Expression profiles of lncRNAs, miRNAs, and mRNAs and interaction analysis indicate their potential involvement during testicular fusion in Spodoptera litura.

Testicular fusion of Spodoptera litura occures during metamorphosis, which benefits sperms development. Previous research identified involvement of ECM-integrin interaction pathways, MMPs in testicular fusion, but the regulatory mechanism remains unclear. RNA-seq was performed to analyze long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) in testes, aiming to uncover potential regulatory mechanisms of testicular fusion. 2150 lncRNAs, 2742 targeted mRNAs, and 347 miRNAs were identified in testes at three different developmental stages. Up-regulated DElncRNAs and DEmRNAs, as well as down-regulated DEmiRNAs, were observed during testicular fusion, while the opposite expression pattern was observed after fusion. Enrichment analysis of DEmRNAs revealed that cAMP signal pathway, ECM remodeling enzymes, ECM-integrin interaction pathways, and cell adhesion molecules were potentially associated with testicular fusion. The identified DElncRNA-DEmiRNA-DEmRNA regulatory network related to cAMP signal pathway, ECM remodeling enzymes suggests their roles during testicular fusion. Our research will provide new targets for studying the mechanism of testicular fusion.

Recombinant Human Clusterin Seals Damage to the Ocular Surface Barrier in a Mouse Model of Ophthalmic Preservative-Induced Epitheliopathy.

There is a significant unmet need for therapeutics to treat ocular surface barrier damage, also called epitheliopathy, due to dry eye and related diseases. We recently reported that the natural tear glycoprotein CLU (clusterin), a molecular chaperone and matrix metalloproteinase inhibitor, seals and heals epitheliopathy in mice subjected to desiccating stress in a model of aqueous-deficient/evaporative dry eye. Here we investigated CLU sealing using a second model with features of ophthalmic preservative-induced dry eye. The ocular surface was stressed by topical application of the ophthalmic preservative benzalkonium chloride (BAC). Then eyes were treated with CLU and sealing was evaluated immediately by quantification of clinical dye uptake. A commercial recombinant form of human CLU (rhCLU), as well as an rhCLU form produced in our laboratory, designed to be compatible with U.S. Food and Drug Administration guidelines on current Good Manufacturing Practices (cGMP), were as effective as natural plasma-derived human CLU (pCLU) in sealing the damaged ocular surface barrier. In contrast, two other proteins found in tears: TIMP1 and LCN1 (tear lipocalin), exhibited no sealing activity. The efficacy and selectivity of rhCLU for sealing of the damaged ocular surface epithelial barrier suggests that it could be of therapeutic value in treating BAC-induced epitheliopathy and related diseases.

A novel model of a biomechanically induced osteoarthritis-like cartilage for pharmacological in vitro studies.

Excessive pressure or overload induces and aggravates osteoarthritic changes in articular cartilage, but the underlying biomechanical forces are largely ignored in existing pharmacological in vitro models that are used to investigate drugs against osteoarthritis (OA). Here, we introduce a novel in vitro model to perform pathophysiological and pharmacological investigations, in which cartilage explants are subjected to intermittent cyclic pressure, and characterize its ability to mimic OA-like tissue reactivity. Mechanical loading time-dependently increased the biosynthesis, content and retention of fibronectin (Fn), whereas collagen metabolism remained unchanged. This protocol upregulated the production and release of proteoglycans (PGs). The release of PGs from explants was significantly inhibited by a matrix metalloproteinase (MMP) inhibitor, suggesting the involvement of such proteinases in the destruction of the model tissue, similar to what is observed in human OA cartilage. In conclusion, the metabolic alterations in our new biomechanical in vitro model are similar to those of early human OA cartilage, and our pharmacological prevalidation with an MMP-inhibitor supports its value for further in vitro drug studies.

In vitro stem cell modelling demonstrates a proof-of-concept for excess functional mutant TIMP3 as the cause of Sorsby fundus dystrophy.

