Bcl-2 siRNA augments taxol mediated apoptotic death in human glioblastoma U138MG and U251MG cells

Neurochem Res. 2009 Jan;34(1):66-78. doi: 10.1007/s11064-008-9659-z. Epub 2008 Mar 21.

Abstract

The anti-neoplastic drug taxol binds to beta-tubulin to prevent tumor cell division, promoting cell death. However, high dose taxol treatment may induce cell death in normal cells too. The anti-apoptotic molecule Bcl-2 is upregulated in many cancer cells to protect them from apoptosis. In the current study, we knocked down Bcl-2 expression using cognate siRNA during low-dose taxol treatment to induce apoptosis in two human glioblastoma U138MG and U251MG cell lines. The cells were treated with either 100 nM taxol or 100 nM Bcl-2 siRNA or both for 72 h. Immunofluorescent stainings for calpain and active caspase-3 showed increases in expression and co-localization of these proteases in apoptotic cells. Fluorometric assays demonstrated increases in intracellular free [Ca(2+)], calpain, and caspase-3 indicating augmentation of apoptosis. Western blotting demonstrated dramatic increases in the levels of Bax, Bak, tBid, active caspases, DNA fragmentation factor-40 (DFF40), cleaved fragments of lamin, fodrin, and poly(ADP-ribose) polymerase (PARP) during apoptosis. The events related to apoptosis were prominent more in combination therapy than in either treatment alone. Our current study demonstrated that Bcl-2 siRNA significantly augmented taxol mediated apoptosis in different human glioblastoma cells through induction of calpain and caspase proteolytic activities. Thus, combination of taxol and Bcl-2 siRNA offers a novel therapeutic strategy for controlling the malignant growth of human glioblastoma cells.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Apoptosis / drug effects*
  • Apoptosis / physiology*
  • BH3 Interacting Domain Death Agonist Protein / metabolism
  • Calcium / metabolism
  • Calpain / genetics
  • Caspase 3 / genetics
  • Cell Line, Tumor
  • Cytochromes c / metabolism
  • Dose-Response Relationship, Drug
  • Down-Regulation
  • Fluorescent Antibody Technique
  • Glioblastoma / drug therapy
  • Glioblastoma / pathology*
  • Humans
  • In Situ Nick-End Labeling
  • Paclitaxel / pharmacology*
  • Paclitaxel / therapeutic use
  • Proto-Oncogene Proteins c-bcl-2 / genetics*
  • RNA, Small Interfering / pharmacology*
  • RNA, Small Interfering / therapeutic use

Substances

  • BH3 Interacting Domain Death Agonist Protein
  • BID protein, human
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Small Interfering
  • Cytochromes c
  • Calpain
  • Caspase 3
  • Paclitaxel
  • Calcium