Activity Of MMPs Research Articles

There and back again: Intracellular trafficking, release and recycling of matrix metalloproteinases.

Matrix metalloproteinases are a family of zinc-dependent endopeptidases that are involved in a large variety of proteolytic processes in physiological and pathological scenarios, including immune cell surveillance, tissue homeostasis, or tumor cell metastasis. This is based on their ability to cleave a plethora of substrates that include components of the extracellular matrix, but also cell surface-associated and intracellular proteins. Accordingly, a tight regulatory web has evolved that closely regulates spatiotemporal activity of specific MMPs. An often underappreciated mechanism of MMP regulation involves their trafficking to and from specific subcellular sites that require MMP activity only for a certain period. In this review, we focus on the current knowledge of MMP intracellular trafficking, their secretion or surface exposure, as well as their recycling back from the cell surface. We discuss molecular mechanisms that enable these steps, in particular microtubule-dependent motility of vesicles that is driven by molecular motors and directed by vesicle regulatory proteins. Finally, we also point out open questions in the field of MMP motility that may become important in the future.

PGF2α-FP Receptor Ameliorates Senescence of VSMCs in Vascular Remodeling by Src/PAI-1 Signal Pathway.

Senescence in vascular smooth muscle cells (VSMCs) is involved in vascular remodeling of aged mice. ProstaglandinF2α- (PGF2α-) FP receptor plays a critical role in cardiovascular diseases (CVDs), hypertension, and cardiac fibrosis. However, its role in senescence-induced arteriosclerosis is yet to be fully elucidated. In this study, we found that FP receptor expression increased in aged mouse aortas and senescence VSMCs. FP receptor gene silencing can ameliorate vascular aging and inhibit oxidative stress, thereby reducing the expression of PAI-1, inhibiting the activation of MMPs, and ultimately improving the excessive deposition of ECM and delaying the process of vascular fibrosis. FP receptor could promote VSMC senescence by upregulated Src/PAI-1 signal pathway, and inhibited FP receptor/Src/PAI-1 pathway could ameliorate VSMCs aging in vitro, evidenced by the decrease of senescence-related proteins P16, P21, P53, and GLB1 expressions. These results suggested that FP receptor is a promoter of vascular aging, by inducing cellular aging, oxidative stress, and vascular remodeling via Src and PAI-1 upregulation.

The Effect of Alcoholic Essence Satureja Sahandica Bornm L. on the Expression of Metalloproteinases 1, 2, 9 and 13 in a Model Similar to Osteoarthritis

Loss of the extracellular matrix of cartilage is one of the most important and prominent features of osteoarthritis (OA). The ECM has two important components, including the proteoglycan aggrecan and type II collagen, which are the main targets of MMPs and ADAM TSs enzymes. In this paper, the effect of ASSB Compared with dexamethasone and ibuprofen as representatives of steroidal and nonsteroidal anti-inflammatory drugs. One way to prevent irreversible joint damage in OA is to inhibit MMPs. ASSB can improve the condition of patients with osteoarthritis by reducing the expression of the MMPs 9 and 13 gene. <i>Background</i>: OA is a progressive joint disease characterized by cartilage degeneration, cartilage bone regeneration, and synovial membrane inflammation and is exacerbated with age. Given the side effects of conventional OA treatments, including nonsteroidal anti-inflammatory drugs and corticosteroids, it is important to consider new treatments for this disorder. Recently, the effect of matrix metalloproteinases on the pathogenesis of OA has been attracted attention. <i>Objective</i>: The aim of this paper was to evaluate the impact of Alcoholic Satureja Sahandica Bornem L. (ASSB) essence on inhibiting the expression of metalloproteinase gene 1, 2, 9 and 13 in bovine fibroblast-like synoviocytes stimulated by lipopolysaccharide as a model of osteoarthritis. <i>Methods</i>: Synovial cells were isolated from the articular cartilage of the radiocarpal joint of an 8-month-old Holstein cow. After determining the degree of toxicity by MTT Assay, the cells were exposed to stimulation of LPS without or in the presence of dexamethasone, ibuprofen and alcoholic Satureja Sahandica Bornem essence (ASSB). Metalloproteinase gene expression 1, 2, 9 and 13 were measured by RT-PCR and ELISA. The effect of ASSB on migration and cell invasion was investigated by Transwell chambers. <i>Results</i>: The results showed that Satureja Sahandica Bornm (SSB) essential oil significantly reduced the expression of metalloproteinases 9 and 12 in bovine fibroblast-like synovocytes stimulated by lipopolysaccharide. It also suppressed migration and invasion of these cells. However, Satureja Sahandica Bornm L. had no significant impact on the expression of MMPs 1 and 2. <i>Conclusion</i>: Based on our results, ASSB can significantly reduce the activity and inflammatory effect of MMPs 9 and 12 in OA, its potential role as a complement to NSAIDs and common corticosteroids was confirmed. However, cellular modeling does not confirm the beneficial effect of OA in patients.

