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Background and aims: The expression of tissue and serum matrix metalloproteinase-7 (MMP-7) was shown to be elevated both in colon cancer and dysplastic lesions. We aimed to evaluate, for the first time, its role as a diagnostic marker in Lynch syndrome (LS) carriers, a hereditary syndrome with predisposition to colon cancer. Methods: This was a case control study. Baseline serum MMP-7 levels were determined by ELISA in 40 colon cancer patients, 62 LS-carriers and 60 healthy controls. Retrieved data from medical files included demographics, background diseases, clinical data regarding tumor characteristics and genetic data. We assessed the association of serum MMP-7 levels with different variables in the study cohort using linear regression model adjusted for potential confounders. Results: In crude analysis, serum MMP-7 levels were significantly higher in colon cancer group compared to LS-carriers and controls [median (IQR) 4.1 ng/ml (2.7–6.0), 2.3 ng/ml (1.7–3.1), 2.5 ng/ml (1.5–3.7), respectively; p value - p < 0.001) while there was no difference between the two last groups (p value = 0.583). However, after adjusting for age and gender, LS-carriers' patients had 18% higher concentrations of serum MMP-7 compared to healthy controls (p value = 0.037), while colon cancer patients had 50% higher serum MMP-7 level in comparison to healthy controls (p value < 0.001). Additionally, age was positively associated with higher serum MMP-7 levels across all study groups (r = 0.67, p value < 0.001). In contrast, no correlation was observed between serum MMP-7 and either tumor staging and gene mutation. Conclusions: Age-adjusted serum MMP-7 levels in asymptomatic LS carriers are higher than its levels in healthy population. While in colon cancer, MMP-7 higher level probably reflects the tumor burden and may have a prognostic effect, its significance and clinical applicability as a biomarker for tumorigenesis in LS is less clear and should be elucidated. [ABSTRACT FROM AUTHOR]

Background and Aims: Inflammation due to the excess of nutrient intake plays an important role in the pathophysiology of metabolic syndrome (MetS). Here, the potential influence of neutrophils and their degranulation markers on MetS improvement upon dietary and behavioral counselling, has been investigated. Specifically, we aimed at investigating their role as potential predictors of metabolic syndrome improvements.Methods and Results: patients with MetS (n = 127) received behavioral and dietary recommendations before follow-up at 6 months. Serum levels of matrix metalloproteinases (MMP)8, MMP9, myeloperoxidase (MPO), tissue inhibitor of MMP (TIMP)-1, TIMP-2, TIMP-3 and resistin were tested at baseline. In the whole cohort, baseline levels of proinflammatory MMP8, MMP9 and MPO increased together with the number of MetS criteria. Seventy-three (57%) patients experienced a reduction in MetS-defining criteria at follow-up. With respect to those with no improvement, such individuals showed lower weight and waist circumference at enrolment, less frequent smoking habits, higher levels of triglycerides and lower circulating MMP8. At logistic regression analysis, baseline MMP8 showed negative predictive ability (odds ratio (OR) 0.979 [0.961-0.997]; p = 0.025) against MetS improvement. Such findings hold true even when included in the backward stepwise logistic regression model confirming MMP8 as an independent predictor (OR 0.970 [0.949-0.993]; p = 0.009). Receiver operating characteristic (ROC) curve confirmed the predictive ability of MMP8 combined in a model including baseline MetS criteria and waist circumference. Bootstrap resampling analysis internally validated our findings.Conclusion: Improvement of MetS is independently associated with baseline low MMP-8 levels, suggesting a pivotal role for inflammation in metabolic alteration. [ABSTRACT FROM AUTHOR]

Platismatia glauca is a popular lichen traditionally used as a spice and possesses significant anti‐cancer potential, whose anti‐migratory/anti‐invasive properties were mostly disregarded. Migration/invasion of cancer cells is processed in cancer metastasis and targeting their markers is an important strategy in anti‐cancer treatment. We examined the anti‐migratory/anti‐invasive properties of P. glauca extract on two colorectal carcinoma cell lines (HCT‐116 and SW‐480) and elucidated possible mechanisms underlying these properties. Cell migration was evaluated by wound healing and RTCA methods. Immunofluorescent assay was used for the analysis of protein, while qRT‐PCR for gene expression of migratory/invasive markers. ELISA assay was applied for the determination of MMP‐9 concentration. P. glauca extract inhibited the motility of tested cells, by reducing pro‐migratory/pro‐invasive markers and potentially retaining intercellular connections. Treatment showed cell‐selective effects, and HCT‐116 cells were more responsive. Our study presents important scientific novelty, thus these lichen properties should be furtherly examined regarding the amelioration of anti‐cancer treatment. Practical applications: Based on the evidence we provided in the present study, we have demonstrated that lichen species Platismatia glauca possess important biological activity, which has not been sufficiently investigated so far. It is of great importance to explore its anti‐cancer potential, not only from a cytotoxic point of view but especially anti‐migratory and anti‐invasive. Herein, we showed that this species expresses significant suppressive effects on migration and invasiveness of colorectal carcinoma cells. This tested lichen has the potential to be used as a natural complementary anti‐cancer treatment, with special reference on the dose applied and type of carcinoma. Our study represents a significant novelty in the field of scientific investigation of lichens and natural products, and further detailed studies are needed on in vitro and in vivo model systems. [ABSTRACT FROM AUTHOR]

Though COVID-19 is primarily characterized by symptoms in the periphery, it can also affect the central nervous system (CNS). This has been established by the association between stroke and COVID-19. However, the molecular mechanisms that cause stroke related to a COVID-19 infection have not been fully explored. More specifically, stroke and COVID-19 exhibit an overlap of molecular mechanisms. These similarities provide a way to better understand COVID-19 related stroke. We propose here that peripheral macrophages upregulate inflammatory proteins such as matrix metalloproteinases (MMPs) in response to SARS-CoV-2 infection. These inflammatory molecules and the SARS-CoV-2 virus have multiple negative effects related to endothelial dysfunction that results in the disruption of the blood–brain barrier (BBB). Finally, we discuss how the endothelial blood–brain barrier injury alters central nervous system function by leading to astrocyte dysfunction and inflammasome activation. Our goal is to elucidate such inflammatory pathways, which could provide insight into therapies to combat the negative neurological effects of COVID-19. [ABSTRACT FROM AUTHOR]

