Effects of Connexin43 Overexpression on U251 Cell Growth, Migration, and Apoptosis

Lu, Jin; Yu, Mingyue; Zhang, Lin; Lue, Su; Zhang, Hao; Zhao, Hang; Xu, Yanyan; Liu, Haiyan

doi:10.12659/msm.905130

Cited by 7 publications

(7 citation statements)

References 13 publications

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“…24 The U251 glioblastoma cell line has been selected due to its endogenous expression of functional Cx43 GJs. 25,26 The assay was validated using previously published GJ inhibitors such as MEF, quinidine, or niflumic acid, 17,27,39,40 whereas carbenoxolone did not display any inhibition in this setup. Those drugs present different mechanisms of action, as for instance carbenoxolone acts by modulating protein kinase Adependent pathway 41 and MEF through interference with loop gating.…”

Section: Discussionmentioning

confidence: 99%

“…24 U251 glioblastoma cells were selected due to their expression of functional Cx43 GJs. 25,26 Screening of the Prestwick Chemical Library Ò (PCL; 1,280 compounds) that mostly contains approved drugs (or compounds under preclinical and clinical development) led to the identification of new classes of Cx43 inhibitors. Some of those newly identified inhibitors were previously described as modulators of microtubules dynamics strongly suggesting that GJs function requires a proper microtubule cytoskeleton network.…”

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confidence: 99%

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High-Content Screening Identifies New Inhibitors of Connexin 43 Gap Junctions

Picoli

Soleilhac

Journet

et al. 2019

ASSAY and Drug Development Technologies

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Gap junctions (GJs) are dynamic structures composed of hexamers of connexins (Cxs), a class of transmembrane proteins enabling channel-mediated direct intercellular communication through cell-cell diffusion of ions and small metabolites. In defined conditions, Cxs also work as hemichannels allowing exchanges between the cytoplasm and the extracellular medium. The most common GJ channel is formed by connexin 43 (Cx43) and plays an important role in physiological and pathological processes in excitable tissues, such as heart and brain. Hence, Cx43 has been largely envisioned as a new therapeutic target in cancer, neurological and psychiatric indications, or cardiovascular diseases. Identifying new pharmacological inhibitors of Cx43 GJs with different mechanisms of action and from diverse chemical classes is thus highly challenging. We present here a high-content screening method, based on the evaluation of fluorescent dye transfer rates between adjacent cells to monitor the function of GJs in U251 glioblastoma cells expressing high levels of Cx43. This assay was validated using well-described pharmacological GJ inhibitors such as mefloquine. The method was adapted to screen a library of 1,280 Food and Drug Administration-and European Medicines Agency-approved drugs that led to the selection of both known and new inhibitors of GJ channel function. We further focused on a specific class of microtubule-targeting agents, confirming that a proper tubulin network is required for functional Cx43 GJ channels.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

High-Content Screening Identifies New Inhibitors of Connexin 43 Gap Junctions

Picoli

Soleilhac

Journet

et al. 2019

ASSAY and Drug Development Technologies

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“…It was previously demonstrated that Cx43 reduces the sensitivity of human LN18 and LN229 glioma cells to TMZ by reducing the Bax/Bcl-2 ratio and the release of cytochrome c ( 29 ). Overexpression of Cx43 promotes the migration of U251 cells and inhibits apoptosis by upregulating Bcl-2 and downregulating the expression of Bax and caspase-3 ( 46 ). Thus, the effect of rExo on the expression of Bcl-2, Bax and cleaved caspase-3 in U251s cells was examined.…”

Section: Discussionmentioning

confidence: 99%

“…There are at least two plausible explanations for the role of exosomal Cx43 in the resistance of glioma cells to TMZ. First, the exosomal Cx43 may enhance TMZ resistance through modulating the expression of the pro-apoptotic and anti-apoptotic proteins and inhibiting the intrinsic apoptosis pathway ( 29 , 46 ). Second, high expression of Cx43 in rExo may contribute to the formation of gap junction channels between exosomes and recipient cells, which facilitates transferring various TMZ resistance-related factors from exosomes to U251s cells.…”