Changes in the levels of MMP-9, MMP-2, and their inhibitors in the serum of dairy cows with laminitis

Laminitis is regarded as an important underlying cause of lameness disorders, yet its specific pathogenesis remains unclear. Consistent histological changes in cows with laminitis consist in the degradation of the basement membrane (BM). The breakdown of BM is accomplished by numerous proteases, especially MMP-9 and MMP-2. This study recruited 45 cows according to veterinary diagnostic criteria and divided them into three groups: subclinical laminitis cows (SCL, n = 15), chronic laminitis cows (CL, n = 15), and healthy cows (CON, n = 15). After blood samples were collected from the jugular vein, the serum was separated and frozen at –80°C. The serum samples were analyzed by gelatin zymography and reverse zymography to evaluate the activities of MMP-9, MMP-2, TIMP-1, and TIMP-2. In the SCL group, the activity of MMP-9 significantly increased (P &lt;0.01) and the activity of TIMP-1 significantly decreased (P &lt;0.05), compared with those in the CON group, while the activities of MMP-2 and TIMP-2 were not significantly different. In the CL group, the activities of MMP-9 (P &lt;0.001) and MMP-2 (P &lt;0.05) significantly increased, and the activities of TIMP-1 (P &lt;0.01) and TIMP-2 (P &lt;0.05) significantly decreased compared with those in the CON group. This is the first study to report changes in the content of MMP-2 and MMP-9 and their inhibitors, TIMP-2 and TIMP-1, in the serum of dairy cows with laminitis. These results indicate that inadequate regulation of the activities of MMPs and TIMPs in serum may play a role in the development of laminitis in dairy cows.

Matrix Metalloproteinase-2 Inhibition in Acute Ischemia-Reperfusion Heart Injury-Cardioprotective Properties of Carvedilol.

Matrix metalloproteinase 2 (MMP-2) is activated in hearts upon ischemia-reperfusion (IR) injury and cleaves sarcomeric proteins. It was shown that carvedilol and nebivolol reduced the activity of different MMPs. Hence, we hypothesized that they could reduce MMPs activation in myocytes, and therefore, protect against cardiac contractile dysfunction related with IR injury. Isolated rat hearts were subjected to either control aerobic perfusion or IR injury: 25 min of aerobic perfusion, followed by 20 min global, no-flow ischemia, and reperfusion for 30 min. The effects of carvedilol, nebivolol, or metoprolol were evaluated in hearts subjected to IR injury. Cardiac mechanical function and MMP-2 activity in the heart homogenates and coronary effluent were assessed along with troponin I content in the former. Only carvedilol improved the recovery of mechanical function at the end of reperfusion compared to IR injury hearts. IR injury induced the activation and release of MMP-2 into the coronary effluent during reperfusion. MMP-2 activity in the coronary effluent increased in the IR injury group and this was prevented by carvedilol. Troponin I levels decreased by 73% in IR hearts and this was abolished by carvedilol. Conclusions: These data suggest that the cardioprotective effect of carvedilol in myocardial IR injury may be mediated by inhibiting MMP-2 activation.