Aims Marfan syndrome is one of the most common inherited disorders of connective tissue caused by fibrillin-1 mutations, characterized by enhanced transcription factor AP-1 DNA binding activity and subsequently abnormally increased expression and activity of matrix-metalloproteinases (MMPs). We aimed to establish a novel adeno-associated virus (AAV)-based strategy for long-term expression of an AP-1 neutralizing RNA hairpin (hp) decoy oligonucleotide (dON) in the aorta to prevent aortic elastolysis in a murine model of Marfan syndrome. Methods and results Using fibrillin-1 hypomorphic mice (mgR/mgR), aortic grafts from young (9 weeks old) donor mgR/mgR mice were transduced ex vivo with AAV vectors and implanted as infrarenal aortic interposition grafts in mgR/mgR mice. Grafts were explanted after 30 days. For in vitro studies, isolated primary aortic smooth muscle cells (SMCs) from mgR/mgR mice were used. Elastica-van-Giesson staining visualized elastolysis, reactive oxygen species (ROS) production was assessed using dihydroethidine staining. RNA F.I.S.H. verified AP-1 hp dON generation in the ex vivo transduced aortic tissue. MMP expression and activity were assessed by western blotting and immunoprecipitation combined with zymography. Transduction resulted in stable therapeutic dON expression in endothelial and SMCs. MMP expression and activity, ROS formation as well as expression of monocyte chemoattractant protein-1 were significantly reduced. Monocyte graft infiltration declined and the integrity of the elastin architecture was maintained. RNAseq analysis confirmed the beneficial effect of AP-1 neutralization on the pro-inflammatory environment in SMCs. Conclusion This novel approach protects from deterioration of aortic stability by sustained delivery of nucleic acids-based therapeutics and further elucidated how to interfere with the mechanism of elastolysis. [ABSTRACT FROM AUTHOR]

Though COVID-19 is primarily characterized by symptoms in the periphery, it can also affect the central nervous system (CNS). This has been established by the association between stroke and COVID-19. However, the molecular mechanisms that cause stroke related to a COVID-19 infection have not been fully explored. More specifically, stroke and COVID-19 exhibit an overlap of molecular mechanisms. These similarities provide a way to better understand COVID-19 related stroke. We propose here that peripheral macrophages upregulate inflammatory proteins such as matrix metalloproteinases (MMPs) in response to SARS-CoV-2 infection. These inflammatory molecules and the SARS-CoV-2 virus have multiple negative effects related to endothelial dysfunction that results in the disruption of the blood–brain barrier (BBB). Finally, we discuss how the endothelial blood–brain barrier injury alters central nervous system function by leading to astrocyte dysfunction and inflammasome activation. Our goal is to elucidate such inflammatory pathways, which could provide insight into therapies to combat the negative neurological effects of COVID-19.

Cancer progression with uncontrolled tumor growth, local invasion, and metastasis depends largely on the proteolytic activity of numerous matrix metalloproteinases (MMPs), which affect tissue integrity, immune cell recruitment, and tissue turnover by degrading extracellular matrix (ECM) components and by releasing matrikines, cell surface-bound cytokines, growth factors, or their receptors. Among the MMPs, MMP-14 is the driving force behind extracellular matrix and tissue destruction during cancer invasion and metastasis. MMP-14 also influences both intercellular as well as cell–matrix communication by regulating the activity of many plasma membrane-anchored and extracellular proteins. Cancer cells and other cells of the tumor stroma, embedded in a common extracellular matrix, interact with their matrix by means of various adhesive structures, of which particularly invadopodia are capable to remodel the matrix through spatially and temporally finely tuned proteolysis. As a deeper understanding of the underlying functional mechanisms is beneficial for the development of new prognostic and predictive markers and for targeted therapies, this review examined the current knowledge of the interplay of the various MMPs in the cancer context on the protein, subcellular, and cellular level with a focus on MMP14.

Introduction: Matrix metalloproteinases (MMP's) and tribbles 3 human homolog (Trb3) are implicated in atherosclerosis. Changes in the concentration of these biomolecules signal the risk of atherosclerosis in type 2 subjects (T2DM), with or without hypertension (HT), at an early stage. Aim: Our aim was to assess the relation between noninvasive arterial stiffness indices and circulating levels of MMP2, MMP9 and Trb3. Methodology: The study included 144 participants divided into 4 groups: T2DM > 5 years + HT, DM + HT (n = 55), T2DM < 2 years, DM (n = 28), HT (n = 31), and healthy controls (HC) (n = 30). Anthropometric measurements and blood biochemistry profiles were established using standard protocols. MMP2, MMP9 and Trb3 were estimated using ELISA. Pulse wave velocities (PWV) and arterial stiffness indices (ASI) were measured using PeriScopeTM. Results were analysed using SPSS 21. Results: MMP2, average Brachial Stiffness Index (ba ASI) (Pearson's r = 0.235, p = 0.005) and Ankle-Brachial Index (ABI) (Pearson's r = 0.225, p = 0.007) were positively correlated. Average Ankle ASI (Ank ASI) was positively correlated to Trb3 (Pearson's r = 0.184, p = 0.028), but negatively to MMP9 (Pearson's r = − 0.184, p = 0.027). In multiple linear regression, MMP2 influenced ba ASI [adjusted R2 = 0.038; F (3) = 2.862, p = 0.039] and ABI [adjusted R2 = 0.033; F (3) = 2.642, p = 0.052]. MMP9 influenced Ank ASI [adjusted R2 = 0.058; F (3) = 3.912, p = 0.01]. Conclusion: Arterial stiffness indices and matrix metalloproteinases conform early risk of atherosclerosis in diabetic subjects. [ABSTRACT FROM AUTHOR]

