Matrix Metalloproteinase Inhibitor Research Articles

Metalloproteinase inhibitors in the adhesion process dental

Matrix metalloproteinases (MMPs) are enzymes involved in tooth physiological development, caries processes, and hybrid layer degradation, in addition to being associated with dentin collagen breakdown. The interplay of MMP activity, masticatory forces, and biofilm action may, over time, compromise composite resin restorations. To reduce hybrid layer degradation by endogenous proteases and extend the longevity of resin restorations, MMP inhibitors and collagen cross-linking agents have been extensively studied. This work aims to identify, through a literature review, agents capable of inactivating MMPs at the dentin adhesive interface. This integrative review, based on studies retrieved from PUBMED/MEDLINE, LILACS, BBO, and VHL databases, identified 19 agents able to inhibit MMP activity at the adhesive-dentin interface in studies published between January 2018 and June 2023. It can be concluded that several agents can partially or completely block MMP activity, thereby enhancing restoration longevity. However, further studies are required to facilitate the clinical use of these agents, beyond chlorhexidine, in dental practice, with the application techniques suited to dentists’ daily clinical routines.

Muscle Tissue Response to Vipera berus berus and Vipera berus nikolskii Venoms: A Study on Matrix Metalloproteinases and Cytokine Modulation

Vipera berus, also known as the common adder, is a snake species widely distributed in Europe and parts of Asia. Its venom is a mix of enzymatic and non-enzymatic components causing both local and systemic effects including edema, hemorrhage, myonecrosis, gastrointestinal disorders, and rarely – rhabdomyolysis, coagulopathy and hypovolemic shock. Matrix metalloproteinases (MMPs) and cytokines, being inflammatory mediators, participate both in the development of these reactions and in tissue regeneration after acute damage. The current study was aimed at analyzing the concentrations of important mediators of tissue damage, inflammation and regeneration, namely, MMP-1, -2, -3, -8, -10, TIMP-1 (a tissue inhibitor of MMPs), and pro- and anti-inflammatory cytokines, in rat muscle after intraperitoneal injection of venoms from two subspecies of adders – V. berus berus and V. berus nikolskii — to clarify their role in the local response of skeletal muscle to the studied venoms. Changes in muscle tissue were revealed, namely, significant increase in almost all MMPs including MMP-2, as well as TIMP-1, IFN-γ and anti-inflammatory IL-4 and IL-10 content. Moreover, a tendency for some pro-inflammatory cytokines levels to decline 24 h after venom administration was observed, which may be a sign of inflammation suppression and the beginning of reparative processes associated with extracellular matrix remodeling and formation of an optimal environment for muscle regeneration. However, to establish the exact mechanisms of the changes we found, further studies are needed. The ability of V. berus nikolskii venom to induce more pronounced changes was also shown. Our results may be useful for the development of differentiated and more effective treatment strategies for the consequences of bites by these snakes, which include myonecrosis

MMP Inhibition, Marginal Integrity and Cytotoxicity of Zinc-Releasing GIC

ObjectivesThe degradation of the restorative-dentin interface due to endogenous dentin enzymes, such as matrix metalloproteinases (MMPs), is a significant issue that accelerates the deterioration of the dentin matrix and leads to the failure of restorative treatments. Caredyne Restore (CR), a novel glass ionomer cement (GIC) with zinc ions in its formulation, represents the latest effort to mitigate this issue. This investigation aimed to evaluate the MMPs inhibitory effect, marginal integrity, and cytotoxicity of CR compared to a conventional GIC, Fuji IX (FIX). MethodsThe inhibitory effect of CR on MMPs activity was evaluated using in-situ zymography to visualize the endogenous gelatinolytic activity in the GIC-dentin interface. Additionally, CR's sealing properties were investigated using dye-assisted confocal laser microscopy (CLSM) to assess marginal leakage across the GIC-dentin interface. Both in-situ zymography and CLSM observations were conducted 1 day and seven days after pH cycling. Finally, the cytotoxicity of the GIC eluates was examined on rat dental pulp cells (RPC-C2A). Assay measurements were performed after 24 and 48 h of incubation with test solutions prepared using various GIC eluate concentrations. ResultsThe MMPs activity of the CR specimens was significantly lower than that of FIX specimens after seven days of pH cycling. Signs of marginal leakage were also lower in the CR specimens. Comparison of CR and FIX eluates at the same concentration showed no significant difference in terms of biocompatibility. ConclusionsCaredyne restore is capable of inhibiting endogenous enzyme activity and improving sealing properties, while maintaining sufficient biocompatibility. Clinical SignificanceZinc ions released from Caredyne Restore offer a safe way to improve the quality of dental restorations and promote minimally invasive treatment, especially in lesions where the dentin matrix is susceptible to enzymatic degradation and recurrent caries.

