Matrix Metalloproteinase Activity Research Articles

Anti-photoaging effect and mechanism of flexible liposomes co-loaded with apigenin and doxycycline

Prolonged exposure to UV light can lead to photo-ageing of the skin. Therefore, the development and application of anti-photoaging drugs is urgent. In this study, we co-loaded apigenin (Apn) and doxycycline (Doc), a broad-spectrum inhibitor of matrix metalloproteinases (MMPs), into flexible liposomes to exert anti-photoaging effects by combating oxidative stress, anti-inflammation, reducing the activation of MMPs and preventing collagen loss. The results showed that we prepared a flexible liposome (A/D-FLip) containing Apn and Doc. Its appearance, particle size and Zeta potential were normal and it had good encapsulation efficiency, drug loading, in vitro release and transdermal efficiency. In cellular experiments, A/D-FLip could inhibit oxidative stress damage, reduce inflammatory factors and decrease the activation of MMPs in Human immortalized keratinocytes (HaCaT) cells; in animal experiments, A/D-FLip could inhibit skin damage and reduce skin collagen loss by decreasing the activation of MMPs, thus inhibiting skin photoaging in mice. In conclusion, A/D-FLip has good anti-photoaging effects and it has the potential to become an effective skin care product or drug against UV damage and skin photoaging in the future.

ETV2 Enhances CXCL5 Secretion from Endothelial Cells, Leading to the Promotion of Vascular Smooth Muscle Cell Migration.

Abnormal communication between endothelial cells (ECs) and vascular smooth muscle cells (VSMCs) promotes vascular diseases, including atherogenesis. ETS variant transcription factor 2 (ETV2) plays a substantial role in pathological angiogenesis and the reprogramming of ECs; however, the role of ETV2 in the communication between ECs and VSMCs has not been revealed. To investigate the interactive role of ETV2 in the EC to VSMC phenotype, we first showed that treatment with a conditioned medium from ETV2-overexpressed ECs (Ad-ETV2 CM) significantly increased VSMC migration. The cytokine array showed altered levels of several cytokines in Ad-ETV2 CM compared with those in normal CM. We found that C-X-C motif chemokine 5 (CXCL5) promoted VSMC migration using the Boyden chamber and wound healing assays. In addition, an inhibitor of C-X-C motif chemokine receptor 2 (CXCR2) (the receptor for CXCL5) significantly inhibited this process. Gelatin zymography showed that the activities of matrix metalloproteinase (MMP)-2 and MMP-9 increased in the media of VSMCs treated with Ad-ETV2 CM. Western blotting revealed a positive correlation between Akt/p38/c-Jun phosphorylation and CXCL5 concentration. The inhibition of Akt and p38-c-Jun effectively blocked CXCL5-induced VSMC migration. In conclusion, CXCL5 from ECs induced by ETV2 promotes VSMC migration via MMP upregulation and the activation of Akt and p38/c-Jun.

Functional roles of tissue factor and EMMPRIN/CD147 present on bladder urothelial carcinoma-derived extracellular vesicles.

e16557 Background: Extracellular vesicles are involved in the homeostatic functions and extracellular matrix remodeling in the body. We previously found that the antithrombotic drug Tinzaparin, at clinical concentrations, induces the sufficient release of the tissue factor pathway inhibitor TFPI to functionally inhibit the prothrombotic effector tissue factor expressed on the vesicles released by malignant tumor cells. Treatment of in vitro cancer cells with Tinzaparin results in the inhibition of the tissue factor/ERK pathway. Our hypothesis is that vesicles derived from metastatic urinary tumors are enriched in tissue factor; therefore, their elusive functioning is affected by the suppressed tissue factor caused by TFPI. Here, we show that this observation may have major clinical relevance. Methods: To test the contribution of extracellular vesicles to the mechanism and extent of cell invasion, we assessed two human adherent bladder urothelial carcinoma cell lines, RT4 and J82, which represent non-muscle and muscle-invasive diseases, respectively. Urine samples were obtained from a cohort of 75 outpatients who underwent transurethral resection subdivided by the presence of cancer or muscle invasion. Vesicles were isolated using sequential centrifugation and further validated for their size, nanoparticle tracking analysis, and surface markers using flow cytometry. The tissue factor activity index levels of the vesicles and the potential for in vitro induction of matrix metalloproteinases (MMPs) were quantified using colorimetric enzymatic methods. The levels of phosphorylated and total Erk1/2 were measured using an in vitro kinase assay and western blot. Results: Vesicles derived from the invasive cells J82 displayed higher levels of the MMP inducer EMMPRIN/CD147. The distinct pattern of surface markers in muscle-invasive-derived vesicles, featuring enhanced potential for inducing Erk1/2 phosphorylation and in vitro MMP activation as key processes of extracellular matrix degradation, was drastically suppressed by a potent, specific small molecule CD147 inhibitor. There was higher tissue factor activity, a key initiator of coagulation, among the patients with bladder urothelial carcinoma than those without. In the subgroup analysis, the tissue factor activity index and CD147 were higher in patients with muscle-invasive disease than in patients with non-muscle-invasive disease, showing high accuracy in distinguishing patients regarding invasion potential. Conclusions: The tissue factor activity index of urine extracellular vesicles may be considered a marker of bladder urothelial carcinoma. In addition to tissue factor expression, CD147 is crucial in promoting the potential of extracellular vesicles to induce MMP-mediated extracellular matrix degradation and, consequently, cell invasion, thereby allowing metastatic disease progression.

