Increased Expression Of TIMP-1 Research Articles

Gene expression associated with vaginal bleeding in women using the 52-mg levonorgestrel hormonal intrauterine device: A prospective study.

To evaluate gene expression associated with vaginal bleeding in the 52-mg hormonal intrauterine device (IUD) users. We conducted a prospective study involving 100 women seeking to use the 52-mg hormonal IUD for contraception. We excluded women with a history or current condition of abnormal uterine bleeding and who were unable to attend a 1-year follow up. Women who expelled the device, removed it for reasons unrelated to vaginal bleeding, or were lost to follow up were discontinued. We collected endometrial biopsies immediately before IUD placement and assessed 20 selected genes using reverse transcription quantitative polymerase chain reaction. Users maintained a uterine bleeding diary for 12 months following IUD insertion. For statistical analysis, participants were categorized into groups with or without vaginal bleeding at 3 and 12 months. Women with elevated CXCL9 expression had an 8.15-fold higher likelihood of experiencing vaginal bleeding at 3 months (odds ratio [OR] 8.15, 95% confidence interval [CI] 2.24-29.61, P = 0.001). At 12 months of follow up, women with increased TIMP1 expression had a 2.74-fold higher chance of experiencing vaginal bleeding (OR 2.74, 95% CI 1.08-6.95, P = 0.033). CXCL9 ≥ 1.5 and IL17A ≥ 0.68 were associated with a higher probability of vaginal bleeding at 3 months, while TIMP1 levels ≥0.943 were linked to an increased risk of bleeding at 12 months. Users of the 52-mg hormonal IUD with elevated relative CXCL9 expression face an increased risk of vaginal bleeding at 3-month follow up, whereas those with heightened TIMP1 expression are more likely to experience vaginal bleeding at 12 months.

Sonicated Extract from the Aril of Momordica Cochinchinensis Inhibits Cell Proliferation and Migration in Aggressive Prostate Cancer Cells.

Momordica cochinchinensis or gac fruit has been reported to have several biological activities, including antioxidation, anti-inflammatory, and anticancer activities. However, the effect on cancer cell metastasis has not been extensively studied. With this aim, the extract from the aril part was selected and investigated for prostate cancer cell migration. The aril extracts were prepared as boiled extract, sonicated extract, ethanol extract, and HAE (hexane:acetone:ethanol; 2 : 1 : 1) extract, while the prostate cancer cell models were PC-3 and LNCaP cells. An MTT assay was performed to compare the antiproliferative effect between prostate cancer cells and normal Vero cells. As a result, the sonicated extract had the highest efficiency in PC-3 cells, with IC50 values of 2 mg/mL and 0.59 mg/mL for 48 and 72 h, respectively, while it had less of an effect in LNCaP cells and was not toxic to normal cells. Cell damage was further confirmed using LDH and cell cycle analysis. As a result, the sonicated extract did not cause cell damage or death and only inhibited cell proliferation. The effect on cancer metastasis was further examined by wound healing, transwell migration assays, and western blotting. The results demonstrated that the sonicated extract inhibited PC-3 cell migration and decreased MMP-9 but increased TIMP-1 expression. All these results support that gac fruit is a valuable source for further development as an anticancer agent for prostate cancer patients.

The impact of Sambucus nigra L. extract on inflammation, oxidative stress and tissue remodeling in a rat model of lipopolysaccharide-induced subacute rhinosinusitis

Rhinosinusitis is a common disorder related to inflammation of paranasal sinuses and nasal cavity mucosa. Herbal medicinescould be an option in the treatment of rhinosinusitis due to their anti-inflammatory and anti-oxidative properties. The study aims to investigate the effect of intranasal Sambucus nigra L. subsp. nigra (SN) extract against inflammation, oxidative stress, and tissue remodeling in nasal and sinus mucosa, but also in serum, lungs, and brain, in Wistar rat model of subacute sinonasal inflammation induced by local administration of lipopolysaccharides (LPS), from Escherichia Coli. The cytokines (TNF-α, IL-1β, IL-6) and oxidative stress (malondialdehyde) in nasal mucosa, blood, lungs, and brain were analyzed. In addition, a histopathological examination was performed, and NF-kB, MMP2, MMP9, TIMP1 expressions were also evaluated in nasal mucosa. Both doses of LPS increased the production of cytokines in all the investigated tissues, especially in the nasal mucosa and blood (p < 0.01 and p < 0.05), and stimulated their secretion in the lungs, and partially in the brain. Malondialdehyde increased in all the investigated tissues (p < 0.01 and p < 0.05). In parallel, upregulation of NF-kB and MMP2 expressions with downregulation of TIMP1, particularly at high dose of LPS, was observed. SN extract reduced the local inflammatory response, maintained low levels of IL-6, TNF-α, and IL-1β. In lungs, SN reduced all cytokines levels while in the brain, the protective effect was noticed only on IL-6. Additionally, SN diminished lipid peroxidation and downregulated NF-kB in animals exposed to a low dose of LPS, with increased TIMP1 expression, while in animals treated with a high dose of LPS, SN increased NF-kB, MMP2, and MMP9 levels. In conclusion, SN extract diminished the inflammatory response, reduced generation of reactive oxygen species (ROS) and, influenced MMPs expressions, suggesting the benficial effect of SN extract on tissue remodeling in subacute rhinosinusitis and on systemic inflammatory response.

