Abstract
Glioblastoma multiforme (GBM) is described as an invasive astrocytic tumor in adults. Despite current standard treatment approaches, the outcome of GBM remains unfavorable. The downregulation of connexin 43 (Cx43) expression is one of the molecular transformations in GBM cells. The Cx43 levels and subsequently gap junctional intercellular communication (GJIC) have an important role in the efficient transfer of cytotoxic drugs to whole tumor cells. As shown in our previous study, the stimulation of the β2-adrenergic receptor (β2-AR) leads to the modulation of Cx43 expression level in the GBM cell line. Here we further examine the effect of clenbuterol hydrochloride as a selective β2-AR agonist on the Cx43 expression in human GBM-derived astrocyte cells and human olfactory ensheathing cells (OECs) as a potent vector for future gene therapy. In this experiment, first we established a primary culture of astrocytes from GBM samples and verified the purity using immunocytofluorescent staining. Western blot analysis was performed to evaluate the Cx43 protein level. Our western blot findings reveal that clenbuterol hydrochloride upregulates the Cx43 protein level in both primary human astrocyte cells and human OECs. Conversely, ICI 118551 as a β2-AR antagonist inhibits these effects. Moreover, clenbuterol hydrochloride increases the Cx43 expression in primary human astrocyte cells and OECs co-culture systems, and ICI 118551 reverses these effects. To confirm the western blot results, immunocytofluorescent staining was performed to evaluate the β2-AR agonist effect on Cx43 expression. Our immunocytofluorescent results supported western blot analysis in primary human astrocyte cells and the OECs co-culture system. The results of this study suggest that the activation of β2-AR with regard to Cx43 protein levels enhancement in GBM cells and OECs might be a promising approach for GBM treatment in the future.
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Acknowledgments
This study was technically supported by the Brain and Spinal Cord Injury Research Center, Neuroscience Institute, Tehran University of Medical Sciences, Tehran, Iran. We are grateful to the Brain and Spinal Cord Injury Research Center, Neuroscience Institute, Tehran University of Medical Sciences, Tehran, Iran.
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Saereh Hosseindoost: Project administration, Investigation, writing the manuscript, analyzed the data. Shiva Hashemizadeh and Zeinab Gharaylou: performed experiments. Ahmad Reza Dehpour: Interpretation of data and review of the manuscript. Seyed Amir Hossein Javadi: Prepared the GBM samples and review of the manuscript. Babak Arjmand: Isolation and Preparation of OECs and review of the manuscript. Mahmoudreza Hadjighassem: Designed and directed the research and devised the main conceptual ideas. All authors read and approved the final manuscript.
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All experiments were performed according to the guidelines of Tehran University of Medical Sciences. Ethical approval for this study was obtained from the Tehran University of Medical Sciences ethics committee.
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Hosseindoost, S., Hashemizadeh, S., Gharaylou, Z. et al. β2-Adrenergic Receptor Stimulation Upregulates Cx43 Expression on Glioblastoma Multiforme and Olfactory Ensheathing Cells. J Mol Neurosci 70, 1451–1460 (2020). https://doi.org/10.1007/s12031-020-01542-7
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DOI: https://doi.org/10.1007/s12031-020-01542-7