PECULIARITIES OF CHANGES IN THE BALANCE OF THE ANGIOGENESIS REGULATORS AND ACTIVITIES OF MATRIX METALOPROTEINASES -2, -9 IN CHRONIC ULCERS IN DIABETIC PATIENTS

Abstract. Aim. Chronic diabetic foot ulcers are serious complications of diabetes mellitus, which account for 85 % of purulent-necrotic lesions of the lower extremities. This study was conducted to assess the levels of protein regulators of angiogenesis (vascular endothelial growth factor, or VEGF, hypoxia-inducible factor-1α and angiostatins) and to assess the activity of matrix metalloproteinases (MMPs) (gelatinases MMP-2 and -9) in chronic wound tissue of diabetic patients.
 Methods and materials: VEGF and angiostatin levels were analyzed by western blot, MMP activities were assessed by gelatin zymography. We found that the tissue of diabetic wounds is characterized by a reduced level of VEGF (by 2.5 times compared to acute wound tissue P<0.01) and increased levels of angiostatin, which are not detected in non-diabetic wounds. In the tissues of diabetic wounds, there is an approximately 5-fold increase in the activity of MMP-2 and -9 compared to intact skin tissue. The expression of the central regulator of hypoxia-related processes HIF-1α was increased by 4.4 times in diabetic wounds compared to the this value in acute wounds (P<0.01). Conclusions. We established an inverse correlation between the levels of HIF-1α and VEGF in dermal biopsies collected from chronic skin lesions. The obtained results indicate that increased production of angiogenic inhibitor, angiostatin, can counteract VEGF-induced proangiogenic signaling and, together with MMP hyperactivation, may contribute to poor ischemic ulcer healing.

Antitumor Properties of Matrikines of Different Origins: Prospects and Problems of Their Application.

Matrikines (MKs) can be a rich source of functional nutrition components and additional therapy, thereby contributing to human health care and reducing the risk of developing serious diseases, including cancer. Currently, functionally active MKs as products of enzymatic transformation by matrix metalloproteinases (MMPs) are used for various biomedical purposes. Due to the absence of toxic side effects, low species specificity, relatively small size, and presence of various targets at the cell membranes, MKs often exhibit antitumor properties and, therefore, are promising agents for antitumor combination therapy. This review summarizes and analyzes the current data on the antitumor activity of MKs of different origins, discusses the problems and prospects for their therapeutic use, and evaluates the experimental results of studying the antitumor properties of MKs from different echinoderm species generated with the help of a complex of proteolytic enzymes from red king crab Paralithodes camtschatica. Special attention is paid to the analysis of possible mechanisms of the antitumor action of various functionally active MKs, products of the enzymatic activity of various MMPs, and the existing problems for their use in antitumor therapy.

Signal-On Mass Spectrometric Biosensing of Multiplex Matrix Metalloproteinases with a Phospholipid-Structured Mass-Encoded Microplate.