Fisetin suppresses cigarette smoke extract-induced epithelial to mesenchymal transition of airway epithelial cells through regulating COX-2/MMPs/β-catenin pathway

Cigarette smoke exposure leads to upregulation of cyclooxygenase‐2 (COX‐2), an inducible enzyme that synthesizes prostaglandin E2 (PGE2) and promotes airway inflammation. COX-2 overexpression is frequently implicated in inflammation, invasion, metastasis, and epithelial-mesenchymal transition (EMT). However, its detailed molecular mechanism in cigarette smoke induced EMT is not clear. Further, fisetin, a bioflavonoid, exhibits antioxidant and anti-inflammatory properties, but its effect in modulating COX-2-mediated inflammation and downstream sequelae remains unexplored. Therefore, we have investigated the mechanism of cigarette smoke-induced COX-2-mediated EMT in airway epithelial cells and examined the role of fisetin in controlling this aberration. MTT, trypan blue staining, gelatin zymography, Western blotting, invasion, wound healing, and tumor sphere formation assays in cigarette smoke extract (CSE) and/or fisetin treated airway epithelial cells, and in-silico molecular docking studies were performed. Results revealed that CSE exposure increased the expression and activity of COX-2, MMP-2/9, and β-catenin and also enhanced expression of EMT markers leading to higher migration and invasion potential of airway epithelial cells. A specific COX-2 inhibitor NS-398 as well as fisetin treatment reversed the expression of EMT biomarkers, reduced the activity of MMP-2/9, and blocked the migration and invasion potential induced by CSE. Further, PGE2 also increased MMPs activity, invasion, and migration potential similar to CSE, which were significantly reversed by fisetin. In-silico studies showed a high binding affinity of fisetin to key EMT associated proteins, validating its anti-EMT potential. Thus, our study firstly unearths the mechanism of CSE-induced EMT in airway epithelial cells via COX-2/MMP/β-catenin pathway, and secondly, it reveals that fisetin could significantly reverse CSE-induced EMT by inhibiting COX-2, indicating that fisetin could be an effective drug candidate for cigarette smoke-induced lung dysfunction.

STAT3-induced NCK1 elevation promotes migration of triple-negative breast cancer cells via regulating ERK1/2 signaling.

Noncatalytic region of tyrosine kinase 1 (NCK1) plays a key role in extracellular matrix degradation, which is required for the metastasis of triple-negative breast cancer (TNBC). However, the role NCK1 plays in the metastatic progression of TNBC is unknown. Based on online databases, NCK1 was found to be highly expressed in TNBC as compared to normal breast-like subjects, which was also confirmed using TNBC cells and a tissue microarray. NCK1 expression gradually decreased with increased tumor stage. High NCK1 expression displayed a poor prognosis in lymph node-positive metastatic TNBC patients, but not in lymph node-negative patients. Using transwell assays and immunoblotting, we confirmed that NCK1 overexpression promoted, while NCK1 downregulation inhibited migration capabilities, as well as the expression of matrix metalloproteinases (MMP2/9), uridylyl phosphate adenosine, and plasminogen activator inhibitor-1 in TNBC cells. Mechanistically, NCK1 upregulation mediated the activation of MMP2/9 through ERK1/2 activity. Signal transducer and activator of transcription 3 (STAT3) was positively correlated with NCK1. STAT3 could directly bind to the promoter region of NCK1 to promote its expression and was accompanied by the elevation of MMP2/9 and ERK1/2 signaling, which were partially abolished by the knockdown of NCK1 in TNBC cells. NCK1 may serve as a diagnostic and prognostic marker of metastatic TNBC. STAT3 upregulation promoted the expression of NCK1, which subsequently induced the migration and activity of MMPs in a ERK1/2 signaling-dependent manner in TNBC cells. NCK1 is a promising target for improving TNBC migration.

Astragaloside IV Attenuates Ocular Hypertension in a Mouse Model of TGFβ2 Induced Primary Open Angle Glaucoma

