High MMP-9 activity characterizes pleural tuberculosis correlating with granuloma formation

Tuberculosis (TB) pleural disease is complicated by extensive tissue destruction. Matrix metalloproteinase (MMP)-1 and -9 are implicated in immunopathology of pulmonary and central nervous system TB. There are few data on MMP activity in TB pleurisy. The present study investigated MMP-1, -2 and -9 and their specific inhibitors (tissue inhibitor of metalloproteinase (TIMP)-1 and -2) in tuberculous effusions, and correlated these with clinical and histopathological features. Clinical data, routine blood tests, and pleural fluid/biopsy material were obtained from 89 patients presenting with pleural effusions in a TB-endemic area. MMP-1, -2 and -9 were measured by zymography or western blot, and TIMP-1 and -2 by ELISA. Pleural biopsies were examined microscopically, cultured for acid–alcohol fast bacilli and immunostained for MMP-9. Tuberculous pleural effusions contained the highest concentrations of MMP-9 compared with malignant effusions or heart failure transudates. MMP-9 concentrations were highest in effusions from patients with granulomatous biopsies: median (interquartile range) 108 (61–218) pg·mL-1 versus 43 (12–83) pg·mL-1 in those with nongranulomatous pleural biopsies. MMP-1 and -2 were not upregulated in tuberculous pleural fluid. The ratio of MMP-9:TIMP-1 was significantly higher in TB effusions. Tuberculous pleurisy is characterised by a specific pattern of matrix metalloproteinase-9 upregulation, correlating with the presence of granulomas and suggesting a specific role for matrix metalloproteinase-9 in inflammatory responses in tuberculous pleural disease.

STAT3, p38 MAP Kinase and NF-{kappa}B Drive Unopposed Monocyte-dependent Fibroblast MMP-1 Secretion in Tuberculosis.

Tissue destruction characterizes infection with Mycobacterium tuberculosis (Mtb). Type I collagen provides the lung's tensile strength, is extremely resistant to degradation, but is cleaved by matrix metalloproteinase (MMP)-1. Fibroblasts potentially secrete quantitatively more MMP-1 than other lung cells. We investigated mechanisms regulating Mtb-induced collagenolytic activity in fibroblasts in vitro and in patients. Lung fibroblasts were stimulated with conditioned media from Mtb-infected monocytes (CoMTb). CoMTb induced sustained increased MMP-1 (74 versus 16 ng/ml) and decreased tissue inhibitor of metalloproteinase (TIMP)-1 (8.6 versus 22.3 ng/ml) protein secretion. CoMTb induced a 2.7-fold increase in MMP-1 promoter activation and a 2.5-fold reduction in TIMP-1 promoter activation at 24 hours (P = 0.01). Consistent with this, TIMP-1 did not co-localize with fibroblasts in patient granulomas. MMP-1 up-regulation and TIMP-1 down-regulation were p38 (but not extracellular signal–regulated kinase or c-Jun N-terminal kinase) mitogen-activated protein kinase–dependent. STAT3 phosphorylation was detected in fibroblasts in vitro and in tuberculous granulomas.STAT3 inhibition reduced fibroblast MMP-1 secretion by 60% (P = 0.046). Deletion of the MMP-1 promoter NF-B–binding site abrogated promoter induction in response to CoMTb. TNF-, IL-1ß, or Oncostatin M inhibition in CoMTb decreased MMP-1 secretion by 65, 63, and 25%, respectively. This cytokine cocktail activated the same signaling pathways in fibroblasts and induced MMP-1 secretion similar to that induced by CoMTb. This study demonstrates in a cellular model and in patients with tuberculosis that in addition to p38 and NF-B, STAT3 has a key role in driving fibroblast-dependent unopposed MMP-1 production that may be key in tissue destruction in patients.

Salbutamol up-regulates matrix metalloproteinase-9 in the alveolar space in the acute respiratory distress syndrome.

Objectives: Acute respiratory distress syndrome (ARDS) is characterized by alveolar-capillary barrier damage. Matrix metalloproteinases (MMPs) are implicated in the pathogenesis of ARDS. In the Beta Agonists in Acute Lung Injury Trial, intravenous salbutamol reduced extravascular lung water (EVLW) in patients with ARDS at day 4 but not inflammatory cytokines or neutrophil recruitment. We hypothesized that salbutamol reduces MMP activity in ARDS.