The detection of matrix metalloproteinases (MMPs) is of great importance for diagnosis and staging of cancer. This work proposed a signal-on mass spectrometric biosensing strategy with a phospholipid-structured mass-encoded microplate for assessment of multiplex MMP activities. The designed substrate and internal standard peptides were subsequently labeled with the reagents of isobaric tags for relative and absolute quantification (iTRAQ), and DSPE-PEG(2000)maleimide was embedded on the surface of a 96-well glass bottom plate to fabricate the phospholipid-structured mass-encoded microplate, which offered a simulated environment of the extracellular space for enzyme reactions between MMPs and the substrates. The strategy achieved multiplex MMP activity assays by dropping the sample in the well for enzyme cleavages, followed by adding trypsin to release the coding regions for ultrahigh performance liquid chromatography-tandem mass spectrometric (UHPLC-MS/MS) analysis. The peak area ratios of released coding regions and their respective internal standard (IS) peptides exhibited satisfied linear ranges of 0.05-50, 0.1-250, and 0.1-100 ng mL-1 with the detection limits of 0.017, 0.046, and 0.032 ng mL-1 for MMP-2, MMP-7, and MMP-3, respectively. The proposed strategy demonstrated good practicability in inhibition analysis and detections of multiplex MMP activities in serum samples. It is of great potential for clinical applications and can be expanded for multiplex enzyme assays.

RETRACTED ARTICLE: IL-1RA inhibits esophageal carcinogenesis and lymphangiogenesis via downregulating VEGF-C and MMP9

Interleukin-1 receptor antagonist (IL-1RA) has been shown to play an important role in cancer progression. However, its pathogenic effects and molecular mechanism in the malignant progression of esophageal squamous cell carcinoma (ESCC) remain largely unknown. This study was designed to explore the function of IL-1RA in ESCC and determine the relationship between IL-1RA and lymph node metastasis in ESCC patients. The clinical relevance of IL-1RA in relation to the clinicopathological features and prognosis of 100 ESCC patients was analyzed. The function and underlying mechanisms of IL-1RA in the growth, invasion, and lymphatic metastasis in ESCC were explored both in vitro and in vivo. The therapeutic effect of anakinra, an IL-1 receptor antagonist, on ESCC was also evaluated in animal experiments. Downregulation of IL-1RA was observed in ESCC tissues and cells and was found to be strongly correlated with pathological stage (P = 0.034) and lymphatic metastasis (P = 0.038). Functional assays demonstrated that upregulation of IL-1RA reduced cell proliferation, migration, and lymphangiogenesis both in vitro and in vivo. Mechanistic studies revealed that overexpression of IL-1RA activated the epithelial-to-mesenchymal transition (EMT) in the ESCC cells through activation of MMP9 and regulation of the expression and secretion of VEGF-C through the PI3K/NF-κB pathway. Anakinra treatment resulted in significant inhibition of tumor growth, lymphangiogenesis, and metastasis. IL-1RA inhibits lymph node metastasis of ESCC by regulating the EMT through activation of matrix metalloproteinase 9(MMP9) and lymphangiogenesis, driven by VEGF-C and the NF-κB signaling pathway. Anakinra may be an effective drug for the inhibition of ESCC tumor formation and lymph node metastasis.

Matrix metalloproteinases in arthritis: towards precision medicine

Proteolysis of structural molecules of the extracellular matrix (ECM) is an irreversible post-translational modification in all arthropathies. Common joint disorders, including osteoarthritis and rheumatoid arthritis, have been associated with increased levels of matrix remodelling enzymes, including matrix metalloproteinases (MMPs). MMPs, in concert with other host proteinases and glycanases, destroy proteoglycans, collagens and other ECM molecules. MMPs may also control joint remodelling indirectly by signalling through cell-surface receptors or by proteolysis of cytokines and receptor molecules. After synthesis as pro-forms, MMPs can be activated by various types of post-translational modifications, including proteolysis. Once activated, MMPs are controlled by general and specific tissue inhibitors of metalloproteinases (TIMPs). In rheumatoid arthritis, proteolysis of the ECM results in so-called remnant epitopes that enhance and perpetuate autoimmune processes in susceptible hosts. In osteoarthritis, the considerable production of MMP-13 by chondrocytes, often concurrent with mechanical overload, is a key event. Hence, information about the regulation, timing, localization and activities of MMPs in specific disease phases and arthritic entities will help to develop better diagnostics. Insights into beneficial and detrimental effects of MMPs on joint tissue inflammation are also necessary to plan and execute (pre)clinical studies for better therapy and precision medicine with MMP inhibitors. With the advances in proteomics and single-cell transcriptomics, two critical points need attention: neglected neutrophil MMP biology, and the analysis of net proteolytic activities as the result of balances between MMPs and their inhibitors.

