The effect of chemotherapy and Wilms' Tumor 1 silencing on glioblastoma cell death in-vitro.

摘要

Glioblastoma multiform, the most common type of primary brain tumor, is incurable and progresses rapidly. In terms of extending patient survival, progress over the last few decades has been measured in months. Further improvement in neurosurgical techniques is not expected to result in a cure. The remaining frontier, where heavy expectations lie, is molecular medicine.; One gene target of particular interest is Wilms' Tumor 1 (WT1). It is expressed in the majority of glioblastomas and absent from normal brain. WT1 inhibition reportedly causes glioblastoma apoptosis. Initially, we hypothesized that in glioblastoma cell lines, silencing WT1 would decrease expression of the WT1 transcriptional target Bcl-2, thereby enhancing apoptosis and chemosensitivity.; WT1 was silenced using RNA interference. This decreased survival and chemoresistance to BCNU and cisplatin, but not through the modulation of Bcl-2 mRNA. In fact, the mechanism of WT1 mediated cell death was not apoptosis. We then examined other causes for decreased survival including cell cycle arrest, senescence and autophagy. Our results indicated that different mechanisms account for cell death caused by WT1 downregulation and chemotherapy treatment.; WT1 silencing was found to have minimal influence on DNA damage, cell cycle arrest, senescence, apoptosis and autophagy. Interestingly, WT1 silencing was associated with IGF-1R upregulation. It may seem paradoxical that IGF-1R upregulation decreased cell survival. However, the role of IGF-1R in a cell is complex, and increased IGF-1R activity is linked with brain differentiation and a newly described form of programmed cell death termed "paraptosis."; Surprisingly, treatment with BCNU or cisplatin induced a striking amount of autophagy, a result not previously reported with any cell type. Both drugs also dramatically increased the activity of DNA damage repair proteins. Cisplatin, not BCNU, significantly increased DNA fragmentation. Neither agent caused remarkable senescence.; In conclusion, WT1 silencing and chemotherapeutic agents synergistically decreased cell viability through distinct cell death programs. The mechanism by which WT1 diminishes survival in unclear, but it maybe related to IGF-1R. BCNU and cisplatin activated autophagy which we suspect is an underappreciated form of cell death in glioblastomas.