Methods: MMP-1/-2/-3/-7/-8/-9/-12/-13 was measured in supernatants of distal lung epithelial cells, type II alveolar cells, and bronchoalveolar lavage (BAL) fluid from patients in the Beta Agonists in Acute Lung Injury study by multiplex bead array and tissue inhibitors of metalloproteinases (TIMPs)-1/-2 by enzyme-linked immunosorbent assay. MMP-9 protein and activity levels were further measured by gelatin zymography and fluorokine assay.

Measurements and Main Results: BAL fluid MMP-1/-2/-3 declined by day 4, whereas total MMP-9 tended to increase. Unexpectedly, salbutamol augmented MMP-9 activity. Salbutamol induced 33.7- and 13.2-fold upregulation in total and lipocalin-associated MMP-9, respectively at day 4, compared with 2.0- and 1.3-fold increase in the placebo group, p < 0.03. Salbutamol did not affect BAL fluid TIMP-1/-2. Net active MMP-9 was higher in the salbutamol group (4222 pg/mL, interquartile range: 513-7551) at day 4 compared with placebo (151 pg/mL, 124-2108), p = 0.012. Subjects with an increase in BAL fluid MMP-9 during the 4-day period had lower EVLW measurements than those in whom MMP-9 fell (10 vs. 17 mL/kg, p = 0.004): change in lung water correlated inversely with change in MMP-9, r = -.54, p = 0.0296. Salbutamol up-regulated MMP-9 and down-regulated TIMP-1/-2 secretion in vitro by distal lung epithelial cells. Inhibition of MMP-9 activity in cultures of type II alveolar epithelial cells reduced wound healing.

Conclusions: Salbutamol specifically up-regulates MMP-9 in vitro and in vivo in patients with ARDS. Up-regulated MMP-9 is associated with a reduction in EVLW. MMP-9 activity is required for alveolar epithelial wound healing in vitro. Data suggest MMP-9 may have a previously unrecognized beneficial role in reducing pulmonary edema in ARDS by improving alveolar epithelial healing.

Oxidative stress and increased type-IV collagenase levels in bronchoalveolar lavage fluid from newborn babies

Oxidative stress may increase lung permeability by upregulation of matrix-metalloproteinase-9 (MMP-8), a type-IV collagenase that can disrupt alveolar basement membranes. We have compared a marker of oxidative stress (protein carbonyl residues) with levels of MMP-8 and its inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1), in bronchoalveolar lavage samples from newborn babies. Bronchoalveolar lavage samples (n = 87, two from each time point) were taken in the first 6 postnatal days from 41 ventilated babies: 18 of

Comprehensive profiling and localisation of the matrix metalloproteinases in urothelial carcinoma

The matrix metalloproteinases (MMPs) are endopeptidases which break down the extracellular matrix and regulate cytokine and growth factor activity. Several MMPs have been implicated in the promotion of invasion and metastasis in a broad range of tumours including urothelial carcinoma. In this study, RNA from 132 normal bladder and urothelial carcinoma specimens was profiled for each of the 24 human MMPs, the four endogenous tissue inhibitors of MMPs (TIMPs) and several key growth factors and their receptors using quantitative real time RT-PCR. Laser capture microdissection (LCM) of RNA from 22 tumour and 11 normal frozen sections was performed allowing accurate RNA extraction from either stromal or epithelial compartments. This study confirms the over expression in bladder tumour tissue of well-documented MMPs and highlights a range of MMPs which have not previously been implicated in the development of urothelial cancer. In summary, MMP-2, MT1-MMP and the previously unreported MMP-28 were very highly expressed in tumour samples while MMPs 1, 7, 9, 11, 15, 19 and 23 were highly expressed. There was a significant positive correlation between transcript expression and tumour grade for MMPs 1, 2, 8, 10, 11, 12, 13, 14, 15 and 28 (P < 0.001). At the same confidence interval, TIMP-1 and TIMP-3 also correlated with increasing tumour grade. LCM revealed that most highly expressed MMPs are located primarily within the stromal compartment except MMP-13 which localised to the epithelial compartment. This work forms the basis for further functional studies, which will help to confirm the MMPs as potential diagnostic and therapeutic targets in early bladder cancer.

