Investigation of adaptive responses in bystander cells in 3D Cultures containing tritium-labeled and unlabeled normal human fibroblasts

Massimo Pinto, Edouard I. Azzam, Roger W. Howell

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Abstract

The study of radiation-induced bystander effects in normal human cells maintained in three-dimensional (3D) architecture provides more in vivo-like conditions and is relevant to human risk assessment. Linear energy transfer, dose and dose rate have been considered as critical factors in propagating radiation-induced effects. This investigation uses an in vitro 3D tissue culture model in which normal AG1522 human fibroblasts are grown in a carbon scaffold to investigate induction of a G1 arrest in bystander cells that neighbor radiolabeled cells. Cell cultures were co-pulse-labeled with 3Hdeoxycytidine (3HdC) to selectively irradiate a minor fraction of cells with 15 keV/m β particles and bromodeoxyuridine (BrdU) to identify the radiolabeled cells using immunofluorescence. The induction of a G1 arrest was measured specifically in unlabeled cells (i.e. bystander cells) using a flow cytometry-based version of the cumulative labeling index assay. To investigate the relationship between bystander effects and adaptive responses, cells were challenged with an acute 4 Gy γ-radiation dose after they had been kept under the bystander conditions described above for several hours, and the regulation of the radiation-induced G1 arrest was measured selectively in bystander cells. When the average dose rate in 3HdC-labeled cells (<16 of population) was 0.040.37 Gy/h (average accumulated dose 0.1410 Gy), no statistically significant stressful bystander effects or adaptive bystander effects were observed as measured by magnitude of the G1 arrest, micronucleus formation, or changes in mitochondrial membrane potential. Higher dose rates and/or higher LET may be required to observe stressful bystander effects in this experimental system, whereas lower dose rates and challenge doses may be required to detect adaptive bystander responses. © 2010 by Radiation Research Society.
Original languageEnglish
Pages (from-to)216 - 227
Number of pages12
JournalRadiation Research
Volume174
Issue number2
DOIs
Publication statusPublished - 2010
Externally publishedYes

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All Science Journal Classification (ASJC) codes

  • Radiology Nuclear Medicine and imaging
  • Biophysics
  • Radiation
  • Medicine(all)

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