The Deep Proteomics Approach Identified Extracellular Vesicular Proteins Correlated to Extracellular Matrix in Type One and Two Endometrial Cancer.

Among gynecological cancers, endometrial cancer is the most common in developed countries. Extracellular vesicles (EVs) are cell-derived membrane-surrounded vesicles that contain proteins involved in immune response and apoptosis. A deep proteomic approach can help to identify dysregulated extracellular matrix (ECM) proteins in EVs correlated to key pathways for tumor development. In this study, we used a proteomics approach correlating the two acquisitions-data-dependent acquisition (DDA) and data-independent acquisition (DIA)-on EVs from the conditioned medium of four cell lines identifying 428 ECM proteins. After protein quantification and statistical analysis, we found significant changes in the abundance (p < 0.05) of 67 proteins. Our bioinformatic analysis identified 26 pathways associated with the ECM. Western blotting analysis on 13 patients with type 1 and type 2 EC and 13 endometrial samples confirmed an altered abundance of MMP2. Our proteomics analysis identified the dysregulated ECM proteins involved in cancer growth. Our data can open the path to other studies for understanding the interaction among cancer cells and the rearrangement of the ECM.

[Knockdown mitochondrial transcription factor A (TFAM) inhibits autophagy, proliferation, invasion and migration in human cervical cancer and osteosarcoma cells].

Objective To investigate the effects of mitochondrial transcription factor A (TFAM) on mitochondrial function, autophagy, proliferation, invasion, and migration in cervical cancer HeLa cells and osteosarcoma U2OS cells. Methods TFAM small-interfering RNA (si-TFAM) was transfected to HeLa and U2OS cells for downregulating TFAM expression. Mito-Tracker Red CMXRos staining combined with laser confocal microscopy was used to detect mitochondrial membrane potential (MMP). MitoSOXTM Red labeling was used to test mitochondrial reactive oxygen species (mtROS) levels. The expression of mitochondrial DNA (mtDNA) was detected by real-time quantitative PCR. Changes in the number of autophagosomes were detected by immunofluorescence cytochemistry. Western blot analysis was used to detect the expressions of TFAM, autophagy microtubule associated protein 1 light chain 3A/B (LC3A/B), autophagy associated protein 2A (ATG2A), ATG2B, ATG9A, zinc finger transcription factor Snail, matrix metalloproteinase 2 (MMP2) and MMP9. CCK-8 assay and plate clony formation assay were used to detect cell proliferation, while TranswellTM assay and scratch healing assay were used to detect changes in cell invasion and migration. Results The downregulation of TFAM expression resulted in a decrease in MMP and mtDNA copy number, but an increase in mtROS production. The protein content of LC3A/B decreased significantly compared to the control group and the number of autophagosomes in the cytoplasm decreased significantly. The expressions of ATG2B and ATG9A in the early stage of autophagy were significantly reduced. The expressions of Snail, MMP2 and MMP9 proteins in HeLa and U2OS cells were also decreased. The proliferation, invasion and migration ability of HeLa and U2OS cells were inhibited after being interfered with TFAM expression. Conclusion Downregulation of TFAM expression inhibits mitochondrial function, delays autophagy process and reduces the proliferation, invasion and migration ability of cervical cancer cells and osteosarcoma cells.

[The Miao medicine Sidaxue alleviates rheumatoid arthritis in rats possibly by downregulating matrix metalloproteinases].