Stroke is one of the leading causes of death and disability worldwide. However, treatment options for ischemic stroke remain limited. Matrix-metalloproteinases (MMPs) contribute to brain damage during ischemic strokes by disrupting the blood-brain barrier (BBB) and causing brain edemas. Carnosine, an endogenous dipeptide, was found by us and others to be protective against ischemic brain injury. In this study, we investigated whether carnosine influences MMP activity. Brain MMP levels and activity were measured by gelatin zymography after permanent occlusion of the middle cerebral artery (pMCAO) in rats and in vitro enzyme assays. Carnosine significantly reduced infarct volume and edema. Gelatin zymography and in vitro enzyme assays showed that carnosine inhibited brain MMPs. We showed that carnosine inhibited both MMP-2 and MMP-9 activity by chelating zinc. Carnosine also reduced the ischemia-mediated degradation of the tight junction proteins that comprise the BBB. In summary, our findings show that carnosine inhibits MMP activity by chelating zinc, an essential MMP co-factor, resulting in the reduction of edema and brain injury. We believe that our findings shed new light on the neuroprotective mechanism of carnosine against ischemic brain damage.

Planarians have been long known for their regenerative ability, which hinges on pluripotency. Recently, however, the planarian model has been successfully established for routine toxicological screens aimed to assess overproliferation, mutagenicity and tumorigenesis. In this study, we focused on planarian tumor suppressor genes (TSGs) and their role during chemically induced carcinogenic stress in Schmidtea mediterranea. Combining in silico and proteomic screens with exposure to human carcinogen type 1A agent cadmium (Cd), we showed that many TSGs have a function in stem cells and that, in general, exposure to Cd accelerated the onset and increased the severity of the observed phenotype. This suggested that the interaction between environmental and genetic factors plays an important role in tumor development in S. mediterranea. Therefore, we further focused on the synergistic effects of Cd exposure and p53 knockdown (KD) at the cellular and molecular levels. Cd also produced a specific proteomic landscape in homeostatic animals, with 172 proteins differentially expressed, 43 of which were downregulated. Several of these proteins have tumor suppressor function in human and other animals, namely Wilms Tumor 1 Associated Protein (WT1), Heat Shock Protein 90 (HSP90), Glioma Pathogenesis-Related Protein 1 (GLIPR1) and Matrix Metalloproteinase B (Smed-MMPB). Both Glipr1 and MmpB KD produced large outgrowths, epidermal lesions and epidermal blisters. The epidermal blisters that formed as a consequence of Smed-MmpB KD were populated by smedwi1+ cells, many of which were actively proliferating, while large outgrowths contained ectopically differentiated structures, such as photoreceptors, nervous tissue and a small pharynx. In conclusion, Smed-MmpB is a planarian TSG that prevents stem cell proliferation and differentiation outside the proper milieu.

Activation of vascular smooth muscle cells (VSMCs) represents a key event of atherosclerosis. It is triggered e.g. by the action of platelet-derived growth factor (PDGF). The polyphenol (+)-episesamin (ES) is known to block the TNF-α-induced mitogenic VSMC response. Aiming to develop novel therapeutic options e.g. by dietary supplements, we studied if ES and its stereoisomer sesamin (SE) reduce PDGF-BB-induced growth and migration, which both promote atherosclerosis. ES and SE reduced basal and PDGF-BB-induced proliferation and migration of human, murine and rat VSMC by impaired activation of MAPK and PI3K pathways and reduced oxidative stress by induction of haem oxygenase-1 expression. Moreover, ES and SE blocked PDGF-BB-induced activation of NF-κB with subsequently reduced expression and secretion of the gelatinases matrix-metalloproteinase-(MMP)-2 and MMP-9. In conjunction with their natural origin and well-tolerable properties, these data suggest ES and SE as potential candidates for a complemental treatment of VSMC specific vascular diseases like atherosclerosis.

Objective. Class 3 semaphorins regulate diverse cellular processes relevant to the pathology of RA, including immune modulation, angiogenesis, apoptosis and invasive cell migration. Therefore, we analysed the potential role of class 3 semaphorins in the pathology of RA. Methods. Protein and mRNA expression in RA synovial tissue, SF and fibroblast-like synoviocytes (FLS) were determined by immunoblotting and quantitative PCR (qPCR). RA FLS migration and invasion were determined using wound closure and transwell invasion assays, respectively. PlexinA1, neuropilin-1 and neuropilin-2 expression was knocked down using small interfering RNA (siRNA). Activation of FLS intracellular signalling pathways was assessed by immunoblotting. Results. mRNA expression of semaphorins (Sema)3B, Sema3C, Sema3F and Sema3G was significantly lower in the synovial tissue of early arthritis patients at baseline who developed persistent disease compared with patients with self-limiting disease after 2 years follow-up. Sema3B and Sema3F expression was significantly lower in arthritis patients fulfilling classification criteria for RA compared with those who did not. FLS expression of Sema3A was induced after stimulation with TNF, IL-1β or lipopolysaccharides (LPS), while Sema3B and Sema3F expression was downregulated. Exogenously applied Sema3A induced the migration and invasive capacity of FLS, while stimulation with Sema3B or Sema3F reduced spontaneous FLS migration, and platelet-derived growth factor induced cell invasion, effects associated with differential regulation of MMP expression and mediated by the PlexinA1 and neuropilin-1 and -2 receptors. Conclusion. Our data suggest that modulation of class 3 semaphorin signaling could be a novel therapeutic strategy for modulating the invasive behaviour of FLS in RA. [ABSTRACT FROM AUTHOR]

An imbalance of regeneration and destruction of the extracellular matrix due to a plethora of chemo- and cytokines, elevated matrix metalloproteinases, bacterial contamination and repetitive painless tissue damage can lead the chronicity of a wound, especially in diabetic foot ulcers (DFU). Along general lines, wound healing and cancer development are similar. Therefore chronic wounds prepare a breeding ground for cancer development. Several characteristics such as increase in size, verrucous everted margins and contact bleeding are suspicious for malignant growth in a chronic wound. While previously the term Marjolin’s ulcer was attributed to a malignant tumor in (burn) scars, it is nowadays used for every malignant tumor in chronic wounds. Furthermore, chronic ulcers in diabetic feet are susceptible for malignant transformation. We describe two cases of squamous cell carcinoma in patients with DFU—a 71 year-old woman and a 67 year old man. Both received total tumor excision and split-skin grafts with good short-time results.