The role of LL-37 in periodontal wound healing

Introduction: As a result of chronic inflammation during periodontal disease the junctional epithelium becomes micro-ulcerated. The inflammatory process is mediated by both bacterial and host cell products. Host defence peptides such as defensins, secretory leucocyte protease inhibitor (SLPI) and the sole human cathelicidin, LL-37, are secreted by both periodontal cells and neutrophils into gingival crevicular fluid (GCF). They have the ability to modulate the immune response in periodontitis and are thought to have a potential role in periodontal wound healing. Objectives: The aims of this study were to determine the role of LL-37 in the production of Interleukin (IL)-8, IL-6, hepatocyte growth factor (HGF) and basic-fibroblast growth factor (bFGF) by gingival fibroblasts. The role of LL-37 in modulating total matrix metalloproteinase (MMP) activity and expression of tissue inhibitors of metalloproteinase (TIMP)-1 and -2 by gingival fibroblasts was also investigated. Methods: Primary gingival fibroblasts were co-cultured with concentrations of LL-37 (1, 5 and 10µg/ml) for 24 hours and their supernatants tested for levels of IL-8 and IL-6, HGF, bFGF, TIMP-1 and TIMP-2 by ELISA. Rates of MMP turnover in the supernatants were tested by fluorogenic assay using fluorescence resonance energy transfer (FRET) peptide substrates. Cytotoxicity was measured by MTT assay. Statistical significance was measured using the independent t-test and p<0.05 was considered significant. Results: LL-37 significantly upregulated levels of IL-8, IL-6, HGF, bFGF and TIMP-1 (p<0.05) in a dose-dependent fashion. LL-37 significantly decreased the total MMP activity (p<0.05). None of the LL-37 concentrations tested were cytotoxic to gingival fibroblasts. Conclusion: These results indicate that LL-37 is involved in periodontal wound healing. LL-37 increased levels of proinflammatory cytokines and increased levels of growth factors involved in re-epithelialisation. LL-37 has the ability to regulate remodelling of the periodontium by controlling MMP overactivity both directly and by stimulating production of inhibitors by gingival fibroblasts.

Metabolically-inactive glucagon-like peptide-1(9-36)amide confers selective protective actions against post-myocardial infarction remodelling

BACKGROUND: Glucagon-like peptide-1 (GLP-1) therapies are routinely used for glycaemic control in diabetes and their emerging cardiovascular actions have been a major recent research focus. In addition to GLP-1 receptor activation, the metabolically-inactive breakdown product, GLP-1(9-36)amide, also appears to exert notable cardiovascular effects, including protection against acute cardiac ischaemia. Here, we specifically studied the influence of GLP-1(9-36)amide on chronic post-myocardial infarction (MI) remodelling, which is a major driver of heart failure progression.

METHODS: Adult female C57BL/6 J mice were subjected to permanent coronary artery ligation or sham surgery prior to continuous infusion with GLP-1(9-36)amide or vehicle control for 4 weeks.

RESULTS: Infarct size was similar between groups with no effect of GLP-1(9-36)amide on MI-induced cardiac hypertrophy, although modest reduction of in vitro phenylephrine-induced H9c2 cardiomyoblast hypertrophy was observed. Whilst echocardiographic systolic dysfunction post-MI remained unchanged, diastolic dysfunction (decreased mitral valve E/A ratio, increased E wave deceleration rate) was improved by GLP-1(9-36)amide treatment. This was associated with modulation of genes related to extracellular matrix turnover (MMP-2, MMP-9, TIMP-2), although interstitial fibrosis and pro-fibrotic gene expression were unaltered by GLP-1(9-36)amide. Cardiac macrophage infiltration was also reduced by GLP-1(9-36)amide together with pro-inflammatory cytokine expression (IL-1β, IL-6, MCP-1), whilst in vitro studies using RAW264.7 macrophages revealed global potentiation of basal pro-inflammatory and tissue protective cytokines (e.g. IL-1β, TNF-α, IL-10, Fizz1) in the presence of GLP-1(9-36)amide versus exendin-4.

CONCLUSIONS: These data suggest that GLP-1(9-36)amide confers selective protection against post-MI remodelling via preferential preservation of diastolic function, most likely due to modulation of infiltrating macrophages, indicating that this often overlooked GLP-1 breakdown product may exert significant actions in this setting which should be considered in the context of GLP-1 therapy in patients with cardiovascular disease.

