Crypt-restricted metallothionein immunopositivity in murine colon: validation of a model for studies of somatic stem cell mutation

The ability to visualize the cellular effects of a somatic mutation is rele vant to studies of cell kinetics and carcinogenesis. In the colon, mutagen administration leads to scattered crypt-restricted loss of activity of the X-linked enzyme glucose-6-phosphate dehydrogenase (G6PD); it has been shown that this is due to somatic mutation in the G6PD gene. Mutagen-induced cry pt-restricted immunopositivity for metallothionein (MT) has been reported i n one study in the mouse colon; if this is also due to somatic mutation, it provides a simple method for studying the phenomenon which could be carrie d out on paraffin sections. This study shows that, as in the G6PD model, th e frequency of crypt-restricted immunopositivity for MT is very low in untr eated animals, but increases proportionately with the dose of mutagen admin istered. There is a good overall correlation of a range of MT-positive cryp t frequencies with those derived from studies using G6PD. As with the G6PD model, the MT-positive crypt phenotype evolves over time after mutagen admi nistration; initially individual crypts include both positive and negative phenotype cells, but later almost all involved crypts are composed entirely of MT-positive cells. The frequency of MT-positive crypts stabilizes after a few weeks and remains at the same level 6 months later. All these observ ations are qualitatively identical to those found using the G6PD model and provide strong evidence that stable, crypt-restricted immunopositivity for MT results from a mutation affecting expression of the metallothionein gene in a colonic stem cell. This model will provide a useful tool to study fac tors influencing stem cell mutation frequency and cell kinetics in the colo n. Copyright (C) 2000 John Wiley & Sons, Ltd.