Downregulation Of MMP-9 Expression Research Articles

Preliminary investigation on the mechanism of baicalein regulating the effects of Nischarin on invasion and apoptosis of human breast cancer cells MCF-7 through Wnt3α/β-catenin pathway

BackgroundBreast cancer (BC) remains the leading cause of cancer-related mortality in women. Here, we investigate the anti-tumor effects of baicalein on human BC cells (MCF-7 cells) and explore if it regulates the Nischarin protein via Wnt3α/β-catenin signaling pathway. MethodsWe employed Wnt3α and DKK-1 to activate and inhibit the Wnt/β-catenin signaling pathway, respectively. We used CCK-8 cell viability, flow cytometry apoptosis, wound-healing and transwell migration/invasion assays. Further, using western blotting and real-time quantitative PCR (q-PCR) we analyzed expression levels of Nischarin, MMP-9, Wnt/β-catenin pathway (β-catenin, Axin 1), and apoptotic pathway (Bax, Bcl-2) proteins and their mRNAs. ResultsWe found that baicalein inhibits MCF-7 cell viability and promotes apoptosis (evidenced by increased Bax and decreased Bcl-2 expressions) in a concentration-dependent manner. It also inhibits TPA-induced migration and invasion, and downregulates MMP-9 expression. Baicalein reverses the increase in cell viability caused by Wnt3α-induced Wnt/β-catenin pathway activation. Conversely, baicalein counteracts the increase in apoptosis caused by DKK-1 mediated inhibition of the Wnt/β-catenin pathway. Additionally, baicalein upregulates Nischarin expression via modulating the Wnt/β-catenin pathway as indicated by the antagonistic effects of Wnt3α and DKK-1 on this effect of baicalein. ConclusionBaicalein exerts anti-tumor effects on MCF-7 cells through the Wnt3α/β-catenin signaling pathway, and promotes apoptosis and inhibits migration and invasion. The upregulation of Nischarin by baicalein further suggests a potential therapeutic target for BC treatment.

Downregulation of MMP-9 by epicatechin can improve the radiosensitivity of non-small cell lung cancer.

Radiation therapy is a crucial treatment for nonsmall cell lung cancer (NSCLC), but its effectiveness is limited by the resistance of tumor cells to radiation. This study aimed to evaluate the effect of epicatechin (EC) on radiosensitivity in NSCLC and to determine its relationships with matrix metalloproteinase (MMP)-9. MMP-9 expression was detected by Western blotting, and the expression of the DNA damage marker protein was detected by immunofluorescence. Cell viability was assessed using the CCK-8 assay, and cell proliferation was evaluated using the clonogenesis assay. Flow cytometry was used to determine the cell apoptosis, whereas cell migration and invasion were detected using the transwell assays. The cells were treated with ionizing radiation (IR) and EC to verify the sensitizing effect of EC on radiation therapy. MMP-9 expression was elevated in the NSCLC cells and tissues. DNA damage and cell apoptosis were increased, whereas cell vigor, proliferation, migration, and invasion were significantly decreased after IR. MMP-9 knockdown strengthened the impact of IR on the biological behaviors of the cells. EC + IR had the best effect on promoting DNA damage and the biological behaviors of the NSCLC cells; alternatively, the overexpression of MMP-9 weakened the role of EC. This study shows that EC can downregulate MMP-9 expression, promote DNA damage, reduce cell viability, proliferation, migration, and invasion, and facilitate cell apoptosis, thus, showing potential as a radiosensitizer for NSCLC.

GATA4 regulates the transcription of MMP9 to suppress the invasion and migration of breast cancer cells via HDAC1-mediated p65 deacetylation

GATA-binding protein 4 (GATA4) is recognized for its significant roles in embryogenesis and various cancers. Through bioinformatics and clinical data, it appears that GATA4 plays a role in breast cancer development. Yet, the specific roles and mechanisms of GATA4 in breast cancer progression remain elusive. In this study, we identify GATA4 as a tumor suppressor in the invasion and migration of breast cancer. Functionally, GATA4 significantly reduces the transcription of MMP9. On a mechanistic level, GATA4 diminishes MMP9 transcription by interacting with p65 at the NF-κB binding site on the MMP9 promoter. Additionally, GATA4 promotes the recruitment of HDAC1, amplifying the bond between p65 and HDAC1. This leads to decreased acetylation of p65, thus inhibiting p65’s transcriptional activity on the MMP9 promoter. Moreover, GATA4 hampers the metastasis of breast cancer in vivo mouse model. In summary, our research unveils a novel mechanism wherein GATA4 curtails breast cancer cell metastasis by downregulating MMP9 expression, suggesting a potential therapeutic avenue for breast cancer metastasis.