Diabetes mellitus exacerbates inflammation in a murine model of ligature-induced peri-implantitis: A histological and microtomographic study.

To investigate the influence of diabetes mellitus (DM) in a murine model of peri-implantitis (PI). Twenty-seven 4-week-old C57BL/6J male mice had their first and second maxillary left molars extracted. Eight weeks later, one machined implant was placed in each mouse. Four weeks after osseointegration, the mice were divided into three groups: (a) control (C), (b) PI and (c) DM + PI. DM was induced by streptozotocin (STZ) administration. After DM induction, PI was induced using ligatures for 2 weeks. The hemimaxillae were collected for micro-CT and histological analyses. The primary outcomes consisted of linear (mm) and volumetric (mm3) bone loss. Secondary outcomes were based on histological analysis and included inflammatory infiltrate, osteoclastic activity, matrix organization, composition and remodelling. Data are presented as means ± SEM. Statistical analyses were performed using one-way ANOVA, followed by Tukey's test. Gingival tissue oedema was detected in the PI and DM + PI groups. Micro-CT showed significantly increased linear and volumetric bone loss in the DM + PI group compared to the C and PI groups. H&E staining showed greater inflammatory response and bone resorption in the PI and DM + PI groups than in the C group. The DM + PI group had significantly higher osteoclast numbers than the C and PI groups. Picrosirius red stained less for types I and III collagen in the PI and DM + PI groups than in the C group. There was a significant increase in monocyte/macrophage (CD-11b) counts and matrix metalloproteinases (MMP-2 and MMP-8) marker levels and a significant decrease in the matrix metalloproteinases inhibition marker (TIMP-2) levels in the DM + PI group compared to the C and PI groups. DM exacerbates PI-induced soft-tissue inflammation, matrix degradation and bone loss.

Genetic removal of Nlrp3 protects against sporadic and R345W Efemp1-induced basal laminar deposit formation.

Chronic, unresolved inflammation has long been speculated to serve as an initiating and propagating factor in numerous neurodegenerative diseases, including a leading cause of irreversible blindness in the elderly, age-related macular degeneration (AMD). Intracellular multiprotein complexes called inflammasomes in combination with activated caspases facilitate production of pro-inflammatory cytokines such as interleukin 1 beta. Specifically, the nucleotide-binding oligomerization (NOD)-like receptor protein 3 (NLRP3) has received heightened attention due to the wide range of stimuli to which it can respond and its potential involvement in AMD. In this study, we directly tested the role of Nlrp3 and its downstream effector, caspase 1 (Casp1) in mediating early AMD-like pathology (i.e., basal laminar deposits [BLamDs]) in wild-type (WT) mice and the Malattia Leventinese/Doyne honeycomb retinal dystrophy (ML/DHRD) mouse model (p.R345W mutation in Efemp1). Compared to aged-matched controls, R345W +/+ knockin mice demonstrated increased Muller cell gliosis, subretinal Iba-1 + microglial cells, higher Nlrp3 immunoreactivity in the retina, as well as significant transcriptional upregulation of complement component 3, Nlrp3, pro-Il1b, pro-caspase-1, and tissue inhibitor of matrix metalloproteinase 3 in the retinal pigmented epithelium (RPE)/choroid. These findings were accompanied by an age-related increase in BLamD formation in the R345W +/+ mice. Genetic elimination of either Nlrp3 or Casp1 significantly reduced both the size and coverage of BLamDs in the R345W +/+ background, highlighting an important and underappreciated pathway that could affect ML/DHRD onset and progression. Moreover, Nlrp3 knockout reduced spontaneous, idiopathic BLamDs in WT mice, suggesting translatability of our findings not only to rare inherited retinal dystrophies, but also potentially to AMD itself.