Background. Chronic obstructive lung disease (COPD) is characterised by partially reversible usually progressive airflow limitation caused by inflammation and remodelling. Stromelysin-1 (MMP-3) has regulatory activity on other matrix-metalloproteinases. Altered MMP-3 activity has been described in different diseases. We investigated the role of a promoter MMP-3 polymorphism in determining susceptibility and severity of COPD. Methods. We studied 147 patients with COPD in stable conditions and distinguished two groups based on FEV1 values. In 100 patients functional modifications across a two-year period were noted. 133 healthy subjects were used as controls. Genotyping for the –1171 5A/6A MMP-3 polymorphism was performed using nucleotide sequencing. Results. No difference was noted in the genotype distribution between COPD patients and controls. However, among patients with severe disease 6A/6A genotype and 6A allelic frequency were significantly more represented than among mild-moderate patients (p < 0.05). The 6A/6A genotype was also associated with a higher FEV1 decline over time. Conclusions. Our data suggests that –1171 6A allele does not represent a risk factor for the development of COPD while it is associated with more severe disease and different functional decline. We hypothesise that a disregulation of MMP-3, possibly caused by the –1171 5A/6A polymorphism or other linked variants, may lead to different progression in COPD.

Objective To evaluate cytotoxicity and effect on protease activity of epigallocatechin-gallate extracted from experimental restorative dental copolymers in comparison to the control compound chlorhexidine. Methods Copolymer disks were prepared from bis-GMA/TEGDMA (70/30 mol%) containing no compound (control) or 1% w/w of either epigallocatechin-gallate or chlorhexidine. MDPC-23 odontoblast-like cells were seeded with the copolymer extracts leached out into deionized water. Cell metabolic activity was quantified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay at 24, 48, 72 h. Inhibition of protease activity by resin extracts was measured by a collagenolytic/genatinolytic enzyme activity assay and gelatin zymography. Data for MTT and protease inhibition were analyzed using two-way ANOVA followed by Tukey or Bonferroni post hoc tests ( α = 0.05). Results The MTT revealed that at 72 h, extracts from control (16.7%) and chlorhexidine (22.3%) copolymers induced significant reduction in cell metabolism ( p < 0.05). All copolymer extracts caused enzymatic inhibition in a dose dependent manner ( p < 0.01). Even when highly diluted, epigallocatechin-gallate extract had a significant antiproteolytic activity ( p < 0.05). Zymograms showed that all extracts reduced activity of MMP-2 and MMP-9 (pro- and active forms), with MMP-9 exhibiting the highest percentage inhibition revealed by densitometry. Conclusions Epigallocatechin-gallate and chlorhexidine extracts did not exert cytotoxicity on evaluated cells when compared to control extracts. Both compounds retained antiproteolytic activity after extraction from a dental copolymer. Clinical significance Once extracted from a dental copolymer, epigallocatechin-gallate is not cytotoxic and retains antiproteolytic activity. These results may allow incorporation of epigallocatechin-gallate as a natural-safe alternative to chlorhexidine in functionalized restorative materials. [ABSTRACT FROM AUTHOR]

Adverse outcome in bacterial meningitis is associated with the breakdown of the blood–brain barrier (BBB). Matrix-metalloproteinases (MMPs) facilitate this process by degradation of components of the BBB. This in turn results in acute complications of bacterial meningitis including edema formation, increased intracranial pressure and subsequent ischemia. We determined the parenchymal balance of MMP-9 and TIMP-1 (tissue inhibitor of MMP) and the structural integrity of the BBB in relation to cortical damage in an infant rat model of pneumococcal meningitis. The data demonstrate that the extent of cortical damage is significantly associated with parenchymal gelatinolytic activity and collagen type IV degradation. The increased gelatinolysis was found to be associated with a brain parenchymal imbalance of MMP-9/TIMP-1. These findings provide support to the concept that MMPs mediated disruption of the BBB contributes to the pathogenesis of bacterial meningitis and that protection of the vascular unit may have neuroprotective potential.

Activation of vascular smooth muscle cells (VSMCs) represents a key event of atherosclerosis. It is triggered e.g. by the action of platelet-derived growth factor (PDGF). The polyphenol (+)-episesamin (ES) is known to block the TNF-α-induced mitogenic VSMC response. Aiming to develop novel therapeutic options e.g. by dietary supplements, we studied if ES and its stereoisomer sesamin (SE) reduce PDGF-BB-induced growth and migration, which both promote atherosclerosis. ES and SE reduced basal and PDGF-BB-induced proliferation and migration of human, murine and rat VSMC by impaired activation of MAPK and PI3K pathways and reduced oxidative stress by induction of haem oxygenase-1 expression. Moreover, ES and SE blocked PDGF-BB-induced activation of NF-κB with subsequently reduced expression and secretion of the gelatinases matrix-metalloproteinase-(MMP)-2 and MMP-9. In conjunction with their natural origin and well-tolerable properties, these data suggest ES and SE as potential candidates for a complemental treatment of VSMC specific vascular diseases like atherosclerosis. [ABSTRACT FROM AUTHOR]