La Rapamycine inhibe l’expression de l’ARNm de l’ADAMTS-4 induit par les cytokines dans les chondrocytes articulaires

Introduction: Durant la pathogenèse d’ostéoarthrose (OA), les cytokines pro-inflammatoires IL-1β (Interleukin-1 beta) et TNF-α (Tumor necrosis factor alpha) stimulent la dégradation des agrécanes par l’aggrécanase-1 ou ADAMTS-4 (a disintegrin and metalloproteinase with thrombospondin motif). Ces cytokines peuvent stimuler plusieurs voies de signalisation conduisant ainsi à l’augmentation de l’expression des ADAMTS dans les chondrocytes humains. Les TIMPs (tissue inhibitor of metalloproteinases) présentent des inhibiteurs endogènes de l’ADAMTS. Nous avons démontré que la Rapamycine (un immunosuppresseur et un inhibiteur du mamalian target of Rapamycin (mTOR)) peut avoir des effets bénéfiques dans cette pathologie. Notre étude examine l’effet de la Rapamycine sur l’expression de l’ADAMTS-4 induit par les cytokines, son implication dans certaines voies de signalisation, et son effet sur l’expression du TIMP-3. Méthodes: Des chondrocytes normaux sont traités avec la Rapamycine seule ou stimulés aussi avec l’IL-1β et le TNF-α. Les effets de la Rapamycine sur l’expression de l’ADAMTS-4 et du TIMP-3 ont été étudiés par l’analyse RT-PCR et l’activité enzymatique a été étudiée par la technique d’ELISA. Les effets de la Rapamycine sur certaines voies de signalisation ont été étudiés par le Western blot. Résultats: Nous avons trouvé que la Rapamycine inhibe l’expression de l’ARNm de l’ADAMTS-4 induit par les cytokines pro-inflammatoires dans les chondrocytes humains. L’activité enzymatique de l’ADAMTS-4 induit par l’IL-1β a été légèrement diminuée par la Rapamycine. En plus, cette dernière a montré de différents effets sur plusieurs voies de signalisation stimulées par l’IL-1β et le TNF-α telles que les voies des MAPKs (Mitogen activated protein kinase), de l’AKT, et de la p70 S6 kinase. La Rapamycine a inhibé partiellement l’activation de la phosphorylation de l’ERK1/2 MAPK (extracellular signal-regulated protein kinase MAPK) en présence du TNF-α seulement. En outre, la Rapamycine a inhibé la phosphorylation des protéines p38 MAPK, JNK (c-Jun N-terminal kinase), et AKT activée par l’IL-1β seulement. En plus, la phosphorylation de la protéine p70 S6K stimulée par l’IL-1β et le TNF-α a été inhibée par la Rapamycine. D’autre part, nous avons démontré que le niveau du TIMP-3 a été augmenté en présence de la Rapamycine. Conclusion: Ces résultats suggèrent que la Rapamycine peut bloquer l’action de l’ADAMTS-4 via l’inhibition de l’activation des MAPKs, de l’AKT, et de la p70 S6K. La Rapamycine pourrait ainsi être considérée pour la prévention de la perte du cartilage chez les patients ostéoarthritiques.

Étude in vitro de l’implication des cytokines de type Th17 dans la fibrose hépatique

Introduction: L’activation des cellules stellaires hépatiques (CSHs) est un point clé du processus de fibrose hépatique. Les lymphocytes T CD4+ intra-hépatiques sont une source majeure de cytokines anti-inflammatoires comme l’IL-10 et pro-inflammatoire (IL-17A), hépatoprotectrice (IL-22) produites par les Th17. Les Th17 sont impliqués dans de nombreuses pathologies inflammatoires mais l’effet de ces cellules sur les CSHs n’est pas encore élucidé. Objectif: Comprendre le rôle des cytokines de type Th17 dans le processus d’activation des CSHs. Méthodes: La lignée de CSHs humaine LX2 a été stimulée par l’IL-17A ou l’IL-22 puis comparée à des cellules traitées par le TGF-b et le tampon phosphate salin (PBS). L’activation des CSHs a été évaluée en examinant les molécules profibrotique alpha-smooth muscle actin (a-SMA), collagène de type I (COL1A1) et inhibiteur produits par les tissus des métalloprotéases matricielles I (TIMP-I) par q-PCR. L’expression protéique a été validée par immunobuvardage ou coloration au rouge de picro Sirius. L’expression membranaire de l’IL-10Rb, du TGF-b-RII et de l’IL-17RA a été mesurée par cytométrie en flux. Résultats: L’IL-17A et l’IL-22 n’activent pas les cellules LX2, car aucune induction d’a-SMA, de COL1A1 et de TIMP-I n’a été observée. Cependant, l’IL-17A et l’IL-22 sensibilisent les CSHs à l’action du TGF-b, tel que démontré par une forte expression et production d’a-SMA, collagène type I et TIMP-I. L’IL-17A, mais pas l’IL-22, induit la surexpression à la surface cellulaire du TGF-b-RII et inhibe partiellement la baisse d’expression du TGF--RII après stimulation au TGF-b. Conclusion: Nos résultats démontrent une fonction pro-fibrotique de l’IL-17A et de l’IL-22, car les deux cytokines sensibilisent les CSHs à l’action du TGF-b. L’IL-17A agit via la surexpression et la stabilisation du TGF-b-RII tandis que l’IL-22 agit probablement par des mécanismes intracellulaires.