Clinical and morphological features of the myometrium in patients with placental adherent and invasive pathology

BACKGROUND: The fast increase in the frequency of placental adherent and invasive pathology worldwide accounts for the growing interest in studying the pathogenesis of placenta accreta. According to the literature, the clinical and morphological features of the myometrium from the placental attachment area in placental adherent and invasive pathology are described in single articles. Study of morphological features using proteolytic markers in the myometrium from the placenta accreta area could help in understanding the pathogenesis of placental adherent and invasive pathology. For the first time in this study, the clinical and morphological features of the myometrium from the placental attachment area in patients with uterine scar and placental adherent and invasive pathology are compared with the myometrium of women with uterine scar without placenta accreta and intact myometrium. AIM: The aim of this study was to evaluate the expression of matrix metalloproteinase 2 and tissue inhibitors of matrix metalloproteinases 1 and 2 in myometrial biopsies in placental adherent and invasive pathology. MATERIALS AND METHODS: This study included 15 myometrial biopsies from the placental site, which were divided into three groups according to the clinical diagnosis of the patients: group 1 (main group), with a uterine scar after cesarean section and placental adherent and invasive pathology (n = 5); group 2 (comparison group), with a uterine scar after cesarean section (n = 5); group 3 (control group), with a normal pregnancy without a uterine scar (n = 5). Histological examination was carried out using the standard procedure. Immunohistochemical study was performed using antibodies to matrix metalloproteinase 2 and tissue inhibitors of matrix metalloproteinases 1 and 2 (Abcam, USA). Morphometry was carried out using the VideoTesT-Morphology 5.2 program (Videotest Ltd., Russia). The statistical analysis was performed using the IBM SPSS Statistics 26.0 software. RESULTS: In the biopsies of the main group, we verified terminal chorionic villi with hypervascularization and uneven plethora of the vascular bed among hypertrophied muscle fibers, a basal plate with lumen ectasia of unevenly full-blooded vessels, and the absence of a decidual membrane, in contrast to groups 2 and 3. The matrix metalloproteinase 2 expression area in the main group was higher than in the comparison and control groups (p = 0.008; p1–2 = 0.049*, p1–3 = 0.011), the matrix metalloproteinase 2 optical density not differing between the groups (p = 0.122). The tissue inhibitors of matrix metalloproteinases 1 expression area was higher in the comparison group compared to the main group (p = 0.035; p2–3 = 0.032), and the tissue inhibitors of matrix metalloproteinases 1 and 2 optical density was higher in the comparison group compared to control (p = 0.008; p2–3 = 0.005). CONCLUSIONS: In myometrial biopsies of patients with a uterine scar and placental adherent and invasive pathology, we verified pathology of the basal plate of the placenta, increased matrix metalloproteinase 2 expression and decreased tissue inhibitors of matrix metalloproteinases 1 expression, which may indicate the peculiarities of the inflammatory response in the myometrium in placental adherent and invasive pathology.

Elevated MMP-9, Survivin, TGB1 and Downregulated Tissue Inhibitor of TIMP-1, Caspase-3 Activities are Independent of the Low Levels miR-183 in Endometriosis.