Platelet rich fibrin effect in MMP-9 and TIMP-1 expression as bacterial keratitis therapy

Bacterial keratitis is an ocular emergency and is one of the leading causes of blindness worldwide. Pseudomonas aeruginosa is the most frequent causative agent of keratitis. The infectious process of P. aeruginosa is highly invasive and can rapidly lead to corneal ulcers, ocular infections (endophthalmitis), and corneal perforation. Delayed or inadequate treatment may result in complications such as corneal fibrosis and neovascularization. Platelet-rich fibrin (PRF), which contains high levels of platelets, can aid in hemostasis, cell migration, and proliferation to accelerate the wound healing process. PRF contains several growth factors, such as Platelet-derived growth factor (PDGF), Vascular endothelial growth factor (VEGF), and Transforming growth factor (TGF- β), which are released in a time-dependent manner during the wound healing process, ranging from 7 to 28 days. TGF- β regulates the expression and activity of matrix metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-9) involved in wound remodeling and healing, where MMPs are regulated or balanced with tissue inhibitors of metalloproteinases (TIMP-1, TIMP-2). In previous studies, PRF was found to be effective in managing bacterial keratitis in cats. In this discussion, we explore the molecular effects of PRF as an adjuvant therapy for bacterial keratitis

Induction of MMP-3 and MMP-9 expression during Helicobacter pylori infection via MAPK signaling pathways.

Helicobacter pylori (H. pylori)-induced gastric pathology involves remodeling of extracellular matrix mediated by aberrant activity of matrix metalloproteinases (MMPs). We have previously shown that in vitro H. pylori infection leads to MMP-3 and MMP-9 overexpression, associated with phosphorylation of bacterial oncoprotein CagA. We extended these findings in an in vivo model of H. pylori infection and further assessed the involvement of MAPK pathways in MMP expression. C57BL/6 mice were infected with H. pylori strains HPARE, HPARE ΔCagA, and SS1, for 6 and 9 months. Transcriptional expression of Mmp-3 and Mmp-9 was evaluated via qPCR while respective protein levels in the gastric mucosa were determined immunohistochemically. Epithelial cell lines AGS and GES-1 were infected with H. pylori strain P12 in the presence of chemical inhibitors of JNK, ERK1/2, and p38 pathways, for 24 h. mRNA and protein expression of MMP-3 and MMP-9 were determined via qPCR and Western blot, respectively. We observed transcriptional activation of Mmp-3 and Mmp-9 as well as aberrant MMP-3 and MMP-9 protein expression in murine gastric tissue following H. pylori infection. CagA expression was associated with MMP upregulation, particularly during the early time points of infection. We found that inhibition of ERK1/2 resulted in reduced mRNA and protein expression of MMP-3 and MMP-9 during H. pylori infection, in both cell lines. Expressed protein levels of both MMPs were also found reduced in the presence of JNK pathway inhibitors in both cell lines. However, p38 inhibition resulted in a more complex effect, probably attributed to the accumulation of phospho-p38 and increased phospho-ERK1/2 activity due to crosstalk between MAPK pathways. H. pylori colonization leads to the upregulation of MMP-3 and MMP-9 in vivo, which primarily involves ERK1/2 and JNK pathways. Therefore, their inhibition may potentially offer a protective effect against gastric carcinogenesis and metastasis.