Sorsby fundus dystrophy (SFD) is a rare autosomal dominant disease of the macula that leads to bilateral loss of central vision and is caused by mutations in the TIMP3 gene. However, the mechanisms by which TIMP3 mutations cause SFD are poorly understood. Here, we generated human induced pluripotent stem cell-derived retinal pigmented epithelial (hiPSC-RPE) cells from three SFD patients carrying TIMP3 p.(Ser204Cys) and three non-affected controls to study disease-related structural and functional differences in the RPE. SFD-hiPSC-RPE exhibited characteristic RPE structure and physiology but showed significantly reduced transepithelial electrical resistance associated with enriched expression of cytoskeletal remodelling proteins. SFD-hiPSC-RPE exhibited basolateral accumulation of TIMP3 monomers, despite no change in TIMP3 gene expression. TIMP3 dimers were observed in both SFD and control hiPSC-RPE, suggesting that mutant TIMP3 dimerisation does not drive SFD pathology. Furthermore, mutant TIMP3 retained matrix metalloproteinase activity. Proteomic profiling showed increased expression of ECM proteins, endothelial cell interactions and angiogenesis-related pathways in SFD-hiPSC-RPE. By contrast, there were no changes in VEGF secretion. However, SFD-hiPSC-RPE secreted higher levels of monocyte chemoattractant protein 1, PDGF and angiogenin. Our findings provide a proof-of-concept that SFD patient-derived hiPSC-RPE mimic mature RPE cells and support the hypothesis that excess accumulation of mutant TIMP3, rather than an absence or deficiency of functional TIMP3, drives ECM and angiogenesis-related changes in SFD. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Amphiphilic peptide-based MMP3 inhibitors for intra-articular treatment of knee OA.

Since 2007, Metalloproteases (MMPs) have been considered potential targets for treating osteoarthritis (OA), for which the primary pathogenic event is the extensive degeneration of articular cartilage. MMP3 is an enzyme critical for these degenerative changes. However, problems of selectivity, low bioavailability and poor metabolic profile during clinical trials of MMPs inhibitors (MMPIs) led to limited beneficial effect and thus did not justify further pursuit of the clinical studies. In a previous work, a new alkyl derivative of hyaluronic acid (HA), HYADD4®, previously approved as intra-articular treatment for knee OA, was studied in vitro and in vivo as MMP3I. Molecular simulation studies confirmed the interaction between the alkyl side chain of this HA derivative and the additional S1' pocket of MMP3. However, the high MW and the polar HA backbone of HYADD4® imply a high desolvation energy cost, which can potentially decrease its inhibitory potency. In this study, a new class of MMP3Is based on a small peptide backbone (CGV) chemically derivatized with an alkyl chain was developed through interactive cycles of design, synthesis and screening, accompanied by computational evaluation and optimization. Two MMP3Is, e(I) and l(II), were selected because of their effective inhibitory activity (3.2 and 10.2 µM, respectively) and water solubility. Both MMPIs showed a broad range of inhibitory effects against almost all the MMPs tested. In an in vitro model of inflammatory OA, e(I) was the most effective MMPI: at the concentration of 93 µM, it reversed inflammatory outcomes. Moreover, because of its amphiphilic structure, the e(I) MMPI promoted stable micellar formulation at concentrations higher than 0.2 mg/mL in water. The findings were confirmed by TEM and Nile red staining analysis. Based on these results, the e(I) MMPI can be considered a good candidate for the intra-articular treatment of OA, and the micellar formulation of this peptide in an aqueous buffer can potentially increase the bioavailability and, thus, the efficacy of the MMPIs.

Methylation degree of metalloproteinase inhibitor RECK gene: Links to RECK protein level and hepatocellular carcinoma in chronic HCV infection patients.

The RECK gene, a tumor suppressor gene, inhibits angiogenesis, invasion, and tumor metastasis. Epigenetic regulation of the RECK gene constitutes a potent approach to the molecular basis of liver malignancy. This study aims to evaluate the promoter methylation status of the RECK gene and its serum level in patients with hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC) and the potential association of RECK gene methylation with clinical criteria of HCC. One hundred and fifty-five subjects were included (healthy control [55], chronic HCV patients [55], HCV-related HCC patients [45]). The methylation status of the RECK gene promoter and serum RECK level were investigated by methylation-specific PCR and enzyme-linked immunosorbent assay techniques, respectively. RECK gene promoter hypermethylation was recorded in 46.7% of HCC patients, and 10.9% of HCV patients, but not in control subjects (0%). It was related to RECK protein level, varices, edema, ascites, lymph node metastasis, vascular invasion, and the largest diameter of focal lesions. Meanwhile, it was not associated with focal lesion number nor distant metastasis of HCC. In conclusion, RECK gene promoter hypermethylation is linked to HCV genotype-4-related HCC. Moreover, different degrees of RECK gene promoter methylation are associated with serum RECK level, lymph node metastasis, and vascular invasion, which could prove its pathogenic role in hepatocarcinogenesis in chronic HCV-infected patients.