OBJECTIVE: To explore the inhibitory effect of Sidaxue, a traditional Miao herbal medicine formula, on articular bone and cartilage destruction and synovial neovascularization in rats with collagen-induced arthritis (CIA). METHODS: In a SD rat model of CIA, we tested the effects of daily gavage of Sidaxue at low, moderate and high doses (10, 20, and 40 g/kg, respectively) for 21 days, with Tripterygium glycosides (GTW) as the positive control, on swelling in the hind limb plantar regions by arthritis index scoring. Pathologies in joint synovial membrane of the rats were observed with HE staining, and serum TNF-α and IL-1ß levels were detected with ELISA. The expressions of NF-κB p65, matrix metalloproteinase 1 (MMP1), MMP2 and MMP9 at the mRNA and protein levels in the synovial tissues were detected using real-time PCR and Western blotting. Network pharmacology analysis was conducted to identify the important target proteins in the pathways correlated with the therapeutic effects of topical Sidaxue treatment for RA, and the core target proteins were screened by topological analysis. RESULTS: Treatment with GTW and Sidaxue at the 3 doses all significantly alleviated plantar swelling, lowered arthritis index scores, improved cartilage and bone damage and reduced neovascularization in CIA rats (P<0.05), and the effects of Sidaxue showed a dose dependence. Both GTW and Sidaxue treatments significantly lowered TNF-α, IL-1ß, NF-κB p65, MMP1, MMP2, and MMP9 mRNA and protein expressions in the synovial tissues of CIA rats (P<0.05). Network pharmacological analysis identified MMPs as the core proteins associated with topical Sidaxue treatment of RA. CONCLUSION: Sidaxue alleviates articular bone and cartilage damages and reduces synovial neovascularization in CIA rats possibly by downregulating MMPs via the TNF-α/IL-1ß/NF-κB-MMP1, 2, 9 signaling pathway, and MMPs probably plays a key role in mediating the effect of Sidaxue though the therapeutic pathways other than oral administration.

The Potential of the Fibronectin Inhibitor Arg-Gly-Asp-Ser in the Development of Therapies for Glioblastoma.

Glioblastoma (GBM) is the most lethal and common malignant primary brain tumor in adults. An important feature that supports GBM aggressiveness is the unique composition of its extracellular matrix (ECM). Particularly, fibronectin plays an important role in cancer cell adhesion, differentiation, proliferation, and chemoresistance. Thus, herein, a hydrogel with mechanical properties compatible with the brain and the ability to disrupt the dynamic and reciprocal interaction between fibronectin and tumor cells was produced. High-molecular-weight hyaluronic acid (HMW-HA) functionalized with the inhibitory fibronectin peptide Arg-Gly-Asp-Ser (RGDS) was used to produce the polymeric matrix. Liposomes encapsulating doxorubicin (DOX) were also included in the hydrogel to kill GBM cells. The resulting hydrogel containing liposomes with therapeutic DOX concentrations presented rheological properties like a healthy brain. In vitro assays demonstrated that unmodified HMW-HA hydrogels only caused GBM cell killing after DOX incorporation. Conversely, RGDS-functionalized hydrogels displayed per se cytotoxicity. As GBM cells produce several proteolytic enzymes capable of disrupting the peptide-HA bond, we selected MMP-2 to illustrate this phenomenon. Therefore, RGDS internalization can induce GBM cell apoptosis. Importantly, RGDS-functionalized hydrogel incorporating DOX efficiently damaged GBM cells without affecting astrocyte viability, proving its safety. Overall, the results demonstrate the potential of the RGDS-functionalized hydrogel to develop safe and effective GBM treatments.

Luteolin Inhibits Lung Cancer Cell Migration by Negatively Regulating TWIST1 and MMP2 Through Upregulation of miR-106a-5p.

BACKGROUND: Luteolin, a common dietary flavonoid found in plants, has been shown to have anti-cancer properties. However, its exact mechanisms of action in non-small cell lung cancer (NSCLC) are still not fully understood, particularly its role in regulating broader genomic networks and specific gene targets. In this study, we aimed to elucidate the role of microRNAs (miRNAs) in NSCLC treated with luteolin, using A549 cells as a model system. MATERIALS AND METHODS: miRNA profiling was conducted on luteolin-treated A549 cells using Exiqon microarrays, with validation of selected miRNAs by qRT-PCR. Bioinformatic analysis identified the regulatory roles of miRNAs in biological processes and pathways following luteolin treatment. Computational algorithms were employed to identify potential target genes. A549 cells were transfected with miR-106a-5p mimic and inhibitor or their corresponding controls. The expression levels of 2 genes, twist basic helix-loop-helix transcription factor 1 (TWIST1) and matrix metallopeptidase 2 (MMP2), and cell migration were assessed. RESULTS: miRNA profiling identified 341 miRNAs, with 18 exhibiting significantly altered expression (P < 0.05). Subsequent qRT-PCR analysis confirmed altered expression of 6 selected miRNAs. KEGG and GO analyses revealed significant alterations in pathways and biological processes crucial for tumor biology. TWIST1 and MMP2, which both contain conserved miR-106a-5p binding sites, exhibited an inverse correlation with the expression levels of miR-106a-5p. Dual-luciferase reporter assays confirmed TWIST1 and MMP2 as direct targets of miR-106a-5p. Luteolin treatment led to a reduction in A549 cell migration, and this reduction was further amplified by the overexpression of miR-106a-5p. CONCLUSION: Luteolin inhibits A549 cell migration by modulating the miRNA landscape, shedding light on its mechanisms and laying the foundation for miRNA-based therapeutic approaches for NSCLC.