Avaliação do potencial terapêutico de inibidores da Galectina-3 em modelo experimental de lesão hepática crônica

As lesões crônicas do fígado são resultantes de agressões persistentes, onde a desorganização e destruição do tecido podem desencadear processos de regeneração e fibrose. Para que a integridade e homeostase do órgão sejam restauradas, várias vias intracelulares e intercelulares são ativadas. Uma delas é a através da liberação de moléculas pró-fibrogênicas, a exemplo da lectina solúvel bgalactosídea, a galectina-3 (Gal-3). A alta expressão dessa lectina tem sido associada a fibrogênese no fígado. A descoberta de moléculas capazes de se ligar à Gal-3 e inibir a sua ação são importantes no desenvolvimento de terapias antifibrosantes. A pectina cítrica modificada (PCM) e a N-acetilactosamina (LacNAc) demonstraram ação benéfica no tratamento de doenças fibróticas, incluindo renais e cardíacas, contudo, pouco se sabe sobre suas eficácias na fibrose hepática. Diante disso, o objetivo deste estudo foi investigar os efeitos das administrações da PCM e LacNAc quanto aos níveis de Gal-3 e fibrose em modelo experimental de lesão hepática crônica. Inicialmente, a fibrose hepática foi induzida em camundongos C57BL/6 pela administração de tetracloreto de carbono a 20 por cento diluído em azeite de oliva. Grupos de camundongos com dois ou quatro meses de lesão foram tratados com PCM (1 por cento e 5 por cento, fornecida ad libitum) e com LacNAc, por via intraperitoneal. Adicionalmente, camundongos knockouts para o gene da Gal-3 (Gal-3-/-) foram utilizados como um controle. Subsequentemente às intervenções, análises morfométricas, bioquímicas, imunológicas e de biologia molecular foram realizadas. Na análise morfométrica, não se verificou alteração no percentual de tecido fibroso entre os grupos tratados com PCM (1 por cento e 5 por cento) ou LacNAc quando comparados com seus respectivos controles, o que foi confirmado pela dosagem dos níveis de hidroxiprolina. Adicionalmente, não foi observada alteração dos níveis de Gal-3, MMP-9 e TIMP-1 após tratamento com os inibidores. Os resultados obtidos indicaram que a PCM e a LacNAc não foram capazes de inibir a Gal-3 no fígado, e portanto, não interferiram na deposição de tecido fibroso nesse modelo experimental

Daidzein, R-(+)equol and S-(-)equol inhibit the invasion of MDA-MB-231 breast cancer cells potentially via the down-regulation of matrix metalloproteinase-2

