Astrocytes promote glioma invasion via the gap junction protein connexin43

Abstract

Reactive astrocytes are integral to the glioma microenvironment. Connexin43 (Cx43) is a major gap junction protein in astrocytes and its expression is enhanced significantly in glioma-associated astrocytes, especially at the peri-tumoral region. Although downregulation of Cx43-mediated intercellular communication is associated with increased malignancy in tumor cells, the role of Cx43 in stromal cells in glioma progression is not defined. Using a mouse model consisting of syngeneic intracranial implantation of GL261 glioma cells into Nestin-Cre:Cx43(fl/fl) mice where Cx43 was eliminated in astrocytes, we demonstrate a role of astrocytic Cx43 in the dissemination of glioma cells from the tumor core. To determine whether heterocellular communication between astrocytes and glioma cells is essential for reduced invasion in the absence of astrocytic Cx43, we abolished channel formation between glioma cells and astrocytes by either knocking down Cx43 in glioma cells with short hairpin RNA (shRNA) or overexpressing a dominant-negative channel-defective Cx43-T154A mutant in these cells. Although Cx43shRNA in glioma cells reduced invasion, expression of Cx43-T154A had no effect on glioma invasion, suggesting tumoral Cx43 may influence motility independently from its channel function. Alteration in astrocytic Cx43 function, such as by replacing the wild-type allele with a C-terminal truncated Cx43 mutant exhibiting reduced intercellular coupling, is sufficient to reduce glioma spreading into the brain parenchyma. Our results reveal a novel role of astrocytic Cx43 in the formation of an invasive niche and raise the possibility to control glioma progression by manipulating the microenvironment.

Novel approach to temozolomide resistance in malignant glioma: connexin43-directed therapeutics

“…Expression of Cx43 is significantly enhanced in tumor-associated astrocytes especially at the region adjacent to the tumor core [11 •• ]. Recent reports suggest that astrocytic Cx43 may contribute to TMZ resistance in addition to enhancing GBM cell proliferation and migration.…”

“…In a co-culture system of human astrocytes and tumor cells designed to simulate the tumor microenvironment, siRNA knockdown of astrocytic Cx43 abolished GJIC between glioma cells and astrocyte and increased TMZ-induced apoptosis of GBM cells [42]. Furthermore, in a mouse model consisting of syngeneic intracranial implantation of GL261 malignant glioma cells into Nestin-Cre:Cx43 fl/fl mice, in which Cx43 is selectively eliminated in astrocytes, it was demonstrated that reduction of astrocytic Cx43 decreases the invasion of tumor cells from the tumor core in the presence of an intact immune system [11 •• ]. The importance of astrocytic Cx43 in the regulation of GBM resistance and in tumor invasiveness strongly supports the manipulation of Cx43 in the GBM microenvironment as a promising therapeutic strategy in controlling glioma.…”

“…An added layer of complexity is afforded through channel-independent mechanisms via the stabilization or destabilization of multiprotein complexes mediated through protein binding sites on the connexin protein. Connexin43 (Cx43) is the most ubiquitous connexin and in addition to having critical regulatory and developmental roles in many tissues, has been implicated as a driver of tumor invasion [11 •• ], a marker of tumor progression [12], and an inducer of TMZ resistance in GBM cells [6 •• ,13–15,16 •• ]. Furthermore, the expression and function of connexins in the microenvironment of malignant glioma is an evolving field, where recent studies investigating the function of Cx43 in the tumor microenvironment reveal that Cx43 in peritumoral cells regulates gliomagenesis, tumor progression, and treatment resistance.…”

The Role of Astrocytes in Tumor Growth and Progression

“…Cx43 is widely expressed in adult astrocytes and exhibits increased expression in reactive astrocytes induced by various brain pathologies and intercellular calcium signaling [67][68][69][70][71][72][73]. Also, Cx43-mediated intercellular communication between astrocytes plays an important role in the invasion of glioma cells in the brain [63].…”

“…The proliferative dysfunction and invasion of gliomas are associated with changes in gap junction communication [62,63]. In metastatic brain tumors, reactive astrocytes protect melanoma cells from chemotherapy induced cell death by sequestering intracellular calcium through gap junctions [64].…”