Inhibition of MMP-2 and MMP-9 by Dietary Antioxidants in THP-1 Macrophages and Sera from Patients with Breast Cancer.

Polyphenols, the main antioxidants of diet, have shown anti-inflammatory, antioxidant and anticarcinogenic activities. Here, we compared the effects of four polyphenolic compounds on ROS production and on the levels of matrix metalloproteinase (MMP)-2 and -9, which represent important pathogenetic factors of breast cancer. THP-1 differentiated macrophages were activated by LPS and simultaneously treated with different doses of a green tea extract (GTE), resveratrol (RSV), curcumin (CRC) and an olive fruit extract (oliplus). By using the 2,2-Diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay, we found that all of the tested compounds showed antioxidant activity in vitro. In addition, GTE, RSV and CRC were able to counteract ROS production induced by H2O2 in THP-1 cells. As assessed by a zymographic analysis of THP-1 supernatants and by an "in-gel zymography" of a pool of sera from patients with breast cancer, the antioxidant compounds used in this study inhibited both the activity and expression of MMP-2 and MMP-9 through different mechanisms related to their structures and to their ability to scavenge ROS. The results of this study suggest that the used antioxidants could be promising agents for the prevention and complementary treatment of breast cancer and other diseases in which MMPs play a pivotal role.

Clinical value of KiSS-1 and MMP-2 expression levels in breast cancer tissue in evaluating prognosis of elderly breast cancer patients after modified radical mastectomy.

We attempted to clarify clinical value of KiSS-1 and MMP-2 levels in breast cancer (BC) tissue in evaluating prognosis of elderly BC patients after modified radical mastectomy (MCM). The data of 192 elderly female BC patients receiving MCM in our hospital from January 2018 to December 2022 were collected. According to prognosis, patients received division into poor prognosis group (n = 43) and good prognosis group (n = 149). The serum CEA level and KiSS-1 and MMP-2 levels in BC tissue received measurement in both groups. The predictive value of KiSS-1 and MMP-2 alone and jointly in adverse prognosis of elderly BC patients after MCM received assessment. Results showed that No statistical significance was exhibited between both groups in general data (P > 0.05). The serum CEA level and MMP-2 expression in BC tissue in poor prognosis group exhibited elevation relative to those in good prognosis group, and KiSS-1 expression in BC tissue in poor prognosis group exhibited depletion relative to that in good prognosis group, indicating statistical significance (P < 0.05). The high-level KiSS-1 might be a protective element for adverse prognosis of elderly BC patients after MCM, and high-level CEA and MMP-2 might be an independent risk element for adverse prognosis of elderly BC patients after MCM (P < 0.05). KiSS-1 and MMP-2 alone and jointly predicted AUC of adverse prognosis in elderly BC patients after MCM were 0.93, 0.802 and 0.958, with certain predictive values; when cutoff values of KiSS-1 and MMP-2 were 6.15 and 2.26, the predictive value was the best. In conclusion, KiSS-1 and MMP-2 levels in BC tissue possess relation to adverse prognosis of MCM. KiSS-1 and MMP-2 levels in elderly BC patients before surgery may be detected in the future to assist in prognosis evaluation of elderly BC patients after MCM.

Irradiated riboflavin over nonradiated one: Potent antimigratory, antiproliferative and cytotoxic effects on glioblastoma cells.

