Dataset Information

COX?2 promotes epithelial?mesenchymal transition and migration in osteosarcoma MG?63 cells via PI3K/AKT/NF??B signaling.

ABSTRACT: The present study aimed to investigate the mechanism by which cyclooxygenase?2 (COX?2) promotes the metastasis of MG?63 osteosarcoma cells through the PI3K/AKT/NF??B pathway. To achieve this, a recombinant lentivirus containing the COX?2 gene was constructed in order to overexpress COX?2; a recombinant lentivirus containing a control sequence was also constructed. A Transwell chamber migration assay was performed to quantify the migration of the COX?2?transduced cells, and of cells treated with a COX?2 inhibitor (NS398) or a PI3K inhibitor (LY294002). Immunofluorescence assays were performed to determine changes in E?cadherin, vimentin and NF??B expression levels. ELISAs were performed to quantify the levels of matrix metallopeptidase (MMP)?2, MMP?9 and vascular endothelial growth factor (VEGF) in the culture medium. Western blot analysis was conducted to measure the protein expression levels of MMP?2, MMP?9, PI3K, phosphorylated (p?) PI3K, AKT, p?AKT, inhibitor of NF??? kinase (IKK) and p?IKK. The results demonstrated that the migration ability of the COX?2?overexpressing MG?63 cells was significantly increased compared with the control cells. The migration ability of cells treated with NS398 or LY294002 was significantly decreased. Compared with the control cells, E?cadherin expression was significantly decreased in COX?2?overexpressing cells, while the expression levels of vimentin, MMP?2, MMP?9, VEGF, p?PI3K, p?AKT and p?IKK were significantly increased. Compared with the control cells, E?cadherin expression was significantly increased in cells treated with NS398 or LY294002, while the expression levels of vimentin, MMP?2, MMP?9, VEGF, p?PI3K, p?AKT, and p?IKK were significantly decreased. The total protein levels of PI3K, AKT and IKK were not changed among the treatment groups. In summary, COX?2 overexpression decreased the expression levels of the epithelial protein E?cadherin and increased the expression levels of the mesenchymal proteins vimentin, MMP?2 and MMP?9, as well as promoted cell migration, by activating the PI3K/AKT/NF??B signaling pathway.

SUBMITTER: Zhang X

PROVIDER: S-EPMC6755176 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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COX‑2 promotes epithelial‑mesenchymal transition and migration in osteosarcoma MG‑63 cells via PI3K/AKT/NF‑κB signaling.

Zhang Xueliang X Qu Peng P Zhao Hui H Zhao Tong T Cao Nong N

Molecular medicine reports 20190821 4

The present study aimed to investigate the mechanism by which cyclooxygenase‑2 (COX‑2) promotes the metastasis of MG‑63 osteosarcoma cells through the PI3K/AKT/NF‑κB pathway. To achieve this, a recombinant lentivirus containing the COX‑2 gene was constructed in order to overexpress COX‑2; a recombinant lentivirus containing a control sequence was also constructed. A Transwell chamber migration assay was performed to quantify the migration of the COX‑2‑transduced cells, and of cells treated with ...[more]

PMID: 31485669

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Project description:BackgroundPhytoestrogens such as genistein, the most prominent isoflavone from soy, show concentration-dependent anti-estrogenic or estrogenic effects. High genistein concentrations (>10 ?M) also promote proliferation of bone cancer cells in vitro. On the other hand, the most active component of the vitamin D family, calcitriol, has been shown to be tumor protective in vitro and in vivo. The purpose of this study was to examine a putative synergism of genistein and calcitriol in two osteosarcoma cell lines MG-63 (early osteoblast), Saos-2 (mature osteoblast) and primary osteoblasts.MethodsThus, an initial screening based on cell cycle phase alterations, estrogen (ER) and vitamin D receptor (VDR) expression, live cell metabolic monitoring, and metabolomics were performed.ResultsExposure to the combination of 100 ?M genistein and 10 nM calcitriol reduced the number of proliferative cells to control levels, increased ERß and VDR expression, and reduced extracellular acidification (40%) as well as respiratory activity (70%), primarily in MG-63 cells. In order to identify the underlying cellular mechanisms in the MG-63 cell line, metabolic profiling via GC/MS technology was conducted. Combined treatment significantly influenced lipids and amino acids preferably, whereas metabolites of the energy metabolism were not altered. The comparative analysis of the log2-ratios revealed that after combined treatment only the metabolite ethanolamine was highly up-regulated. This is the result: a strong overexpression (350%) of the enzyme sphingosine-1-phosphate lyase (SGPL1), which irreversibly degrades sphingosine-1-phosphate (S1P), thereby, generating ethanolamine. S1P production and secretion is associated with an increased capability of migration and invasion of cancer cells.ConclusionFrom these results can be concluded that the tumor promoting effect of high concentrations of genistein in immature osteosarcoma cells is reduced by the co-administration of calcitriol, primarily by the breakdown of S1P. It should be tested whether this anti-metastatic pathway can be stimulated by combined treatment also in metastatic xenograft mice models.

