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.

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.

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.

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.

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.

Harmaline induces apoptosis and inhibits migration in A2780 ovarian cancer cells in vitro.

Ovarian cancer is one of the most prevalent malignancies in women. Harmaline is reported to have powerful anticancer properties. We aimed to investigate the apoptotic and antimetastatic properties of harmaline in A2780 ovarian cancer cells. Cell viability, apoptosis, migration, and invasion were investigated in cells treated with harmaline. Reactive oxygen species (ROS) production, mRNA expression of apoptosis-associated genes, MMP-2, and MMP-9 were measured. Harmaline attenuated the viability of A2780 ovarian cancer cells in a dose- and time-dependent way. Furthermore, compared to NIH/3T3 mouse normal cell line (IC50 = 417 µM), the malignant A2080 cells were more sensitive to harmaline (IC50 = 300 µM after 24 h). Harmaline increased the production of ROS, raised the mRNA expression of p53 and the Bax/Bcl2 ratio. Harmaline also increased the proportion of cells in the late apoptotic and necrotic phases. MMP-2 and MMP-9's mRNA expression, gelatinase activity, and migration of A2780 cells also decreased by harmaline. These findings suggest that harmaline may have the potential to be a therapeutic drug for ovarian cancer by triggering apoptosis and suppressing invasion and migration.

Osteosarcoma cells exhibit functional interactions with stromal cells, fostering a lung microenvironment conducive to the establishment of metastatic tumor cells.

BACKGROUND: Osteosarcoma (OS) stands out as the most common bone tumor, with approximately 20% of the patients receiving a diagnosis of metastatic OS at their initial assessment. A significant challenge lies in the frequent existence of undetected metastases during the initial diagnosis. Mesenchymal stem cells (MSCs) possess unique abilities that facilitate tumor growth, and their interaction with OS cells is crucial for metastatic spread. METHODS AND RESULTS: We demonstrated that, in vitro, MSCs exhibited a heightened migration response toward the secretome of non-metastatic OS cells. When challenged to a secretome derived from lungs preloaded with OS cells, MSCs exhibited greater migration toward lungs colonized with metastatic OS cells. Moreover, in vivo, MSCs displayed preferential migratory and homing behavior toward lungs colonized by metastatic OS cells. Metastatic OS cells, in turn, demonstrated an increased migratory response to the MSCs' secretome. This behavior was associated with heightened cathepsin D (CTSD) expression and the release of active metalloproteinase 2 (MMP2) by metastatic OS cells. CONCLUSIONS: Our assessment focused on two complementary tumor capabilities crucial to metastatic spread, emphasizing the significance of inherent cell features. The findings underscore the pivotal role of signaling integration within the niche, with a complex interplay of migratory responses among established OS cells in the lungs, prometastatic OS cells in the primary tumor, and circulating MSCs. Pulmonary metastases continue to be a significant factor contributing to OS mortality. Understanding these mechanisms and identifying differentially expressed genes is essential for pinpointing markers and targets to manage metastatic spread and improve outcomes for patients with OS.

Minocycline induced apoptosis and suppressed expression of matrix metalloproteinases 2 and 9 in the breast cancer MCF-7 cells.

BACKGROUND: In women, breast cancer is the second most frequent type of cancer. Looking for new and effective cancer-specific therapies with little to no adverse effects on healthy cells is critical. OBJECTIVE: Minocycline, a second-generation tetracycline, has shown anticancer effects by targeting multiple pathways in various cancers. This study aimed to determine minocycline effects on the cell proliferation, apoptosis, and invasion of the human MCF-7 cells. METHODS: MTT assay was used to evaluate the cytotoxicity of minocycline on the cells. Flow cytometry was performed to investigate the induction of apoptosis and the cell cycle progression. The expression levels of apoptotic and migration proteins and genes were assessed by western blotting and qRT-PCR. The scratch test was performed to evaluate the anti-migration effect of the drug. RESULTS: The results indicated that the IC50 value of minocycline for MCF-7 cells was 36.10 µM. Minocycline treatment caused sub-G1 cell accumulation, indicating a significant apoptotic effect on the MCF-7 cells. Annexin-V/PI staining revealed a significant rise in early and late apoptotic cell percentages. Minocycline up-regulated Bax and Caspase-3 expression and down-regulated Bcl-2 and Pro-Cas3. The scratch test revealed significant anti-migration effects for minocycline. Furthermore, it caused down-regulation of MMP-2 and MMP-9 in a concentration-dependent method. CONCLUSION: These findings further confirmed the anticancer effect of minocycline and highlighted that minocycline maybe considered as potential therapeutic agent for breast cancer treatment.