Riboflavin is a water-soluble yellowish vitamin and is controversial regarding its effect on tumour cells. Riboflavin is a powerful photosensitizer that upon exposure to radiation, undergoes an intersystem conversion with molecular oxygen, leading to the production of ROS. In the current study, we sought to ascertain the impact of irradiated riboflavin on C6 glioblastoma cells regarding proliferation, cell death, oxidative stress and migration. First, we compared the proliferative behaviour of cells following nonradiated and radiated riboflavin. Next, we performed apoptotic assays including Annexin V and caspase 3, 7 and 9 assays. Then we checked on oxidative stress and status by flow cytometry and ELISA kits. Finally, we examined inflammatory change and levels of MMP2 and SIRT1 proteins. We caught a clear antiproliferative and cytotoxic effect of irradiated riboflavin compared to nonradiated one. Therefore, we proceeded with our experiments using radiated riboflavin. In all apoptotic assays, we observed a dose-dependent increase. Additionally, the levels of oxidants were found to increase, while antioxidant levels decreased following riboflavin treatment. In the inflammation analysis, we observed elevated levels of both pro-inflammatory and anti-inflammatory cytokines. Additionally, after treatment, we observed reduced levels of MMP2 and SIRT. In conclusion, radiated riboflavin clearly demonstrates superior antiproliferative and apoptotic effects on C6 cells at lower doses compared to nonradiated riboflavin.

Effects of bone marrow mesenchymal stem cell-conditioned medium on the proliferation and migration of liposarcoma cells.

INTRODUCTION: Liposarcoma constitutes a prevalent subtype of soft tissue sarcoma, represents approximately 20% of all sarcomas. However, conventional chemotherapeutic agents have shown restricted effectiveness in treating liposarcoma patients. Accumulating evidence indicates that mesenchymal stem cells (MSCs) have the characteristic of migration to tumor site, promote or suppress tumors. How human bone marrow mesenchymal stem cells (BMSCs) contribute to liposarcoma phenotype remains poorly understood. This study aims to investigate the effects of human bone marrow mesenchymal stem cell-conditioned medium (BMSC-CM) on the proliferation and migration of liposarcoma cell lines 93T449 and SW872, as well as explore potential underlying mechanisms of BMSC-CM action on these cells. MATERIALS AND METHODS: We transfected BMSCs with lentiviral constructs to knock down the transcriptional co-activator Yes-associated protein 1 (YAP1), conditioned medium (CM) obtained from BMSCs and shYAP1-BMSC, respectively. Liposarcoma cell lines 93T449 and SW872 were co-cultured with BMSC-CM or shYAP1-BMSC-CM. Cell proliferation ability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell apoptosis was evaluated using flow cytometric assay. A wound healing assay was used to analyze cell migration. The expression levels of YAP1, Bcl-2, and matrix metalloproteinase-2 (MMP-2) were determined by western blot assay. RESULTS: Co-culturing liposarcoma cell lines 93T449 and SW872 with BMSC-CM promoted tumor cell proliferation, while shYAP1-BMSC-CM significantly inhibited cell viability and migration, induced apoptosis, and downregulated Bcl-2 and MMP-2 expression. CONCLUSIONS: These findings provide new insights into the impact of BMSC-CM on liposarcoma and suggest its possible involvement in liposarcoma cell growth.

Cold atmospheric plasma attenuates skin cancer via ROS induced apoptosis.

BACKGROUND: Cold atmospheric plasma (CAP) has been widely used in biomedical research, especially in vitro cancer therapy. Cutaneous squamous cell carcinoma (CSCC) is a malignant tumor originating from epidermal keratinocytes. However, the mechanism of CAP therapy on CSCC remains unclear. METHODS AND RESULTS: The animal models of CSCC induced by 7,12-dimethylbenz(a) anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA) were constructed. For the CAP treatment group, after each TPA application, CAP was administered for 3 min twice weekly after drying. HE staining were used to detect the pathological status of tumor tissue in each group. The levels of PCNA, Bcl-2, Bax, MMP2 and MMP9 were evaluated by western blot and qPCR. TUNEL staining were used to detect apoptosis in tumor tissues. In vivo, serum samples were used for ELISA of total ROS. MTT assay was used to detect the viability of A431 cells. Western blot and qPCR were used to detect the levels of PCNA, Bcl-2, Bax, MMP2 and MMP9 in A431 cells. A431 cell proliferation was examined by colony formation assay. The proportions of apoptosis of A431 cells were detected by flow cytometry. Transwell assessed the ability of A431 cells migration and proliferation. We found that CAP could induce skin cancer cells apoptosis and inhibit the progress of skin cancer. Through experiments in vitro, reactive oxygen species (ROS) generated by N-acetylcysteine (NAC) and CAP inhibited the proliferation and migration of A431 skin cancer cells while promoting apoptosis. CONCLUSIONS: These evidences suggest the protective effect of CAP in CSCC, and CAP has the potential clinical application of CSCC.