A type IV collagenase inhibitor, N-hydroxy-3-phenyl-2-(4-phenylbenzenesulfonamido) propanamide (BiPS), suppresses skin injury induced by sulfur mustard.

Sulfur mustard (SM) is a highly toxic blistering agent thought to mediate its action, in part, by activating matrix metalloproteinases (MMPs) in the skin and disrupting components of the basement membrane zone (BMZ). Type IV collagenases (MMP-9) degrade type IV collagen in the skin, a major component of the BMZ at the dermal-epidermal junction. In the present studies, a type IV collagenase inhibitor, N-hydroxy-3-phenyl-2-(4-phenylbenzenesulfonamido) propanamide (BiPS), was tested for its ability to protect the skin against injury induced by SM in the mouse ear vesicant model. SM induced inflammation, epidermal hyperplasia and microblistering at the dermal/epidermal junction of mouse ears 24-168 h post-exposure. This was associated with upregulation of MMP-9 mRNA and protein in the skin. Dual immunofluorescence labeling showed increases in MMP-9 in the epidermis and in the adjacent dermal matrix of the SM injured skin, as well as breakdown of type IV collagen in the basement membrane. Pretreatment of the skin with BiPS reduced signs of SM-induced cutaneous toxicity; expression of MMP-9 mRNA and protein was also downregulated in the skin by BiPS. Following BiPS pretreatment, type IV collagen appeared intact and was similar to control skin. These results demonstrate that inhibiting type IV collagenases in the skin improves basement membrane integrity after exposure to SM. BiPS may hold promise as a potential protective agent to mitigate SM induced skin injury.

Matrix metalloproteinase: An upcoming therapeutic approach for idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a debilitating condition where excess collagen deposition occurs in the extracellular matrix. At first sight, it is expected that the level of different kinds of matrix metalloproteinases might be downregulated in IPF as it is a matrix degrading collagenase. However, the role of some matrix metalloproteinases (MMPs) is profibrotic where others have anti-fibrotic functions. These profibrotic MMPs effectively promote fibrosis development by stimulating the process of epithelial to mesenchymal transition. These profibrotic groups also induce macrophage polarization and fibrocyte migration. All of these events ultimately disrupt the balance between profibrotic and antifibrotic mediators, resulting aberrant repair process. Therefore, inhibition of these matrix metalloproteinases functions in IPF is a potential therapeutic approach. In addition to the use of synthetic inhibitor, various natural compounds, gene silencing act as potential natural MMP inhibitor to recover IPF.

Matrix metalloproteinase inhibitor and sunscreen effective compounds from Rumex crispus L.: isolation, identification, bioactivity and molecular docking study.

INTRODUCTION: Rumex (Polygonaceae) species are widely grown worldwide and their main compounds are anthranoids, tannins, naphthalenes and flavonoids. Matrix metalloproteinase (MMP) enzymes that cause the breakdown of the extracellular matrix and harmful ultraviolet (UV) rays are two important causes of skin ageing. Although there have been many studies on anthraquinones, there are not many studies on their anti-ageing effects. OBJECTIVE: The aim of this study was to determine the anti-ageing effects of pure compounds isolated from Rumex crispus L. by measuring their MMP inhibitory and sunscreen activities. METHODOLOGY: Molecular docking studies on anthranoid, flavonoid, tannin and naphthalene skeletons with MMP enzymes were applied for the first time. Various chromatographic methods were used for isolation studies. The structure elucidation was performed by spectroscopic [UV, infrared (IR), electrospray ionisation mass spectrometry (ESI-MS), nuclear magnetic resonance (NMR)] methods. To determine the anti-ageing capacity of compounds, MMP-1, MMP-8, MMP-13 enzyme inhibitory effects and sun protection factors (SPFs) were investigated for the first time. RESULTS: Molecular docking results showed that skeletons had low binding energy to the target and were likely to inhibit MMP enzymes. Then, eight anthraquinones and a naphthalene glycoside were isolated from the underground parts of Rumex crispus L. According to the results, MMP inhibitory effects of compounds were found to be high. In vitro and molecular docking study results confirmed each other. All anthraquinones showed high sun protection and emodin showed highest UV capability with SPF of 30.59. CONCLUSION: This study shows that anthraquinones may be evaluated as new potential MMP inhibitors, sun protective and anti-ageing agents.