Abstract 378: Loss Of Nuclear Receptor Subfamily 4 Group A 1 (NR4A1) Results In Impaired Post-thrombotic Vein Wall Remodeling By Promoting A Dysregulated Antifibrinolytic Phenotype In Monocytes

Objectives: Nuclear Receptor Subfamily 4 Group A1 (NR4A1) is a suppressor of monocyte/macrophage (MO/Mϕ) inflammatory output that antagonizes TGFβ-signaling. As NR4A1 inhibits epithelial-to-mesenchymal transition and tissue fibrosis, we sought to determine the role of NR4A1 in MO/Mϕ mediated post-thrombotic vein wall remodeling, which when exaggerated, can result in post-thrombotic syndrome (PTS). Methods: To assess the role of MO/Mϕ NR4A1 in post-thrombotic vein wall remodeling, we employed a knockout-rescue schema which featured whole mouse loss of function ( Nr4a1 -/- ) models and adoptive bone marrow derived MO/Mϕtransfer from Nr4a1 replete mice (WT-BMDMs). Stasis venous thrombosis was induced by the inferior vena cava (IVC) ligation. Characterization of vein wall remodeling/endothelialization was performed using Masson Trichrome staining and CD31 immunostaining, respectively. Quantitation of mRNA and protein levels of factors critical for MO/Mϕ-mediated vein wall remodeling was performed using qRT-PCR and ELISA, and matrix metalloproteinase (MMP) activity was measured using zymography. Results: We observed that Nr4a1 -/- mice exhibited impaired vein wall remodeling after 8- and 14-days post IVC ligation as indicated by thicker vein wall measurements and impaired luminal endothelialization. Adoptive transfer of WT-BMDMs into Nr4a1 -/- mice rescued this phenotype (Fig. 1A-C). Nr4a1 -/- mice did not exhibit any differences in thrombus burden (data not shown). Finally, although we observed induced mRNA levels of Mmp2 and Mmp9 in the vein walls of Nr4a1 -/- mice that underwent adoptive transfer, we noted suppressed MMP2 enzyme activity despite no compensatory changes in MMP9 activity or TGFβ/IL6 levels (Fig. 1D, data not shown). Conclusions: NR4A1 loss results in impaired post-thrombotic vein wall remodeling and agonism of NR4A1 may be a potential therapeutic strategy to combat PTS.

Cytoprotective Properties of Citronella Oil (Cymbopogon nardus (L.) Rendl.) and Lemongrass Oil (Cymbopogon citratus (DC.) Stapf) through Attenuation of Senescent-Induced Chemotherapeutic Agent Doxorubicin on Vero and NIH-3T3 Cells.

This study aimed to determine the cytoprotective potentials of citronella (Cymbopogon nardus (L.) Rendl.) essential oil (CO) and lemongrass (Cymbopogon citratus (DC.) Stapf) essential oil (LO). The essential oils from citronella and lemongrass were obtained by steam-water distillation, then analyzed using Gas Chromatography-Mass Spectrophotometry (GC-MS) to determine the chemical constituents. The antioxidant activity of CO and LO was compared using a total antioxidant capacity kit. The viability of normal kidney epithelial cells Vero and fibroblast NIH-3T3 as the cell models were tested using a trypan blue exclusion assay. The effect of cellular senescence inhibition on both cell models was measured using senescence-associated β-galactosidase (SA-β-gal) staining. The mechanism of action of CO and LO in the protection of cellular damage against doxorubicin was also confirmed through 2',7'-dichlorofluorescin diacetate (DCFDA) staining to discover the ability to decrease reactive oxygen species (ROS) levels and a gelatin zymography assay to observe the activity of matrix metalloproteinases (MMPs). The major marker components of CO and LO were citronellal and citral, respectively. Both oils showed low cytotoxic activity against Vero and NIH-3T3 cells, with IC50 values of over 40 µg/mL. LO exhibited higher antioxidant capacity than CO, but there was no effect on the intracellular ROS level of both oils on Vero and NIH-3T3 cells. However, CO and LO decreased cellular senescence induced by doxorubicin exposure on both cells, as well as suppressed MMP-2 expression. Conclusion: Both CO and LO decrease the cellular senescence and MMP-2 expression with less cytotoxic effects on normal cells independently from their antioxidant capacities. The results were expected to support the use of CO and LO as tissue protective and anti-aging agents in maintaining the body's cellular health against chemotherapeutics or cellular damaging agents.

