The micronucleus test (MNT) has shown increased micronuclei (MN) frequencies in BRCA associated and sporadic breast cancer patients, Ataxia telangiectasia and Nijmegen Breakage Syndrome patients, demonstrating a common cellular phenotype of increased radiosensitivity. Some genes, causative of these diseases, have also recently been associated with prostate cancer. In order to investigate if prostate cancer exhibits the cellular phenotype of increased radiosensitivity, we performed MNT analysis on 22 sporadic prostate cancer patients and 43 male controls. We determined the baseline MN frequency, in order to see in vivo chromosomal damage without radiation, and induced (after irradiation with 2 Gy) frequency of MN, both in binucleated cells (BNC) obtained from cultured peripheral blood lymphocytes. An automated image analysis system was used to score the MN employing two different classifiers (Classifier A and B) for detection of BNC. The mean baseline frequencies were 48/43 MN/1000 BNC (A/B) for the controls and 42/50 (A/B) for prostate cancer patients. The induced MN frequencies amounted to 107/111 MN/1000 BNC (A/B) for controls and 111/114 MN/1000 BNC (A/B) for prostate cancer patients. The obtained MN frequencies did not result in a statistically significant difference between unselected cases and controls. However, restricting the analysis to young patients (50-60 years, N = 7) and age-matched controls (N = 7) revealed marginally significant higher MN frequencies in patients. We conclude that increased radiosensitivity is not a property of prostate cancer patients in general.

The quantification of DNA damage, both in vivo and in vitro, can be very time consuming, since large amounts of samples need to be scored. Additional uncertainties may arise due to the lack of documentation or by scoring biases. Image analysis automation is a possible strategy to cope with these difficulties and to generate a new quality of reproducibility. In this communication we collected some recent results obtained with the automated scanning platform Metafer, covering applications that are being used in radiation research, biological dosimetry, DNA repair research and environmental mutagenesis studies. We can show that the automated scoring for dicentric chromosomes, for micronuclei, and for Comet assay cells produce reliable and reproducible results, which prove the usability of automated scanning in the above mentioned research fields.

The micronucleus assay (MNT) in human lymphocytes is frequently used to assess chromosomal damage as a consequence of environmental mutagen exposure, to assess the effect of mutagens or to search for reduced DNA repair capacity after a mutagenic challenge. We have established an automated scoring procedure for the cytokinesis blocked MNT based on computerized image analysis (Metasystems Metafer 4 version 2.12). To evaluate the results we used the reproducibility of counts, established a dose-response curve for gamma-irradiation and used the ability of the system to differentiate between breast cancer patients and controls as a biological reference, a difference which we had observed before by visual counting. Blood cultures were irradiated with gamma-rays (2 Gy) at the beginning and treated with cytochalasin B during the last 24 h. The slides were stained with Giemsa for visual counting and with DAPI for automated analysis. Our test sample consisted of 73 persons (27 with breast cancer and 26 female and 20 male controls). A comparison between visual counting (controls, mean MN frequency 313) and automated counting (mean MN frequency 106) in slides from the same culture revealed a large drop for the automated counts. However, the automated counts were as reproducible as the visual counts [coefficient of variation (CV) on the sample approximately 20%; CV on repeated counts of the same slides approximately 5%] and both counts were highly correlated. Furthermore, the discrimination between cases and controls improved for automated counting of slides from the same cultures [visual odds rato (OR) < or = 4.0, P = 0.009; automated OR > 16, P < 0.0001], with a strong dependence on the set of parameters used. This improvement was confirmed in a validation sample of an additional 21 controls and 20 cases (OR = 11, P = 0.0018) performed as a prospective or diagnostic test.