Project description:BackgroundAberrant expression of Aldo-Keto reductase family 1 member B10 (AKR1B10) was associated with tumor size and metastasis of breast cancer in our published preliminary studies. However, little is known about the detailed function and underlying molecular mechanism of AKR1B10 in the pathological process of breast cancer.MethodsThe relationship between elevated AKR1B10 expression and the overall survival and disease-free survival of breast cancer patients was analyzed by Kaplan-Meier Plotter database. Breast cancer cell lines overexpressing AKR1B10 (MCF-7/AKR1B10) and breast cancer cell lines with knockdown of AKR1B10 (BT-20/shAKR1B10) were constructed to analyze the impact of AKR1B10 expression on cell proliferation and migration of breast cancer. The expression levels of AKR1B10 were detected and compared in the breast cancer cell lines and tissues by RT-qPCR, western blot and immunohistochemistry. The proliferation of breast cancer cells was monitored by CCK8 cell proliferation assay, and the migration and invasion of breast cancer cells was observed by cell scratch test and transwell assay. The proliferation- and EMT-related proteins including cyclinD1, c-myc, Survivin, Twist, SNAI1, SLUG, ZEB1, E-cadherin, PI3K, p-PI3K, AKT, p-AKT, IKBα, p-IKBα, NF-κB p65, p-NF-κB p65 were detected by western blot in breast cancer cells. MCF-7/AKR1B10 cells were treated with LY294002, a PI3K inhibitor, to consider the impact of AKR1B10 overexpression on the PI3K/AKT/NF-κB signal cascade and the presence of NF-κB p65 in nuclear. In vivo tumor xenograft experiments were used to observe the role of AKR1B10 in breast cancer growth in mice.ResultsAKR1B10 expression was significantly greater in breast cancer tissue compared to paired non-cancerous tissue. The expression of AKR1B10 positively correlated with lymph node metastasis, tumor size, Ki67 expression, and p53 expression, but inversely correlated with overall and disease-free survival rates. Gene Ontology analysis showed that AKR1B10 activity contributes to cell proliferation. Overexpression of AKR1B10 facilitated the proliferation of MCF-7 cells, and induced the migration and invasion of MCF-7 cells in vitro in association with induction of epithelial-mesenchymal transition (EMT). Conversely, knockdown of AKR1B10 inhibited these effects in BT-20 cells. Mechanistically, AKR1B10 activated PI3K, AKT, and NF-κB p65, and induced nuclear translocation of NF-κB p65, and expression of proliferation-related proteins including c-myc, cyclinD1, Survivin, and EMT-related proteins including ZEB1, SLUG, Twist, but downregulated E-cadherin expression in MCF-7 cells. AKR1B10 silencing reduced the phosphorylation of PI3K, AKT, and NF-κB p65, the nuclear translocation of NF-κB p65, and the expression of proliferation- and migration-related proteins in BT-20 cells. LY294002, a PI3K inhibitor, attenuated the phosphorylation of PI3K, AKT, and NF-κB p65, and the nuclear translocation of NF-κB p65. In vivo tumor xenograft experiments confirmed that AKR1B10 promoted breast cancer growth in mice.ConclusionsAKR1B10 promotes the proliferation, migration and invasion of breast cancer cells via the PI3K/AKT/NF-κB signaling pathway and represents a novel prognostic indicator as well as a potential therapeutic target in breast cancer.

Dataset Information

COX?2 promotes epithelial?mesenchymal transition and migration in osteosarcoma MG?63 cells via PI3K/AKT/NF??B signaling.

Publications

COX‑2 promotes epithelial‑mesenchymal transition and migration in osteosarcoma MG‑63 cells via PI3K/AKT/NF‑κB signaling.

Similar Datasets

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