Inhibition of melanoma cell migration and invasion by natural coumarin auraptene through regulating EMT markers and reducing MMP-2 and MMP-9 activity.

Melanoma, the most invasive form of skin cancer, shows a rising incidence trend in industrial countries. Since the main reason for the failure of current therapeutic approaches against melanoma is metastasis, there is a great interest in introducing effective natural agents to combat melanoma cell migration and invasion. Auraptene (AUR) is the most abundant coumarin derivative in nature with valuable pharmaceutical effects. In this study, we aimed to investigate whether AUR could induce inhibitory effects on the migration and invasion of melanoma cells. B16F10 melanoma cells were treated with different concentrations of AUR and the viability of cells was evaluated by alamarBlue assay. Then, cells were treated with 20 µM AUR, and wound healing, invasion, and adhesion assays were carried out. In addition, the activity of matrix metalloproteinase-2 (MMP-2) and MMP-9 was assessed by gelatin zymography and the expression of genes related to epithelial-mesenchymal transition (EMT) was investigated by qPCR. Finally, the interactions between AUR and MMPs were stimulated by molecular docking. Findings revealed that AUR significantly reduced the migration and invasion of B16F10 cells while improved their adhesion. Furthermore, results of gelatin zymography indicated that AUR suppressed the activity of MMP-2 and MMP-9, and qPCR revealed negative regulatory effect of AUR on the expression of mesenchymal markers including fibronectin and N-cadherin. In addition, molecular docking verified the interactions between AUR and the active sites of wild-type and mutant MMP-2 and MMP-9. Accordingly, AUR could be considered as a potential natural agent with inhibitory effects on the migration and invasion of melanoma cells for future preclinical studies.

METTL3 Promotes Osteosarcoma Metastasis via an m6A-dependent Epigenetic Activity of CBX4.

BACKGROUND: Osteosarcoma cells are prone to metastasis, and the mechanism of N6-methyladenosine (m6A) methylation modification in this process is still unclear. Methylation modification of m6A plays an important role in the development of osteosarcoma, which is mainly due to abnormal expression of enzymes related to methylation modification of m6A, which in turn leads to changes in the methylation level of downstream target genes messenger RNA (mRNA) leading to tumor development. METHODS: We analyzed the expression levels of m6A methylation modification-related enzyme genes in GSE12865 whole-genome sequencing data. And we used shRNA (short hairpin RNA) lentiviral interference to interfere with METTL3 (Methyltransferase 3) expression in osteosarcoma cells. We studied the cytological function of METTL3 by Cell Counting Kit-8 (CCK8), flow cytometry, migration and other experiments, and the molecular mechanism of METTL3 by RIP (RNA binding protein immunoprecipitation), Western blot and other experiments. RESULTS: We found that METTL3 is abnormally highly expressed in osteosarcoma and interferes with METTL3 expression in osteosarcoma cells to inhibit metastasis, proliferation, and apoptosis of osteosarcoma cells. We subsequently found that METTL3 binds to the mRNA of CBX4 (chromobox homolog 4), a very important regulatory protein in osteosarcoma metastasis, and METTL3 regulates the mRNA and protein expression of CBX4. Further studies revealed that METTL3 inhibited metastasis of osteosarcoma cells by regulating CBX4. METTL3 has been found to be involved in osteosarcoma cells metastasis by CBX4 affecting the protein expression of matrix metalloproteinase 2 (MMP2), MMP9, E-Cadherin and N-Cadherin associated with osteosarcoma cells metastasis. CONCLUSIONS: These results suggest that the combined action of METTL3 and CBX4 plays an important role in the regulation of metastasis of osteosarcoma, and therefore, the METTL3-CBX4 axis pathway may be a new potential therapeutic target for osteosarcoma.

Inhibiting IL11RA to mitigate hepatic metastasis in skin cutaneous melanoma: Comprehensive insights from in vitro and in vivo investigations.

