 without stimulation. Under the cultivation on a fibronectin-coated dish, rhIL-1alpha increased the secretion of proMMP-9 from the epithelial cells in a dose-dependent manner. Moreover, rhIL-1alpha induced the secretion of proMMP-3 and plasminogen activator urokinase (u-PA) from the epithelial cells, and converted the secreted proMMP-3 to the active form in the presence of plasminogen. The secreted proMMP-9 was also activated in the presence of rhIL-1alpha and plasminogen. Hence, our results suggest that IL-1alpha may up-regulate not only proMMP-9 secretion but also proMMP-9 activation by inducing proMMP-3 and u-PA production in the cyst epithelial cells by autocrine/paracrine regulatory mechanisms.
The concentrations of doxycycline and 4-de-dimethylaminotetracycline required to inhibit 50% of collagenase activity were
found to be 15 to 30 microM for human neutrophil and gingival crevicular fluid collagenases. Fibroblast collagenase was relatively
resistant to inhibition by tetracyclines; the 50% inhibitory concentrations of doxycycline and 4-de-dimethylaminotetracycline
were 280 and 510 microM, respectively.
Matrix metalloproteinases are an important group of zinc enzymes responsible for degradation of the extracellular matrix components such as collagen and proteoglycans in normal embryogenesis and remodeling and in many disease processes such as arthritis, cancer, periodontitis, and osteoporosis. A matrixin family is defined, comprising at least seven members that range in size from Mr 28,000 to 92,000 and are related in gene sequence to collagenase. All family members are secreted as zymogens that lose peptides of about 10,000 daltons upon activation. Latency is due to a conserved cysteine that binds to zinc at the active center. Latency is overcome by physical (chaotropic agents), chemical (HOCl, mercurials), and enzymatic (trypsin, plasmin) treatments that separate the cysteine residue from the zinc. Expression of the metalloproteinases is switched on by a variety of agents acting through regulatory elements of the gene, particularly the AP-1 binding site. A family of protein inhibitors of Mr 28,500 or less binds strongly and stoichiometrically in noncovalent fashion to inhibit members of the family. The serum protein alpha 2-macroglobulin and relatives are also strongly inhibitory.
Collagenase in human neutrophils is found within intracellular granules which can be stimulated to be secreted with phorbol myristic acetate. This extracellular secreted form of neutrophil collagenase was isolated by immunoaffinity chromatography using a monoclonal antibody previously shown to specifically recognize neutrophil collagenase. The enzyme efficiently bound to this column and was eluted with NaSCN as three major species of 75, 57, and 22 kDa, respectively. These proteins were closely related immunologically since, after radiolabeling and separation by gel filtration, each of the three proteins was precipitated by the monoclonal antibody. Also, the 75- and 57-kDa proteins exhibited collagenase activity after elution from polyacrylamide gels run under nondenaturing conditions. Further, the 57-kDa protein autodegraded into a 22-kDa protein with time. Polyclonal antibody, prepared to the 57-kDa enzyme, also recognized the 75- and 22-kDa proteins using an immunoblot technique. When crude neutrophil supernatants containing latent collagenase were immunoblotted, both the 75- and the 57-kDa enzymes were present. Our immunoaffinity purified active enzymes, although activated during the course of purification, resemble the latent enzymes in crude neutrophil supernatants. The multiple forms of secreted collagenase from degranulated leukocytes may resemble more closely that seen in inflammation.
A simple and efficient procedure was employed for the electrophoretic transfer of proteins from sodium dodecyl sulfate-polyacrylamide gels to sheets of pure, unmodified nitrocellulose. Immobilized proteins could then be radiographically visualized in situ by reaction with specific antibody and the subsequent binding of radioiodinated Staphylococcus protein A to the immune complexes. The detection of murine leukemia virus antigens in complex cellular lysates was used to demonstrate the efficacy of this technique.