This study aimed to measure the correlation between miR-183 and gene expression that regulates apoptosis and adhesion mechanism that may be linked to the pathogenesis of endometriosis. Forty-four subjects, including 22 control subjects, participated in this study. We collected ectopic endometriosis and endometrial samples. For the control, the sample was taken from endometrial tissue through pipelle biopsy. RNA was extracted from all tissues using RNA mini kit, and the expression was assessed using quantitative-real time PCR. Relative mRNA and miRNA expression were presented using the formula of the Livak method. The data were statistically analyzed using GraphPad Prism 8. The expression of Caspase-3, Survivin, Integrin β1 (ITGB1), matrix metalloproteinase-9 (MMP-9), and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) (adhesion- and apoptosis-related gene) were calculated using the relative expression method. We found significant differences in Caspase-3, Survivin, ITGB1, MMP-9, and TIMP-1 expression between ectopic endometriosis tissues of women with endometriosis compared to healthy endometrium. MMP-9, Survivin, and ITGB1 was significantly increased in the endometriosis group, while Caspase-3, TIMP-1, and miR-183 were significantly reduced in the endometriosis group. No correlation was found between the expression level of miR-183 and Caspase3, Survivin, ITGB1, and Cadherin in both tissue types. Despite the difference in expression levels of miR-183 and associated adhesion- and apoptosis-related genes, there was no significant association between miR-183 with specific adhesion and apoptosis genes in endometriosis tissue.

Matrix Metalloproteinase Inhibition in Rat Abdominal Aortic Aneurysm

Matrix Metalloproteinase Inhibition in Rat Abdominal Aortic Aneurysm

Engineered TIMP2 with narrow MMP-9 specificity is an effective inhibitor of invasion and proliferation of triple-negative breast cancer cells

Matrix metalloproteinases (MMPs) are a family of endopeptidases that degrade extracellular matrix (ECM) proteins, functioning in various physiological processes such as tissue remodeling, embryogenesis, and morphogenesis. Dysregulation of these enzymes is linked to multiple diseases. Specific inhibition of particular MMPs is crucial for anti-MMP drug development as some MMPs have shown anti-disease properties. In this study, we aimed to design a highly specific inhibitor of MMP-9, that plays a crucial role in cell invasion and metastasis, using tissue inhibitor of metalloproteinases 2 (TIMP2), an endogenous broad-family MMP inhibitor, as a prototype. In our earlier work, we were able to narrow down the specificity of the N-terminal domain of TIMP2 (N-TIMP2) toward MMP-9, yet at the expense of lowering its affinity to MMP-9. In this study, a library of N-TIMP2 mutants based on previous design with randomized additional positions was sorted for binding to MMP-9 using yeast surface display. Two selected N-TIMP2 mutants were expressed, purified and their inhibitory activity against a panel of MMPs was measured. The best engineered N-TIMP2 mutant (REY) exhibited a 2-fold higher affinity to MMP-9 compared to that of the WT N-TIMP2, and 6- to 1.1x104-fold increase in binding specificity toward MMP-9 compared to five alternative MMPs. Moreover, REY demonstrated a significant increase in inhibition of cell invasion and proliferation compared to the WT N-TIMP2 in MDA-MB-231 breast cancer cells. Therefore, our engineered N-TIMP2 mutant emerges as a promising candidate for future therapeutic development, offering precise targeting of MMP-9 in MMP-9-driven diseases.

Determining key residues of engineered scFv antibody variants with improved MMP-9 binding using deep sequencing and machine learning

Given the crucial role of specific matrix metalloproteinases (MMPs) in the extracellular matrix, an imbalance in the regulation of activation of matrix metalloproteinase-9 (MMP-9) zymogen and inhibition of the enzyme can result in various diseases, such as cancer, neurodegenerative, and gynecological diseases. Thus, developing novel therapeutics that target MMP-9 with single-chain antibody fragments (scFvs) is a promising approach. We used fluorescent-activated cell sorting (FACS) to screen a synthetic scFv antibody library displayed on yeast for enhanced binding to MMP-9. The screened scFv mutants demonstrated improved binding to MMP-9 compared to the natural inhibitor of MMPs, tissue inhibitor of metalloproteinases (TIMPs). To identify the molecular determinants of these engineered scFv variants that affect binding to MMP-9, we used next-generation DNA sequencing and computational protein structure analysis. Additionally, a deep-learning language model was trained on the screened scFv library of variants to predict the binding affinities of scFv variants based on their CDR-H3 sequences.

Protein Kinase C and Matrix Metalloproteinases Expression Using Phorbol Myristate Acetate in Degenerative Intervertebral Disc Cells.