Role of matrix metalloproteinase MMP-2, MMP-9 and tissue inhibitor of metalloproteinase (TIMP-1) in the clinical progression of pediatric acute lymphoblastic leukemia

ABSTRACT Background Matrix metalloproteinases (MMPs) play a crucial role in cancer progression and metastasis, however their role in pediatric Acute lymphoblastic leukemia (ALL) is still unrevealed. Methods The diagnostic, prognostic and predictive value of tissue inhibitor of metalloproteinase (TIMP-1), MMP-2, MMP-9 and CD34+CD38− cancer stem cells (CSCs) were assessed in bone marrow (BM) samples of 76 ALL children using Flow Cytometry analysis. Results There was a significant increase in TIMP-1 [1.52 (0.41–10) versus 0.91(0.6–1.12); respectively, p < 0.001], and CSCs CD34+CD38− [1 (0.03–18.6) versus 0.3 (0.01–1.1), p < 0.001] expression in ALL patients compared to controls. While there were no significant differences regarding MMP-2 and MMP-9 expression between the two groups. The sensitivity, specificity, area under curve (AUC) of MMP-2 were (80.3%, 53.3% and 0.568, p = 0.404), and of MMP-9 were (53.9%, 40% and 0.660, p = 0.053). While that of TIMP-1 were (78.9%, 100% and 0.892, p < 0.001), and that of CD34+CD38− CSCs were (78.9%, 73.3% and 0.855, p < 0.001). Increased TIMP-1 expression associated with the high-risk disease (p < 0.001). CD34+CD38− CSCs and MMP-2 overexpression associated with MRD at day-15, increased BM blast cell count at diagnosis and at day-15 ( p < 0.05). TIMP-1 overexpression is associated with shorter DFS and OS rates ( p = 0.009 and p = 0.048). Multivariate logistic regression analysis showed that both TIMP-1 [OR: 4.224, p = 0.046], and CD34+CD38− CSCs [OR: 6.873, p = 0.005] could be potential independent diagnostic factors for pediatric ALL. Conclusion TIMP-1 and CD34+CD38− CSCs could be possible useful diagnostic markers for pediatric ALL. Also, TIMP-1 is a promising prognostic marker for poor outcome of the patients.

Effect of Lapachol on the Inhibition of Matrix Metalloproteinase Related to the Invasion of Human Fibrosarcoma Cells.

Anti-cancer effect of lapachol contained in Tabebuia avellandae has been poorly understood until now. The aim of this study was to investigate the inhibitory effect of lapachol on MMPs related to cell invasion. Its action mechanism was elucidated by analyzing the activity and the expression of MMPs and the proteins involved in the signaling pathway of cell invasion. The cytotoxicity of lapachol was evaluated by MTT assay in HT1080 cells. The effects of lapachol on the expression and the activation of MMPs were analyzed by western blot, immunofluorescence staining, and gelatin zymography assays. Their gene expression was analyzed by RT-PCR, and metastasis was evaluated by cell invasion assay. Lapachol below 2 μM showed no cytotoxicity. It was observed that lapachol above 0.5 μM inhibited the activation of MMP-2 and MMP-9 stimulated by PMA. In particular, the protein and gene expression levels of MMP-2 stimulated by PMA were remarkably decreased in the presence of lapachol at 1 μM compared with the PMA treatment group. In addition, lapachol increased the expression level of TIMP-1 compared with the PMA treatment group. Moreover, lapachol decreased the expression level of p-p38 among MAPKs compared with the PMA treatment group. It was also found that the expression level of p65, a part of NF-kB, in nuclei was reduced in the presence of lapachol above 0.5 μM compared with the PMA treatment group. In addition, lapachol inhibited the invasion of human fibrosarcoma cells stimulated with VEGF. Above results suggest that lapachol could play an important role in the modulation of MMPs related to cell invasion via the increase in TIMP-1 expression as well as the inactivation of p38 through NF-kB transcription factor.