Matrix metalloproteinases (MMPs) are a family of nine or more highly homologous Zn(++)-endopeptidases that collectively cleave most if not all of the constituents of the extracellular matrix. The present review discusses in detail the primary structures and the overlapping yet distinct substrate specificities of MMPs as well as the mode of activation of the unique MMP precursors. The regulation of MMP activity at the transcriptional level and at the extracellular level (precursor activation, inhibition of activated, mature enzymes) is also discussed. A final segment of the review details the current knowledge of the involvement of MMP in specific developmental or pathological conditions, including human periodontal diseases.
High animal collagenase (EC 3.4.24.7) activity was detected together with gelatinolytic activity directly in human radicular cyst fluids and in 4 M urea extracts of cyst wall tissues. Both Both cyst wall and cyst fluid collagenases were essentially in latent form and were most effectively activated by p-aminophenylmercuric acetate (APMA). Both APMA-activated collagenases broke down type I collagen preferentially to type III collagen resembling these of polymorphonuclear leukocyte origin.
Proteolytic potential of matrix metalloproteinases (interstitial collagenase, gelatinase A and gelatinase B) and serin proteinases (cathepsin G and PMN elastase) were examined in 15 interface tissues between bone and implants in loose total hip replacement prostheses. The results were compared to those of 7 samples of non-inflammatory control synovial tissues. Functional enzyme activities of interstitial collagenase, gelatinase A, cathepsin G and PMN elastase were higher in the interface tissues than those in the control tissues. Induction of gelatinase B was found in the interface tissues. These results suggest that elevated proteolytic potential in the periprosthetic tissues causes extracellular matrix degradation around total hip replacement prostheses, which leads to the loosening of these implants.
Mammalian collagenase (EC 3.4.24.7) was extracted from human keratocyst capsule. The ability of human keratocyst capsule collagenase to degrade soluble interstitial collagens, types I, II and III, was studied in order to elucidate the cellular source(s) of collagenase. Clear differences in the susceptibility of interstitial collagens to human keratocyst capsule collagenase was observed, i.e., type I and II collagens were degraded at equal rate and considerably faster than type III collagen, which was only negligibly degraded. The results indicate that collagenase in human keratocyst capsule extract has characteristics closely resembling those exhibited by human polymorphonuclear leukocyte collagenase rather than fibroblast collagenase. Collagenase secreted by polymorphonuclear leukocytes appears to be the main type of collagenase present in human keratocyst capsule, although the presence of a specific keratocyst capsule collagenase with similar substrate specificity cannot be excluded.
Mammalian collagenases (EC 3.4.24.7) have been suggested as playing an essential role in the initiation of the collagen degradation in periodontal diseases. Two distinct types of interstitial collagenases have been characterized in vertebrate tissues. These enzymes, the fibroblast- and the neutrophil-type collagenases, differ in molecular weight and antigenic properties, as well as substrate specificity and mechanism of activation. In order to determine the cellular origin and mode of action of collagenase in periodontal tissue, we studied the molecular size, the substrate specificity and the activation of collagenases partially purified from inflamed human gingival extracts, sulcular fluid, gingival explant culture medium and polymorphonuclear leukocytes (PMN). Types I, II and III collagens used as substrates were purified from bovine tendon, cartilage and amnion membrane, respectively. Apparent molecular weights of 70–75 k were obtained for gingival extract, sulcular fluid and PMN collagenases and 45 k for gingival explant culture collagenase by gel filtration technique. The gingival extract and sulcular fluid collagenases as well as PMN collagenase could be activated by gold thioglucose and gold thiomalate: no activation of gingival explant culture collagenase was noted. The gingival extract collagenase, sulcular fluid collagenase and PMN collagenase degraded preferentially types I and II collagens relative to type-III collagen. In contrast, gingival explant culture collagenase degraded preferentially types I and III collagens relative to type-II collagen. The results indicate that collagenase in extracts of inflamed human gingiva and in sulcular fluid during inflammation is mostly derived from PMN cells. On the other hand, collagenase produced by gingival explants in culture is probably synthesized by fibroblasts.