Network pharmacology analysis and experimental validation to explore the mechanism of kaempferol in the treatment of osteoporosis

Osteoporosis (OP) is a prevalent global disease characterized by bone mass loss and microstructural destruction, resulting in increased bone fragility and fracture susceptibility. Our study aims to investigate the potential of kaempferol in preventing and treating OP through a combination of network pharmacology and molecular experiments. Kaempferol and OP-related targets were retrieved from the public database. A protein–protein interaction (PPI) network of common targets was constructed using the STRING database and visualized with Cytoscape 3.9.1 software. Enrichment analyses for GO and KEGG of potential therapeutic targets were conducted using the Hiplot platform. Molecular docking was performed using Molecular operating environment (MOE) software, and cell experiments were conducted to validate the mechanism of kaempferol in treating OP. Network pharmacology analysis identified 54 overlapping targets between kaempferol and OP, with 10 core targets identified. The primarily enriched pathways included atherosclerosis-related signaling pathways, the AGE/RAGE signaling pathway, and the TNF signaling pathway. Molecular docking results indicated stable binding of kaempferol and two target proteins, AKT1 and MMP9. In vitro cell experiments demonstrated significant upregulation of AKT1 expression in MC3T3-E1 cells (p < 0.001) with kaempferol treatment, along with downregulation of MMP9 expression (p < 0.05) compared to the control group. This study predicted the core targets and pathways of kaempferol in OP treatment using network pharmacology, and validated these findings through in vitro experiments, suggesting a promising avenue for future clinical treatment of OP.

Multifunctional photothermally responsive hydrogel as an effective whole-process management platform to accelerate chronic diabetic wound healing

The management of chronic diabetic wounds is a complex issue that requires wound repair, regulation of inflammatory levels, and intervention to prevent bacterial infection. To address this issue, we developed a multifunctional photothermally responsive hydrogel (PAG-CuS) as an effective platform for managing the entire wound-healing process, including promoting wound healing, providing anti-inflammatory therapy, and performing photothermal sterilization. Constructed through copolymerization of acrylic acid (AA), methacrylic anhydride-modified gelatin (GelMA), and lipoic acid sodium (LAS) coated copper sulfide nanoparticles (CuS@LAS), PAG-CuS possessed a porous three-dimensional structure that promoted cell adhesion and had a substantial water-holding capacity. Additionally, the internal CuS@LAS not only conferred photothermal antibacterial properties to the hydrogel but also served as physical cross-linking agents, thus enhancing its mechanical strength. Under the NIR-induced photothermal effect, the porous hydrogel liberates CuS@LAS, and later CuS@LAS expels LAS via micelle deassembly to eliminate intracellular ROS. This results in the down-regulation of MMP-9 expression, promoting ECM production and facilitating wound healing. Meanwhile, the release of Cu2+ from PAG-CuS could enhance CD31 expression in endothelial cells, promoting microvessel formation, which is crucial for wound healing. In the diabetic wound model of GK rats, the PAG-CuS hydrogel reduced ROS levels, increased microvessel count, improved epithelialization, and enhanced wound healing. Therefore, this versatile photothermal hydrogel has the potential to be applied in sterilization, scavenging free radicals, and promoting angiogenesis, making it an effective and comprehensive solution to manage the challenges of diabetic wounds. Statement of significanceAssessment of functional recovery and timely adjustment of treatment strategy is critical in the management of chronic diabetic wounds. In this work, we prepared PAG-CuS composite hydrogels by integrating in situ reduction, chemical crosslinking, and nanoenhancement techniques. The near-infrared light-induced photothermal effect of PAG-CuS gel rapidly kills bacteria at the lesion site, and the generated heat simultaneously promotes the multilevel release of LAS from the gel, which could regulate the levels of ROS and MMP-9 to promote extracellular matrix formation. In addition, the Cu2+ released from the gel can promote the formation of blood vessels to improve blood oxygenation. Therefore, this project proposes a synergistic solution to realize the whole process of management to accelerate chronic diabetic wound healing.