Abstract 527: Redox Dysregulation Of Vascular Smooth Muscle Sirtuin-1 In Marfan Syndrome

Background: Thoracic aortic aneurysms and dissections (TAAD) are life-threatening aortic conditions and a major cardiovascular complication of Marfan syndrome (MFS). We previously demonstrated that vascular smooth muscle sirtuin-1 (Sirt1), a lysine deacetylase, protects against aortic dissection by suppressing aberrant activation of matrix metalloproteinases (MMP2/9). Objective: In this study, we investigated whether Sirt1’s dysregulation contributed to TAAD in MFS. Approach & Results: Our results showed that oxidative stress markers (3-nitrotyrosine and 4-hydroxy-2-nonenal) were significantly elevated in aortas of MFS patients, compared to healthy controls. Similarly, reversible oxidative post-translational modifications (rOPTM) of protein cysteines, mainly S-glutathionylation, were dramatically increased in aortas of fibrillin-1 hypomorphic mice (Fbn1 mgR/mgR ), an established model of MFS prone to aortic dissection/rupture. We found that Sirt1 was one of these oxidatively modified proteins in Fbn1 mgR/mgR aortas and smooth muscle cells (VSMCs). rOPTM of Sirt1 resulted in decreased Sirt1 activity, measured as levels of acetylated p53 and histone 3, and increased MMP2/9 activity, measured by in gel-zymography and in situ, in VSMCs and aortas. Deletion of glutaredoxin-1 (Glrx), a specific de-glutathionylation enzyme, increased rOPTM of Sirt1, rOPTM-mediated inhibition of Sirt1 activity, and MMP2/9 activity in VSMCs. On the contrary, overexpression of Glrx or of an oxidation-resistant Sirt1 mutant with an AAV prevented these changes in VSMCs. Lastly, VSMC-specific deletion of Sirt1 in Fbn1 mgR/mgR (SMKO- Fbn1 mgR/mgR ) mice significantly worsened TAAD progression leading to aortic rupture in 50% of SMKO-Fbn1 mgR/mgR mice, compared to 25% of Fbn1 mgR/mgR mice. Conclusions: Our results strongly suggest that preventing rOPTM of Sirt1, thereby preserving its activity, may be a novel therapeutic strategy to prevent TAAD in individuals with MFS, for which currently no targeted therapies are available.

The invariant chain CD74 protein is a cell surface binding partner of TIMP-1 in breast cancer cells.

Tissue inhibitor of metalloproteinases-1 (TIMP-1) regulates the proteolytic activity of matrix metalloproteinases (MMPs), playing an important role in the homeostasis of the extracellular matrix. Beyond its well-known role in tissue maintenance, TIMP-1 has been associated with multiple MMP-independent cytokine-like functions. The protein structure of TIMP-1, with two distinct domains, one interacting with MMPs and another able to bind multiple partners, provides a rationale for this multi-functionality. The identification of CD63 as a cell surface receptor for TIMP-1, able to mediate intracellular signaling through the Erk/MAPK axis, provided a molecular basis for the role of TIMP-1 in cellular signaling. However, several lines of evidence suggest that TIMP-1 may be able to associate with many interaction partners, thus attaining multiple functions. To enable the identification of previously unknown interaction partners that may underpin the core cellular functions of TIMP-1, known as well as unknown, we performed a yeast two-hybrid screening using a mammary gland complementary DNA (cDNA) library. We report here the identification of multiple interactors, including MHC class II-associated invariant chain γ (CD74). We verified that CD74 interacts with TIMP-1 in breast cancer cells and that this interaction contributes to cellular internalization of TIMP-1 and mediates intracellular signaling through the Akt signaling axis in breast cancer cells. These data provide new insights into the complex nature of the functions of TIMP-1 and their potential mechanistic basis.

Sevoflurane and isoflurane anesthesia induce redox imbalance, but only sevoflurane impairs vascular contraction.

Volatile anesthetics may cause vascular dysfunction; however, underlying effects are unclear. The aim of the present study was to investigate whether sevoflurane and isoflurane affect vascular function, nitric oxide (NO) bioavailability, and biomarkers of oxidative stress and inflammation. Wistar rats were divided into three experimental groups: Not anesthetized (control group) or submitted to anesthesia with isoflurane (Iso group) or sevoflurane (Sevo group). Hemodynamic parameters were monitored during anesthesia, and blood gas values and biochemical determinants were analyzed. Isometric contractions were recorded in aortic rings. Vasoconstriction induced by potassium chloride (KCl) and phenylephrine (Phe) were measured. No differences in hemodynamic parameters and blood gasses variables were observed. Impaired KCl and Phe-induced contractions were observed in endothelium-intact aorta of Sevo compared to Iso and Control groups. Redox imbalance was found in Sevo and Iso groups. Reduced NO bioavailability and increased activity of matrix metalloproteinase 2 (MMP-2) were observed in Sevo, but not in the Iso group. While reduced IL-10 and IL-1β were observed in Sevo, increases in IL-1β in the Iso group were found. Sevoflurane, but not isoflurane, anesthesia impairs vasocontraction, and reduced NO and cytokines and increased MMP-2 activity may be involved in vascular dysfunction after sevoflurane anesthesia.