OBJECTIVE: This study aimed to investigate the role of Interleukin-11 receptor alpha (IL11RA) in skin cutaneous melanoma (SKCM) metastasis to the liver. METHODS: Human SKCM cell lines (A375, A375-MA2, SK-MEL-28, RPMI-7951) and primary dermal fibroblasts (HDFa) were utilized to assess IL11RA expression. IL11RA siRNA was transfected into RPMI-7951 and A375-MA2 cells for Wound healing and Transwell invasion assays. Il11ra knockout (KO) mice and wild-type (WT) mice were injected with B16-F10 cells into the spleen to evaluate hepatic melanoma metastasis. Correlation between IL11RA and MMP family genes was explored using online databases, including LinkedOmics, TIMER (Tumor Immune Estimation Resource), and GEPIA (Gene Expression Profiling Interactive Analysis). RT-qPCR and Western blotting were performed for expression analysis of Mmp2 and Mmp9 in liver tissues of mice. The impact of IL11RA on the STAT3 pathway was investigated in vitro and in vivo. RESULTS: Elevated expression of IL11RA was observed in SKCM cell lines compared to normal cells. IL11RA downregulation significantly inhibited migratory and invasive capabilities of A375-MA2 and RPMI-7951 in vitro. Il11ra gene knockout in mice demonstrated a substantial reduction in hepatic melanoma metastasis. Correlation analyses revealed associations between IL11RA and MMP2/MMP8. Il11ra gene knockout significantly decreased Mmp2 expression while increasing Mmp8 in liver tissues. IL11RA correlated positively with STAT3, and its inhibition led to a suppressed STAT3 pathway in SKCM cells and mouse liver tissue. CONCLUSION: IL11RA plays a crucial role in SKCM metastasis, affecting migratory and invasive abilities. Targeting IL11RA may offer a promising avenue for therapeutic interventions in cutaneous melanoma progression.

Relationship between the Expression of Matrix Metalloproteinases and Their Tissue Inhibitors in Patients with Brain Tumors.

Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) play critical roles in regulating processes associated with malignant behavior. These endopeptidases selectively degrade components of the extracellular matrix (ECM), growth factors, and their receptors, contributing to cancer cell invasiveness and migratory characteristics by disrupting the basal membrane. However, the expression profile and role of various matrix metalloproteinases remain unclear, and only a few studies have focused on differences between diagnoses of brain tumors. Using quantitative real-time PCR analysis, we identified the expression pattern of ECM modulators (n = 10) in biopsies from glioblastoma (GBM; n = 20), astrocytoma (AST; n = 9), and meningioma (MNG; n = 19) patients. We found eight deregulated genes in the glioblastoma group compared to the benign meningioma group, with only MMP9 (FC = 2.55; p = 0.09) and TIMP4 (7.28; p < 0.0001) upregulated in an aggressive form. The most substantial positive change in fold regulation for all tumors was detected in matrix metalloproteinase 2 (MNG = 30.9, AST = 4.28, and GBM = 4.12). Notably, we observed an influence of TIMP1, demonstrating a positive correlation with MMP8, MMP9, and MMP10 in tumor samples. Subsequently, we examined the protein levels of the investigated MMPs (n = 7) and TIMPs (n = 3) via immunodetection. We confirmed elevated levels of MMPs and TIMPs in GBM patients compared to meningiomas and astrocytomas. Even when correlating glioblastomas versus astrocytomas, we showed a significantly increased level of MMP1, MMP3, MMP13, and TIMP1. The identified metalloproteases may play a key role in the process of gliomagenesis and may represent potential targets for personalized therapy. However, as we have not confirmed the relationship between mRNA expression and protein levels in individual samples, it is therefore natural that the regulation of metalloproteases will be subject to several factors.

A review of matrix metalloproteinase-2-sensitive nanoparticles as a novel drug delivery for tumor therapy.

Matrix metalloproteinase-2 (MMP-2)-responsive nanodrug vehicles have garnered significant attention as antitumor drug delivery systems due to the extensive research on matrix metalloproteinases (MMPs) within the tumor extracellular matrix (ECM). These nanodrug vehicles exhibit stable circulation in the bloodstream and accumulate specifically in tumors through various mechanisms. Upon reaching tumor tissues, their structures are degraded in response to MMP-2 within the ECM, resulting in drug release. This controlled drug release significantly increases drug concentration within tumors, thereby enhancing its antitumor efficacy while minimizing side effects on normal organs. This review provides an overview of MMP-2 characteristics, enzyme-sensitive materials, and current research progress regarding their application as MMP-2-responsive nanodrug delivery system for anti-tumor drugs, as well as considering their future research prospects. In conclusion, MMP-2-sensitive drug delivery carriers have a broad application in all kinds of nanodrug delivery systems and are expected to become one of the main means for the clinical development and application of nanodrug delivery systems in the future.