レーザーマイクロダイセクションを用いたCAGリピートの不安定化機構の研究

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We investigated the molecular mechanism on the meiotic and mitotic instability of CAG repeats causing neurodegenerative disorders such as Huntington disease (HD) and hereditary ataxias.First, employing a laser-captured microdissection (LCM), we investigated the meiotic instability of CAG repeats in the germ-line cells in the Huntington disease (HD) replacement mouse. LCM enables the isolation of single lineage testicular cells for subsequent molecular analysis. We found that CAG repeats in the spermatid are significantly smaller than the spermatocyte in the mouse homozygous for the expanded allele (Mann-Whitney U test, P<0.05). With regard to the mouse examined, the CAG repeats tended to contract with the cell differentiation.Sccond, we investigated the mitotic instability of CAG repeats in the variable brain cell lineage in two patients with Machado-Joseph disease (MJD) using LCM.We found that CAG repeat size in the cells of cerebellar cortex (molecular, Purkinje, and granular cell la … More yers) is significantly smaller than that in the cells of cerebellar white matter (Mann-Whitney U test, P<0.05). There was no significant differences in the CAG repeat size among the cells of the cerebellar cortex.Third, although the intergenerational stability of the CAG repeat number has been considered to be a specific molecular feature of SCA6 compared with other CAG repeat diseases, we showed meiotic instability of the CAG repeats in the SCA6/CACNL1A gene in two Japanese SCA6 families, including de novo expansion. In one family, the CAG _<20> allele expanded to the CAG_<26> one during paternal transmission, and in the other family, the CAG_<19> allele expanded to the CAG_<20> one during maternal transmission. This is the first case of haplotype analysis-proven de novo expansion in SCA6, confirming the derivation of an expanded allele from one normal allele.Finally, we examined whether the postnatal expansion of the CAG repeats in the blood cells for 'CAG repeat diseases' occurs. We analyzed the CAG repeats in the umbilical cord blood (UCB) and peripheral blood (PB) cells with HD and MJD in adulthood. We found that somatic mosaicism in the blood cells of HD and MJD significantly increases over time (Mann-Whitney U test, P<0.005), indicating that somatic instability is continuous throughout the life of patients. This is the first report on the somatic instability of CAG repeats in the blood cells of 'CAG repeat diseases'. Less