PF-04449913 Inhibits Proliferation and Metastasis of Colorectal Cancer Cells by Down-regulating MMP9 Expression through the ERK/p65 Pathway.

Colorectal cancer remains a life-threatening malignancy with increasing morbidity and mortality worldwide. Therefore, new and effective anti-colorectal cancer therapeutics are urgently needed. In this study, we have studied the anti-tumor properties and potential mechanisms of PF-04449913. Colorectal cancer cell viability was reduced by PF-04449913 in a dose-dependent manner. The migration and invasion ability of malignant colon cells were attenuated by the drug, as demonstrated by the Transwell test. Moreover, PF-04449913 repressed the phosphorylation levels of ERK and other proteins, and the expression levels of MMP9. The anti-tumor effects of the drug in vivo were demonstrated in BALB/c-nude mice models, and PF-04449913 inhibited the malignant phenotype of colorectal cancer cells, including reduction of tumor size and promotion of apoptosis. At the molecular level, PF-04449913 induced a significant decrease in ERK and p65 protein phosphorylation levels and inhibited MMP9 protein expression. Both in vivo and in vitro results showed PF-04449913 to demonstrate antitumor effects, which have been proposed to be mediated through blockade of the ERK/p65 signaling pathway, and subsequent repression of MMP9 expression. Our study provides a new perspective on the potential clinical application of PF-04449913 in the treatment of colorectal cancer.

Simultaneous suppression of miR-21 and restoration of miR-145 in gastric cancer cells; a promising strategy for inhibition of cell proliferation and migration.

Gastric cancer (GC) is the third leading cause of cancer-related death worldwide. microRNAs are a group of regulatory non-coding RNAs that are involved in GC progression. miR-145 as a tumor suppressor and miR-21 as an oncomiR were shown to be dysregulated in many cancers including GC. This research aimed to enhance the expression of miR-145 while reducing the expression of miR-21 and examine their impact on the proliferation, apoptosis, and migration of GC cells. KATO III cells with high expression levels of miR-21-5p and low expression of miR-145-5p were selected. These cells were then transfected with either miR-145-5p mimics or anti-miR-21-5p, alone or in combination. Afterward, the cell survival rate was determined using the MTT assay, while apoptosis induction was investigated through V-FITC/PI and DAPI staining. Additionally, cell migration was examined using the wound healing assay, and cell cycle progression was analyzed through flow cytometry. Furthermore, gene expression levels were quantified utilizing the qRT-PCR technique. The study's findings indicated that the co-replacement of miR-145-5p and anti-miR-21-5p led to a decrease in cell viability and the induction of apoptosis in GC cells. This was achieved via modulating the expression of Bax and Bcl-2, major cell survival regulators. Additionally, the combination therapy significantly increased sub-G1 cell cycle arrest and reduced cell migration by downregulating MMP-9 expression as an epithelial-mesenchymal transition marker. This study provides evidence for the therapeutic possibility of the combination of miR-145-5p and anti-miR-21-5p and also suggests that they could inhibit cell proliferation by modulating the PTEN/AKT1 signaling pathway. Our research revealed that utilizing miR-145-5p and anti-miR-21-5p together could be a promising therapeutic approach for treating GC.

#3839 THERAPEUTIC TARGETS OF ARSENIC TRIOXIDE IN LUPUS NEPHRITIS – DATA FROM BIOINFORMATICS ANALYSIS AND IN VITRO STUDIES

Abstract Background and Aims Patients with systemic lupus erythematosus (SLE) are at an increased risk of developing lupus nephritis (LN), which results in high morbidity and mortality. There is preliminary data showing the efficacy of low dose of Arsenic Trioxide (ATO) in reducing renal flares amongst LN patients, but the underlying mechanism for such benefit remains unclear. Method We performed bioinformatic analysis and network pharmacology to elucidate the potential targets from differentially expressed genes (DEGs) of human SLE and LN peripheral blood mononuclear cells (PBMCs) datasets and pharmacological databases. The relationship between the potential targets and immune cells was further analyzed. Results Twelve intersection genes of DEGs in SLE, predicting targets of ATO and immune related genes were identified. KEGG pathway analysis suggested that the mechanisms of ATO in SLE were associated with IL-17 signalling pathway (p = 1.67E-18), TNF signalling pathway (p = 5.77E-11) and NOD-like receptor signalling pathway (p = 1.93E-09). Ten types of immune cells showed significant difference with intersection genes expression in SLE using ssGSEA approach. MMP9 was the most significantly associated with immune cells and showed positive correlations with macrophages and neutrophils. A significant TF-MMP9-miRNA regulatory network was constructed using cytoscape software. Pilot data from our in vitro studies suggested that ATO might downregulate MMP9 expression in PBMCs obtained from LN patients during disease remission (n = 3). Conclusion ATO interacts with immune cells in LN patients via different inflammatory pathways and may downregulate MMP9 expression in PBMCs.