Thesis--University of Helsinki, 1989. Includes bibliographical references (p. 53-62).
The fibrinolytic activity in the tissues of cystic lesions of jaw bones was investigated by the semiquantitative method of Astrup. The specimens examined were ameloblastoma (9 cases), follicular cyst (8 cases) and radicular cyst (14 cases). High fibrinolytic activity was observed in ameloblastoma, while in radicular cyst the activity was variable. It is suggested that in radicular cyst inflammatory episodes play an important role in activating local fibrinolysis, while in ameloblastoma the tissue itself has a great capacity to induce locally activated plasmin.
Degradation of cartilage in rheumatoid arthritis (RA) may be in part due to release of collagenase from specific granules of polymorphonuclear neutrophil leukocytes (PMNs) during degranulation. We decided to study, not synovial fluid (SF) collagenase, but PMN collagenase reserves. PMN were isolated from parallel SF and peripheral blood (PB) samples obtained from 7-arthritis patients. PMNs were stimulated in vitro by tetradecanoyl-phorbol-13-acetate (TPA). Collagenase activity in the supernatant without and with phenylmercuric chloride activation was studied. Compared to PB PMNs, there was no consistent decrease in the total collagenase reserves in the inflammatory SF PMNs. This suggests that the release of collagenase in the inflammatory synovial fluid does not deplete SF PMNs of the collagenase synthesized at the myelocyte stage. The role of PMN collagenase in pathogenesis of cartilage destruction would then seem to be more dependent on local release and autoactivation at cartilage surface by adherent PMNs and not excessive collagenase release from free floating SF PMNs at single cell level. Furthermore, under the experimental conditions used the proportion of collagenase released in active form was higher in SF PMN specimens than in PB PMN specimens (p less than 0.01). The predominant collagenous component of adult cartilage, native type II collagen, was degraded by PMN collagenase as fast as native type I collagen. These findings suggest an important role for this enzyme in destruction of the free cartilage surface in RA.
Latent human fibroblast collagenase (HFC) can be activated by a variety of seemingly disparate means. In addition to the well-characterized activation by trypsin and organomercurial compounds, the enzyme can be activated to various extents by surfactants such as sodium dodecyl sulfate, by chaotropic ions such as SCN-, by disulfide compounds such as oxidized glutathione, by sulfhydryl alkylating agents such as N-ethylmaleimide, and by oxidants such as NaOCl. The underlying basis for these activations is the modification, exposure, or proteolytic release of the Cys73 residue from its habitat in the latent enzyme where it is thought to be complexed to the active-site zinc atom. This residue is not accessible for reaction with small molar excesses of dithionitrobenzoate in native, latent HFC. However, on addition of EDTA, this residue becomes fully exposed and is quantitatively labeled. All modes of activation of latent HFC are believed to involve the dissociation of Cys73 from the active-site zinc atom and its replacement by water, with the concomitant exposure of the active site. This is thought to be the primary event that precedes the well-known autolytic cleavages that are observed following the appearance of collagenase activity. The dissociation of Cys73 from the zinc atom in the latent enzyme "switches" the role of the zinc from a noncatalytic to a catalytic one. This "cysteine switch" mechanism of regulation may be applicable to the entire collagenase gene family.