Cigarette Smoke Extract Stimulates MMP-2 Production in Nasal Fibroblasts via ROS/PI3K, Akt, and NF-κB Signaling Pathways.

Cigarette smoke exposure has been shown to be associated with chronic rhinosinusitis and tissue remodeling. The present study aimed to investigate the effects of cigarette smoke extract (CSE) on matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase (TIMP) production in nasal fibroblasts and to determine the underlying molecular mechanisms. Primary nasal fibroblasts from six patients were isolated and cultured. After the exposure of fibroblasts to CSE, the expression levels of MMP-2, MMP-9, TIMP-1, and TIMP-2 were measured by real-time PCR, ELISA, and immunofluorescence staining. The enzymatic activities of MMP-2 and MMP-9 were measured by gelatin zymography. Reactive oxygen species (ROS) production was analyzed using dichloro-dihydro-fluorescein diacetate and Amplex Red assays. PI3K/Akt phosphorylation and NF-κB activation were determined by Western blotting and luciferase assay. CSE significantly increased MMP-2 expression and inhibited TIMP-2 expression but did not affect MMP-9 and TIMP-1 expression. Furthermore, CSE significantly induced ROS production. However, treatment with ROS scavengers, specific PI3K/Akt inhibitors, NF-κB inhibitor, and glucocorticosteroids significantly decreased MMP-2 expression and increased TIMP-2 expression. Our results suggest that steroids inhibit CSE-regulated MMP-2 and TIMP-2 production and activation through the ROS/ PI3K, Akt, and NF-κB signaling pathways in nasal fibroblasts. CSE may contribute to the pathogenesis of chronic rhinosinusitis by regulating MMP-2 and TIMP-2 expression.

Melatonin Enhances the Usefulness of Ionizing Radiation: Involving the Regulation of Different Steps of the Angiogenic Process.

Radiotherapy is a part of cancer treatment. To improve its efficacy has been combined with radiosensitizers such as antiangiogenic agents. Among the mechanisms of the antitumor action of melatonin are antiangiogenic effects. Our goal was to investigate whether melatonin may modulate the sensitivity of endothelial cells (HUVECs) to ionizing radiation. Melatonin (1 mM) enhanced the inhibition induced by radiation on different steps of the angiogenic process, cell proliferation, migration, and tubular network formation. In relation with the activity and expression of enzymes implicated in estrogen synthesis, in co-cultures HUVECs/MCF-7, radiation down-regulated aromatase mRNA expression, aromatase endothelial-specific promoter I.7, sulfatase activity and expression and 17β-HSD1 activity and expression and melatonin enhanced these effects. Radiation and melatonin induced a significant decrease in VEGF, ANG-1, and ANG-2 mRNA expression. In ANG-2 and VEGF mRNA expression melatonin potentiated the inhibitory effect induced by radiation. In addition, melatonin counteracted the stimulatory effect of radiation on FGFR3, TGFα, JAG1, IGF-1, and KDR mRNA expression and reduced ANPEP expression. In relation with extracellular matrix molecules, radiation increased MMP14 mRNA expression and melatonin counteracted the stimulatory effect of radiation on MMP14 mRNA expression and increased TIMP1 expression, an angiogenesis inhibitor. Melatonin also counteracted the stimulatory effect of radiation on CXCL6, CCL2, ERK1, ERK2, and AKT1 mRNA expression and increased the inhibitory effect of radiation on NOS3 expression. In CAM assay, melatonin enhanced the reduction of the vascular area induced by radiation. Melatonin potentiated the inhibitory effect on the activation of p-AKT and p-ERK exerted by radiation. Antiangiogenic effect of melatonin could be mediated through AKT and ERK pathways, proteins involved in vascular endothelial (VE) cell growth, cell proliferation, survival, migration, and angiogenesis. In addition, radiation increased endothelial cell permeability and melatonin counteracted it by regulating the internalization of VE-cadherin. Radiation has some side effects on angiogenesis that may reduce its effectiveness against tumor growth and melatonin is able to neutralize these negative actions of radiation. Additionally, melatonin potentiated radiation-induced antiangiogenic actions on several steps of the angiogenic process and enhanced its antitumor action. Our findings point to melatonin as a useful molecule as adjuvant to radiotherapy in cancer treatment.