378 RAGE Ablation Attenuates Glioma Progression and Enhances Tumor Immune Responses by Suppressing Galectin-3 Expression

INTRODUCTION: Malignant gliomas consist of heterogenous cellular components that have adopted multiple overlapping escape mechanisms that overcome both targeted and immune-based therapies. The receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily that is activated by diverse proinflammatory ligands present in the tumor microenvironment (TME). Activation of RAGE by its ligands stimulates multiple signaling pathways that are important in tumor growth, invasion, and immune escape. METHODS: We evaluated genetic ablation of RAGE on the tumorigenicity of two syngeneic murine glioma models. RAGE expression was inhibited in the GL261 and K-Luc gliomas by shRNA and CRSPR/Cas9 techniques prior to intracranial implantation. Tumor growth, invasion, and inflammatory responses were examined by immunohistochemistry, qPCR, ELISA, western blotting, NanoString, flow cytometry, and in vivo survival analysis. RESULTS: Intracellular RAGE ablation abrogated glioma growth and invasion by suppressing AKT and ERK1/2 activities and by downregulating MMP9 expression. Interestingly, RAGE inhibition also enhanced tumor inflammatory responses by downregulating the expression of galectin-3, rendered the immunotherapy resistant K-Luc gliomas responsive to immune checkpoint blockade therapy, and significantly increased survival. CONCLUSIONS: This study is the first to link RAGE expression with galactin-3 as an immune modulator in glioblastoma and revealed that the effect of glioma RAGE expression on TME was mediated through downregulation of Gal-3 and subsequent polarization of Tumor-associated macrophages. The therapeutic significance of RAGE downregulation was demonstrated by sensitizing a highly invasive and immune-resistant glioma model to immune checkpoint blockade and strongly supports the development of RAGE ablation as a complementary treatment strategy for malignant gliomas.

Indirect bilirubin impairs invasion of osteosarcoma cells via inhibiting the PI3K/AKT/MMP-2 signaling pathway by suppressing intracellular ROS

BackgroundOsteosarcoma is most prevalently found primary malignant bone tumors, with primary metastatic patients accounting for approximately 25% of all osteosarcoma patients, yet their 5-year OS remains below 30%. Bilirubin plays a key role in oxidative stress-associated events, including malignancies, making the regulation of its serum levels a potential anti-tumor strategy. Herein, we investigated the association of osteosarcoma prognosis with serum levels of TBIL, IBIL and DBIL, and further explored the mechanisms by which bilirubin affects tumor invasion and migration. MethodsROC curve was plotted to assess survival conditions based on the determined optimal cut-off values and the AUC. Then, Kaplan-Meier curves, along with Cox proportional hazards model, was applied for survival analysis. Inhibitory function of IBIL on the malignant properties of osteosarcoma cells was examined using the qRT-PCR, transwell assays, western blotting, and flow cytometry. ResultsWe found that, versus osteosarcoma patients with pre-operative higher IBIL (>8.9 μmol/L), those with low IBIL (≤8.9 μmol/L) had shorter OS and PFS. As indicated by the Cox proportional hazards model, pre-operative IBIL functioned as an independent prognostic factor for OS and PFS in total and gender-stratified osteosarcoma patients (P < 0.05 for all). In vitro experiments further confirmed that IBIL inhibits PI3K/AKT phosphorylation and downregulates MMP-2 expression via reducing intracellular ROS, thereby decreasing the invasion of osteosarcoma cells. ConclusionsIBIL may serve as an independent prognostic predictor for osteosarcoma patients. IBIL impairs invasion of osteosarcoma cells through repressing the PI3K/AKT/MMP-2 pathway by suppressing intracellular ROS, thus inhibiting its metastatic potential.

