β2-Adrenergic receptor agonist enhances the bystander effect of HSV-TK/GCV gene therapy in glioblastoma multiforme via upregulation of connexin 43 expression

Glioblastoma multiforme (GBM) is the most invasive form of primary brain astrocytoma. Gene therapy using the herpes simplex virus thymidine kinase/ganciclovir (HSV-TK/GCV) is a new strategy for GBM treatment. As the connexin 43 (Cx43) levels are downregulated in GBM cells, it seems that the upregulation of Cx43 could improve the efficacy of the gene therapy. This study aims to evaluate the effect of clenbuterol hydrochloride (Cln) as a β2-adrenergic receptor agonist on HSV-TK/GCV gene therapy efficacy in human GBM cells using olfactory ensheathing cells (OECs) as vectors. The lentivirus containing the thymidine kinase gene was transduced to OECs and the effective dose of GCV on cells was measured by MTT assay. We found that Cln upregulated Cx43 expression in human GBM cells and OECs and promoted the cytotoxic effect of GCV on the co-culture cells. Western blot results showed that Cln increased the cleaved caspase-3 expression and the Bax/Bcl2 ratio in the co-culture of GBM cells and OEC-TK. Also, the flow cytometry results revealed that Cln increased apoptosis in the co-culture of GBM cells and OEC-TK cells. This study showed that Cln via upregulation of Cx43 expression could enhance the bystander effect of HSVTK-GCV gene therapy in human GBM cells.