SBDS‐deficiency results in deregulation of reactive oxygen species leading to increased cell death and decreased cell growth

C Ambekar, B Das, H Yeger, Y Dror - Pediatric blood & cancer, 2010 - Wiley Online Library
C Ambekar, B Das, H Yeger, Y Dror
Pediatric blood & cancer, 2010Wiley Online Library
Abstract Background Shwachman–Diamond syndrome (SDS) is characterized by reduced
hematopoietic and exocrine pancreatic cell numbers and a marked propensity for leukemia.
Most patients have mutations in the SBDS gene. We previously reported that SBDS‐
deficient cells overexpress Fas, undergo accelerated spontaneous and Fas‐mediated
apoptosis and grow slowly. However the mechanism of how SBDS regulates apoptosis
remains unknown. Several studies have shown that reactive oxygen species (ROS) regulate …
Background
Shwachman–Diamond syndrome (SDS) is characterized by reduced hematopoietic and exocrine pancreatic cell numbers and a marked propensity for leukemia. Most patients have mutations in the SBDS gene. We previously reported that SBDS‐deficient cells overexpress Fas, undergo accelerated spontaneous and Fas‐mediated apoptosis and grow slowly. However the mechanism of how SBDS regulates apoptosis remains unknown. Several studies have shown that reactive oxygen species (ROS) regulate cell growth and spontaneous and Fas‐mediated cell death. Therefore, we hypothesized that SBDS‐deficiency disrupts ROS regulation and subsequently increases sensitivity to Fas stimulation and reduced cell growth.
Procedure
SBDS was knocked down in HeLa cervical cancer cells and TF‐1 myeloid cells using short hairpin RNA. ROS levels were evaluated by oxidation of 2′,7′‐dichlorodihydrofluorescein diacetate. Apoptosis and cell growth were evaluated with and without antioxidants by annexin V/propidium iodide and 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assays, respectively.
Results
We found that shRNA mediated SBDS‐knockdown resulted in a significant increase in ROS levels compared to control cells. Fas stimulation further increased ROS levels in the SBDS‐knockdown HeLa cells more than in the controls. Importantly, balancing ROS levels by antioxidants rescued SBDS‐deficient cells from spontaneous and Fas‐mediated apoptosis and reduced cell growth.
Conclusions
ROS levels are increased in SBDS‐deficient cells, which leads to increased apoptosis and decreased cell growth. Increased baseline and Fas‐mediated ROS levels in SBDS‐deficient cells can enhance the sensitivity to Fas stimulation. By balancing ROS levels, antioxidants can improve cell growth and survival in SBDS‐deficient cells. Pediatr Blood Cancer. 2010;55:1138–1144. © 2010 Wiley‐Liss, Inc.
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