Effect of Sulforaphane on Bladder Compliance in a Rat Model of Partial Bladder Outlet Obstruction

Aims To investigate the effect of Nrf2 activator sulforaphane (SFN) on bladder compliance and the underlying mechanisms in a rat model of partial bladder outlet obstruction (BOO). Methods Male 8-week-old Sprague-Dawley rats were divided into three groups. BOO rats were given daily 0.5 mg/kg sulforaphane (BOO+SFN) or vehicle (BOO) intraperitoneally for 4 weeks, while sham-operated rats were treated with vehicle (Sham). Bladder compliance, histological alteration, and collagen deposition were evaluated. The expression levels of collagen I, collagen III, MMP-1, and TIMP-1 were measured by immunohistochemistry and western blotting. Results BOO led to a significant decrease in bladder compliance. The change was partially restored by SFN treatment. The expression of MMP-1 was significantly decreased accompanying with increased TIMP-1 expression in BOO rats compared with that in Sham rats, which was ameliorated by SFN treatment. Moreover, the increased collagen I/collagen III ratio in the BOO group was reversed by SFN treatment. Conclusions Sulforaphane suppressed collagen deposition by regulating the MMP-1 and TIMP-1 expression and decreasing the collagen I/III expression ratio in BOO rats and improved bladder compliance.

Role of Nampt-Sirt6 Axis in Renal Proximal Tubules in Extracellular Matrix Deposition in Diabetic Nephropathy

Nicotinamide adenine dinucleotide (NAD+) metabolism plays a critical role in kidneys. We previously reported that decreased secretion of a NAD+ precursor, nicotinamide mononucleotide (NMN), from proximal tubules (PTs) can trigger diabetic albuminuria. In the present study, we investigated the role of NMN-producing enzyme nicotinamide phosphoribosyltransferase (Nampt) in diabetic nephropathy. The expression of Nampt in PTs was downregulated in streptozotocin (STZ)-treated diabetic mice when they exhibited albuminuria. This albuminuria was ameliorated in PT-specific Nampt-overexpressing transgenic (TG) mice. PT-specific Nampt-conditional knockout (Nampt CKO) mice exhibited TBM thickening and collagen deposition, which were associated with the upregulation of the profibrogenic gene TIMP-1. Nampt CKO mice also exhibited the downregulation of sirtuins, particularly in Sirt6. PT-specific Sirt6-knockout mice exhibited enhanced fibrotic phenotype resembling that of Nampt CKO mice with increased Timp1 expression. In conclusion, the Nampt-Sirt6 axis in PTs serves as a key player in fibrogenic extracellular matrix remodeling in diabetic nephropathy.

Anti-metastatic and anti-proliferative activity of eugenol against triple negative and HER2 positive breast cancer cells

BackgroundEugenol is a natural phenolic compound and possesses anticancer and antibacterial activities. Breast cancer is a major global health problem, and most of the chemotherapeutic agents are highly toxic with long-term side effects. Therefore, this study aimed to explore the possibility of using eugenol as an anti-metastatic and anti-proliferative agent against MDA-MB-231 and SK-BR-3 breast cancer cells.MethodsBreast cancer cell lines MDA-MB-231 and SK-BR-3 were treated with eugenol and cell proliferation was measured using a real-time cell electronic sensing system. Annexin V analysis with flow cytometry was used to detect the effect of eugenol on cell death. In MDA-MB-231 and SK-BR-3 cells, metastatic potential after eugenol treatment was examined using a wound-healing assay. Real-time PCR was used to study the effect of eugenol on the expression of anti-metastatic genes such as MMP2, MMP9, and TIMP-1, and genes involved in apoptosis including Caspase3, Caspase7, and Caspase9.ResultsTreatment with 4 μM and 8 μM eugenol for 48 h significantly inhibited cell proliferation of MDA-MB-231, with an inhibition rate of 76.4%, whereas 5 μM and 10 μM of eugenol for 48 h significantly inhibited the proliferation of SK-BR-3 cells with an inhibition rate of 68.1%. Eugenol-treated cells showed significantly decreased MMP2 and MMP9 expression and an insignificant increase in TIMP1 expression in HER2 positive and triple negative breast cancer cells. Eugenol significantly increased the proportion of MDA-MB-231 and SK-BR-3 cells in late apoptosis and increased the expression of Caspase3, Caspase7, and Caspase9.ConclusionTo the best of our knowledge, this is the first study to describe the anti-metastatic effect of eugenol against MDA-MB-231 and SK-BR-3 breast cancer cell lines.