Pioglitazone inhibits oxidative stress, MMP-mediated inflammation and vascular dysfunction in high glucose-induced human saphenous vein grafts

AimsThe aim of this study was to investigate the effects of pioglitazone on reactive oxygen species (ROS), expressions/activities of MMPs and TIMP-2, and VSMC proliferation and vascular reactivity in high glucose (HG)-induced human saphenous vein (HSV) grafts. MethodsHSV grafts (n = 10) obtained from patients undergoing CABG were incubated with 30 mM glucose and/or 10 μM pioglitazone or 0.1 % DMSO for 24 h after endothelium removal. ROS levels were examined by chemiluminescence assay, MMP-2,-9,-14, TIMP-2, and α-SMA expression/activity was determined by gelatine zymography/immunohistochemistry. Vascular reactivity to potassium chloride, noradrenaline, serotonin, prostaglandin F2α and papaverine was assessed in HSVs. ResultsHG induced superoxide anion (SA) (123 %) and other ROS levels (159 %), up-regulated MMP-2 expression (180 %)/activity (79 %), MMP-14 expression (24 %) and MMP-9 activity while down-regulating TIMP-2 expression (27 %). HG elevated total MMP-2/TIMP-2 ratio (483 %) and MMP-14/TIMP-2 ratio (78 %). However, HG plus pioglitazone inhibited SA (30 %) and other ROS levels (29 %), down-regulated MMP-2 expression (76 %)/activity (83 %), MMP-14 expression (38 %) and MMP-9 activity, while reversing TIMP-2 expression (44 %). HG plus pioglitazone decreased total MMP-2/TIMP-2 ratio (91 %) and MMP-14/TIMP-2 ratio (59 %). HG impaired contractions to all agents but pioglitazone improved them. ConclusionsPioglitazone may contribute to the prevention of restenosis and maintaining vascular function in HSV grafts of DM patients undergoing CABG.

RAGE ablation attenuates glioma progression and enhances tumor immune responses by suppressing galectin-3 expression.

Malignant gliomas consist of heterogeneous cellular components that have adopted multiple overlapping escape mechanisms that overcome both targeted and immune-based therapies. The receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily that is activated by diverse proinflammatory ligands present in the tumor microenvironment. Activation of RAGE by its ligands stimulates multiple signaling pathways that are important in tumor growth and invasion. However, treatment strategies that only target the interaction of RAGE with its ligands are ineffective as cancer therapies due to the abundance and diversity of exogenous RAGE ligands in gliomas. As an alternative approach to RAGE ligand inhibition, we evaluated the genetic ablation of RAGE on the tumorigenicity of 2 syngeneic murine glioma models. RAGE expression was inhibited in the GL261 and K-Luc gliomas by shRNA and CRSPR/Cas9 techniques prior to intracranial implantation. Tumor growth, invasion, and inflammatory responses were examined by histology, survival, Nanostring, and flow cytometry. Intracellular RAGE ablation abrogated glioma growth and invasion by suppressing AKT and ERK1/2 activities and by downregulating MMP9 expression. Interestingly, RAGE inhibition in both glioma models enhanced tumor inflammatory responses by downregulating the expression of galectin-3 and potentiated immunotherapy responses to immune checkpoint blockade. We demonstrated that intracellular RAGE ablation suppresses multiple cellular pathways that are important in glioma progression, invasion, and immune escape. These findings strongly support the development of RAGE ablation as a treatment strategy for malignant gliomas.

IMMU-08. RAGE ABLATION ATTENUATES GLIOMA PROGRESSION AND ENHANCES TUMOR IMMUNE RESPONSES BY SUPPRESSING GALECTIN-3 EXPRESSION