Elevated intraocular pressure (IOP) is a major risk factor in developing primary open angle glaucoma (POAG), which is the most common form of glaucoma. Transforming growth factor-beta 2 (TGFβ2) is a pro-fibrotic cytokine that plays an important role in POAG pathogenesis. TGFβ2 induced extracellular matrix (ECM) production, deposition and endoplasmic reticulum (ER) stress in the trabecular meshwork (TM) contribute to increased aqueous humor (AH) outflow resistance and IOP elevation. Drugs which alter the glaucomatous fibrotic changes and ER stress in the TM may be effective in reducing ocular hypertension. Astragaloside IV (AS.IV), a novel saponin isolated from the roots of Astragalus membranaceus, has demonstrated antifibrotic and ER stress lowering effects in various tissues during disease conditions. However, the effect of AS.IV on glaucomatous TM fibrosis, ER stress and ocular hypertension has not been studied. Primary human TM cells treated with AS.IV decreased TGFβ2 induced ECM (FN, Col-I) deposition and ER stress (KDEL, ATF4 and CHOP). Moreover, AS.IV treatment reduced TGFβ2 induced NF-κB activation and αSMA expression in TM cells. We found that AS.IV treatment significantly increased levels of matrix metalloproteases (MMP9 and MMP2) and MMP2 enzymatic activity, indicating that the antifibrotic effects of AS.IV are mediated via inhibition of NF-κB and activation of MMPs. AS.IV treatment also reduced ER stress in TM3 cells stably expressing mutant myocilin. Interestingly, the topical ocular AS.IV eye drops (1 mM) significantly decreased TGFβ2 induced ocular hypertension in mice, and this was associated with a decrease in FN, Col-1 (ECM), KDEL (ER stress) and αSMA in mouse TM tissues. Taken together, the results suggest that AS.IV prevents TGFβ2 induced ocular hypertension by modulating ECM deposition and ER stress in the TM.

Mechanisms of Covid-induced Heart failure

Sars-Cov-2 binds to ACE-2 receptor, resulting in excess amount of ACE-1 dependent angiotensin production. Sars-Cov-2 has four stages of symptoms, 80% if symptomatic patients have been shown to only suffer a “mild” disease course, meanwhile 20% endure stage 3, which is characterized by conditions such as ARDS, shock, and multiorgan failure. The binding of Covid to ACE receptors promotes the conversion of angiotensin 1 to angiotensin 2, constricting blood vessels, leading to thrombosis, lack of oxygen, and Ischemia. In Cardiac injury, the Covid receptor becomes represented in the heart, inducing in a pro-inflammatory increase, high cytokine concentrations, myocyte cardiac apoptosis, cardiac arrythmia, cardiac fibrotic tissue, myocarditis, and Heart failure. Cytokine storms associated with production of S1P, TNF activation of MMPs prompts myocardial depression, dilation of LV, and Heart failure. Covid-induced Sepsis corresponds with dysregulated host immune responses, increased pro-inflammatory mediators, fluid leakage, increase in cGMP, LV dysfunction, and arrythmia. Blood clots in the capillaries surrounding the kidneys generate renal impairment. Progression of renal impairment generates a state of systemic inflammation. Increased sodium content in the body results in elevated plasma blood and uncontrolled hypervolemia. Emotional and physical stresses during Sars-Cov-2 induce blood gas changes, angiotensin converting enzyme imbalance, and immune/inflammatory factors. Overactivation of the SNS induces Takotsubo syndrome. In ARDS, fluid leak in the membrane gas exchange region of the lung results in vascular remodeling. Inducing further vascular remodeling as part of the body’s response to hypoxia. The constant vascular remodeling triggers RHF.

Protective effects of GLHP from Gracilaria lemaneiformis against UVB-induced photodamage in human immortalized keratinocytes cells and BALB/c mice.

Our previous study showed that the water-soluble heteropolysaccharide extracted from Gracilaria lemaneiformis (GLHP) has excellent anti-inflammation and anti-oxidant properties. This study explored the efficacy of GLHP against skin anti-photoaging in human immortalized keratinocytes (HaCaT) cells and BALB/c mice under UVB irradiation. Cell viability, antiapoptotic, reactive oxygen species (ROS) scavenging activity, mitochondrial membrane potential, and cell wound scratch assays were conducted, as well as assessment of inflammation markers and sun protection factors. The in vitro results showed that GLHP pretreatment significantly inhibited UVB-induced apoptosis, reversed the decrease of cell viability via downregulating the expression of apoptosis-related protein caspase-3, accelerated the migration of HaCaT cells, and promoted wound healing. Notably, the protective effect of GLHP may be associated with the scavenging of ROS and the decrease of mitochondrial membrane potential. Moreover, GLHP pretreatment significantly restrained the upregulation of iNOS (UVB-induced inflammation marker), suppressed the expression of P-ERK and NF-κB, and decreased the activity of MMPs, suggesting that it exerts the therapeutic effects by inhibiting the MAPK/NF-κB signal pathway. Results obtained after conducting the in vivo assay confirmed that GLHP could reverse the UVB-induced increase of epidermal thickness in BALB/c mice. In conclusion, this study shows that GLHP can be utilized as a safer resource in the manufacture of anti-aging cosmetics because it exerts excellent anti-photoaging effects.