Effect of the ethanolic extract of pequi (Caryocar brasiliense) peel on acute cardiotoxicity induced by doxorubicin in Wistar rats (Rattus norvegicus albinus)

This study evaluated the activity of the ethanolic extract of pequi peel (EEPP) and immunostaining with matrix metalloproteinases 2 and 9 (MMP2 and MMP9), and tissue inhibitor of metalloproteinases 1 and 2 (TIMP1 and TIMP2) in the myocardium of rats subjected to acute cardiotoxicity using doxorubicin (DOX). Thirty Wistar rats (six groups of five animals) were used as follows: sham group (SG), water and saline; group G1, 16 mg/kg of DOX and 300 mg/kg of EEPP for 17 days; group G2, 16 mg/kg of DOX and 600 mg/kg of EEPP for 17 days; group G3, 16 mg/kg of DOX and 300 mg/kg of EEPP for 10 days; group G4, 16 mg/kg of DOX and 600 mg/kg of EEPP for 10 days; and control group (CG), 16 mg/kg of DOX. Three days after administering DOX (day 17), euthanasia was performed, and samples were collected for anatomopathological analysis. Myocyte vacuolar degeneration, cardiomyocyte disorganization and myofibrillar fragmentation, necrosis, mononuclear inflammatory infiltrate, Anitschkow cells, fibrosis, congestion, and edema were observed in the hearts of DOX recipients. In G1 and G2, EEPP attenuated myocyte vacuolar degeneration whereas in G4, EEPP attenuated cardiomyocyte disorganization. The percentage of cells immunoreactive for TIMP1 was higher in G1. It was concluded that EEPP minimizes the deleterious effects of DOX on rat myocardium. Doses of 300 and 600 mg/kg for 17 days attenuate the vacuolar degeneration of myocytes. The dose of 600 mg/kg for 10 days reduced cardiomyocyte disorganization, and the dose of 300 mg/kg for 17 days increased TIMP1 expression in the myocardium of DOX-treated rats.

UV-induced DNA methyltransferase 1 promotes hypermethylation of tissue inhibitor of metalloproteinase 2 in the human skin

BackgroundUltraviolet (UV) radiation has been reported to influence epigenetic regulation by affecting the expression of genome regulators such as DNA methyltransferase 1 (DNMT1). DNMT1 is a “gene silencer,” that is responsible for the maintenance of DNA methylation and contribution to de novo methylation. Implications of DNMT1′s involvement in the expression of UV-induced proteins have been previously reported. ObjectiveTo investigate for changes in DNA methylation-associated gene expressions by UV and to analyze the role of DNA methylation in the suppression of TIMP2 in UV-irradiated human skin. MethodsThe expression of DNA methylation-associated proteins and TIMP2 were analyzed in UV-irradiated human skin in vivo and in human dermal fibroblasts in vitro. To investigate the relationship between DNMT1 and TIMP2, we assessed the effect of DNMT1 knockdown, inhibition and overexpression on TIMP2 levels in human dermal fibroblasts. Lastly, methylation-specific PCR was used to confirm increased DNA methylation in TIMP2 promoter in response to UV. ResultsDNMT1 expression significantly increased whereas TIMP2 expression decreased in UV-irradiated human skin in vivo and in vitro. Downregulation of DNMT1 by knockdown or inhibition resulted in increased TIMP2 expression, whereas the overexpression of DNMT1 resulted in decreased TIMP2 expression. Lastly, methylation-specific PCR confirmed increased methylation on the CpG island residing in TIMP2 promoter in UV-irradiated human dermal fibroblasts. ConclusionThese findings suggest that UV-induced expression of DNMT1 may be responsible for mediating DNA hypermethylation in TIMP2, and thus, silencing its expression, in UV-exposed human skin.

Gramine inhibits angiogenesis and induces apoptosis via modulation of TGF-β signalling in 7,12 dimethylbenz[a]anthracene (DMBA) induced hamster buccal pouch carcinoma