Pentoxifylline (PTX), a methylxanthine derivative is a non-steroidal immunomodulating agent with unique hemorheologic properties. It is used in the treatment of intermittent claudication as it increases the amount of oxygen reaching tissues by increasing the flexibility of red blood cells. Recently, it has also shown to exhibit anti-metastatic and anti-angiogenic activities in B16F10 melanoma cells both in vitro as well as in vivo. As per the reports, the choice of drug in the treatment of breast cancer is paclitaxel, but the major limitation is its toxicity. However, the effects of PTX on metastatic processes in breast cancer are not currently known. Therefore, in this study, we have examined the effect of PTX in MDA-MB-231 human breast cancer cells. The MTT assay showed dose- and time-dependent decreases in cellular proliferation. The non-toxic concentration of PTX selected were 1, 2.5 and 5 mM for 24 h. PTX induced a G0-G1 cell-cycle arrest leading to apoptosis. Further, it affected adhesion to both the matrigel and collagen type-IV in a time- and dose-dependent manner. The PTX impeded the migration of MDA-MB-231 cells and also decreased the activities of both MMP-2 and MMP-9. Thus, PTX at non-toxic doses affected cellular proliferation, adhesion, migration and invasion. These results demonstrate its anti-metastatic effect on MDA-MB-231 cells, and further studies need to be carried out to understand the mechanism of action. [ABSTRACT FROM AUTHOR]

Objective(s) The aim of the present study was to compare the therapeutic indices of several agents used in treatment of inflammatory conditions which included: Vitamin E, hydro-alcoholic extracts of Glycyrrhiza glabra, Matricaria aurea, dexamethasone, piroxicam and diclofenac using Wehi-164 fibrosarcoma cells. Materials and Methods Cytotoxicity evaluation was based on vital dye exclusion assay. Matrix-metalloproteinases inhibition (MMPI) was assessed by gelatinase zymography method. The collected data were used to estimate the IC50 (50% MMPI concentration), LC50 (50% cytotoxicity concentration) and the therapeutic index (LC50/IC50). Results Among the natural anti-inflammatory agents used, M. aurea was the least toxic and the most effective inhibitor of MMP. Vitamin E not only increased MMP activity, but also was the most toxic of all the agents tested. Next in terms of toxicity to vitamin E was G. glabra. Diclofenac was more toxic than both piroxicam and dexamethasone. Conclusion Findings from this study suggest that medicinal plants reputed to have anti-inflammatory properties are not equally effective and safe. In order to assess the implications of these findings, further in vitro and in vivo studies are needed. [ABSTRACT FROM AUTHOR]

Cancer progression with uncontrolled tumor growth, local invasion, and metastasis depends largely on the proteolytic activity of numerous matrix metalloproteinases (MMPs), which affect tissue integrity, immune cell recruitment, and tissue turnover by degrading extracellular matrix (ECM) components and by releasing matrikines, cell surface-bound cytokines, growth factors, or their receptors. Among the MMPs, MMP-14 is the driving force behind extracellular matrix and tissue destruction during cancer invasion and metastasis. MMP-14 also influences both intercellular as well as cell–matrix communication by regulating the activity of many plasma membrane-anchored and extracellular proteins. Cancer cells and other cells of the tumor stroma, embedded in a common extracellular matrix, interact with their matrix by means of various adhesive structures, of which particularly invadopodia are capable to remodel the matrix through spatially and temporally finely tuned proteolysis. As a deeper understanding of the underlying functional mechanisms is beneficial for the development of new prognostic and predictive markers and for targeted therapies, this review examined the current knowledge of the interplay of the various MMPs in the cancer context on the protein, subcellular, and cellular level with a focus on MMP14. [ABSTRACT FROM AUTHOR]

Summary: Objective: Cartilage degradation in osteoarthritis (OA) generates the type II collagen fragments, Helix-II and CTX-II that can be used as clinical biological markers. Helix-II and C-telopeptide of type II collagen (CTX-II) levels are associated independently with progression of OA suggesting that they may be generated through different collagenolytic pathways. In this study we analyzed the release of Helix-II and CTX-II from human cartilage collagen by the proteinases reported to play a role in cartilage degradation. Methods: In vitro, human articular cartilage extract was incubated with activated human recombinant cathepsins (Cats) and matrix-metalloproteases (MMPs). Next, we analyzed the spontaneous release of Helix-II and CTX-II from cartilage sections of patients with knee OA who were immediately deep frozen after joint replacement to preserve endogenous enzyme activity until assay. Cartilage sections were then incubated for up to 84h in the presence or absence of E-64 and GM6001, inhibitors of cysteine proteases and MMPs, respectively. Results: In vitro, Cats K, L and S generated large amount of Helix-II, but not CTX-II. Cat B generated CTX-II fragment, but destroyed Helix-II immunoreactivity. Cat D was unable to digest intact cartilage. MMPs-1, -3, -7, -9, and -13 efficiently released CTX-II, but only small amount of Helix-II. Neither CTX-II nor Helix-II alone was able to reflect accurately the collagenolytic activity of Cats and MMPs as reflected by the release of hydroxyproline. In OA cartilage explants, E-64 blunted the release of Helix-II whereas the release of CTX-II could be completely abrogated by GM6001 and only partly by E-64. Conclusion: These in vitro and ex vivo experiments of human cartilage suggest that Helix-II and CTX-II could be released in part by different enzymatic pathways. Helix-II and CTX-II alone reflect only partially overall cartilage collagen degradation. These findings may explain why these two biological markers could provide complementary information on disease progression in OA. [Copyright &y& Elsevier]