Abstract INTRODUCTION Malignant gliomas consist of heterogenous cellular components that have adopted multiple overlapping escape mechanisms that overcome both targeted and immune-based therapies. The receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily that is activated by diverse proinflammatory ligands present in the tumor microenvironment (TME). Activation of RAGE by its ligands stimulates multiple signaling pathways that are important in tumor growth, invasion, and immune escape. OBJECTIVE We aimed to demonstrate that blockade of RAGE expression may tackle resistance pathways by inhibiting GBM progression, invasion, and immune evasion. METHODS We evaluated genetic ablation of RAGE on the tumorigenicity of two syngeneic murine glioma models. RAGE expression was inhibited in the GL261 and K-Luc gliomas by shRNA and CRSPR/Cas9 techniques prior to intracranial implantation. Tumor growth, invasion, and inflammatory responses were examined by immunohistochemistry, qPCR, ELISA, western blotting, NanoString, flow cytometry, and in vivo survival analysis. RESULTS Intracellular RAGE ablation abrogated glioma growth and invasion by suppressing AKT and ERK1/2 activities and by downregulating MMP9 expression. Interestingly, RAGE inhibition also enhanced tumor inflammatory responses by downregulating the expression of galectin-3, rendered the immunotherapy resistant K-Luc gliomas responsive to immune checkpoint blockade therapy, and significantly increased survival. CONCLUSION This study is the first to link RAGE expression with galactin-3 as an immune modulator in glioblastoma and revealed that the effect of glioma RAGE expression on TME was mediated through downregulation of Gal-3 and subsequent polarization of Tumor-associated macrophages. The therapeutic significance of RAGE downregulation was demonstrated by sensitizing a highly invasive and immune-resistant glioma model to immune checkpoint blockade and strongly supports the development of RAGE ablation as a complementary treatment strategy for malignant gliomas.

PRSS2 overexpression relates to poor prognosis and promotes proliferation, migration and invasion in gastric cancer

Serine protease 2 (PRSS2) plays a pivotal role in tum or pathogenesis as a serine protease. In this paper, we investigated the expression and role of PRSS2 in gastric cancer (GC). Quantitative real-time polymerase chain reaction (qRT-PCR), western blotting and immunohistochemistry (IHC) were performed to detect PRSS2 expression in GC tissues. The correlations between PRSS2 and clinicopathological variables and prognosis were analyzed. The effects of PRSS2 on gastric cancer in vitro and in vivo were detected by Cell Counting Kit-8 (CCK-8), colony formation assay, migration and invasion test, and nude mouse experiment. We observed that the PRSS2 mRNA and protein levels were upregulated in GC tissues, and PRSS2 expression was associated with GC N stage. Patients with high PRSS2 expression have a poor prognosis. Finally, the regulatory effect of PRSS2 on the biological behaviors of GC cells was assessed in GC cells transfected with lentiviral interference fragments of PRSS2. The results suggested that PRSS2 knockdown decreased the proliferation, migration and invasion of GC cells and downregulated MMP-9 expression. In summary, PRSS2 overexpression is associated with poor postoperative prognosis in GC patients and can be used as a potential biomarker to assess the prognosis of GC.

Nanog suppression enhanced the chemosensitivity of human non-small-cell lung cancer cells to Cisplatin and inhibited cell migration

Lung cancer is the leading cause of cancer-associated death in the world. As one of the leading transcription factors in controlling stemness features, Nanog was shown to promote cancer progression, metastasis, and drug resistance. Considering that, this research was conducted to evaluate the effect of Nanog suppression using specific siRNA on the chemosensitivity of lung cancer cells to Cisplatin through inhibition of cell proliferation, migration, and stemness as well as apoptosis induction. Then, A549 lung cancer cells were transfected with Nanog siRNA and treated with Cisplatin individually or combined. Subsequently, to investigate cell proliferation and apoptosis induction, MTT assay and Annexin V/PI staining were performed, respectively. Also, colony formation assay was carried out to evaluate cell stemness features, and migration ability of A549 cells was followed using a wound-healing assay. Gene expression was quantified via qRT-PCR. The obtained results illustrated that siRNA-mediated Nanog suppression remarkably increased the chemosensitivity of A549 cells to Cisplatin through apoptosis induction. Consistently, Nanog suppression combined with Cisplatin led to upregulation of Caspase-3 apoptotic gene and Bax/Bcl-2 ratio. Besides, Nanog knockdown, individually or combined with Cisplatin, prevented colony formation ability of A549 cells by downregulating Sox2 and CD44 genes. It was also indicated that the combination therapy remarkably downregulated MMP9 expression and subsequently suppressed A549 cell migration. A significant reduction was also observed in c-Myc and PD-L1 gene expression levels. In conclusion, the findings of the current study demonstrated that silencing Nanog combined with Cisplatin could be a potent treatment approach for lung cancer patients.

Lupeol synergizes with doxorubicin to induce anti-proliferative and apoptotic effects on breast cancer cells.