BackgroundTransforming growth factor-β (TGF-β) and its receptors are considered as a novel target in cancer chemotherapy. Gramine, an indole alkaloid, possesses various pharmacological properties including antiproliferative and anticancer. However, the anti-angiogenic property remains unexplored. PurposeThe present study was designed to evaluate the anti-angiogenic and apoptosis induction properties of gramine through inhibiting TGF-β on DMBA induced oral squamous cell carcinoma (OSCC) in the hamster buccal pouch (HBP). MethodsThe effects of gramine on TGF-β signalling in DMBA induced carcinogenic events such as angiogenesis and apoptosis were analysed by studying the mRNA expression using RT-PCR, protein expression by western blot and histopathological analysis using haematoxylin and eosin (H & E) staining. ResultsGramine significantly inhibited phosphorylation and nuclear translocation of Smad2 and Smad4 by blocking activity of the TGFβ-RII, RI and activation of inhibitory Smad7. Gramine inhibited angiogenic markers such as MMP-2, MMP-9, HIF-1α, VEGF, and VEGF-R2 as well as increased TIMP-2 expression. Furthermore, gramine induced apoptosis in DMBA induced tumour bearing animals by up regulating the pro apoptotic proteins Bax, cytochrome C, apaf-1, caspase-9 caspase-3 and PARP. ConclusionIn this study, we clearly demonstrated that gramine treatment diminishes angiogenesis and induces apoptosis in hamster buccal pouch (HBP) carcinogenesis by modulating TGF-β signals.

The Paracrine Effect of Degenerated Disc Cells on Healthy Human Nucleus Pulposus Cells Is Mediated by MAPK and NF-κB Pathways and Can Be Reduced by TGF-β1.

Inflammation is thought to have a major role in the pathogenesis of disc degeneration. Studies have shown that nucleus pulposus cells (NPCs) respond to one or two specific cytokines by regulating cell proliferation or matrix synthesis. However, the effects of a cocktail of factors secreted by degenerated disc cells on transplanted exogenous healthy NPCs remain unknown. Concentrations of multiple cytokines in degenerated disc tissue-conditioned medium (dCM) were measured using enzyme-linked immunosorbent assay (ELISA). 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and Ki67 immunofluorescence staining were used to evaluate the proliferation of cells in dCM. The function of exogenous NPCs cultured in dCM was evaluated by examining catabolic markers (ADAMTS-4, ADAMTS-5, MMP-1, MMP-3, and MMP-13), anabolic markers (TIMP-1, TIMP-2, and TIMP-3), and the extracellular matrix protein-aggrecan (ACAN) and collagen II (COL2)-expression with real time polymerase chain reaction (RT-PCR). Mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) pathway activation was observed using Western blotting. Finally, we examined the role of transforming growth factor (TGF)-β1 in reducing dCM-mediated exogenous NPC dysfunction. Levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-1α, IL-2, IL-4, IL-6, IL-8, IL-10, IL-17, interferon-γ (IFN-γ), and prostaglandin E2 (PGE2) were higher and TGF-β1 levels were lower in dCM compared with the control medium. Treatment with dCM increased the proliferation of healthy NPCs. NPCs exhibited significantly higher expression of ADAMTS-4, ADAMTS-5, MMP-1, MMP-3, and MMP-13 and decreased TIMP-2, ACAN, and COL2 expression in the dCM group in a dose- and time-dependent manner. Treatment with dCM moderately increased TIMP-1 expression and had no effect on TIMP-3 mRNA levels. The MAPK and NF-κB pathways were implicated in dCM-mediated responses of healthy NPCs. TGF-β1 partially reversed the dCM-mediated NPC dysfunction. Increased levels of inflammatory factors and decreased TGF-β1 levels in dCM suggest an inflammatory environment in degenerated disc tissue. The catabolic effect of dCM on human healthy NPCs is mediated by MAPK and NF-κB pathways and can be reduced by TGF-β1.

Effect of the oral iron chelator deferiprone in diabetic nephropathy rats.

The aim of the present study was to investigate the effects of the iron chelator deferiprone in diabetic nephropathy (DN) rats and the mechanisms involved. Thirty-two male Wistar rats (180-220 g, 6 weeks old) were randomly divided into a control group, a DN group and two DN groups treated with either 50 or 100 mg/kg per day deferiprone. The DN group was established by feeding of a high-carbohydrate-fat diet and injection of 35 mg/kg streptozotocin into the vena caudalis. The duration of deferiprone treatment was 20 weeks. Histopathological changes were detected by hematoxylin-eosin and Masson staining, as well as transmission electron microscopy. Levels of nuclear factor (NF)-κB, monocyte chemotactic protein (MCP)-1, matrix metalloproteinase (MMP)-9, tissue-specific inhibitor of metalloproteinase (TIMP)-1, cyclo-oxygenase (COX)-2, and nitrotyrosine were determined in kidney tissues using reverse transcription-polymerase chain reaction (RT-PCR), western blotting, and immunohistochemistry. Histopathological observations showed that deferiprone treatment alleviated inflammation infiltrates and collagenous fibrosis in DN rats. Results from RT-PCR and western blotting indicated that deferiprone inhibited the expression of NF-κB, MCP-1, COX-2, and nitrotyrosine, which were overexpressed in DN rats. Immunohistochemistry showed that the mechanism of deferiprone action may involve regulation of MMP-9 and TIMP-1. Decreased MMP-9 expression and increased TIMP-1 expression in DN rats were significantly promoted and inhibited by deferiprone, respectively. Iron chelation by oral deferiprone has a renoprotective effect in DN rats by relieving oxidative stress, inflammation, and fibrosis, which is related to the cytokines NF-κB, MCP-1, MMP-9, TIMP-1, COX-2, and nitrotyrosine.