The aim of this study was to determine the prognostic role of matrix metalloproteinases in rectal cancer. Formalin-fixed and paraffin-embedded tissue sections of 94 rectal carcinomas were used for the immunohistochemical analysis of matrix metalloproteinases (MMP)-2, MMP-7, MT1-MMP, and tissue inhibitor of metalloproteinases (TIMP)-2. Inclusion criteria were sporadic rectal adenocarcinoma resected curatively (including total mesorectal excision), adjuvant radiochemotherapy in UICC stages II and III, and complete intra-institutional follow-up. Results of immunohistochemistry were correlated with clinical and histopathologic data from the prospective rectal cancer registry and prognosis. End points of the prognostic analysis were tumor progression caused by local and/or distant recurrence and 5-year survival (disease-free and overall). To assess prognostic significance, statistics included univariate and multivariate analysis ( p<0.05 statistically significant). Of the 94 rectal carcinomas, 35% (33/94) showed an epithelial MMP-2 expression, 77% (72/94) were MMP-2 positive in the stroma. Fifty-four percent (51/94) were MMP-7 positive, and 47% (46/94) were positive for both MT1-MMP and TIMP-2. The stromal MMP-2 staining pattern was correlated with the depth of invasion (pT status, p=0.006) with MMP-7 ( p=0.016) and TIMP-2 expression ( p=0.036). Positive expression of MMP-2 in tumor epithelium was correlated with MMP-7 ( p=0.027), MT1-MMP ( p=0.036), and TIMP-2 expression ( p<0.0001). A positive staining pattern of MMP-7 was significantly correlated with depth of invasion and TIMP-2 ( p<0.01). The positive staining pattern of MT1-MMP was correlated with epithelial MMP-2 ( p=0.036), MMP-7 ( p=0.004), and TIMP-2 expression ( p=0.002). TIMP-2 immunoreactivity correlated with depth of invasion ( p=0.013), epithelial MMP-2 ( p<0.001), stromal MMP-2 ( p=0.036), MMP-7 ( p<0.001), and MT1-MMP ( p=0.002). Neither pattern correlated with age, gender, tumor stage (UICC), grading, preoperative serum carcinoembryonic antigen (CEA) level, or nodal status ( p>0.05). Within a mean follow-up of 46 months, tumor progression, caused by either local recurrence or distant metastasis, occurred in 14 patients (15.4%). There was no significant association between the MMP expression and the incidence of local and/or distant recurrence. In terms of survival, preoperative CEA level (disease-free 5-year survival 46% with increased CEA vs 70% with normal CEA, p=0.01; overall 5-year survival 43 vs 74%, p<0.01) and UICC stage were the only factors to be significantly related to 5-year survival by univariate analysis, whereas the metalloproteinases failed to show a significant association. In multivariate analysis, CEA and UICC stage were not identified as independent factors predictive of survival. MMP-2, MMP-7, MT1-MMP, and TIMP-2 do not appear to be significant predictors of prognosis in a homogenous collective of curatively resected rectal adenocarcinomas. [ABSTRACT FROM AUTHOR]

Abstract: Adverse outcome in bacterial meningitis is associated with the breakdown of the blood–brain barrier (BBB). Matrix-metalloproteinases (MMPs) facilitate this process by degradation of components of the BBB. This in turn results in acute complications of bacterial meningitis including edema formation, increased intracranial pressure and subsequent ischemia. We determined the parenchymal balance of MMP-9 and TIMP-1 (tissue inhibitor of MMP) and the structural integrity of the BBB in relation to cortical damage in an infant rat model of pneumococcal meningitis. The data demonstrate that the extent of cortical damage is significantly associated with parenchymal gelatinolytic activity and collagen type IV degradation. The increased gelatinolysis was found to be associated with a brain parenchymal imbalance of MMP-9/TIMP-1. These findings provide support to the concept that MMPs mediated disruption of the BBB contributes to the pathogenesis of bacterial meningitis and that protection of the vascular unit may have neuroprotective potential. [Copyright &y& Elsevier]

Abstract: Aim of this study was the investigation of systemic biochemical regulation mechanisms of bone regeneration by angiogenic and matrix-degrading enzymes during distraction osteogenesis compared to rigid osteotomy bone healing. Serum samples of 10 otherwise healthy patients with callus distraction for lower limb-lengthening and 10 osteotomy patients undergoing elective axis correction have been collected prospectively in a standardized time schedule before and up to 6 months after the procedure. At the end of the individual investigation period, concentrations of metalloproteinases (MMP-9, -13), tissue inhibitors of metalloproteinases (TIMP-1, TIMP-2) and the angiogenic factors angiogenin and VEGF have been detected by use of commercially available enzyme immunoassays. Results have been compared to our preliminary study on proMMP-1–3. In distraction osteogenesis, significantly elevated serum concentrations compared to baseline could be detected postoperatively for proMMP-1, MMP-9, TIMP-1, angiogenin and VEGF but not for proMMP-2, proMMP-3 or TIMP-2. In patients with rigid osteotomy healing, MMP-9, TIMP-1, TIMP-2, angiogenin and VEGF were significantly increased respectively. Comparison of both patient collectives revealed significantly higher increases of serum proMMP-1, VEGF and TIMP-1 in distraction patients during the lengthening period and significantly higher serum concentrations of TIMP-2 in late fracture healing period in osteotomy patients. Serum levels of MMP-13 were below the lowest standards, and therefore quantitative analysis was not possible. Bone regeneration in distraction osteogenesis and rigid osteotomy healing is accompanied by systemic increase of matrix-degrading and angiogenic factors in a certain time course and quantity. This might reflect biochemical regulation of local bone healing in the circulation. ProMMP-1, VEGF and TIMP-1 seem to be key regulatory factors during distraction osteogenesis. [Copyright &y& Elsevier]

Gürkan A,Çınarcık S, Hüseyinov A. Adjunctive subantimicrobial dose doxycycline: effect on clinical parameters and gingival crevicular fluid transforming growth factor-β 1 levels in severe, generalized chronic periodontitis. J Clin Periodontol 2005; 32: 244–253. doi:10.1111/j.1600-051X.2005.00663.x.© Blackwell Munksgaard, 2005.At present there is limited data concerning the efficacy of non-surgical periodontal therapy supplemented with subantimicrobial dose doxycycline (SDD) in the treatment of severe, generalized periodontitis. The purpose of the present study was to evaluate the effect of adjunctive SDD therapy on clinical periodontal parameters and gingival crevicular fluid (GCF) transforming growth factor-beta1 (TGF-β1) levels in patients with severe, generalized chronic periodontitis over a 6-month period.Thirty-five patients with severe, generalized periodontitis and 11 periodontally healthy subjects were included in the present study. Patients received full-mouth supragingival debridment at baseline and randomized to take either SDD b.i.d. or placebo b.i.d. for 3 months. Patients received root planing and oral hygiene instruction once a week for four consecutive weeks. Clinical measurements including probing depth (PD), clinical attachment level, papilla bleeding index and plaque index and GCF sampling were performed at baseline, 3 and 6 months. The GCF TGF-β1 levels were analysed by enzyme-linked immunosorbent assay.Thirteen patients in both study groups completed the 6-month trial. Following scaling and root planing (SRP) plus SDD and SRP plus placebo therapy significant improvements in clinical periodontal parameters of both groups were observed (p<0.025). In the SDD group a significantly higher percentage (%73.4) of deep pockets resolved (PD reduction⩾3 mm from baseline) when compared with placebo group (%49.7) at 6 months (p<0.05). At baseline there were no significant differences in GCF TGF-β1 levels between three groups. Both total amount and concentration of GCF TGF-β1 in SDD and placebo groups increased when compared with baseline at 3 months. However, only GCF TGF-β1 levels of SDD group was significantly higher than baseline (p<0.025) and placebo group (p<0.017) at 3 months. At 6 months GCF TGF-β1 levels of both groups were similar to baseline levels (p<0.025).These data indicate that combination of SDD with non-surgical therapy improves clinical parameters of periodontal disease and increases GCF TGF-β1 levels together with a decrease in prevalence of residual pockets in patients with severe, generalized chronic periodontitis. Increased GCF TGF-β1 levels following SDD therapy might suggest a novell pleiotrophic mechanism for tetracyclines to inhibit connective tissue breakdown. [ABSTRACT FROM AUTHOR]