Urolithin A affects cellular migration and modulates matrix metalloproteinase expression in colorectal cancer cells.

Colorectal cancer (CRC) is the world's second most common gastrointestinal malignancy. Preventing tumor cell proliferation and dissemination is critical for patient survival. Polyphenols have a variety of health advantages and can help prevent cancer. The current study examined different cellular activities of the gut-microbiota metabolite urolithin A (UA) on several colon cancer cell lines. The results revealed that UA suppressed cell growth in a dose- and time-dependent manner. In the current investigation, UA substantially affected cell migration in the wound-healing experiment and greatly decreased the number of colonies generated in each CRC cell culture. UA decreased cellular migration in CRC cells 48 h after treatment, which was significant (p < .001) compared to the migration rate in untreated cells. When compared to untreated cells, UA slowed the process of colony formation by reducing the number of colonies or altering their morphological shape. The western blot analysis investigation revealed that UA inhibits cellular metastasis by lowering the expression levels of matrix metalloproteinases 1 and 2 (MMP1 and MMP2) by more than 43% and 41% (p < .001) in HT29, 28% and 149% (p < .001) in SW480, and 90% and 74% (p < .001) in SW620, respectively, at a 100 µM dosage of UA compared to the control. Surprisingly, at a 100 µM dosage of UA, the expression levels of the tissue inhibitor of metalloproteinases 1 (TIMP1) were elevated in HT29, SW480, and SW620 cells treated with 100 µM of UA by more than 89%, 57%, and 29%, respectively. Our findings imply that UA has anticancer properties and might be used therapeutically to treat CRC. The findings provided the first indication of the influence of UA on cellular migration and metastasis in colon cancer cells. All of these data showed that UA might be used as an adjuvant therapy in the treatment of various forms of CRC.

Identifying and validating MMP family members (MMP2, MMP9, MMP12, and MMP16) as therapeutic targets and biomarkers in kidney renal clear cell carcinoma (KIRC).

Kidney Renal Clear Cell Carcinoma (KIRC) is a malignant tumor that carries a substantial risk of morbidity and mortality. The MMP family assumes a crucial role in tumor invasion and metastasis. This study aimed to uncover the mechanistic relevance of the MMP gene family as a therapeutic target and diagnostic biomarker in Kidney Renal Clear Cell Carcinoma (KIRC) through a comprehensive approach encompassing both computational and molecular analyses. STRING, Cytoscape, UALCAN, GEPIA, OncoDB, HPA, cBioPortal, GSEA, TIMER, ENCORI, DrugBank, targeted bisulfite sequencing (bisulfite-seq), conventional PCR, Sanger sequencing, and RT-qPCR based analyses were used in the present study to analyze MMP gene family members to accurately determine a few hub genes that can be utilized as both therapeutic targets and diagnostic biomarkers for KIRC. By performing STRING and Cytohubba analyses of the 24 MMP gene family members, MMP2 (matrix metallopeptidase 2), MMP9 (matrix metallopeptidase 9), MMP12 (matrix metallopeptidase 12), and MMP16 (matrix metallopeptidase 16) genes were denoted as hub genes having highest degree scores. After analyzing MMP2, MMP9, MMP12, and MMP16 via various TCGA databases and RT-qPCR technique across clinical samples and KIRC cell lines, interestingly, all these hub genes were found significantly overexpressed at mRNA and protein levels in KIRC samples relative to controls. The notable effect of the up-regulated MMP2, MMP9, MMP12, and MMP16 was also documented on the overall survival (OS) of the KIRC patients. Moreover, targeted bisulfite-sequencing (bisulfite-seq) analysis revealed that promoter hypomethylation pattern was associated with up-regulation of hub genes (MMP2, MMP9, MMP12, and MMP16). In addition to this, hub genes were involved in various diverse oncogenic pathways. The MMP gene family members (MMP2, MMP9, MMP12, and MMP16) may serve as therapeutic targets and prognostic biomarkers in KIRC.