Section: Discussionmentioning

confidence: 99%

Exosomal connexin 43 regulates the resistance of glioma cells to temozolomide

Yang

Zhang

et al. 2021

Oncol Rep

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Glioblastoma is the most common and aggressive brain tumor and it is characterized by a high mortality rate. Temozolomide (TMZ) is an effective chemotherapy drug for glioblastoma, but the resistance to TMZ has come to represent a major clinical problem, and its underlying mechanism has yet to be elucidated. In the present study, the role of exosomal connexin 43 (Cx43) in the resistance of glioma cells to TMZ and cell migration was investigated. First, higher expression levels of Cx43 were detected in TMZ-resistant U251 (U251r) cells compared with those in TMZ-sensitive (U251s) cells. Exosomes from U251s or U251r cells (sExo and rExo, respectively) were isolated. It was found that the expression of Cx43 in rExo was notably higher compared with that in sExo, whereas treatment with rExo increased the expression of Cx43 in U251s cells. Additionally, exosomes stained with dioctadecyloxacarbocyanine (Dio) were used to visualized exosome uptake by glioma cells. It was observed that the uptake of Dio-stained rExo in U251s cells was more prominent compared with that of Dio-stained sExo, while 37,43 Gap27, a gap junction mimetic peptide directed against Cx43, alleviated the rExo uptake by cells. Moreover, rExo increased the IC 50 of U251s to TMZ, colony formation and Bcl-2 expression, but decreased Bax and cleaved caspase-3 expression in U251s cells. 37,43 Gap27 efficiently inhibited these effects of rExo on U251s cells. Finally, the results of the wound healing and Transwell assays revealed that rExo significantly enhanced the migration of U251s cells, whereas 37,43 Gap27 significantly attenuated rExo-induced cell migration. Taken together, these results indicate the crucial role of exosomal Cx43 in chemotherapy resistance and migration of glioma cells, and suggest that Cx43 may hold promise as a therapeutic target for glioblastoma in the future.

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“…U118MG, a WHO grade IV human GBM cell line, derived from highly malignant GBM tumors. U251 glioma cells are derived from a grade II neuroblastoma (39–41). The morphology of U251 cells are epithelial, while the morphology of U118MG cells are mixed and both glioblastoma and astrocytoma cells are present (42, 43).…”

Section: Discussionmentioning

confidence: 99%

CircPCMTD1 Acts as the Sponge of miR-224-5p to Promote Glioma Progression

Zheng

et al. 2019

Front. Oncol.

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Glioma is the most common malignant tumor of the central nervous system with high morbidity and mortality. Circular RNAs (circRNAs) are abundant non-coding RNAs, which contribute to tumor progression by competing with other endogenous RNAs such as microRNA (miRNA). MiRNA are a class of small non-coding RNAs, which interrupt the translation of target mRNAs. CircPCMTD1 (hsa-circ-0001801) is a newly discovered circRNA that was found to be significantly upregulated in glioma. However, its function is unclear. In this study, circPCMTD1 upregulation promoted the cell viability, migration and invasion dramatically, while the inhibition of circPCMTD1 led to a significant reduction of tumor growth in vivo . MiRNAs microarray analyses on circPCMTD1 silencing models in U251 and U118MG cells were performed, and the results suggested that circPCMTD1 knockdown could upregulate the expression of miR-224-5p and downregulate the expression of mTOR, one of miR-224-5p targets, in both cell lines. According to the prediction from circular RNA interactome and Targetscan, there was a complementary sequence in circPCMTD1 for miR-224-5p. Dual-luciferase reporter assay demonstrated that circPCMTD1 were targets of miR-224-5p. RIP assay was also performed to further confirm their directly interaction. Overexpression of miR-224-5p inhibited the viability and proliferation, migration, and invasion of U251 and U118MG glioma cells. In conclusion, circPCMTD1 could contribute to the promotion of glioma progression, and it may serve as the sponge of miR-224-5p to exert its function.

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Effects of Connexin43 Overexpression on U251 Cell Growth, Migration, and Apoptosis

Cited by 7 publications

References 13 publications

High-Content Screening Identifies New Inhibitors of Connexin 43 Gap Junctions

High-Content Screening Identifies New Inhibitors of Connexin 43 Gap Junctions

Exosomal connexin 43 regulates the resistance of glioma cells to temozolomide

CircPCMTD1 Acts as the Sponge of miR-224-5p to Promote Glioma Progression

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