Cathepsin D is a lysosomal acid proteinase which is involved in the malignant progression of breast cancer and other gynecological tumors. Clinical investigations have shown that in breast cancer patients cathepsin D overexpression was significantly correlated with a shorter free-time disease and overall survival, whereas in patients with ovarian or endometrial cancer this phenomenon was associated with tumor aggressiveness and a degree of chemoresistance to various antitumor drugs such as anthracyclines, cis-platinum and vinca alkaloids. Therefore, a lot of research has been undertaken to evaluate the role and the prognostic value of cathepsin D also in other solid neoplasms. However, conflicting results have been generated from these studies. The discrepancies in these results may, in part, be explained with the different methodological approaches used in order to determine the levels of expression of the enzyme in tumor tissues and body fluids. Further investigations using well-standardized techniques may better define the clinical significance of cathepsin D expression in solid tumors. Nevertheless, evidence emerging from these studies indicates that this proteinase seems to facilitate early phases of tumor progression such as cell proliferation and local dissemination. These findings support the concept that cathepsin D may be a useful marker for identifying patients with highly malignant tumor phenotypes who may need more aggressive clinical treatment; this enzyme may also be considered as a potential target for a novel therapeutic approach in the treatment of solid neoplasms. [ABSTRACT FROM AUTHOR]

Purpose: Despite improved surgical techniques, anastomotic leakage is still a serious complication in colorectal surgery, resulting in increased morbidity and mortality. Based on recent studies showing a disturbance of the collagen synthesis in hernia patients, this pilot study was designed to find out whether there are any hints of disorders concerning the collagen metabolism in patients developing anastomotic leakage after colonic surgery. Methods: Ten samples of colonic tissue from patients who developed anastomotic dehiscence after surgery for colonic cancer were compared with 14 specimens from patients with uncomplicated anastomotic healing. All samples were obtained at the primary operation. We performed a sirius red test for the overall collagen content and immunhistochemical studies facing differentiation between collagen I and III and the expression of MMP-1 and MMP-13. Results: In the bowel sections from patients developing anastomotic leakage, decreased levels of collagen I and III, and a smaller amount of overall collagen were found. The ratio of collagen I:III was similar in both groups. While MMP-1 was expressed in all samples, there was a difference concerning the expression of MMP-13. In only 6 of 14 patients with uncomplicated healing was MMP-13 present, while in 9 of 10 patients developing a leakage. Conclusions: Our data may indicate the presence of an individual disturbance of the collagen metabolism in patients developing a leakage after colorectal surgery. Further studies are underway to define more precisely whether a preexisting impairment of wound healing could be a risk factor for anastomotic leakage. [ABSTRACT FROM AUTHOR]

A lot of parallels have been described between invasion of malignant tumor cells and leukocyte movement during inflammatory responses. Concerning these similarities, we investigated the function of cytokine-suppressive anti-inflammatory drugs (CSAIDs), which act via inhibition of stress-activated MAP-kinases, in regulation of expression of proteolytic enzymes and in vitro invasion of malignant melanoma cells. The p38MAPK inhibitor SB203580 reduced matrigel invasion of MeWo cells by 60%, while the MEK-1 inhibitor PD98059 did not have any effect on invasion. Active p38MAPK was detected in MeWo cells by immunoblotting and confocal microscopy. Cells showed a constitutive expression of matrix-metalloproteinase (MMP)-2 as well as tissue inhibitor of metalloproteinases (TIMP)-1 and TIMP-2 mRNAs. Expression of MMP-1 or urokinase-type plasminogen activator (uPA) was not detected by Northern blot. Inhibition of p38MAPK by the specific inhibitor SB203580 resulted in downregulation of MMP-2 mRNA and protein levels as well as gelatinolytic activity, while expression levels of TIMP-1 and TIMP-2 mRNAs were not changed. The specific MEK-1 inhibitor PD98059 did not change expression of MMP-2 or TIMPs. Neither SB203580 nor PD98059 changed proliferation of cells. The results suggest that stress-activated protein kinases like p38MAPK are involved in regulation of expression of MMP-2 as well as in vitro invasion of malignant melanoma cells. Inhibitors of p38MAPK may be promising substances to interfere with a signaling cascade associated with invasion of malignant tumor cells. [ABSTRACT FROM AUTHOR]