Degeneration of nucleus pulposus (NP) cells involves multiple factors. The relationship between the canonical Wnt/β-catenin signaling pathway and matrix metalloproteinases (MMPs) is important in cellular senescence. Protein kinase C (PKC), an intermediate of the non-canonical Wnt pathway stimulated by phorbol myristate acetate (PMA), possibly prevents NP cell senescence, although not yet demonstrated in human-based studies. This study aimed to investigate the effect of PMA stimulation on the non-canonical and canonical Wnt pathways and MMP expression in human NP cells to ascertain its inhibitory effects on the senescence of NP cells. Human disc tissues of Pfirrmann grades 1 and 2 were collected from patients during spinal surgery and subsequently cultured. Protein and ribonucleic acid (RNA) were isolated from NP cells treated with PMA (400 nM) for 24 hours. Expression of MMP1, MMP13, tissue inhibitor of matrix metalloproteinase 1 (TIMP1), a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5), transient receptor potential vanilloid 4 (TRPV4), interleukin-6 (IL-6), and β-catenin were detected using western blot analysis. Messenger RNA (mRNA) expression of type II collagen and glycosaminoglycan (GAG) were analyzed using reverse transcription polymerase chain reaction. IL-6 and prostaglandin E2 (PGE2) levels were measured using enzyme-linked immunosorbent assay. Expression of PKC-δ (intermediate of the non-canonical Wnt pathway) and β-catenin (intermediate of the canonical Wnt pathway) was increased by PMA treatment. The mRNA levels of type II collagen and GAG increased; however, their protein levels were not altered. PMA treatment increased the expression of MMP1, TIMP1, ADAMTS5, IL-6, PGE2, and TRPV4; however, the expression of MMP13 and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) was unaltered. PMA activated PKC-δ, affecting the non-canonical Wnt pathway; however, its effect on β-catenin in the canonical Wnt pathway was limited. β-catenin activation through the TRPV4 channel led to increased expression of MMP1 and ADAMTS5 and that of IL-6 and PGE2 owing to NF-κB expression. Consequently, the degeneration of NP cells was not prevented.

Effect of matrix metalloproteinase inhibitors on microtensile bond strength to dentin using self-etch adhesive - in vivo study

Effect of matrix metalloproteinase inhibitors on microtensile bond strength to dentin using self-etch adhesive - in vivo study

A molecular docking insight: Phyllanthus niruri L. constituents targeting MMP-9 for angiogenesis inhibition and IL-1beta for antiinflammatory action in endometriosis therapy

Endometriosis, characterized by inflammatory lesions resembling endometrium outside the uterine cavity, induces chronic inflammation, anatomical changes and persistent pain. The expression of Interleukin 1beta (IL-1beta) is significantly associated with endometriosis, contributing to inflammatory process and fertility issues. Matrix metalloproteinase 9 (MMP-9) is crucial in the adhesion and angiogenesis of endometrial tissue. This study investigates the potential of Phyllanthus niruri L. in inhibiting MMP-9 and IL-1beta through molecular docking analysis. Molecular docking was performed using Discovery Studio Visualizer, Open Babel, PyRx and AutoDock Vina. The inhibitory activities of bioactive compounds on MMP-9 and IL-1beta were predicted. Biological activity and cytotoxicity were assessed using PASS and CLC-Pred respectively. Kaempferol and quercetin from P. niruri exhibited significant MMP-9 expression inhibitory activity. Search Tool for Interacting Chemicals (STITCH) analysis revealed interactions of quercetin and galangin with specific proteins involved in pathways related to endometriosis. Biological activity analysis indicated that quercetin, kaempferol, herbacetin and galangin show potential as MMP -9 expression inhibitors. CLC-Pred analysis suggested high cytotoxicity of kaempferol against glioma. Molecular docking results showed quercetin's potential as an MMP-9 inhibitor and galangin's potential as an IL-1 beta inhibitor. These findings support the therapeutic potential of P. niruri for endometriosis, providing insights for further research in developing innovative therapies targeting endometriosis-related inflammation and angiogenesis.