Anti-cancer and anti-migration effects of lupeol as a biological pentacyclic triterpenoid were investigated individually and in combination with Doxorubicin (DOX) on MCF-7 and MDA-MB-231 breast cancer cells and human foreskin fibroblasts. To uncover the anticancer effect of lupeol and the impact of its combination with DOX, cell viability and scratch assays and dual acridine-orange apoptotic staining were performed. Moreover, the expression of proapoptotic caspase-3 and metastasis-related MMP-9 at the mRNA and protein levels was analyzed using qPCR and western blot techniques. Lupeol synergistically increased the anti-proliferative effect of DOX with IC50 values of 42.55, 62.24 and 65.9μM on MCF-7, MDA-MB-231 and HFF cells, respectively. Lupeol reduced the cell migration and lowered the DOX-induced cell migration, significantly (p < 0.05). The number of apoptotic cells elevated significantly (p < 0.05) when cancer cells were treated with the combination of lupeol and DOX. Lupeol individually and in combination with DOX up-regulated the expression of caspase-3. The proposed combination therapy synergized (3-4 fold) the down-regulation of MMP-9 expression in MCF-7 and MDA-MB-231 cells. Our results indicate that lupeol could be considered as an anticancer agent and anticancer adjuvant in breast cancer-therapy. The anticancer properties of lupeol attribute to its antiproliferative, antimigrative and apoptotic effects.

A polysaccharide from Lycium barbarum L.: Structure and protective effects against oxidative stress and high-glucose-induced apoptosis in ARPE-19 cells

Lycium barbarum polysaccharides (LBPs) are beneficial for vision; however, relevant research has mainly focused on entire crude polysaccharides, with the basis and exact structure of the polysaccharide rarely explored. In this study, LICP009-3F-2a, a novel polysaccharide from Lycium barbarum L., was separated and then purified using anion-exchange and size-exclusion chromatography. Structural characteristics were investigated using chemical and spectroscopic methods, which revealed that LICP009-3F-2a has an Mw of 13720 Da and is an acidic heteropolysaccharide composed of rhamnose (39.1%), arabinose (7.4%), galactose (22.5%), glucose (8.3%), galacturonic acid (13.7%), and glucuronic acid (4.0%). Linkage and NMR data revealed that LICP009-3F-2a has the following backbone: →2)-α-L-Rha-(1 → 2,4)-α-L-Rha-(1 → 4)-α-D-GalAp-(1 → 3,6)-β-D-Galp-(1 → 3,6)-β-D-Galp-(1 → 6)-β-D-Galp-(1→, with three main branches, including: α-L-Araf-(1 → 5)-α-L-Araf-(1 → 6)-β-D-Glcp-(1 → 2,4)-α-L-Rha-(1→, β-D-Glcp-(1 → 4)-β-D-Glcp-(1 → 3,6)-β-D-Galp-(1→, and β-D-Galp-(1 → 3)-β-D-Galp-(1 → 3,6)-β-D-Galp-(1 → . Differential scanning colorimetry and thermogravimetric analysis showed that LICP009-3F-2a is thermally stable, while X-ray diffractometry showed that LICP009-3F-2a has a semi-crystalline structure. In addition, LICP009-3F-2a protects ARPE-19 cells from H2O2-induced oxidative damage by regulating the expression of antioxidant SOD1 and CAT enzymes and down-regulating MMP2 expression. Moreover, LICP009-3F-2a promotes the proliferation of ARPE-19 cells in a concentration-dependent manner, and protects ARPE-19 cells from hyperglycemia by inhibiting apoptosis.

The β3/5 Integrin-MMP9 Axis Regulates Pulmonary Inflammatory Response and Endothelial Leakage in Acute Lung Injury.