Colorectal cancer is one of the most common malignant tumors and entails a relatively poor prognosis. Clinical outcome depends on the extent of local and metastatic tumor spread. Results of in vivo and in vitro studies suggest that the balance between matrix metalloproteinases (MMPs) and their inhibitors (tissue inhibitors of metalloproteinases TIMPs) is altered in neoplasia, contributing to the invasive and metastatic properties of malignant tumors. We quantified tissue concentrations of MMP-2 and TIMP-2 in 65 malignant colorectal lesions and corresponding normal mucosa by enzyme-linked immunosorbent assay, western blotting, and in situ hybridization. In situ hybridization and western blot analyses demonstrated a clear increase in both stromal expression of MMP-2 transcripts and protein in primary carcinomas. The protein concentration of MMP-2 was higher in all tumor stages, except stage I tumors, than in normal mucosa and adenomas. MMP-2 concentrations were not related to tumor differentiation or to colonic versus rectal location. Surprisingly, the MMP-2 concentration was not increased in metastases. Interestingly, tissue concentrations and epithelial mRNA expression of TIMP-2 decreased significantly in primary colorectal cancer (UICC stages III and IV) but increased in metastases. Therefore an increased ratio of MMP-2 to TIMP-2 is strongly associated with advanced tumor stages, but a decreased ratio was observed in metastases. These findings suggest that the MMP-2:TIMP-2 ratio may prove useful as a marker of local invasion but not of metastasis in colorectal cancer. [ABSTRACT FROM AUTHOR]

Cancer progression with uncontrolled tumor growth, local invasion, and metastasis depends largely on the proteolytic activity of numerous matrix metalloproteinases (MMPs), which affect tissue integrity, immune cell recruitment, and tissue turnover by degrading extracellular matrix (ECM) components and by releasing matrikines, cell surface-bound cytokines, growth factors, or their receptors. Among the MMPs, MMP-14 is the driving force behind extracellular matrix and tissue destruction during cancer invasion and metastasis. MMP-14 also influences both intercellular as well as cell-matrix communication by regulating the activity of many plasma membrane-anchored and extracellular proteins. Cancer cells and other cells of the tumor stroma, embedded in a common extracellular matrix, interact with their matrix by means of various adhesive structures, of which particularly invadopodia are capable to remodel the matrix through spatially and temporally finely tuned proteolysis. As a deeper understanding of the underlying functional mechanisms is beneficial for the development of new prognostic and predictive markers and for targeted therapies, this review examined the current knowledge of the interplay of the various MMPs in the cancer context on the protein, subcellular, and cellular level with a focus on MMP14.

Stroke is one of the leading causes of death and disability worldwide. However, treatment options for ischemic stroke remain limited. Matrix-metalloproteinases (MMPs) contribute to brain damage during ischemic strokes by disrupting the blood-brain barrier (BBB) and causing brain edemas. Carnosine, an endogenous dipeptide, was found by us and others to be protective against ischemic brain injury. In this study, we investigated whether carnosine influences MMP activity. Brain MMP levels and activity were measured by gelatin zymography after permanent occlusion of the middle cerebral artery (pMCAO) in rats and in vitro enzyme assays. Carnosine significantly reduced infarct volume and edema. Gelatin zymography and in vitro enzyme assays showed that carnosine inhibited brain MMPs. We showed that carnosine inhibited both MMP-2 and MMP-9 activity by chelating zinc. Carnosine also reduced the ischemia-mediated degradation of the tight junction proteins that comprise the BBB. In summary, our findings show that carnosine inhibits MMP activity by chelating zinc, an essential MMP co-factor, resulting in the reduction of edema and brain injury. We believe that our findings shed new light on the neuroprotective mechanism of carnosine against ischemic brain damage. [ABSTRACT FROM AUTHOR]

UVA irradiation leads to photoaging including clinical features such as wrinkle formation, reduced recoil capacity and blister formation of the skin. Besides synthesis of the extracellular matrix, its regulated degradation by various matrix-metalloproteinases (MMPs) determines the amount and the composition of the extracellular matrix within the dermis and the basement membrane of the dermo-epidermal junction. In this study we therefore ascertained whether UV irradiation could modulate the synthesis of MMPs with substrate specifities for dermal (collagen I, Ill, V) and basement membrane compounds (collagen IV, VII, proteoglycans, laminin) and whether synthesis of the counteracting tissue inhibitor of metalloproteinases (TIMP-1) was also affected. Following UVA irradiation specific mRNAs of MMPs 1, 2 and 3 were induced concomitantly up to 5-fold compared to mock irradiated controls. In contrast, TIMP-1 mRNA levels remained unaltered. Immunoprecipitation indicated that after UVA irradiation synthesis and secretion of MMPs 1, 2 and 3 into the supernatant increased. Taken together, our data show that UVA irradiation coordinately induced MMPs 1, 2 and 3 implying similar mechanisms in their regulatory pathways, while TIMP-1 synthesis was not altered. Hence, unbalanced synthesis of MMPs potentially contributes to the dissolution of dermal and basement membrane compounds finally leading to blister formation and cutaneous photoaging. [ABSTRACT FROM AUTHOR]

ResumoIntroduçãoAgentes promotores de ligações cruzadas têm sido investigados como inibidores da atividade enzimática da dentina, o que favoreceria a longevidade das restaurações adesivas.ObjetivoAvaliar o efeito do tratamento da dentina com proantocianidina (PA), em curtos períodos de tempo, na inibição da atividade de MMPs in situ.Material e métodoQuarenta espécimes de dentina (1×1×6 mm) foram obtidos de molares hígidos e divididos em quatro grupos (n=10). Os espécimes foram condicionados com ácido fosfórico por 15 s, seguido de lavagem em água deionizada. A dentina condicionada foi tratada com: água, 5% PA por 5 s, 15 s ou 30 s. A atividade de MMP foi analisada colorimetricamente (SensoLyte®) e os dados de absorbância (412 nm) foram submetidos aos testes de ANOVA e Tukey (α=0,05).ResultadoTodos os períodos de tratamento foram capazes de reduzir a atividade de MMPs, sendo que os melhores resultados foram observados para a dentina tratada com PA por 15 s (63,1% redução) e 30 s (70,2%). O tratamento por 5 s foi capaz de inibir 39,9% das MMPs.ConclusãoA aplicação de PA sobre a dentina condicionada foi capaz de reduzir a atividade de MMPs mesmo em períodos de tempo extremamente curtos, como 5 s. No entanto, melhores resultados foram obtidos com os maiores períodos de tratamento.