Melatonin inhibits chondrosarcoma cell proliferation and metastasis by enhancing miR-520f-3p production and suppressing MMP7 expression.

Chondrosarcoma has a high propensity to metastasize and responds poorly to chemotherapy and radiation treatment. The enzymatic activity of matrix metalloproteinases (MMPs) is very important in chondrosarcoma metastasis. Melatonin exhibits anticarcinogenic activity in many types of cancers by suppressing the expression of certain MMP family members, but this has not yet been clearly determined in chondrosarcoma. Our study demonstrates that MMP7 plays an essential role in chondrosarcoma cell proliferation, migration, and anoikis resistance. We also found that MMP7 is highly expressed in chondrosarcomas. Our in vitro and in vivo investigations show that melatonin strongly inhibits chondrosarcoma cell proliferation, migration, and anoikis resistance by directly suppressing MMP7 expression. Melatonin reduced MMP7 synthesis by promoting levels of miR-520f-3p expression, which were downregulated in human chondrosarcoma tissue samples. Pharmacological inhibition of miR-520f-3p markedly reversed the effects of melatonin upon chondrosarcoma proliferation and metastasis. Thus, our study suggests that melatonin has therapeutic potential for reducing the tumorigenesis and metastatic potential of chondrosarcoma via the miR-520f-3p/MMP7 axis.

EDTA-functionalized silica nanoparticles as a conditioning agent for dentin bonding using etch-and-rinse technique

ObjectiveThis study investigated the possibility of using ethylenediaminetetraacetic acid functionalized silica nanoparticles (EDTA-SiO2) as a dentin-conditioning agent using etch-and-rinse technique to promote the durability of dentin bonding. MethodsThe SiO2-EDTA were synthesized by N- [(3- trimethoxysilyl) propyl] ethylenediamine triacetic acid (EDTA-TMS) and SiO2 (50 nm), then characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), X-ray photoelectron spectroscopy (XPS) and transmission electron microscopy (TEM). The capacity of SiO2-EDTA to chelate calcium ions from dentin was examined by inductively coupled plasma-optic emission spectrometry (ICP-OES). The dentin surfaces conditioned with SiO2-EDTA were detected by field emission scanning electron microscopy (SEM), TEM and microhardness testing. For dentin bonding, dentin surfaces were adopted wet- or dry-bonding technique and bonded with adhesive (AdperTM Single Bond2) and applied composite resin (Filtek Z350) on them. The durability of dentin bonding was evaluated by mircotensile bond strength test, in-situ zymography and nanoleakage testing. ResultsFTIR, TGA and XPS results showed that SiO2-EDTA contained N element and carboxyl groups. SEM, TEM and microhardness results indicated that SiO2-EDTA group created extrafibrillar demineralization and retained more intrafibrillar minerals within dentin surface. In the dentin bonding experiment, SiO2-EDTA group achieved acceptable bond strength, and reduced the activity of matrix metalloproteinase and nanoleakage along bonding interface. ConclusionIt was possible to generate a feasible dentin conditioning agent (SiO2-EDTA), which could create dentin extrafibrillar demineralization and improve dentin bond durability. Clinical significanceThis study introduces a new dentin conditioning scheme based on SiO2-EDTA to create extrafibrillar demineralization for dentin bonding. This strategy has the potential to be used in clinic to promote the life of restoration bonding.

