n Medical Technology SA - The leukocyte apoptosis assay : a clinical predictor of radiosensitivity

Volume 19, Issue 1
  • ISSN : 1011-5528



Approximately 6% of patients receiving radiotherapy show signs of late toxicity in normal tissue many months after treatment has been completed. The Leukocyte Apoptosis Assay (LAA) has been developed to predict intrinsic radiosensitivity of normal tissue based on the radiation-induced cytotoxic response of CD4 and CD8 lymphocytes. Apoptosis, or programmed cell death, can be observed after radiation exposure and can be evaluated using flow cytometry. The purpose of this study was to determine the reliability of the LAA assay and to standardise it in South Africa. Briefly, heparinised blood was collected and exposed in vitro to 0Gy (control), 2Gy and 8Gy gamma radiation. One sample was exposed to 0Gy, 2Gy, 4gy and 8gy gamma radiation and compared to 0Gy, 1Gy, 2Gy, 3Gy and 4Gy neutron radiation. After 48 hours, lymphocytes were collected and prepared for flow cytometric analysis using FITC-conjugated anti-CD4 and anti-CD8 monoclonal antibodies. Apoptosis was assessed by measuring internucleosomal DNA degradation using propidium iodide (PI) staining. Percentage radiation-induced apoptosis was determined for each sample. Results showed a clear dose response curve for both gamma and neutron exposure and apoptosis after neutron exposure was higher than at iso-equivalent gamma doses. Although inter donor variation was observed, intra donor variation was low where the mean SD was 1.61 for CD4 and 2.16 for CD8. Results were consistent across cell type and the study therefore yielded enough data to base radiosensitivity measurements on CD4 and CD8 lymphocytes. In the clinical setting, patients showing a low apoptotic response should be considered at risk for developing late effects whereas patients who do not present as radiosensitive may receive increased doses of radiation. The LAA would be able to identify radiosensitive patients, thereby enabling the oncologist to stratify each patient to plan an effective treatment regime.

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