BackgroundAcute lung injury (ALI) is a severe respiratory disease with high rates of morbidity and mortality. Many mediators regarding endogenous or exogenous are involved in the pathophysiology of ALI. Here, we have uncovered the involvement of integrins and matrix metalloproteinases, as critical determinants of excessive inflammation and endothelial permeability, in the regulation of ALI.MethodsInflammatory cytokines were measured by quantitative real-time PCR for mRNA levels and ELISA for secretion levels. Endothelial permeability assay was detected by the passage of rhodamine B isothiocyanate-dextran. Mice lung permeability was assayed by Evans blue albumin (EBA). Western blot was used for protein level measurements. The intracellular reactive oxygen species (ROS) were evaluated using a cell-permeable probe, DCFH-DA. Intratracheal injection of lipopolysaccharide (LPS) into mice was conducted to establish the lung injury model.ResultsExogenous MMP-9 significantly aggravated the inflammatory response and permeability in mouse pulmonary microvascular endothelial cells (PMVECs) treated by LPS, whereas knockdown of MMP-9 exhibited the opposite phenotypes. Knockdown of integrin β3 or β5 in LPS-treated PMVECs significantly downregulated MMP-9 expression and decreased inflammatory response and permeability in the presence or absence of exogenous MMP-9. Additionally, the interaction of MMP-9 and integrin β5 was impaired by a ROS scavenger, which further decreased the pro-inflammatory cytokines production and endothelial leakage in PMVECs subjected to co-treatment (LPS with exogenous MMP-9). In vivo studies, exogenous MMP-9 treatment or knockdown β3 integrin significantly decreased survival in ALI mice. Notably, knockdown of β5 integrin alone had no remarkable effect on survival, but which combined with anti-MMP-9 treatment significantly improved the survival by ameliorating excessive lung inflammation and permeability in ALI mice.ConclusionThese findings support the β3/5 integrin-MMP-9 axis as an endogenous signal that could play a pivotal role in regulating inflammatory response and alveolar-capillary permeability in ALI.

Role of NADPH Oxidase-Induced Hypoxia-Induced Factor-1α Increase in Blood-Brain Barrier Disruption after 2-Hour Focal Ischemic Stroke in Rat.

We recently showed that inhibition of hypoxia-induced factor-1α (HIF-1α) decreased acute ischemic stroke-induced blood-brain barrier (BBB) damage. However, factors that induce the upregulation of HIF-1α expression remain unclear. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase played a critical role in reperfusion-induced BBB damage after stroke. However, the role of NADPH oxidase in BBB injury during the acute ischemia stage remains unclear. This study is aimed at investigating the role of NADPH oxidase in BBB injury and the expression of HIF-1α after acute ischemic stroke. A sutured middle cerebral artery occlusion (MCAO) model was used to mimic ischemic stroke in rats. Our results show that the inhibition of NADPH oxidase by apocynin can significantly reduce the BBB damage caused by 2 h ischemic stroke accompanied by reducing the degradation of tight junction protein occludin. In addition, treatment with apocynin significantly decreased the upregulation of HIF-1α induced by 2 h MCAO. More importantly, apocynin could also inhibit the MMP-2 upregulation. Of note, HIF-1α was not colocalized with a bigger blood vessel. Taken together, our results showed that inhibition of NADPH oxidase-mediated HIF-1α upregulation reduced BBB damage accompanied by downregulating MMP-2 expression and occludin degradation after 2 h ischemia stroke. These results explored the mechanism of BBB damage after acute ischemic stroke and may help reduce the associated cerebral hemorrhage transformation after thrombolysis and endovascular treatment after ischemic stroke.

Targeting NF-κB mediated cell signaling pathway and inflammatory mediators by 1,2-diazole in A549 cells in vitro

Lung cancer is the leading cause of cancer deaths globally. The objective of this study was to investigate the effect of 1,2-diazole (pyrazole) as an anti-cancer drug on human non-small cell lung carcinoma A549 cells. We attempt to examine the expression level of pro-inflammatory proteins such as TNF-α, NF-κB-p65, MMP-2 and E-Cadherin which are commonly associated with an inflammatory response in epithelial cells and apoptosis in A549 cells. The LPS-induced cytokines and inflammatory mediators include TNF-α, IL-6, iNOS and COX-2 levels in A549 cells and the effect of pyrazole was studied. The present study reveals that, pyrazole inhibits A549 cells by suppressing TNF-α induced MMP-2 expression, thereby inhibiting the nuclear translocation of NF-κB-p65. Pyrazole significantly up-regulate the E-cadherin level and down-regulated MMP-2 expression that could probably preventing A549 cancer cells to invade. The study further substantiated the anti-cancer property of pyrazole by regulating the above mentioned level of LPS-induced cytokines and inflammatory mediators. The observations of the present study open a possibility for the development of an effective therapeutic agent that targets inflammatory and signaling pathway mediators to challenge human non-small cell lung carcinoma.