Avenanthramide C as a novel candidate to alleviate osteoarthritic pathogenesis

Osteoarthritis (OA) is a degenerative disorder that can result in the loss of articular cartilage. No effective treatment against OA is currently available. Thus, interest in natural health products to relieve OA symptoms is increasing. However, their qualities such as efficacy, toxicity, and mechanism are poorly understood. In this study, we determined the efficacy of avenanthramide (Avn)-C extracted from oats as a promising candidate to prevent OA progression and its mechanism of action to prevent the expression of matrix-metalloproteinases (MMPs) in OA pathogenesis. Interleukin-1 beta (IL-1β), a proinflammatory cytokine as a main causing factor of cartilage destruction, was used to induce OA-like condition of chondrocytes in vitro. Avn-C restrained IL-1β-mediated expression and activity of MMPs, such as MMP-3, -12, and -13 in mouse articular chondrocytes. Moreover, Avn-C alleviated cartilage destruction in experimental OA mouse model induced by destabilization of the medial meniscus (DMM) surgery. However, Avn-C did not affect the expression of inflammatory mediators (Ptgs2 and Nos) or anabolic factors (Col2a1, Aggrecan, and Sox9), although expression levels of these genes were upregulated or downregulated by IL-1β, respectively. The inhibition of MMP expression by Avn-C in articular chondrocytes was mediated by p38 kinase and c-Jun N-terminal kinase (JNK) signaling, but not by ERK or NF-κB. Interestingly, Avn-C added with SB203580 and SP600125 as specific inhibitors of p38 kinase and JNK, respectively, enhanced its inhibitory effect on the expression of MMPs in IL-1β treated chondrocytes. Taken together, these results suggest that Avn-C is an effective candidate to prevent OA progression and a natural health product to relieve OA pathogenesis.

Cold helium plasma jet does not stimulate collagen remodeling in a 3D human dermal substitute

Cold Atmospheric Plasma (CAP) is an emerging physical approach displaying encouraging antitumor and wound healing effects both in vitro and in vivo. In this study, we assessed the potential of direct CAP to remodel skin collagens using an original tissue-engineered human dermal substitute model rich in endogenous extracellular matrix (ECM) covered with 600 µl of culture medium and treated with CAP for 30 and 120 s. Our results indicated that Reactive Oxygen and Nitrogen Species (RONS) such as H2O2, NO3– and NO2– were produced in the medium during treatment. It appeared that in the CAP-treated dermal substitutes 1) cell viability was not altered, 2) pro-collagen I secretion was not modified over 48 h of culture after treatment, 3) global activity of matrix metalloproteinases MMPs was not modulated over 48 h after treatment, and 4) no change in hydroxyproline content was observed over 5 days after treatment. In order to confirm the efficiency of our device, we showed that the plasma-activated culture medium induced cell apoptosis and growth delay using a 3D human tumor spheroid model. In conclusion, no effect of direct CAP treatment was monitored on dermal ECM production and degradation, indicating that CAP does not stimulate collagen remodeling at the tissue scale.

Inflammatory Modulation of Polyethylene Glycol-AuNP for Regulation of the Neural Differentiation Capacity of Mesenchymal Stem Cells.