Cardioprotective Effect of Novel Matrix Metalloproteinase Inhibitors.

BACKGROUND: We recently developed novel matrix metalloproteinase-2 (MMP-2) inhibitor small molecules for cardioprotection against ischemia/reperfusion injury and validated their efficacy in ischemia/reperfusion injury in cardiac myocytes. The aim of the present study was to test our lead compounds for cardioprotection in vivo in a rat model of acute myocardial infarction (AMI) in the presence or absence of hypercholesterolemia, one of the major comorbidities affecting cardioprotection. METHODS: Normocholesterolemic adult male Wistar rats were subjected to 30 min of coronary occlusion followed by 120 min of reperfusion to induce AMI. MMP inhibitors (MMPI)-1154 and -1260 at 0.3, 1, and 3 µmol/kg, MMPI-1248 at 1, 3, and 10 µmol/kg were administered at the 25th min of ischemia intravenously. In separate groups, hypercholesterolemia was induced by a 12-week diet (2% cholesterol, 0.25% cholic acid), then the rats were subjected to the same AMI protocol and single doses of the MMPIs that showed the most efficacy in normocholesterolemic animals were tested in the hypercholesterolemic animals. Infarct size/area at risk was assessed at the end of reperfusion in all groups by standard Evans blue and 2,3,5-triphenyltetrazolium chloride (TTC) staining, and myocardial microvascular obstruction (MVO) was determined by thioflavine-S staining. RESULTS: MMPI-1154 at 1 µmol/kg, MMPI-1260 at 3 µmol/kg and ischemic preconditioning (IPC) as the positive control reduced infarct size significantly; however, this effect was not seen in hypercholesterolemic animals. MVO in hypercholesterolemic animals decreased by IPC only. CONCLUSIONS: This is the first demonstration that MMPI-1154 and MMPI-1260 showed a dose-dependent infarct size reduction in an in vivo rat AMI model; however, single doses that showed the most efficacy in normocholesterolemic animals were abolished by hypercholesterolemia. The further development of these promising cardioprotective MMPIs should be continued with different dose ranges in the study of hypercholesterolemia and other comorbidities.

[Application value of urinary IGFBP7 and TIMP-2 in acute kidney injury with decompensated hepatitis B virus-related liver cirrhosis].

Objective: To investigate the application value of new urinary biomarkers insulin-like growth factor binding protein 7 (IGFBP7) and tissue matrix metalloproteinase inhibitor-2 (TIMP-2) in acute kidney injury with decompensated hepatitis B virus-related liver cirrhosis. Methods: 45 newly hospitalized cases with decompensated hepatitis B virus-related liver cirrhosis were selected. Among them, 19 cases were combined with AKI on admission (cirrhosis-AKI group), 26 cases without AKI (cirrhosis-non-AKI group), and 12 healthy cases (normal control group). First-morning urine samples were collected and IGFBP7 and TIMP-2 were detected by enzyme-linked immunosorbent assay (ELISA). Urinary IGFBP7 and serum creatinine (SCr) were dynamically monitored after hospitalization in cirrhosis-non-AKI group. Normally distributed measurement data were compared by t-test, and non-normally distributed measurement data were compared by rank sum test. The receiver operating characteristic (ROC) curve and area under the curve (AUC) were used to evaluate the diagnostic accuracy of the indicators. Results: Urinary IGFBP7, IGFBP7 with TIMP-2 (IGFBP7×TIMP-2) in cirrhosis-AKI group (n = 19) were equally higher than that of the cirrhosis-non-AKI group (P < 0.05). Urinary IGFBP7, TIMP-2 and IGFBP7×TIMP-2 in cirrhosis-AKI group or cirrhosis-non-AKI group were significantly higher than those of the normal control group (P < 0.01). The AUC of urinary IGFBP7 and urinary IGFBP7×TIMP-2 for diagnosis of AKI were 0.703 (95% CI 0.547-0.860) and 0.700 (95% CI 0.541-0.859), respectively. In the liver cirrhosis-non-AKI group (n = 26), 5 cases of AKI were newly diagnosed according to the changes in SCr during hospitalization (progressive group). Urinary IGFBP7 was significantly increased 2 days before the diagnosis of AKI. The concentration of urinary IGFBP7 at admission in the progressive group (n = 5) was higher than that of the non-progressive group (n = 21) (P < 0.05). Conclusion: Urinary IGFBP7 and TIMP-2 concentrations were significantly increased in patients with decompensated hepatitis B virus-related liver cirrhosis. When AKI occurred, urinary IGFBP7 and IGFBP7×TIMP-2 was further increased. Urinary IGFBP7 is valuable for early AKI diagnosis, and may play a role in predicting AKI occurrence.