TIM3 activates the ERK1/2 pathway to promote invasion and migration of thyroid tumors.

BACKGROUND: This study aims to study the possible action mechanism of T-cell immunoglobulin and mucin domain 3 (TIM3) on the migratory and invasive abilities of thyroid carcinoma (TC) cells. METHODS: GSE104005 and GSE138198 datasets were downloaded from the GEO database for identifying differentially expressed genes (DEGs). Functional enrichment analysis and protein-protein interaction (PPI) analysis were performed on the common DEGs in GSE104005 and GSE138198 datasets. Subsequently, in order to understand the effect of a common DEG (TIM3) on TC cells, we performed in vitro experiments using FRO cells. The migratory and invasive abilities of FRO cells were detected by wound scratch assay and Transwell assay. Proteins expression levels of the phosphorylated (p)-extracellular signal-regulated kinase (ERK)1/2, matrix metalloproteinase-2 (MMP-2) and MMP-9 were determined via Western blotting after ERK1/2 inhibition in TIM3-NC group and TIM3-mimic group. RESULTS: 316 common DEGs were identified in GSE104005 and GSE138198 datasets. These DEGs were involved in the biological process of ERK1 and ERK2 cascade. TIM3 was significantly up-regulated in TC. In vitro cell experiments showed that TIM3 could promote migration and invasion of TC cells. Moreover, TIM3 may affect the migration, invasive abilities of TC cells by activating the ERK1/2 pathway. CONCLUSION: The above results indicate that TIM3 may affect the migratory and invasive of TC cells by activating the ERK1/2 pathway.

LIFU/MMP-2 dual-responsive release of repurposed drug disulfiram from nanodroplets for inhibiting vasculogenic mimicry and lung metastasis in triple-negative breast cancer.

BACKGROUND: Vasculogenic mimicry (VM), when microvascular channels are formed by cancer cells independent of endothelial cells, often occurs in deep hypoxic areas of tumors and contributes to the aggressiveness and metastasis of triple-negative breast cancer (TNBC) cells. However, well-developed VM inhibitors exhibit inadequate efficacy due to their low drug utilization rate and limited deep penetration. Thus, a cost-effective VM inhibition strategy needs to be designed for TNBC treatment. RESULTS: Herein, we designed a low-intensity focused ultrasound (LIFU) and matrix metalloproteinase-2 (MMP-2) dual-responsive nanoplatform termed PFP@PDM-PEG for the cost-effective and efficient utilization of the drug disulfiram (DSF) as a VM inhibitor. The PFP@PDM-PEG nanodroplets effectively penetrated tumors and exhibited substantial accumulation facilitated by PEG deshielding in a LIFU-mediated and MMP-2-sensitive manner. Furthermore, upon exposure to LIFU irradiation, DSF was released controllably under ultrasound imaging guidance. This secure and controllable dual-response DSF delivery platform reduced VM formation by inhibiting COL1/pro-MMP-2 activity, thereby significantly inhibiting tumor progression and metastasis. CONCLUSIONS: Considering the safety of the raw materials, controlled treatment process, and reliable repurposing of DSF, this dual-responsive nanoplatform represents a novel and effective VM-based therapeutic strategy for TNBC in clinical settings.

Determination of anti-cancer effects of Nigella sativa seed oil on MCF7 breast and AGS gastric cancer cells.

BACKGROUND: This study aimed to investigate the cytotoxic, apoptotic, invasion, metastasis, and heat shock proteins (HSPs) effects of N. sativa oil on breast and gastric cancer cells. METHODS: We assessed the cytotoxic and apoptotic effects of various concentrations of N. sativa oil (10-50-100-200 µg/mL) on MCF7 breast cancer and AGS, an adenocarcinoma of the gastric cell line, at 24, 48 and 72 h using the MTT test. Additionally, the expression of the Caspase-3, BCL2/Bax, MMP2-9 and HSP60-70 gene was examined using RT-PCR in cell lines treating with N. sativa. RESULTS: The MTT experiments demonstrate that N. sativa has a time and dose-dependent inhibitory effect on the proliferation of MCF7 and AGS cancer cells. The vitality rates of MCF7 and AGS cells treated with N. sativa were 77.04-67.50% at 24 h, 65.28-39.14% at 48 h, and 48.95-32.31% at 72 h. The doses of 100 and 200 µg/mL were shown to be the most effective on both cancer cells. RT-PCR analysis revealed that N. sativa oil extract increased caspase-3 levels in both cell lines at higher concentrations and suppressed BCL2/Bax levels. Exposure of MCF7 and AGS cell lines to N. sativa caused a significant decrease in the expression of MMP2-9 and HSP60-70 genes over time, particularly at a dosage of 200 µg/mL compared to the control group (p < 0.05). CONCLUSIONS: Our findings indicate that N. sativa oil has a dose-dependent effect on cytotoxicity and the expression of apoptotic, heat shock proteins, and matrix metalloproteinases genes in breast and gastric cancer.