A nanocomposite composed of polyethylene glycol (PEG) incorporated with various concentrations (~17.4, ~43.5, ~174 ppm) of gold nanoparticles (Au) was created to investigate its biocompatibility and biological performance in vitro and in vivo. First, surface topography and chemical composition was determined through UV-visible spectroscopy (UV-Vis), Fourier-transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), scanning electron microscopy (SEM), free radical scavenging ability, and water contact angle measurement. Additionally, the diameters of the PEG-Au nanocomposites were also evaluated through dynamic light scattering (DLS) assay. According to the results, PEG containing 43.5 ppm of Au demonstrated superior biocompatibility and biological properties for mesenchymal stem cells (MSCs), as well as superior osteogenic differentiation, adipocyte differentiation, and, particularly, neuronal differentiation. Indeed, PEG-Au 43.5 ppm induced better cell adhesion, proliferation and migration in MSCs. The higher expression of the SDF-1α/CXCR4 axis may be associated with MMPs activation and may have also promoted the differentiation capacity of MSCs. Moreover, it also prevented MSCs from apoptosis and inhibited macrophage and platelet activation, as well as reactive oxygen species (ROS) generation. Furthermore, the anti-inflammatory, biocompatibility, and endothelialization capacity of PEG-Au was measured in a rat model. After implanting the nanocomposites into rats subcutaneously for 4 weeks, PEG-Au 43.5 ppm was able to enhance the anti-immune response through inhibiting CD86 expression (M1 polarization), while also reducing leukocyte infiltration (CD45). Moreover, PEG-Au 43.5 ppm facilitated CD31 expression and anti-fibrosis ability. Above all, the PEG-Au nanocomposite was evidenced to strengthen the differentiation of MSCs into various cells, including fat, vessel, and bone tissue and, particularly, nerve cells. This research has elucidated that PEG combined with the appropriate amount of Au nanoparticles could become a potential biomaterial able to cooperate with MSCs for tissue regeneration engineering.

Dehydrated human amniotic membrane regulates tenocyte expression and angiogenesis in vitro: Implications for a therapeutic treatment of tendinopathy.

Tendon injuries are among the most common ailments of the musculoskeletal system. Prolonged inflammation and persistent vasculature are common complications associated with poor healing. Damaged tendon, replaced with scar tissue, never completely regains the native structural or biomechanical properties. This study evaluated the effects of micronized dehydrated human amnion/chorion membrane (μdHACM) on the inflammatory environment and hypervascularity associated with tendinopathy. Stimulation of human tenocytes with interleukin‐1 beta (IL1β) induced the expression of inflammatory and catabolic markers, resulting in secretion of active MMPs and type 3 collagen that is associated with a degenerative phenotype. Treatment with μdHACM diminished the effects of IL1β, reducing the expression of inflammatory genes, proteases, and extracellular matrix components, and decreasing the presence of active MMP and type 3 collagen. Additionally, a co‐culture model was developed to evaluate the effects of μdHACM on angiogenesis associated with tendinopathy. Micronized dHACM differentially regulated angiogenesis depending upon the cellular environment in which it was placed. This phenomenon can be explained in part through the detection of both angiogenic protagonists and antagonists in μdHACM. Observations from this study identify a mechanism by which μdHACM regulates inflammatory processes and angiogenesis in vitro, two key pathways implicated in tendinopathic injuries.

HMG-CoA Reductase Inhibitor, Simvastatin Is Effective in Decreasing Degree of Myocarditis by Inhibiting Metalloproteinases Activation.

Background: Acute myocarditis often progresses to heart failure because there is no effective, etiology-targeted therapy of this disease. Simvastatin has been shown to be cardioprotective by decreasing matrix metalloproteinases’ (MMPs) activity. The study was designed to determine whether simvastatin inhibits MMPs activity, decreases the severity of inflammation and contractile dysfunction of the heart in experimental autoimmune myocarditis (EAM). Methods: Simvastatin (3 or 30 mg/kg/day) was given to experimental rats with EAM by gastric gavage for 21 days. Then transthoracic echocardiography was performed, MMPs activity and troponin I level were determined and tissue samples were assessed under a light and transmission electron microscope. Results: Hearts treated with simvastatin did not show left ventricular enlargement. As a result of EAM, there was an enhanced activation of MMP-9, which was significantly reduced in the high-dose simvastatin group compared to the low-dose group. It was accompanied by prevention of myofilaments degradation and reduction of severity of inflammation. Conclusions: The cardioprotective effects of simvastatin in the acute phase of EAM are, at least in part, due to its ability to decrease MMP-9 activity and subsequent decline in myofilaments degradation and suppression of inflammation. These effects were achieved in doses equivalent to therapeutic doses in humans.

