New Assay Methods for Monitoring Radiation Effects.

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A newly appearing p41 protein after X-irradiation of MOLT4 cells was characterized to access the mechanism(s) and feasibility as a marker of radiation effect. Amount of p41 was radiation dose and time dependent. Newly prepared polyclonal antibody against p41 also stained p42 on western blotting analysis of various tumor cell lines. Amino acid analysis of p41 and p42 showed that p41 was a cleavage product of p42. P42 was generated in vitro by protein synthesis and used as a substrate to identify a particular caspase (unpublished data) activated after X-irradiation of MOLT4 cells.Expression of MUC1 mucin of HT-29 cells after X-irradiation of 1, 3, 6, 10 Gy were analyzed for several days during cell death. Glycosylated MUC1 mucins (500 kDa and 390 kDa) and underglycosylated MUC1 mucins (350 kDa and 240 kDa) were found increased in dose-dependent manner reaching maximum level in 4 days. Transcriptional activity of MUC1 gene was analyzed by CAT assay containing the 5'-flanking sequence of MUC1 gene. Induction of CAT activity in HT-29 cells was found elevated from day 2 to 4 of X-irradiation. These findings with increased mRNA determined by RT-PCR indicate that transcriptional activity of MUC1 gene in HT-29 cells was upregulated by X-irradiation.Further characterization is required prior to practical use of these markers to monitor radiation effect, although the present results indicate such possibility.