Deciphering the role of wound healing genes in skin cutaneous melanoma: Insights into expression, methylation, mutations, and therapeutic implications.

Skin Cutaneous Melanoma (SKCM) is a form of cancer that originates in the pigment-producing cells, known as melanocytes, of the skin. Delay wound healing is often correlated with the occurrence of and progression of SKCM. In this comprehensive study, we investigated the intricate roles of two important wound healing genes in SKCM, including Matrix Metalloproteinase-2 (MMP2) and Matrix Metalloproteinase-9 (MMP9). Through a multi-faceted approach, we collected clinical samples, conducted molecular experiments, including RT-qPCR, bisulphite sequencing, cell culture, cell Counting Kit-8, colony formation, and wound healing assays. Beside this, we also used various other databases/tools/approaches for additional analysis including, UALCAN, GEPIA, HPA, MEXPRESS, cBioPortal, KM plotter, DrugBank, and molecular docking. Our results revealed a significant up-regulation of MMP2 and MMP9 in SKCM tissues compared to normal counterparts. Moreover, promoter methylation analysis suggested an epigenetic regulatory mechanism. Validations using TCGA datasets and immunohistochemistry emphasized the clinical relevance of MMP2 and MMP9 dysregulation. Functional assays demonstrated their synergistic impact on proliferation and migration in SKCM cells. Furthermore, we identified potential therapeutic candidates, Estradiol and Calcitriol, through drug prediction and molecular docking analyses. These compounds exhibited binding affinities, suggesting their potential as MMP2/MMP9 inhibitors. Overall, our study elucidates the diagnostic, prognostic, and therapeutic implications of MMP2 and MMP9 in SKCM, shedding light on their complex interplay in SKCM occurrence and progression.

Direct electrochemical reduction of graphene oxide thin film for aptamer-based selective and highly sensitive detection of matrix metalloproteinase 2.

Simple and low-cost biosensing solutions are suitable for point-of-care applications aiming to overcome the gap between scientific concepts and technological production. To compete with sensitivity and selectivity of golden standards, such as liquid chromatography, the functionalization of biosensors is continuously optimized to enhance the signal and improve their performance, often leading to complex chemical assay development. In this research, the efforts are made on optimizing the methodology for electrochemical reduction of graphene oxide to produce thin film-modified gold electrodes. Under the employed specific conditions, 20 cycles of cyclic voltammetry (CV) are shown to be optimal for superior electrical activation of graphene oxide into electrochemically reduced graphene oxide (ERGO). This platform is further used to develop a matrix metalloproteinase 2 (MMP-2) biosensor, where specific anti-MMP2 aptamers are utilized as a biorecognition element. MMP-2 is a protein which is typically overexpressed in tumor tissues, with important roles in tumor invasion, metastasis as well as in tumor angiogenesis. Based on impedimetric measurements, we were able to detect as low as 3.32 pg mL-1 of MMP-2 in PBS with a dynamic range of 10 pg mL-1 - 10 ng mL-1. Further experiments with real blood samples revealed a promising potential of the developed sensor for direct measurement of MMP-2 in complex media. High specificity of detection is demonstrated - even to the closely related enzyme MMP-9. Finally, the potential of reuse was demonstrated by signal restoration after experimental detection of MMP-2.

Functional relevance of MMP2 promoter variants in gallbladder cancer: A case-control study in an Eastern Indian Population.