Matrix metalloproteinse-9 expression in fetal membranes and maternal serum in cases of premature rupture of membranes: Impact of subclinical chorioamnionitis

Background: Matrix metalloproteinase-9 (MMP-9) is the most active MMPs in the fetomaternal membranes. It plays an important role in the breakdown and remodeling of these membranes. Thus, increased knowledge of its expression in cases of premature rupture of membranes (PROM) with and without chorioamnionitis (CA) is essential for developing effective treatment strategies. Patients and Methods: expression of MMP-9 in fetomaternal membranes and maternal serum were performed in 70 full-term pregnant women; 20 cases were with intact membranes (group I), 35 with PROM (group IIA) and 15 with PROM and CA (group IIB). Results: A significant difference in MMMP-9 expression was detected between the studied groups. The highest MMP-9 expression was reached in women with PROM accompanied by histological (subclinical) CA followed by women with PROM without CA, compared to the control group (with intact membranes) at the time of delivery. Immunohistochemical expression was associated with a concomitant increase in serum MMP-9. Conclusion: MMP-9 could predict women liable for premature rupture of membranes, and the determination of its serum level may be an effective tool for PROM detection.

Distinct effects of polyphenols and solvents on dentin collagen crosslinking interactions and biostability

ObjectiveTo evaluate the effects of different polyphenols and solvents on dentin collagen’s crosslinking interactions and biostabilization against MMPs and collagenase degradation. MethodsTwo polyphenols [proanthocyanidin (PA) and quercetin (QC)] with different water solubility were prepared as treatment solutions using ethanol (EtOH) or dimethyl sulfoxide (DMSO) as solvents. 6-um-thick dentin films were microtomed from dentin slabs of third molars. Following demineralization, films or slabs were subject to 60-s treatment (PA or QC) or no treatment (control) with subsequent extended-rinse with original solvent (EtOH or DMSO) or distilled water (DW). Collagen crosslinking interactions were assessed by FTIR. Biostability was assessed through endogenous MMPs activity via confocal laser scanning microscopy, and exogenous collagenase degradation via weight loss, hydroxyproline release and SEM. Finally, direct collagenase inactivation was also evaluated. Data were analyzed by three-way ANOVA and post-hoc tests (α=0.05%). ResultsDistinct effects of two polyphenols and solvents on collagen crosslinking and biostabilization were observed. Higher crosslinking and biostability efficacy occurred with PA than QC (p<0.001) that demonstrated negligible collagen interactions. With DMSO solvent, efficacy results were significantly reduced with both polyphenols (p<0.05). DMSO-rinse further weakened interactions of PA with collagen, diminishing biostability (p<0.05). Low biostability was detected with QC and DW-rinse, suggesting direct enzymatic inhibition due to physical presence in collagen. SignificanceCollagen crosslinking interactions and biostability depend on polyphenol chemical characteristics. Treatment-solution solvents may affect interactions between polyphenols and collagen, specifically, DMSO showed detrimental effects on collagen crosslinking and biostability and should be used with caution.

Triamcinolone acetonide modulates TGF‑β2‑induced angiogenic and tissue‑remodeling effects in cultured human retinal pigment epithelial cells

Transforming growth factor-β2 (TGF-β2) has been implicated in the pathogenesis of proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR), due to its ability to stimulate the overproduction of pro-angiogenic factors, such as vascular endothelial growth factor (VEGF), and remodeling of the extracellular matrix (ECM). Although intravitreal triamcinolone acetonide (TA) is clinically useful in the treatment of PVR and PDR, its molecular mechanism has yet to be fully elucidated. The present study investigated whether TA treatment altered TGF-β2-driven biological effects on the behavior of cultured human retinal pigment epithelial (RPE) cells, in order to determine which signaling pathway may be essential for the pharmacological action of TA. The R-50 human RPE cell line was treated with TA in the presence of TGF-β2, followed by analyses of cell viability and contraction using cell viability and collagen gel contraction assays. VEGF mRNA expression and protein production were measured using reverse transcription-quantitative PCR and ELISA, respectively. The phosphorylation status of signaling mediators and the protein expression of type I collagen (COL1A1), α-smooth muscle actin (α-SMA), and ECM-remodeling enzymes, including MMP-2 and MMP-9, were analyzed using western blotting. The gelatinolytic activity of MMPs was detected using gelatin zymography. TA treatment exhibited no prominent cytotoxicity but markedly antagonized TGF-β2-induced cytostatic effects on RPE cell viability and TGF-β2-enhanced contractility in collagen gels. In the context of TGF-β2-related signaling, TA significantly attenuated TGF-β2-elicited Smad2, extracellular-regulated kinase (ERK)1/2 and p38 mitogen-activated protein kinase (MAPK) phosphorylation. Moreover, TA markedly mitigated TGF-β2-induced VEGF upregulation through ablation of p38 signaling activity. TA also partially attenuated TGF-β2-elicted expression of COL1A1, α-SMA, MMP-2, and MMP-9, but only suppressed TGF-β2-induced MMP-9 gelatinolytic activity. Mechanistically, the MEK/ERK signaling pathway may have a critical role in the TGF-β2-induced upregulation of COL1A1, α-SMA and MMP-9. In conclusion, TA may be considered a useful therapeutic agent for treating TGF-β2-associated intraocular angiogenesis and tissue remodeling, the underlying mechanism of which may involve the ERK and p38 MAPK signaling pathways.