TIMP-1 expression induced by IL-32 is mediated through activation of AP-1 signal pathway

Hepatic fibrosis is a necessarily stage from the progression of chronic liver diseases to cirrhosis, even hepatocellular carcinoma (HCC). Hepatic fibrosis is characterized by the progressive accumulation of extracellular matrix (ECM). The balance between ECM production and degradation is mediated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). TIMP-1 is an important regulator in the synthesis and degradation of ECM. IL-32, a multi-function cytokine, could induce IL-1β, IL-6, IL-8 and other cytokine expressions by activating AP-1, NF-κβ, p38MAPK signal pathways. IL-32 expression is increased in liver tissues of hepatic fibrosis. However, the role of IL-32 in the pathogenesis of liver fibrosis is not thoroughly clear. Recently, it is demonstrated that TIMP-1 expression is induced by the activation of AP-1 signal pathway. So we assayed the effect of IL-32 on TIMP-1 expression by LX-2 cells (one of HSCs cell lines) in the present study. We found that IL-32 could induce TIMP-1 expression by LX-2 cells at a dose-dependent manner. IL-32 could increase TIMP-1 promoter activity and induce TIMP-1 expression by activating AP-1 signal pathway. Moreover, the increase of TIMP-1 expression could promote the migration of LX-2 cells. In conclusion, we believe that IL-32 might be involved in the pathogenesis of hepatic fibrosis by inducing TIMP-1 expression.

Effect of Nitric Oxide on the Expression of Matrix Metalloproteinase and Its Association with Migration of Cultured Trabecular Meshwork Cells

PurposeTo determine the effect of exogenous nitric oxide (NO) on the migration of trabecular meshwork (TM) cells and its association with expression of matrix metalloproteinases (MMPs).MethodsPrimary human TM cells treated with 1 or 10 µM S-nitroso-N-acetyl-penicillamine (SNAP) and examined for changes in adherence. TM cells were seeded onto transwell culture inserts, and changes in their migratory activity were quantified. Reverse transcription polymerase chain reaction was performed to determine the relative changes in mRNA expression of MMPs and tissue inhibitor of metalloproteinases (TIMPs).ResultsTreatment with SNAP did not significantly suppress TM cell adhesion or migration (p > 0.05). Treatment of TM cells with 10 µM SNAP decreased expression of MMP-2 and increased expression of membrane type MMP-1 and TIMP-2. Treatment with interleukin-1α triggered MMP-3 expression but did not exert significant effects on MMP-3 activation in response to SNAP.ConclusionsThese data suggest that NO revealed no significant effect on the migration of TM cells because NO decreased MMP-2 and increased TIMP-2 expression. Although expression of certain MMPs and TIMPs change in response to NO donors, NO may modulate trabecular outflow by changing the cellular production of extracellular matrix without having a significant effect on the migration of TM cells.

Expression and regulation of tissue inhibitors of metalloproteinases (TIMP1 and TIMP3) in goat oviduct

Tissue inhibitors of metalloproteinases (TIMPs) are associated with several reproductive processes, such as mammalian follicular growth, ovulation, CL formation, and embryonic development. However, the expression and function of TIMPs in goat oviducts remain unclear. This work aimed to identify TIMP1 and TIMP3 expression in the goat oviduct during the estrous cycle via immunohistochemistry, real-time polymerase chain reaction (PCR), and functional studies in cultured goat oviductal epithelial cells. Real-time PCR results demonstrated that TIMP1 and TIMP3 messenger RNAs were expressed in all goat oviductal regions at all stages of the estrous cycle. TIMP1 and TIMP3 proteins were also highly expressed in oviductal epithelial cells with very limited expression in other cell types. Oviductal epithelial cells were treated in vitro with various estradiol concentrations (1–100 nM) for 24 hours. The findings showed that TIMP1 expression increased up to 20 nM but then gradually decreased, whereas no significant effects existed among TIMP3 messenger RNA levels. Time-course studies indicated that estradiol significantly increased TIMP1 expression in a time-dependent manner from 8 hours to 24 hours. By contrast, TIMP3 expression was transiently induced in oviductal epithelial cells at 2 and 4 hours after estradiol treatment. Furthermore, treatment with TIMP1 functionally increased the viability of cultured oviductal epithelial cells. Overall, the results suggested that the differential regulation and function between TIMP1 and TIMP3 might be associated with their unique roles in fertilization and early embryonic development.