[Surgical treatment of internal carotid artery kinking following fibromuscular dysplasia]. / Rezul'taty khirurgicheskogo lecheniya patologicheskoi izvitosti vnutrennei sonnoi arterii na fone fibrozno-myshechnoi displazii.

OBJECTIVE: To evaluate the results of surgical treatment of internal carotid artery kinking following fibromuscular dysplasia. MATERIAL AND METHODS: There were 32 patients who underwent surgical treatment of internal carotid artery kinking following fibromuscular dysplasia. Structural changes of carotid artery wall were analyzed using immunohistochemical survey. Considering destructive changes revealed, we divided all patients into 2 groups in order to assess long-term postoperative outcomes: 1 - ICA resection followed by anastomosis in end-to-end fashion; 2 - ICA replacement. Postoperative analysis included incidence of stroke, thrombosis and deformities of anastomosis zone, regression of cerebrovascular insufficiency. RESULTS: The main «phenotype¼ of arterial wall in patients with ICA kinking following fibromuscular dysplasia is a large number of smooth muscle cells releasing matrix matelloproteinases-2 and -9 and low level of their tissue inhibitor type 1. Postoperative deformities are more common within a year after surgery. Maximum incidence is observed after 12 months. Both ICA resection and replacement are followed by similar incidence of deformity later. No severe deformities were diagnosed. Resection of ICA kinking on the background of fibromuscular dysplasia is followed by comparable results with ICA replacement regarding the incidence stroke, thrombosis and regression of cerebrovascular insufficiency. CONCLUSION: Despite degradation of extracellular matrix, destruction of elastic fibers and their fragmentation, no significant deformities are observed in long-term postoperative period in patients with ICA kinking and fibromuscular dysplasia.

Chlorhexidine, a Matrix Metalloproteinase Inhibitor and the Development of Secondary Caries Wall Lesions in a Microcosm Biofilm Model.

This study investigated the role of a matrix metalloproteinase (MMP) inhibitor (CHX 2%) in the development of secondary caries wall lesions in different interface conditions with small (run 1) and wider gaps (run 2). Dentin discs were restored and pretreated with or without CHX 2%. In run 1, interfaces were made with gaps of 30, 60, or 90 µm. Interfaces with composite placed directly onto the dentin were either bonded (Adper Single Bond 2) or not bonded. In run 2, interfaces were made with gaps of 100 µm, with or without adhesive on the composite side (CLEARFIL SE Bond). Interfaces were either bonded or not bonded, as in run 1. Microcosm biofilms were grown on dentin-composite samples for 14 days. Caries lesion outcomes were analyzed by transversal wavelength-independent microradiography at 3 locations: the outer surface, and the interface wall at a distance of 200 and 500 µm from the gap entrance. Linear regression analyses showed that pretreatment with MMP inhibitor did not influence progression of the wall lesion at any location (p ≥ 0.218). Interfaces with intentional gaps showed positive and significant effect on the wall lesion progression at 200 µm from the gap entrance (p ≤ 0.005). A small trend of increase in wall lesion development was observed at the 200-µm location when bonding was present on the composite side. In conclusion, the dentin pretreatment with CHX 2% was not able to slow down the development of secondary caries wall lesions in small and wide gaps in this biofilm model.