PURPOSE: Soy isoflavones may inhibit tumor cell invasion and metastasis via their effects on matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs). The current study investigates the effects of daidzein, R- and S-equol on the invasion of MDA-MB-231 human breast cancer cells and the effects of these compounds on MMP/TIMP expression at the mRNA level. METHODS: The anti-invasive effects of daidzein, R- and S-equol (0, 2.5, 10, 50 μM) on MDA-MB-231 cells were determined using the Matrigel invasion assay following 48-h exposure. Effects on MMP-2, MMP-9, TIMP-1 and TIMP-2 expression were assessed using real-time PCR. Chiral HPLC analysis was used to determine intracellular concentrations of R- and S-equol. RESULTS: The invasive capacity of MDA-MB-231 cells was significantly reduced (by approximately 50-60 %) following treatment with 50 μM daidzein, R- or S-equol. Anti-invasive effects were also observed with R-equol at 2.5 and 10 μM though overall equipotent effects were induced by all compounds. Inhibition of invasion induced by all three compounds at 50 μM was associated with the down-regulation of MMP-2, while none of the compounds tested significantly affected the expression levels of MMP-9, TIMP-1 or TIMP-2 at this concentration. Following exposure to media containing 50 μM R- or S-equol for 48-h intracellular concentrations of R- and S-equol were 4.38 ± 1.17 and 3.22 ± 0.47 nM, respectively. CONCLUSION: Daidzein, R- and S-equol inhibit the invasion of MDA-MB-231 human breast cancer cells in part via the down-regulation of MMP-2 expression, with equipotent effects observed for the parent isoflavone daidzein and the equol enantiomers.

Matrix metalloproteinases, TIMPs and growth factors regulating ameloblastoma behaviour

Aims: Ameloblastoma is an odontogenic neoplasm with local invasiveness and recurrence. We have previously suggested that growth factors and matrix metalloproteinases (MMPs) influence ameloblastoma invasiveness(1). The aim was to study expression of MMPs, tissue inhibitor of metalloproteinases (TIMPs) and growth factors in ameloblastoma. Methods and results: Thirteen cases of solid/multicystic ameloblastoma were examined. As a control, calcifying cystic odontogenic tumour (CCOT), a non-invasive odontogenic neoplasm with ameloblastomatous epithelium was also studied. Immunohistochemistry detected MMPs, TIMPs and growth factors in ameloblastoma and CCOT. The labelling index (LI) of MMP-9 and TIMP-2 was significantly higher in ameloblastoma compared with CCOT. The LI of epidermal growth factor (EGF), transforming growth factor (TGF)-alpha and epidermal growth factor receptor (EGFR) was also increased in ameloblastoma. This neoplasm showed greater expression of MMPs, TIMPs and growth factors compared with CCOT. We then analysed these molecules in ameloblastoma cells and stroma. Ameloblastoma cells exhibited increased LI of MMP-1, -2 and EGFR. We found a positive correlation between EGF and TIMP-1, and between TGF-alpha and TIMP-2. It is known that signals generated by growth factors are transduced by the ERK pathway. Ameloblastoma stroma exhibited the phosphorylated (activated) form of ERK. Conclusions: These results suggest an interplay involving growth factors MMPs and TIMPs that may contribute to ameloblastoma behaviour. Signals generated by this molecular network would be transduced by ERK 1/2 pathway.

Evidences of the cooperative role of the chemokines CCL3, CCL4 and CCL5 and its receptors CCR1+and CCR5+in RANKL+ cell migration throughout experimental periodontitis in mice

Periodontal disease (PD) is characterized by the inflammatory bone resorption in response to the bacterial challenge, in a host response that involves a series of chemokines supposed to control cell influx into periodontal tissues and determine disease outcome. In this study, we investigated the role of chemokines and its receptors in the immunoregulation of experimental PD in mice. Aggregatibacter actinomycetemcomitans-infected C57BI/6 (WT) mice developed an intense inflammatory reaction and severe alveolar bone resorption, associated with a high expression of CCL3 and the migration of CCR5+, CCR1+ and RANKL+ cells to periodontal tissues. However, CCL3KO-infected mice developed a similar disease phenotype than WT strain, characterized by the similar expression of cytokines (TNF-alpha, IFN-gamma and IL-10), osteoclastogenic factors (RANKL and OPG) and MMPs (MMP-1, MMP-2, MMP-3, TIMP-1 and TIMP-3), and similar patterns of CCR1+, CCR5+ and RANKL+ cell migration. The apparent lack of function for CCL3 is possible due the relative redundancy of chemokine system, since chemokines such as CCL4 and CCL5, which share the receptors CCR1 and CCR5 with CCL3, present a similar kinetics of expression than CCL3. Accordingly, CCL4 and CCL5 kinetics of expression after experimental periodontal infection remain unaltered regardless the presence/absence of CCL3. Conversely, the individual absence of CCR1 and CCR5 resulted in a decrease of leukocyte infiltration and alveolar bone loss. When CCR1 and CCR5 were simultaneously inhibited by met-RANTES treatment a significantly more effective attenuation of periodontitis progression was verified, associated with lower values of bone loss and decreased counts of leukocytes in periodontal tissues. Our results suggest that the absence of CCL3 does not affect the development of experimental PD in mice, probably due to the presence of homologous chemokines CCL4 and CCL5 that overcome the absence of this chemokine. In addition, our data demonstrate that the absence of chemokine receptors CCR1+ and CCR5+ attenuate of inflammatory bone resorption. Finally, our data shows data the simultaneous blockade of CCR1 and CCR5 with MetRANTEs presents a more pronounced effect in the arrest of disease progression, demonstrating the cooperative role of such receptors in the inflammatory bone resorption process throughout experimental PD. (C) 2009 Elsevier Inc. All rights reserved.