Histopathological, biochemical and molecular studies on a model of systemic autoimmune disease induced by sodium silicate in non-genetically prone rats

Sodium silicate (Na2SiO3) is an inorganic silica salt used in many products. Few studies reported autoimmune diseases (AIDs) due to Na2SiO3 exposure. This study investigates the role of Na2SiO3 exposure by different routes and doses in AID development in rats. We assigned 40 female rats to four groups: G1 control group, G2 rats were subcutaneously injected with 5 mg Na2SiO3 suspension, and G3 and G4 rats were orally administered 5 mg and 7 mg Na2SiO3 suspension, respectively. Na2SiO3 was administered weekly for 20 weeks. Serum anti-nuclear antibodies (ANA) detection, histopathology of kidney, brain, lung, liver, and heart, oxidative stress biomarkers (MDA and GSH) in tissues, Matrix metalloproteinase activity in serum, TNF-α, and Bcl-2 expression in tissues were performed. ANA was significantly increased in silicate groups, especially G2. Creatinine was significantly increased in silicate groups. Histopathology revealed vasculitis and fibrinoid degeneration of blood vessels, a picture of immune-mediated glomerulonephritis in the kidneys, and chronic interstitial pneumonia with medial hypertrophy of pulmonary blood vessels. The activity of gelatinases (MMP-2 and MMP-9) and collagenase (MMP-13), which play role in inflammation, remodeling, and immune complex degradation, were significantly increased in the silicate-exposed groups. Bcl-2 was significantly decreased, indicating apoptosis. Therefore, oral administration and subcutaneous injection of Na2SiO3 induced immune-mediated glomerulonephritis with elevated ANA levels and overexpression of TNF-α in rats.

The in vitro anti-cancer synergy of neurokinin-1 receptor antagonist, aprepitant, and 5-aminolevulinic acid in glioblastoma.

Glioblastoma multiforme (GBM) is the most malignant type of cerebral neoplasm in adults with a poor prognosis. Currently, combination therapy with different anti-cancer agents is at the forefront of GBM research. Hence, this study aims to evaluate the potential anti-cancer synergy of a clinically approved neurokinin-1 receptor antagonist, aprepitant, and 5-aminolevulinic acid (5-ALA), a prodrug that elicits fluorescent porphyrins in gliomas on U-87 human GBM cells. We found that aprepitant and 5-ALA effectively inhibited GBM cell viability. The combinatorial treatment of these drugs exerted potent synergistic growth inhibitory effects on GBM cells. Moreover, aprepitant and 5-ALA induced apoptosis and altered the levels of apoptotic genes (up-regulation of Bax and P53 along with downregulation of Bcl-2). Furthermore, aprepitant and 5-ALA increased the accumulation of protoporphyrin IX, a highly pro-apoptotic and fluorescent photosensitizer. Aprepitant and 5-ALA significantly inhibited GBM cell migration and reduced matrix metalloproteinases (MMP-2 and MMP-9) activities. Importantly, all these effects were more prominent following aprepitant-5-ALA combination treatment than either drug alone. Collectively, the combination of aprepitant and 5-ALA leads to considerable synergistic anti-proliferative, pro-apoptotic, and anti-migratory effects on GBM cells and provides a firm basis for further evaluation of this combination as a novel therapeutic approach for GBM.

Cyclic Stretch-Induced Mechanical Stress Applied at 1 Hz Frequency Can Alter the Metastatic Potential Properties of SAOS-2 Osteosarcoma Cells.

Recently, there has been an increasing focus on cellular morphology and mechanical behavior in order to gain a better understanding of the modulation of cell malignancy. This study used uniaxial-stretching technology to select a mechanical regimen able to elevate SAOS-2 cell migration, which is crucial in osteosarcoma cell pathology. Using confocal and atomic force microscopy, we demonstrated that a 24 h 0.5% cyclic elongation applied at 1 Hz induces morphological changes in cells. Following mechanical stimulation, the cell area enlarged, developing a more elongated shape, which disrupted the initial nuclear-to-cytoplasm ratio. The peripheral cell surface also increased its roughness. Cell-based biochemical assays and real-time PCR quantification showed that these morphologically induced changes are unrelated to the osteoblastic differentiative grade. Interestingly, two essential cell-motility properties in the modulation of the metastatic process changed following the 24 h 1 Hz mechanical stimulation. These were cell adhesion and cell migration, which, in fact, were dampened and enhanced, respectively. Notably, our results showed that the stretch-induced up-regulation of cell motility occurs through a mechanism that does not depend on matrix metalloproteinase (MMP) activity, while the inhibition of ion-stretch channels could counteract it. Overall, our results suggest that further research on mechanobiology could represent an alternative approach for the identification of novel molecular targets of osteosarcoma cell malignancy.