Zhang Wei, DeMattia Joseph A, Song Hua, Couldwell William T
Department of Neurosurgery, New York Medical College, Valhalla, New York, USA.
J Neurosurg. 2003 Apr;98(4):846-53. doi: 10.3171/jns.2003.98.4.0846.
Extensive invasion and angiogenesis are hallmark features of malignant gliomas. Communication between malignant glioma cells and surrounding astrocytes occurs, resulting in transformation of the astrocytic phenotype. In the present study, the authors examined whether malignant glioma cells and vascular endothelial cells (VECs) communicate through the formation of gap junctions and whether this communication influences angiogenesis.
Connexin43 (Cx43), a gap junction protein expressed in glioma cells, was identified in human umbilical VECs (HUVECs). Immunocytochemical staining for Cx43 demonstrated immunoreactive plaques at areas of cell-cell contact among HUVECs as well as between HUVECs and Cx43-expressing malignant glioma cells. Dye transfer, performed using the gap junction-permeable dye dicarboxy-dichlorofluorescein diacetate (CDCF), among these cocultures indicated that these were functional communications. Calcium signaling also occurred from malignant glioma cells to HUVECs. Tube formation by HUVECs cocultured with Cx43-transfected T98G malignant glioma cells (T98G-Cx43 cells) or with U87MG malignant glioma cells, which naturally express Cx43, was significantly increased compared with tube formation by HUVECs alone. The difference in tube formation by HUVECs cocultured with empty vector-transfected T98G glioma cells (T98G-mock cells) or with Cx43-deficient U373MG malignant glioma cells and tube formation by HUVECs alone was not statistically significant. Furthermore, the concentration of vascular endothelial growth factor (VEGF), an angiogenic factor important for the induction of angiogenesis and blood vessel formation, was significantly higher in medium harvested from cultures of T98G-Cx43 cells than in that harvested from cultures of control T98G-mock cells. Human malignant glioma U87MG cells also secreted increased concentrations of VEGF as compared with HUVECs alone. Nevertheless, there was no statistically significant difference in tube formation by HUVECs cultured in medium conditioned by either Cx43-expressing or Cx43-deficient glioma cells, suggesting that the direct gap junction communication between glioma cells and HUVECs may play a much more significant role than the increased VEGF secretion in vascular tube formation in this assay.
These results indicate that functional gap junction formation between human malignant glioma cells and VECs occurs. This communication appears to influence tumor angiogenesis. Targeting gap junction signaling may offer a potential mechanism for therapy in patients with these tumors.
广泛侵袭和血管生成是恶性胶质瘤的标志性特征。恶性胶质瘤细胞与周围星形胶质细胞之间存在通讯,导致星形胶质细胞表型发生转化。在本研究中,作者检测了恶性胶质瘤细胞与血管内皮细胞(VECs)是否通过形成缝隙连接进行通讯,以及这种通讯是否影响血管生成。
在人脐静脉内皮细胞(HUVECs)中鉴定出缝隙连接蛋白连接蛋白43(Cx43),其在胶质瘤细胞中表达。对Cx43进行免疫细胞化学染色显示,在HUVECs之间以及HUVECs与表达Cx43的恶性胶质瘤细胞之间的细胞 - 细胞接触区域存在免疫反应性斑块。在这些共培养物中使用缝隙连接可通透染料二羧基 - 二氯荧光素二乙酸酯(CDCF)进行的染料转移表明这些是功能性通讯。钙信号也从恶性胶质瘤细胞传递至HUVECs。与单独培养的HUVECs相比,与转染Cx43的T98G恶性胶质瘤细胞(T98G - Cx43细胞)或天然表达Cx43的U87MG恶性胶质瘤细胞共培养的HUVECs形成的管显著增加。与单独培养的HUVECs相比,与空载体转染的T98G胶质瘤细胞(T98G - mock细胞)或Cx43缺陷型U373MG恶性胶质瘤细胞共培养的HUVECs形成的管之间的差异无统计学意义。此外,血管内皮生长因子(VEGF)是诱导血管生成和血管形成的重要血管生成因子,从T98G - Cx43细胞培养物中收获的培养基中VEGF的浓度明显高于从对照T98G - mock细胞培养物中收获的培养基中的浓度。与单独的HUVECs相比,人恶性胶质瘤U87MG细胞也分泌更高浓度的VEGF。然而,在由表达Cx43或Cx43缺陷型胶质瘤细胞条件培养基培养的HUVECs形成的管之间没有统计学上的显著差异,这表明在该实验中,胶质瘤细胞与HUVECs之间的直接缝隙连接通讯在血管管形成中可能比增加的VEGF分泌起更重要的作用。
这些结果表明人恶性胶质瘤细胞与VECs之间形成了功能性缝隙连接。这种通讯似乎影响肿瘤血管生成。靶向缝隙连接信号传导可能为这些肿瘤患者提供一种潜在的治疗机制。