Inhibition of matrix metalloproteinase activity in the chick sclera and its effect on myopia development

To investigate the contribution of matrix degradation in the two-layer avian sclera to the development of myopia. Tissue inhibitor of metalloproteinase-2 (TIMP-2) was used to inhibit chick scleral collagen degradation with (3)H-proline, a marker for this effect. Ex vivo scleral culture experiments confirmed TIMP-2 doses for in vivo experimentation. Ocular growth and refractive response to exogenous TIMP-2 (11.25, 2.25, and 0.45 picomoles, plus vehicle only) were monitored in 7-day-old chicks during the induction of myopia over 4 days with a translucent occluder. Collagen degradation was assessed, in whole sclera and in separated scleral layers by using the same paradigm (11.25 picomoles TIMP-2; vehicle only).Approximately 60% of collagen degradation was inhibited with low (2 nM) doses of TIMP-2 in the ex vivo sclera. Degradative activity in the in vivo chick sclera increased significantly (46%) during myopia development, with all the altered activity confined to the fibrous layer. Addition of TIMP-2 significantly reduced (by 46%) this accelerated scleral collagen degradation, also by acting in the fibrous layer. TIMP-2 had no significant effect on (3)H-proline incorporated in the cartilaginous scleral layer and cornea. Despite inhibiting collagen degradation TIMP-2 had no significant effect on myopia development. Increased collagen degradation is a feature of scleral remodeling in chick myopia development, but is confined to the fibrous scleral layer. Significant inhibition of this collagenolytic activity with TIMP-2 has little effect on refractive error development, suggesting that collagen degradation in the sclera contributes little to the development of myopia in the chick.

Estudo da expressão imuno-histoquímica das MMPs -2, -7, -9 e -26 e TIMPs -1 e -2 em adenomas pleomórficos e carcinomas adenóides císticos de glândulas salivares menores

The balance between the expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) has been related to various physiological and pathological processes, including salivary gland morphogenesis and tumor invasion and metastasis processes. Pleomorphic adenoma (PA) and adenoid cystic carcinoma (ACC) respectively represent benign and malignant neoplasias of salivary glands. Although they share the same cell origin, they present distinct biological behavior. The aim of this study was to compare the immunohistochemical expression of MMPs -2, -7, -9 and -26, and of TIMPs -1 and -2, in cases of PA and ACC of minor salivary glands. Twenty cases of PA and twenty cases of ACC were assessed according to the presence, intensity and location of MMPs and TIMPs in the tumor parenchyma. Most of the PAs and ACCs presented a high expression of MMP -2, -7, -9 and -26 and of TIMP -1 and -2, predominantly located in tumor cells. There was no significant difference in the expression of MMPs -2 (p=0.359), -7 (p=0.081) and -26 (p=0.553), as well as of TIMPs -1 (p=0.657) and -2 (p=0.248), between the parenchyma of PAs and ACCs. However, MMP-9 showed a significant difference of expression between the two tumors, with the ACC showing more intense marking for this gelatinase (p=0.041). The strong expression of MMP-9 observed in the parenchyma suggests that this gelatinase may play an important role in the biological behavior of these tumors. On the other hand, although there was no significant difference between the marking of MMP -2, 7 and 26 in the studied tumors, the data, when analyzed as a whole, suggest that these proteases may take part in the process of tissue remodeling in both tumors, but do not present a direct relation with the pattern of aggressiveness of ACC. Nonetheless, matrilisins may indirectly influence the behavior of this tumor due to their capacity of activating MMP-9, strongly expressed in the parenchyma of ACC