Circulating Matrix Metalloproteinases for Prediction of Aortic Dilatation in Children with Bicuspid Aortic Valve: A Single-Center, Observational Study

Circulating biomarkers have been proposed for early identification of aortic dilatation progression associated with bicuspid aortic valve (BAV), but matrix metalloproteinases (MMPs) are distinguished as signatures of increased extracellular matrix degradation, a landmark of aneurysm formation. The current study aims to identify the role of MMP-1, MMP-2, MMP-9, and the MMP inhibitor, TIMP-1, in identifying aortic dilation in children with BAV. We conducted a study on 73 children divided into two study groups, depending on the presence of aortic dilatation (group 1–43 BAV controls and group 2–30 children with BAV and aortic dilatation). Each patient underwent a cardiac ultrasound and, in each case, serum MMP-1, MMP-2, MMP-9, and TIMP-1 were quantified using xMAP technology. Comparison of the MMPs between the two study groups revealed significantly higher values only in the case of TIMP-1, among BAV controls. Moreover, the same TIMP-1 inversely correlated with aortic annulus absolute size and z score, as well as with ascending aorta z score. No particular correlation between the aortic phenotype and the presence of aortic dilatation was found. Future longitudinal research starting at pediatric ages could show the significance of MMPs screening in BAV individuals as predictors of aortic aneurysm formation.

Alpha(α)-mangostin (Xanthone of Garcinia mangostana L.): Augmenting Macrophages Activity for an Effective Diabetic Wound Healing

Diabetic wounds are particularly difficult to treat medically because they heal at a slower pace than regular wounds. Macrophages are essential in all stages of normal wound healing, including inflammation, proliferation and rem odelling. When wound healing is affected, macrophages can reduce the level of growth factor and increase the level of interleukin-6 (IL-6), disrupt the balance between tissue inhibitors of metalloproteinase (TIMPs) and matrix metalloproteinase (MMPs), which can slow down healing. Alpha(α)-mangostin, a natural xanthone derived from the pericarp of the mangosteen, has gained considerable attention due to its anti-inflammatory properties, suggesting its potential to promote wound healing. However, its exact role in healing diabetic foot ulcers, common in diabetes, remains unclear. Hence, this study aims to explore how α-mangostin might affect diabetic wound healing by evaluating its impact on PDGF, CTGF, BFGF, VEGF, TGF-β, MMP-9, TIMP-2 and IL-6 secretion in macrophage cells. Human monocytic macrophages (THP-1) were incubated with a 35 mM glucose solution for 72 h to create a glucose-enriched medium. The cells were then incubated with α-mangostin (0.15, 2.5 and 5 µg/mL) together with 35 mM glucose. Carboxymethyl cellulose (CMC) served as positive controls; glucose-enriched media and media-alone served as negative controls. Protein expression was measured using ELISA. α-mangostin (2.5 µg/mL) increased the levels of PDGF and VEGF and decreased the level of MMP-9 compared to glucose controls. There was no significant difference in other growth factors, TIMP-2 and IL-6 protein levels across any of the treatment groups compared to glucose controls. In conclusion, α-mangostin particularly at 2.5 µg/mL demonstrated a significant increase in PDGF and VEGF levels while simultaneously reducing MMP-9 in macrophage cells under glucose-induced conditions. These findings suggest that α-mangostin holds the potential for enhancing the healing of chronic wounds in diabetic conditions. HIGHLIGHTS Treatment of macrophages (THP-1 cells) with 2.5 µg/mL Alpha (α)-mangostin in a high glucose medium led to a significant increase in PDGF and VEGF secretion compared to glucose control. Treatment of the cells with 2.5 µg/mL Alpha (α)-mangostin in a high glucose medium led to a significant reduction of MMP-9 secretion compared to glucose control. There is no significant effects of Alpha (α)-mangostin on IL-6 and TIMP secretion compared to controls This study suggests that Alpha (α)-mangostin may be beneficial in promoting diabetic wound healing via stimulation of PDGF and VEGF secretion and reducing MMP-9 levels. GRAPHICAL ABSTRACT

Matrix metalloproteinases mediate influenza A-associated shedding of the alveolar epithelial glycocalyx.

