Role of Hsp-70 in triptolide-mediated cell death of neuroblastoma

Mara B. Antonoff, Rohit Chugh, Steven J. Skube, Vikas Dudeja, Daniel Borja-Cacho, Kimberly A. Clawson, Selwyn M. Vickers, Ashok K. Saluja

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

Background: Our recent work demonstrated that treatment of neurobastoma with triptolide causes apoptotic cell death in vitro and decreases tumor size in vivo. Triptolide therapy has been associated with reduced expression of Hsp-70, suggesting a mechanism of cell killing involving Hsp-70 inhibition. The principal objective of this study was to investigate the role of Hsp-70 in triptolide-mediated cell death in neuroblastoma. Materials and Methods: Neuroblastoma cells were transfected with Hsp-70-specific siRNA. Viability, caspase activity, and phosphatidylserine externalization were subsequently measured. An orthotopic, syngeneic murine tumor model was developed, and randomized mice received daily injections of triptolide or vehicle. At 21 d, mice were sacrificed. Immunohistochemisty was used to characterize Hsp-70 levels in residual tumors, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) was performed to identify cells undergoing apoptosis. Results: Targeted silencing of Hsp-70 with siRNA significantly decreased cellular viability, augmented caspase-3 activity, and resulted in increased annexin-V staining. These effects parallel those findings obtained following treatment with triptolide. Residual tumors from triptolide-treated mice showed minimal staining with Hsp-70 immunohistochemistry, while control tumors stained prominently. Tumors from treated mice demonstrated marked staining with the TUNEL assay, while control tumors showed no evidence of apoptosis. Conclusions: Use of siRNA to suppress Hsp-70 expression in neuroblastoma resulted in apoptotic cell death, similar to the effects of triptolide. Residual tumors from triptolide-treated mice expressed decreased levels of Hsp-70 and demonstrated significant apoptosis. These findings support the hypothesis that Hsp-70 inhibition plays a significant role in triptolide-mediated neuroblastoma cell death.

Original languageEnglish (US)
Pages (from-to)72-78
Number of pages7
JournalJournal of Surgical Research
Volume163
Issue number1
DOIs
StatePublished - Sep 1 2010

Keywords

  • Hsp-70
  • apoptosis
  • neuroblastoma, heat shock protein
  • triptolide

ASJC Scopus subject areas

  • Surgery

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