ジストロフィン遺伝子部分欠失を示すDMD/BMD患者における骨格筋のジストロフィンmRNAの分析

Duchenne型(DMD)およびBecker型(BMD)筋ジストロフィー患者におけるジストロフィン遺伝子部分欠失がmRNAレベルではどのようになっているのかを明らかにするため、RT-PCRを用いて,欠失を示すDMD11例およびBMD6例について骨格筋のジストロフィンmRNAの分析を行つた.さらに,DMD3例とBMD5例について,semi-nestedPCRによりジストロフィンmRNAを定量した.コントロールに比較して,DMD/BMD患者のジストロフィンmRNAはいずれも有意に減少していたが(p
In order to elucidate the effects of dystrophin gene deletions on the mRNA levels in the Duchenne and Becker muscular dystrophy, we analyzed human muscle dystrophin mRNA from eleven Duchenne muscular dystrophy (DMD) and six Becker muscular dystrophy (BMD) patients, with internal deletions of the dystrophin gene, using RT-PCR. We also quantified the resultant transcripts by means of semi-nested PCR to estimate the dystrophin mRNA in four BMD and three DMD cases. We found that the quantities of muscle dystrophin mRNA were drastically decreased in DMD and BMD patients as compared with in control subjects (p < 0.05), but that there was no significant difference between DMD and BMD. In each case, three regions of the DMD gene (the deletion-containing region, and the upstream and downstream regions) were amplified, and we focused on the deletion boundary regions by sequencing the PCR products. Truncated transcripts were found in 16 cases, while in the remaining case (DMD), no RT-PCR product was amplified for either muscle or lymphocytes, and thus transcription was assumed to be disturbed. We detected a difference between the muscle mRNA and lymphocyte DNA deletions in a patient who died of a rhabdomyosarcoma. In this case, there was an apparent deletion of exon 48 in the lymphocyte DNA, while only 15 by of the 3' region of exon 48 was deleted from the muscle mRNA. This raises the possibility that DNA diagnosis can miss a deletion. Another DMD case showed two distinct deletion patterns in muscle versus lymphocyte mRNA due to somatic mosaicism. Thus, it is important to analyze mRNA to clarify the molecular mechanisms of DMD/BMD.