Inhibitory effects of ascorbic acid toward snake venom metalloproteinase (SVMP) from Indian Echis carinatus venom: Insights from molecular modeling and binding studies.

Classical antivenom therapy is unable to shield complications of viper bite and has limitations such as anaphylaxis and serum sickness. Snake venom metalloproteinases are responsible for local tissue damage and hemorrhage at the bitten site in viper envenomation, and this has led to a persistent search for metalloproteinase inhibitors. Here, we report the inhibitory effects of ascorbic acid against metalloproteinase from Echis carinatus venom both in-silico and in-vitro. Ascorbic acid effectively inhibited the proteolytic activity of E. carinatus venom in a dose-dependent manner. Interaction studies of ascorbic acid with purified ecarin using isothermal titration calorimetry showed favorable binding energy and energetics. The molecular docking of ascorbic acid with ecarin revealed important interactions with residues at the active site pocket of ecarin. It was observed that the ligand behaves as a chelating inhibitor. Thus, the backbone structural scaffold of ascorbic acid can find potential use as building blocks in designing drug-like molecules for viper bite management.

Platelet glycoproteins associated with aspirin-treatment upon platelet activation.

Platelet glycoproteins are known to play central roles in hemostasis and vascular integrity and have pathologic roles in vascular occlusive diseases such as myocardial infarction and stroke. Characterizing glycoproteins within and secreted by platelets can provide insight into the mechanisms that underlie vascular pathologies and the therapeutic benefits or failure of anti-platelet agents. To study the impact of aspirin, which is commonly prescribed for primary and secondary cardiovascular prevention, on the platelet glycoproteome, we evaluated washed platelets from ten donors. The platelet glycoproteome, was studied using an iTRAQ in resting and stimulated states and with and without aspirin treatment. Using solid phase extraction of glycosite-containing peptides (SPEG), we were able to identify 799 unique N-linked glycosylation sites (glycosites) in platelets, representing the largest and the most comprehensive analysis to date. We were able to identity a number of glycoproteins impacted by aspirin treatment, which we validated using global proteomics analysis of platelets and their secreted proteins. In our analyses, metallopeptidase inhibitor 1 (TIMP1) was the single most significantly affected glycoprotein by aspirin treatment. ELISA assays confirmed proteomic results and validated our strategy. Functional analysis demonstrated that TIMP1 levels were highly correlated with platelet reactivity in vitro, with a correlation coefficient of -0.5. The release of TIMP1 from platelets, which was previously unknown to be affected by aspirin treatment, may play important roles in hemostasis and/or vascular integrity. If validated, our findings may be useful for developing assays that assess platelet response to aspirin or other anti-platelet therapies.

Safety of Using Matrix Metalloproteinase Inhibitor in Experimental Glaucoma Filtration Surgery.

We evaluated the safety of matrix metalloproteinase (MMP) inhibitor in experimental glaucoma filtration surgery in an animal model. Fifteen New Zealand white rabbits underwent an experimental trabeculectomy and were randomly allocated into 3 groups according to the adjuvant agent: no treatment group (n = 5), 0.02% mitomycin C (MMC) soaking group (n = 5), and MMP inhibitor (ilomastat) subconjunctival injection group (n = 5). Slit lamp examination with Seidel testing, pachymetry, and specular microscopy was performed preoperatively and postoperatively. The conjunctiva and ciliary body toxicity were evaluated with scores according to the pathologic grading systems. Electron microscopy was used to examine the structural changes in cornea, conjunctiva, and ciliary body. In the ilomastat-treated group, there was no statistically significant change in central corneal thickness preoperatively and at 28 days postoperatively (P = 0.655). There were also no significant changes in specular microscopy findings over the duration of the study in the ilomastat-treated group. The conjunctival toxicity score was 1 in the control group, 1.5 in the ilomastat-treated group, and 2 in the MMC-treated group. When assessing ciliary body toxicity scores, the ilomastat-treated group score was 0.5 and the MMC-treated group score was 1.5. Transmission electron microscopy did not show structural changes in the cornea and ciliary body whereas the structural changes were noticed in MMC group. A single subconjunctival injection of MMP inhibitor during the experimental trabeculectomy showed a less toxic affect in the rabbit cornea, conjunctiva, and ciliary body compared to MMC.