A flavanone from Baccharis retusa (Asteraceae) prevents elastase-induced emphysema in mice by regulating NF-κB, oxidative stress and metalloproteinases.

BackgroundPulmonary emphysema is characterized by irreversible airflow obstruction, inflammation, oxidative stress imbalance and lung remodeling, resulting in reduced lung function and a lower quality of life. Flavonoids are plant compounds with potential anti-inflammatory and antioxidant effects that have been used in folk medicine. Our aim was to determine whether treatment with sakuranetin, a flavonoid extracted from the aerial parts of Baccharis retusa, interferes with the development of lung emphysema.MethodsIntranasal saline or elastase was administered to mice; the animals were then treated with sakuranetin or vehicle 2 h later and again on days 7, 14 and 28. We evaluated lung function and the inflammatory profile in bronchoalveolar lavage fluid (BALF). The lungs were removed to evaluate alveolar enlargement, extracellular matrix fibers and the expression of MMP-9, MMP-12, TIMP-1, 8-iso-PGF-2α and p65-NF-κB in the fixed tissues as well as to evaluate cytokine levels and p65-NF-κB protein expression.ResultsIn the elastase-treated animals, sakuranetin treatment reduced the alveolar enlargement, collagen and elastic fiber deposition and the number of MMP-9- and MMP-12-positive cells but increased TIMP-1 expression. In addition, sakuranetin treatment decreased the inflammation and the levels of TNF-α, IL-1β and M-CSF in the BALF as well as the levels of NF-κB and 8-iso-PGF-2α in the lungs of the elastase-treated animals. However, this treatment did not affect the changes in lung function.ConclusionThese data emphasize the importance of oxidative stress and metalloproteinase imbalance in the development of emphysema and suggest that sakuranetin is a potent candidate that should be further investigated as an emphysema treatment. This compound may be useful for counteracting lung remodeling and oxidative stress and thus attenuating the development of emphysema.Electronic supplementary materialThe online version of this article (doi:10.1186/s12931-015-0233-3) contains supplementary material, which is available to authorized users.

Laminin α4 (LAMA4) expression promotes trophoblast cell invasion, migration, and angiogenesis, and is lowered in preeclamptic placentas

IntroductionThe laminin α4 subunit (LAMA4) has been shown to promote migration, proliferation, and survival of various cell types. This study investigated LAMA4's role in trophoblast cells during placental development. MethodsLAMA4 expression was immunohistochemically assessed in the first trimester and term human placentas. LAMA4 siRNA was applied to silence LAMA4 expression in extravillous explants and HTR8/SVneo cells. Hypoxia–reoxygenation (H/R) conditions were applied to mimic preeclampsia. LAMA4 expression and trophoblast cell invasion, migration, and tube formation (a measure of angiogenesis) were assessed in HTR8/SVneo cells. The p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 was used to study the mechanism underlying LAMA4 activity. LAMA4 promoter methylation was assessed by bisulfite-sequencing polymerase chain reaction (PCR) or methylation-specific PCR. ResultsLAMA4 levels in preeclamptic placentas were significantly lower than those in controls. LAMA4 silencing significantly inhibited extravillous explant outgrowth as well as HTR8/SVneo cell invasion and migration. H/R conditions significantly lowered LAMA4 expression. Application of either H/R conditions or LAMA4 silencing both significantly decreased HTR8/SVneo cell invasion, migration, and tube formation, decreased MMP2 and MMP9 expression, and increased TIMP2 expression. SB203580 significantly reduced LAMA4 expression. LAMA4 silencing significantly decreased p-p38, p-c-Jun N-terminal kinase (JNK), and p-extracellular signal-regulated kinase (ERK) expressions; by contrast, H/R conditions induced significant upregulation of p-p38 and p-ERK but decreased p-JNK. LAMA4 promoter methylation was not significantly altered in preeclamptic placentas compared to controls. ConclusionsLAMA4 expression is lowered in preeclamptic placentas and promotes trophoblast cell invasion, migration, and angiogenesis. H/R conditions decrease LAMA4 expression and appear to decouple the positive relationship between LAMA4 expression and p38 and ERK activation.