Liau L M, Fakhrai H, Black K L
Division of Neurosurgery, UCLA School of Medicine 90095, USA.
Neurol Res. 1998 Dec;20(8):742-7. doi: 10.1080/01616412.1998.11740594.
Using an intracranial rat C6 glioma model, we tested the hypothesis that gene modification of glioma cells to block the expression of the immunosuppressive cytokine TGF-beta (transforming growth factor beta) may enhance anti-tumor immune responses and thereby prolong survival of tumor-bearing animals. The cDNA for simian TGF-beta 2 was ligated in antisense orientation into the episomal plasmid mammalian expression vector pCEP-4. This TGF-beta-antisense vector was transfected into C6 glioma cells by standard electroporation techniques. PCR was used to determine that the rat C6 clones were successfully transfected with the antisense-TGF beta construct. Twenty-nine adult female Wistar rats harboring 7-day-old intracranial C6 tumors were then subcutaneously injected with either saline (n = 9), unmodified C6 glioma cells (n = 10), or TGF-beta-antisense-modified C6 cells (n = 10). Animals were followed for survival, and Fisher's exact method was used to interpret the significance of difference between experimental groups. The survival of tumor-bearing rats injected with TGF-beta-antisense-modified C6 cells was significantly prolonged, relative to the survival of rats receiving injections of saline or unmodified C6 cells alone. Six of the ten (60%) TGF-beta-antisense treated animals survived for 12 weeks, whereas none of the nine (0%) animals treated with saline and none of ten (0%) of those treated with C6 cells alone survived past 5 weeks. These results indicate that the genetic inhibition of immunosuppressive cytokines (such as TGF-beta) may reverse the phenotypic immunosuppression caused by such factors, and thereby prolong the survival of C6 tumor-bearing animals. Future investigations using cytokine gene modifications in other brain tumor models are warranted.
利用颅内大鼠C6胶质瘤模型，我们检验了以下假设：对胶质瘤细胞进行基因改造以阻断免疫抑制细胞因子转化生长因子β（TGF-β）的表达，可能增强抗肿瘤免疫反应，从而延长荷瘤动物的生存期。将猿猴TGF-β2的cDNA以反义方向连接到游离质粒哺乳动物表达载体pCEP-4中。通过标准电穿孔技术将该TGF-β反义载体转染到C6胶质瘤细胞中。采用聚合酶链反应（PCR）确定大鼠C6克隆已成功用反义-TGF-β构建体转染。然后，对29只患有7日龄颅内C6肿瘤的成年雌性Wistar大鼠皮下注射生理盐水（n = 9）、未修饰的C6胶质瘤细胞（n = 10）或TGF-β反义修饰的C6细胞（n = 10）。观察动物的生存期，并采用Fisher精确检验法分析各实验组之间差异的显著性。相对于单独注射生理盐水或未修饰C6细胞的大鼠，注射TGF-β反义修饰C6细胞的荷瘤大鼠生存期显著延长。10只接受TGF-β反义治疗的动物中有6只（60%）存活了12周，而9只接受生理盐水治疗的动物中无一存活超过5周，10只单独接受C6细胞治疗的动物中也无一存活超过5周。这些结果表明，对免疫抑制细胞因子（如TGF-β）进行基因抑制可能逆转此类因子引起的表型免疫抑制，从而延长荷C6肿瘤动物的生存期。有必要在其他脑肿瘤模型中开展利用细胞因子基因改造的进一步研究。
