title: Early Reporting of Apoptosis by Real-time Imaging of Cancer Cells Labeled with Green Fluorescent Protein in the Nucleus and Red Fluorescent Protein in the Cytoplasm.
abstract: OBJECTIVE
  We previously developed PC-3 human prostate cancer cells expressing red fluorescent protein (RFP) in the cytoplasm and green fluorescent protein (GFP) linked to histone H2B expressed in the nucleus. We demonstrate in the present report the use of these dual-color cells for early detection of apoptosis in the presence of cancer chemotherapy agents.
  
  
  METHODS
  Induction of apoptosis was observed by real-time imaging of cytoplasmic and nuclear size and shape changes and nuclear fragmentation using fluorescence microscopy. Apoptosis was also detected by measuring DNA fragmentation. The cancer chemotherapy agents paclitaxel and vinblastine were used for induction of apoptosis.
  
  
  RESULTS
  When the PC-3 dual-color cells were treated with paclitaxel or vinblastine, cytoplasmic and nuclear size and shape changes and nuclear fragmentation were observed by 24 hours. The paclitaxel-treated PC-3 dual-color cells exhibited ring-like structures formed by the fragmented nuclei, which could be brightly visualized by H2B-GFP fluorescence. Apoptosis was also detected by the dual-color PC-3 cells by 24 hours when treated with vinblastine. However, no nuclear ring-like structures were formed in the PC-3 cells by vinblastine treatment. In contrast, DNA fragmentation could not be observed in PC-3 cells until 48 hours after exposure to paclitaxel.
  
  
  CONCLUSIONS
  Dual-color PC-3 cells can serve as a simple real-time early reporter of apoptosis and as a screen for novel cancer therapeutics or genotoxic agents. The dual-color cell real-time imaging assay is a more sensitive and earlier reporter for apoptosis than the DNA fragmentation assay.
authors.affiliation: AntiCancer, Inc., San Diego, CA, U.S.A. AntiCancer Biotech (Beijing) Co., Ltd., Beijing, P.R.China.
authors.forename: Meng
authors.initials: M
authors.lastname: Yang
authors.affiliation: AntiCancer, Inc., San Diego, CA, U.S.A.
authors.forename: Ping
authors.initials: P
authors.lastname: Jiang
authors.affiliation: AntiCancer, Inc., San Diego, CA, U.S.A. Department of Surgery, University of California, San Diego, CA, U.S.A. all@anticancer.com.
authors.forename: Robert M
authors.initials: RM
authors.lastname: Hoffman
mesh_terms: D017209:Apoptosis / Q000235:genetics*; D045744:Cell Line, Tumor; D058948:Cell Tracking / Q000379:methods*; D003593:Cytoplasm / Q000187:drug effects / Q000473:pathology; D053938:DNA Fragmentation / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics; D006801:Humans; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics; D008297:Male; D008856:Microscopy, Fluorescence; D057054:Molecular Imaging; D017239:Paclitaxel / Q000008:administration & dosage; D011471:Prostatic Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000473:pathology*; D000097573:Red Fluorescent Protein
chemical_list: D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D017239:Paclitaxel; D000097573:Red Fluorescent Protein
keywords: DNA fragmentation; GFP; RFP; apoptosis; fusion gene; histone H2B; imaging; nuclear fragmentation; paclitaxel; real-time; vinblastine