The alveolar epithelium is protected by a heparan sulfate-rich, glycosaminoglycan layer called the epithelial glycocalyx. It is cleaved in patients with acute respiratory distress syndrome (ARDS) and in murine models of influenza A (IAV) infection, shedding fragments into the airspace from the cell surface. Glycocalyx shedding results in increased permeability of the alveolar-capillary barrier, amplifying acute lung injury. The mechanisms underlying alveolar epithelial glycocalyx shedding in IAV infection are unknown. We hypothesized that induction of host sheddases such as matrix metalloproteinases (MMPs) during IAV infection results in glycocalyx shedding and increased lung injury. We measured glycocalyx shedding and lung injury during IAV infection with and without treatment with the pan-MMP inhibitor Ilomastat (ILO) and in an MMP-7 knock out (MMP-7KO) mouse. C57BL/6 or MMP-7KO male and female mice were given IAV A/PR/8/34 (H1N1) at 30,000 PFU/mouse or PBS intratracheally. For some experiments, C56BL/6 mice were infected in the presence of ILO (100mg/kg) or vehicle given daily by IP injection. Bronchoalveolar lavage (BAL) and lung tissue were collected on day 1, 3, and 7 for analysis of glycocalyx shedding (BAL Syndecan-1) and lung injury (histology, BAL protein, BAL cytokines, BAL immune cell infiltrates, BAL RAGE). Expression and localization of the sheddase MMP-7 and its inhibitor TIMP-1 was examined by RNAScope. For in vitro experiments, MLE-12 mouse lung epithelial cells were cultured and treated with active or heat-inactivated heparinase (2.5 U/mL) prior to infection with IAV (MOI 1) and viral load and MMP-7 and TIMP-1 expression analyzed. IAV infection caused shedding of the epithelial glycocalyx into the BAL. Inhibition of MMPs with ILO reduced glycocalyx shedding by 36% (p = 0.0051) and reduced lung epithelial injury by 40% (p = 0.0404). ILO also reduced viral load by 68% (p = 0.027), despite having no significant effect on lung cytokine production. Both MMP-7 and its inhibitor TIMP-1 were upregulated in IAV infected mice: MMP-7 colocalized with IAV, while TIMP-1 was limited to cells adjacent to infection. However, MMP-7KO mice had similar glycocalyx shedding, epithelial injury, and viral load compared to WT littermates, suggesting redundancy in MMP sheddase function in the lung. In vitro, heparinase treatment before infection led to a 52% increase in viral load (p = 0.0038) without altering MMP-7 or TIMP-1 protein levels. Glycocalyx shedding and MMPs play key roles in IAV-induced epithelial injury, with significant impact on IAV viral load. Further studies are needed to understand which specific MMPs regulate lung epithelial glycocalyx shedding.

Improving Circulation Half-Life of Therapeutic Candidate N-TIMP2 by Unfolded Peptide Extension.

Matrix metalloproteinases (MMPs) are significant drivers of many diseases, including cancer, and are established targets for drug development. Tissue inhibitors of metalloproteinases (TIMPs) are endogenous MMP inhibitors and are being pursued for the development of anti-MMP therapeutics. TIMPs possess many attractive properties for drug candidates, such as complete MMP inhibition, low toxicity, low immunogenicity, and high tissue permeability. However, a major challenge with TIMPs is their rapid clearance from the bloodstream due to their small size. This study explores a method for extending the plasma half-life of the N-terminal domain of TIMP2 (N-TIMP2) by appending it with a long, intrinsically unfolded tail containing Pro, Ala, and Thr (PATylation). We designed and produced two PATylated N-TIMP2 constructs with tail lengths of 100 and 200 amino acids (N-TIMP2-PAT100 and N-TIMP2-PAT200). Both constructs demonstrated higher apparent molecular weights and retained high inhibitory activity against MMP-9. N-TIMP2-PAT200 significantly increased plasma half-life in mice compared to the non-PATylated variant, enhancing its therapeutic potential. PATylation offers distinct advantages for half-life extension, such as fully genetic encoding, monodispersion, and biodegradability. It can be easily applied to N-TIMP2 variants engineered for high affinity and selectivity toward individual MMPs, creating promising candidates for drug development against MMP-related diseases.

Cancer Prevention and Treatment with Polyphenols: Type IV Collagenase-Mediated Mechanisms.