Danhong injection enhances the therapeutic effect of mannitol on hemispheric ischemic stroke by ameliorating blood-brain barrier disruption

Mannitol, a representative of hyperosmolar therapy, is indispensable for the treatment of malignant cerebral infarction, but its therapeutic effect is limited by its exacerbation of blood-brain barrier (BBB) disruption. This study was to explore whether Danhong injection (DHI), a standardized product extracted from Salvia miltiorrhiza Bunge and Carthamus tinctorius L., inhibits the destructive effect of mannitol on BBB and thus enhancing the treatment of hemispheric ischemic stroke. SD rats were subjected to pMCAO followed by intravenous bolus injections of mannitol with/without DHI intervention. Neurological deficit score, brain edema, infarct volume at 24 h after MCAO and histopathology, microvascular ultrastructure, immunohistochemistry and immunofluorescence staining of endothelial cell junctions, energy metabolism in the ischemic penumbra were assessed. Intravenous mannitol after MCAO resulted in a decrease in 24 h mortality and cerebral edema, whereas no significant benefit on neurological deficits, infarct volume and microvascular ultrastructure. Moreover, mannitol led to the loss of endothelial integrity, manifested by the decreased expression of occludin, junctional adhesion molecule-1 (JAM-1) and zonula occluden-1 (ZO-1) and the discontinuity of occludin staining around the periphery of endothelial cells. Meanwhile, after mannitol treatment, energy-dependent vimentin and F-actin, ATP content, and ATP5D expression were down-regulated, while MMP2 and MMP9 expression increased in the ischemic penumbra. All the insults after mannitol treatment were attenuated by addition of intravenous DHI. The results suggest DHI as a potential remedy to attenuate mannitol-related BBB disruption, and the potential of DHI to upregulate energy metabolism and inhibit the activity of MMPs is likely attributable to its effects observed.

Endogenous Enzymatic Activity in Dentin Treated with a Chitosan Primer.

The aim of this study was to evaluate the effect of different concentrations of chitosan polymer on dentinal enzymatic activity by means of gelatin and in situ zymography. Human dentin was frozen and ground in a miller. Dentin powder aliquots were demineralized with phosphoric acid and treated with three different concentrations of lyophilized chitosan polymer (1, 0.5 and 0.1 wt%) dissolved in distilled water. Dentin proteins were extracted from each experimental group and electrophoresed under non-reducing conditions in 10% SDS-PAGE containing fluorescein-labeled gelatin. After 48 h in the incubation buffer at 37 °C, proteolytic activity was registered under long-wave UV light scanner and quantified by using Image J software. Furthermore, additional teeth (n = 4) were prepared for the in situ zymographic analysis in unrestored as well as restored dentin pretreated with the same chitosan primers. The registered enzymatic activity was directly proportional to the chitosan concentration and higher in the restored dentin groups (p < 0.05), except for the 0.1% chitosan primer. Chitosan 0.1% only showed faint expression of enzymatic activity compared to 1% and 0.5% concentrations. Chitosan 0.1% dissolved in water can produce significant reduction in MMPs activity and could possibly contribute to bond strength preservation over time.