Gallbladder cancer (GBC) is a prevalent and deadly form of bile duct cancer, associated with poor prognosis. This study aimed to investigate the genetic factors contributing to the high incidence of GBC in certain geographical regions, particularly in the Northern and Eastern parts of India. The present case-control study focused on MMP2, a gene involved in tumor progression and metastasis, as a potential candidate in GBC pathogenesis. We scanned MMP2 promoter for twelve SNPs using Sanger's sequencing and carried out a case-control study in 300 cases and 300 control samples. We found five rare variants (rs1961998763, rs1961996235, rs1391392808, rs1488656253, and rs17859816) and one nonpolymorphic SNP (rs17859817). Our results revealed a significant association between GBC and MMP2 promoter SNPs, rs243865 (Allelic-Padjusted = 0.0353) and g.55477735G > A (Allelic-Padjusted = 9.22E-05). Moreover, the haplotype "C-C-A-C-C" exhibited a significant association with GBC (P = 4.23E-05). Genotype-phenotype correlation for variant rs243865, in the GBC patient tissue samples, established that 'T' risk allele carriers had higher expression levels of MMP2. Additionally, luciferase reporter assay in HEK293T cells revealed the probable regulatory role of rs243865 variant allele 'T' in MMP2 expression. Our study uncovers the association of MMP2 promoter SNPs with GBC and their role in regulating its expression.

Comparative analysis of chemical profiles, antioxidant, antibacterial, and anticancer effects of essential oils of two Thymus species from Montenegro.

The essential oils of Thymus vulgaris (TVEO) and Thymus serpyllum (TSEO) show different biological activities. The aim of the study was to evaluate the biological activities of TVEO and TSEO from Montenegro. The main components of TVEO were p-cymene (29.52%), thymol (22.8%) and linalool (4.73%) while the main components of TSEO were p-cymene (19.04%), geraniol (11,09%), linalool (9.16%), geranyl acetate (6.49%) and borneol (5.24%). Antioxidant activity determined via DPPH for TVEO was 4.49 and FRAP 1130.27, while for TSEO it was estimated that DPPH was 4.88 µL/mL and FRAP was 701.25 µmol FRAP/L. Both essential oils were active against all tested bacteria, with the highest level of sensitivity of E. coli with MIC of 1.5625 µL/mL. Essential oils showed strong cytotoxic effects on human cancer cell lines, with IC50 values ranging from 0.20 to 0.24 µL/mL for TVEO and from 0.32 to 0.49 µL/mL for TSEO. TVEO caused apoptosis in cervical adenocarcinoma HeLa cells through activation of caspase-3 and caspase-8, while TSEO caused apoptosis through caspase-3. EOs decreased levels of oxidative stress in normal MRC-5 cells. HeLa cells treated with TVEO had reduced MMP2 expression levels, while cells treated with TSEO had lowered MMP2 and MMP9 levels. The treatment of HeLa cells with TVEO increased the levels of miR-16 and miR-34a, indicating potential tumor-suppressive properties. Our findings suggest that Thymus essential oils may be considered as good candidates for further investigation as cancer-chemopreventive and cancer-therapeutic agents.

HSP27 promotes vasculogenic mimicry formation in human salivary adenoid cystic carcinoma via the AKT-MMP-2/9 pathway.

PURPOSE: To explore the role and mechanism of heat shock protein 27 (HSP27) in SACC VM formation. STUDY DESIGN: Immunohistochemistry and double staining with cluster of differentiation 31 (CD31) and periodic acid-Schiff (PAS) were used to detect HSP27 expression and VM in 70 SACC tissue samples separately. Quantitative real-time polymerase chain reaction (qRT-PCR), western blot analysis, and immunofluorescence were used to detect gene and protein expression. HSP27 in SACC cells were overexpression or downregulated by transfecting HSP27 or short hairpin RNA target HSP27 (sh-HSP27). The migration and invasion abilities of SACC cells were detected using wound healing and Transwell invasion assays. The VM formation ability of the cells in vitro was detected using a Matrigel 3-dimensional culture. RESULTS: HSP27 expression was positively correlated with VM formation and affected the prognosis of patients. In vitro, HSP27 upregulation engendered VM formation and the invasion and migration of SACC cells. Mechanistically, HSP27 upregulation increased Akt phosphorylation and subsequently increased downstream matrix metalloproteinase 2 and 9 expressions. CONCLUSION: HSP27 may plays an important role in VM formation in SACC via the AKT-MMP-2/9 signalling pathway.