Development of a Non-Hydroxamate Dual Matrix Metalloproteinase (MMP)-7/-13 Inhibitor.

Matrix metalloproteinase 7 (MMP-7) is a member of the MMP superfamily and is able to degrade extracellular matrix proteins such as casein, gelatin, fibronectin and proteoglycan. MMP-7 is a validated target for the development of small molecule drugs against cancer. MMP-13 is within the enzyme class the most efficient contributor to type II collagen degeneration and is a validated target in arthritis and cancer. We have developed the dual MMP-7/-13 inhibitor ZHAWOC6941 with IC50-values of 2.2 µM (MMP-7) and 1.2 µM (MMP-13) that is selective over a broad range of MMP isoforms. It spares MMP-1, -2, -3, -8, -9, -12 and -14, making it a valuable modulator for targeted polypharmacology approaches.

[Metalloproteases and their inhibitors: role in pathogenesis of selected examples]. / Metaloproteinazy i ich inhibitory: rola w patogenezie na wybranych przykladach.

Proteolytic enzymes and their inhibitors are crucial in host-pathogen interaction. Metalloproteases secreted by pathogenic microbes play an important role in destroying not only host tissues but also their immune proteins. Metalloproteinase inhibitors, in contrast, may serve as effective therapeutic agents, which is especially important because of the increasing number of microorganisms resistant to known antibiotics. The role of metalloproteases produced by the bacterium Pseudomonas aeruginosa in the colonization of the host organism is described. Attention has also been paid to the role of inhibitors of these enzymes in defense responses and underlined their potential role in inhibiting the development of infection.

Matrix metalloproteinases (MMPs), the main extracellular matrix (ECM) enzymes in collagen degradation, as a target for anticancer drugs.

The main group of enzymes responsible for the collagen and other protein degradation in extracellular matrix (ECM) are matrix metalloproteinases (MMPs). Collagen is the main structural component of connective tissue and its degradation is a very important process in the development, morphogenesis, tissue remodeling, and repair. Typical structure of MMPs consists of several distinct domains. MMP family can be divided into six groups: collagenases, gelatinases, stromelysins, matrilysins, membrane-type MMPs, and other non-classified MMPs. MMPs and their inhibitors have multiple biological functions in all stages of cancer development: from initiation to outgrowth of clinically relevant metastases and likewise in apoptosis and angiogenesis. MMPs and their inhibitors are extensively examined as potential anticancer drugs. MMP inhibitors can be divided into two main groups: synthetic and natural inhibitors. Selected synthetic inhibitors are in clinical trials on humans, e.g. synthetic peptides, non-peptidic molecules, chemically modified tetracyclines, and bisphosphonates. Natural MMP inhibitors are mainly isoflavonoids and shark cartilage.

Experimental Dissection of Metalloproteinase Inhibition-Mediated and Toxic Effects of Phenanthroline on Zebrafish Development.

Metalloproteinases are zinc-dependent endopeptidases that function as primary effectors of tissue remodeling, cell-signaling, and many other roles. Their regulation is ferociously complex, and is exquisitely sensitive to their molecular milieu, making in vivo studies challenging. Phenanthroline (PhN) is an inexpensive, broad-spectrum inhibitor of metalloproteinases that functions by chelating the catalytic zinc ion, however its use in vivo has been limited due to suspected off-target effects. PhN is very similar in structure to phenanthrene (PhE), a well-studied poly aromatic hydrocarbon (PAH) known to cause toxicity in aquatic animals by activating the aryl hydrocarbon receptor (AhR). We show that zebrafish are more sensitive to PhN than PhE, and that PhN causes a superset of the effects caused by PhE. Morpholino knock-down of the AhR rescues the effects of PhN that are shared with PhE, suggesting these are due to PAH toxicity. The effects of PhN that are not shared with PhE (specifically disruption of neural crest development and angiogenesis) involve processes known to depend on metalloproteinase activity. Furthermore these PhN-specific effects are not rescued by AhR knock-down, suggesting that these are bona fide effects of metalloproteinase inhibition, and that PhN can be used as a broad spectrum metalloproteinase inhibitor for studies with zebrafish in vivo.