阳离子脂质体介导HSV-TK基因治疗人脑胶质瘤的实验研究

HSV - TK Gene Therapy for Human Glioma Mediated by Cationic Liposome

颅内恶性肿瘤以胶质瘤最为多见,胶质瘤的预后较差,一直困扰着神经外科医师。本文旨在观察阳离子脂质体介导的HSV-TK/ACV系统对人脑胶质瘤细胞的杀伤作用。方法:本文将新近克隆并构建的含HSV-TK基因真核表达载体用阳离子脂质体介导转染至人脑胶质瘤细胞中,筛选出阳性克隆,通过MTT、FCM及AgNOR检测转染了HSV-TK基因的FJ905胶质瘤细胞对不同浓度的ACV的反应情况。结果:转染了TK基因的胶质瘤细胞在给予ACV后,细胞发生明显的肿胀、变性,以致形成碎片,而未转染TK基因的胶质瘤细胞变化不明显。转染TK基因和未转染TK的肿瘤细胞经ACV作用后,DNA的含量分别为65.81和109.43,两者之间差异显著(P<0.05)。AgNOR嗜银染色测定结果,转染了TK基因和未转染TK基因的肿瘤细胞经ACV作用后,两者测定结果分别为6.67±1.03和14.83±1.47,两者之间差异极显著(P<0.01)。结论:转染了TK基因的胶质瘤细胞对ACV的敏感性明显提高,ACV能明显抑制转染TK基因细胞的DNA合成;HSV-TK/ACV治疗后,人脑胶质瘤细胞的增殖活性降低了。提示HSV-TK/ACV系统可为胶质瘤的基因治疗提供一种可探索的方法。

BACKGROUND & OBJECTIVE: Glioma is the most common primary malignant brain tumors in human. Neurosurgeons feel frustrated by the poor prognosis associated with glioma. The aim of this study is to assess the mechanism and effectiveness of HSV-TK/ACV system in killing human glioma transferred with cationic liposome. METH-ODS: The constructed eukaryotic expressing vector pCR3-TK containing the TK gene was transferred to TJ905 glioma cells by using cationic liposome. The positive clones containing TK gene were selected. The killing effects of various concen-tration of ACV on TJ905 glioma cells transfected with HSV-TK were examined using MTT, FCM and AgNOR. RE-SULTS: Swelling, degeneration or fragmentation was observed in the TJ905 glioma cells tranfected with TK gene af-ter administration of ACV. These changes were not seen in TJ905 cells without TK gene transfection. The DNA concentration index of glioma cells with and with-out TK gene transfection were 65.81 and 109.43 as determined by FCM, respectively (P <0. 05=. The AgNOR granules of these two kinds of cells were 6.67±1.03 and 14.83± 1.47 respectively ( P<0.01=. CONCLU-SIONS: The TK gene-transfected glioma cells were significantly more sensitive to ACV than those without TK gene-transfection. FCM indicated that ACV could suppress the formation of DNA in TK gene transfect-ed glioma cells. The proliferation activity of TK gene transfected glioma cells were found to be suppressed as showed with silver stained AgNOR. This indicated that HSV-TK/ACV gene therapy may be further explored us an innovative method for glioma therapy.