The ability of various reactive oxygen species and serine proteases to activate latent collagenase (matrix metalloproteinase-1) purified from human neutrophils was examined. Latent 70-75 kD human neutrophil collagenase (HNC) was efficiently activated by known non-proteolytic activators phenylmercuric chloride (an organomercurial compound) and gold thioglucose (Au(I)-salt). Corresponding degree of activation was achieved by reactive oxygen species including hypochlorous acid (HOCl), hydrogen peroxide (H2O2) and hydroxyl radical generated by hypoxanthine/xanthine oxidase (HX/XAO). The presence of trace amounts of iron and EDTA were necessary and even enhanced H2O2 induced activation of latent HNC. This activation could be abolished by an iron chelator desferrioxamine and a hydroxyl radical scavenger mannitol. HOCl induced activation of latent HNC was not affected by desferrioxamine and mannitol. Thus, these compounds do not inhibit the active/activated form of HNC. Latent HNC could also be activated by trypsin and chymotrypsin but not by plasmin and plasma kallikrein. The ability of mannitol and desferrioxamine to inhibit the H2O2-induced activation of HNC suggests the transition metal dependent Fenton reaction to be responsible for localized and/or site-specific generation of hydroxyl radical/hydroxyl radical -like oxidants to act as the activating oxygen species. Our results support the ability of myeloperoxidase derived HOCl to act as a direct oxidative activator of HNC and further suggest the existence of a new/alternative oxidative activation pathway of HNC involving hydroxyl radical.
Inactivation of the plasma serine-proteinase inhibitor alpha 1-antitrypsin (alpha 1-AT) by neutrophil metalloproteinases has been reported [Vissers, George, Bathurst, Brennan & Winterbourn (1987) Fed. Proc. Fed. Am. Soc. Exp. Biol. 46, 1390a; (1988) J. Clin. Invest. 82, 706-711; Desrochers & Weiss (1988) J. Clin. Invest. 81, 1646-1650]. To identify the enzyme responsible, supernatant from neutrophils stimulated with phorbol 12-myristate 13-acetate was subjected to preparative SDS/PAGE, both with and without activation of latent metalloproteinases with HgCl2. The lanes were subsequently sliced into pieces, the slices incubated with equimolar amounts of type I collagen and alpha 1-AT in the presence of HgCl2, and the reaction products separated by SDS/PAGE. With the latent supernatant, the characteristic collagen-cleavage products and cleaved alpha 1-AT were present in the same slices, corresponding to an Mr of 80,000-85,000. On treatment with HgCl2 both degradative activities underwent the same molecular-mass shift to a position corresponding to Mr 60,000-65,000. Western blots of neutrophil supernatants, using a polyclonal antibody to purified collagenase, showed Mr values of 83,000 for the latent enzyme and 63,000 for the HgCl2-activated enzyme. Neutrophil collagenase was purified to homogeneity and shown also to exist in a second latent form with Mr 70,000. When activated to the Mr-63,000 form by HgCl2 and incubated with equimolar amounts of collagen and alpha 1-AT, collagenase cleaved alpha 1-AT at almost twice the rate at which collagen was cleaved. alpha 1-AT cleavage was inhibited by 1,10-phenanthroline and by high concentrations of collagen. That the purified collagenase did not contain a contaminant proteinase such as stromelysin was indicated by inability of the preparation to cleave casein. Taken together these results lead us to conclude that neutrophil collagenase is capable of degrading alpha 1-AT. Neutrophil gelatinase also cleaved alpha 1-AT, but cleavage was slow when compared with its activity against gelatin.
Odontogenic cyst capsules were cultured in vitro and the culture media analysed for bone-resorption and interleukin 1-like activity. Five cysts synthesised a non-dialysable bone resorbing factor with significant interleukin 1-like activity. One specimen thought to be a cyst with little interleukin 1 activity proved to be antral lining. The results indicate that interleukin 1 may play an important role in cyst expansion by its direct effects on fibroblast proliferation and bone resorption and by stimulating prostaglandin synthesis in stromal fibroblasts of the cyst capsule.
This chapter discusses the role of plasminogen activators in various biological processes. In specific, it describes two types of plasminogen activators—namely, the urokinase-type plasminogen activator (u-PA) and the tissue-type plasminogen activator (t-PA), which are essentially different gene products. The amino acid sequences of these activators and nucleotide sequences of the corresponding cDNAs have largely been determined, and the cDNAs have been cloned using recombinant techniques. A variety of enzymatic as well as immunological assay and detection methods have also been developed that allows a precise quantification of the activators, a distinction between u-PA and t-PA, determination of whether an activator is present in its active or zymogen form, analysis of the kinetics of different steps of the cascade reaction, and immunocytochemical identification of u-PA and t-PA in tissue sections. Much of the studies on plasminogen activators and cancer has been guided by the hypothesis that proteolysis of the components of extracellular matrix, initiated by the release of plasminogen activator from the cancer cells, plays a decisive role for the degradation of normal tissue, and thereby for invasive growth and metastases.
