| Objective:Mutations in the DMD gene (Xp21) lead to a spectrum of myopathic phenotypes broadly termed the dystrophinopathies, due to absence or dysfunction of the muscle protein dystrophin, including Duchenne muscular dystrophy (DMD), Becker muscular dystrophy (BMD), or DMD-associated dilated cardiomyopathy. Given the X-linked inheritance of this disorder, most heterozygous female carriers of DMD mutations are asymptomatic; however, about8%of these carriers are manifesting carriers who develop symptoms ranging from mild muscle weakness to a rapidly progressive DMD-like muscular dystrophy.2%of them can have no family history.7cases who are diagnosed as female dystrophinopathy by muscle biopsies are collected in this study. We performed protein analysis of dystrophin to exam the expression of dystrophin in female dystrophinopathy cases and compared them with male patients. We also performed karyotype analysis and DMD mutation analysis to clarify the mechanism of female dystrophinopathy cases.Material and methods:We examined7patients’ muscles obtained from the Peripheral Nerve and Muscular Disease Institute of Shandong Qilu Hospital, and1normal,1DMD and1BMD as control. Expression of dystrophin protein was observed using immunohistochemistry (IHC) staining and Western blotting (WB). The biopsied skeletal muscle specimens were flash-frozen in isopentane chilled with liquid nitrogen, and then were made into4um section. We used antibodies for dystrophin C-terminal、dystrophin Rod^dystrophin N-terminal for IHC. We chose6patients’s muscles from above and performed WB using the total protein extracting from the muscle tissue. Karyotype analysis and DMD mutation analysis were performed in the2cases (case1, case2) and we also performed the DMD mutation analysis in another one case (case7).Result:IHC:Among the7female dystrophinopathy cases, there were2absence in dystrophin and5showing a mosaic pattern.WB:For4female dystrophinopathy (2absent and2mosaic),1male DMD and1normal control, WB was performed. The2female DMD and the1male DMD showed that the protein was absent completely, coincident with IHC.Karyotype and DMD mutation analysis:For Case1, DMD multiplex ligation-dependent probe amplification (MLPA) was normal but the karyotype analysis showed46,x,t(x;10)(p22.1;pter). For Case2, Karyotype analysis and DMD MLPA were normal while a heterozygous mutation c.10223+1G>A and a heterozygous genovariation:c.2149G>T were detected by next generation sequencing (NGS). The X-chromosone inactivation analysis showed a ratio of90:10considered "highly skewed" pattern. Microarray Comparatice Genomic Hybridization (aCGH) was normal.Conclusion:1. Among the7female dystrophinopathy,2are consistent with DMD in IHC and the other5show a mosaic pattern.2. The mechanism for female symptomic mechanism include following6cases, that is, deletion in Xp21.2, Turner syndrome with a dystrophin mutation on the remaining X chromosome, skewed X inactivation either in the female DMD mutation carriers or balanced X-autosome translocation patients, uniparental disomy of the entire X chromosome with mutations, co-occurrence of mutations in both dystrophin and androgen-receptor genes, and double dystrophin mutations on both X chromosomes. The mechanism of the2female DMD in this study belongs to2of them.3. For the7cases, clinical manifestations are not related to IHC. The2absent cases have higher levels of serum CK than the other5ones. The severity of clinical manifestations is not related to IHC as well.4. When came across female patients, dystrophinopathy should be considered with other diseases and muscle biopsy may be necessary to clarify the diagnosis. As for carriers, the incidence of DCM is higher than normal people, the diagnosis of female dystrophinopathy is helpful to early diagnosis and treatment of DCM.5. As dystrophinopathy is an X-linked inherited disease. Once diagnosed, it would be helpful in the genetic consultation and the birth rate of dystrophinopathy babies would decrease as well. |
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