Polyphenols are natural compounds found in many plants and their products. Their high structural diversity bestows upon them a range of anti-inflammatory, anti-oxidant, proapoptotic, anti-angiogenic, and anti-metastatic properties, and a growing body of research indicates that a polyphenol-rich diet can inhibit cancer development in humans. Polyphenolic compounds may modulate the expression, secretion, or activity of compounds that play a significant role in carcinogenesis, including type IV collagenases, such as matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9), by suppressing cellular signaling pathways such as nuclear factor-kappa B. These enzymes are responsible for the degradation of the extracellular matrix, thus promoting the progression of cancer. This review discusses the current state of knowledge concerning the anti-cancer activity of polyphenols, particularly curcumin, resveratrol, epigallocatechin-3-gallate, genistein, and quercetin, with a specific focus on their anti-invasive and anti-metastatic potential, based on the most recent in vitro and in vivo studies. It appears that polyphenols may be valuable options for the chemoprevention and treatment of cancer via the inhibition of MMP-2 and MMP-9 and the suppression of signaling pathways regulating their expression and activity.

The Effect of Tissue Inhibitor of Metalloproteinases on Scar Formation after Spinal Cord Injury.

Spinal cord injury (SCI) often results in permanent loss of motor and sensory function. After SCI, the blood-spinal cord barrier (BSCB) is disrupted, causing the infiltration of neutrophils and macrophages, which secrete several kinds of cytokines, as well as matrix metalloproteinases (MMPs). MMPs are proteases capable of degrading various extracellular matrix (ECM) proteins, as well as many non-matrix substrates. The tissue inhibitor of MMPs (TIMP)-1 is significantly upregulated post-SCI and operates via MMP-dependent and MMP-independent pathways. Through the MMP-dependent pathway, TIMP-1 directly reduces inflammation and destruction of the ECM by binding and blocking the catalytic domains of MMPs. Thus, TIMP-1 helps preserve the BSCB and reduces immune cell infiltration. The MMP-independent pathway involves TIMP-1's cytokine-like functions, in which it binds specific TIMP surface receptors. Through receptor binding, TIMP-1 can stimulate the proliferation of several types of cells, including keratinocytes, aortic smooth muscle cells, skin epithelial cells, corneal epithelial cells, and astrocytes. TIMP-1 induces astrocyte proliferation, modulates microglia activation, and increases myelination and neurite extension in the central nervous system (CNS). In addition, TIMP-1 also regulates apoptosis and promotes cell survival through direct signaling. This review provides a comprehensive assessment of TIMP-1, specifically regarding its contribution to inflammation, ECM remodeling, and scar formation after SCI.

Inhibition of osteosarcoma metastasis in vivo by targeted downregulation of MMP1 and MMP9

Osteosarcoma (OS) mortality stems from lung metastases. Matrix metalloproteinases (MMPs) facilitate metastatic dissemination by degrading extracellular matrix components. Herein we studied the impact of targeted MMP downregulation on OS metastasis. Differential gene expression analysis of human OS cell lines revealed high MMP9 expression in the majority of OS cell lines. Furthermore, 143B, a metastatic OS cell line, exhibited increased MMP1 and MMP9 mRNA levels. Gene set enrichment analysis on metastatic and non-metastatic OS patient specimens indicated epithelial-mesenchymal transition as the most enriched gene set, with MMP9 displaying strong association to genes in this network. Using the same dataset, Kaplan-Meier analysis revealed a correlation between MMP1 expression and dismal patient survival. Hence, we undertook targeted suppression of MMP1 and MMP9 gene expression in OS cell lines. The ability of OS cells to migrate and form colonies was markedly reduced upon MMP1 and MMP9 downregulation, whereas their cell proliferation capacity remained intact. MMP9 downregulation decreased tumor growth and lung metastases area in an orthotopic mouse OS model. Consistently, human OS lung metastasis specimens displayed marked MMP9 protein expression. Our findings highlight the role of MMP1 and MMP9 in OS metastasis, warranting further exploration of simultaneous inhibition of MMPs for future OS therapeutics.