The synthesis of prostaglandins by dental squamous cell cysts was studied. Radiothinlayer-chromatography (RTLC) showed C14-labelled arachidonic acid to be converted into the prostaglandins PGE2, 6-oxo-PGF1 alpha, PGF2 alpha and PDG2. In addition, hydroxyfatty acid synthesis in excess of total prostaglandin production was observed. The substances mentioned are thought to be causally involved in the osteolytic activity of dental cysts.
If prostaglandins play a role in bone resorption by dental cysts they must presumably be released at a site accessible to the bone. Bone resorption is not significantly less when the calvaria are incubated with cyst capsule rather than with full thickness cyst wall. The bone resorbing factors therefore seem to be released by the outer capsule adjacent to the bone rather than by the lining epithelium. These observations support the possibility that prostaglandins (perhaps mainly PGE2) produced by dental cyst capsule are responsible for bone resorption by cysts within the jaws.
Collagenolytic activity in human carious and non-carious dentine matrix was compared. Results confirmed the presence of latent collagenase in demineralized dentine and indicated a slow rate of degradation of collagen substrate. Collagenolytic activity was enhanced with the addition of trypsin-TPCK to the demineralized dentine. More activity was observed in the carious dentine, suggesting the presence of collagenase activators or partial enzymic destruction of the inhibitor in the collagen-collagenase-inhibitor complex. It seems that during dentine development collagenase-inhibitor complex is secreted and bound to collagen-dentine matrix, and the enzyme can be activated during the process of dental caries.
We studied the in vivo effect of long-term doxycycline treatment combined with NSAID on human interstitial collagenases, other matrix metalloproteinases, serine proteinases, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) and lactoferrin from saliva and serum during the course of acute reactive arthritis (ReA). Collagenase activity and serine proteases (elastase-like, cathepsin G-like and trypsin-like activities) of saliva (n = 10) and gelatinase, lactoferrin and TIMP-1 of saliva (n = 10) and serum (n = 10) samples before and after 2 months doxycycline treatment, combined with NSAID, were studied by quantitative SDS-PAGE assay, ELISA assay and by spectrophotometric assay. The cellular source and molecular forms of salivary collagenase were characterized by immunoblotting using specific antisera. We found that activities of total and endogenously active interstitial collagenase reduced significantly. The salivary collagenase was found to originate from neutrophils. No fragmentation of either pro 75-kD and active 65-kD MMP-8 was detected after 2 months doxycycline treatment. However, during 2 months doxycycline and NSAID treatment no reduction of salivary and serum gelatinase, lactoferrin and TIMP-1-levels and salivary serine protease activities were detected. The in vivo inhibition of collagenase (MMP-8) activity during long-term doxycycline therapy in human saliva containing inflammatory exudate of ReA patients may contribute to the reduced tissue destruction observed in recent clinical and animal model studies in arthritides during long-term doxycycline/tetracycline treatment.
Matrix metalloproteinases (MMPs) are a family of nine or more highly homologous Zn++endopeptidases that collectively cleave most if not all of the constituents of the extracellular matrix. The present review discusses in detail the primary structures and the overlapping yet distinct substrate specificities of MMPs as well as the mode of activation of the unique MMP precursors. The regulation of MMP activity at the transcriptional level and at the extracellular level (precursor activation, inhibition of activated, mature enzymes) is also discussed. A final segment of the review details the current knowledge of the involvement of MMP in specific developmental or pathological conditions, including human periodontal diseases.
