Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DMD”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 91 records · Page 5Linked to original sources
PubMedDate not supplied

Protein truncation test (PTT) to rapidly screen the DMD gene for translation terminating mutations.

We have developed a rapid and sensitive method to screen the Duchenne muscular dystrophy (DMD) mRNA for translation terminating mutations by a combination of RT-PCR (Reverse Transcription and Polymerase Chain Reaction) and in vitro transcription/translation applied to white blood cell mRNA. This technique was termed the protein truncation test (PTT). Here we demonstrate the detection of a point mutation in a DMD patient and his mother, a carrier. The PTT can also be used for carrier detection when no patient material is available, or in the case of spontaneous mutations. We developed a protocol to screen the total coding region of the DMD gene in 5-10 PTT reactions. Furthermore, PTT could be of diagnostic value in any disease where premature terminations form a substantial part of the total mutation spectrum.

Base Sequence↗
PubMed2006

Novel mutations of dystrophin gene in DMD patients detected by rapid scanning in biplex exons DHPLC analysis.

Mutations in the dystrophin gene result in both Duchenne and Becher muscular dystrophies (DMD and BMD). Approximately 65% of all mutations causing DMD are deletions (60%) or duplications (5%) of large segments of this gene, spanning one exon or more. Due to the large size of the dystrophin gene (79 exons), finding point mutations has been prohibitively expensive and laborious. Recent studies confirm the utility of pre-screening methods, as denaturing high-performance liquid chromatography (DHPLC) analysis in the identification of point mutations in the dystrophin gene, with an increment of mutation detection rate from 65% to more than 92%. Here we suggest an alternative and convenient method of DHPLC analysis in order to find mutations in a more rapid and less expensive way by introducing the analysis of 16 couples of dystrophin amplicons, in biplex exons DHPLC runs. Using this new protocol of biplex exons DHPLC screening, new mutations were identified in four male patients affected by DMD who had tested negative for large DNA rearrangements.

Adult↗
PubMed2004

Early onset of lipofuscin accumulation in dystrophin-deficient skeletal muscles of DMD patients and mdx mice.

Lipofuscin, the so-called ageing pigment, is formed by the oxidative degradation of cellular macromolecules by oxygen-derived free radicals and redox-active metal ions. Usually it accumulates in post-mitotic, long-lived cells such as neurons and cardiac muscle cells. In contrast, it is rarely seen in either normal or diseased skeletal muscle fibres. In this paper, we report that lipofuscin accumulates at an early age in both human and murine dystrophic muscles. Autofluorescent lipofuscin granules were localized, using confocal laser scanning microscopy and electron microscopy, in dystrophin-deficient skeletal muscles of X chromosome-linked young Duchenne muscular dystrophy (DMD) patients and of mdx mice at various ages after birth. Age-matched normal controls were studied similarly. Autofluorescent lipofuscin granules were observed in dystrophic biceps brachii muscles of 2-7-year-old DMD patients where degeneration and regeneration of myofibres are active, but they were rarely seen in age-matched normal controls. In normal mice, lipofuscin first appears in diaphragm muscles nearly 20 weeks after birth but in mdx muscles it occurs much earlier, 4 weeks after birth, when the primary degeneration of dystrophin-deficient myofibres is at a peak. Lipofuscin accumulation increases with age in both mdx and normal controls and is always higher in dystrophic muscles than in age-matched normal controls. At the electron microscopical level, it was confirmed that the localisation of autofluorescent granules observed by light microscopy in dystrophin-deficient skeletal muscles coincided with lipofuscin granules in myofibres and myosatellite cells, and in macrophages accumulating around myofibres and in interstitial connective tissue. Our results agree with previous biochemical and histochemical data implying increased oxidative damages in DMD and mdx muscles. They indicate that dystrophin-deficient myofibres are either more susceptible to oxidative stress, or are subjected to higher intra- or extracellular oxidative stress than normal controls, or both.

Animals↗
PubMed2003

De novo mutations in sporadic deletional Duchenne muscular dystrophy (DMD) cases.

Dinucleotide repeat polymorphism based genetic analysis is a powerful approach to gain insight into rare genetic events like germline mosaicism and de novo mutations. The loss of heterozygosity of polymorphic dinucleotide loci at "deletional hotspot" of dystrophin gene can provide direct evidence of carrier status in female relatives of affected DMD patients with overlapped exonic deletions. We have used 4 STR loci of the central deletional hotspot of the dystrophin gene for genetic analysis in sporadic unrelated DMD families. Twenty-nine mothers of sporadic deletional cases were analysed and their carrier status was determined. Eighteen of them showed heterozygosity in the deleted loci suggesting the occurrence of de novo mutations. In 9 cases, the carrier status was indeterminate while 2 showed germline mosaicism. Our observations reiterated the importance of STR analysis in determining the status of mothers of sporadic deletional DMD cases in order to provide proper genetic counselling.

DNA Mutational Analysis↗
PubMed2004

Recombinant adeno-associated viral (rAAV) vectors as therapeutic tools for Duchenne muscular dystrophy (DMD).

Duchenne muscular dystrophy (DMD) is a lethal genetic muscle disorder caused by recessive mutations in the dystrophin gene. The size of the gene (2.4 Mb) and mRNA (14 kb) in addition to immunogenicity problems and inefficient transduction of mature myofibres by currently available vector systems are formidable obstacles to the development of efficient gene therapy approaches. Adeno-associated viral (AAV) vectors overcome many of the problems associated with other vector systems (nonpathogenicity and minimal immunogenicity, extensive cell and tissue tropism) but accommodate limited transgene capacity (<5 kb). As a result of these observations, a number of laboratories worldwide have engineered a series of microdystrophin cDNAs based on genotype-phenotype relationship in Duchenne (DMD) and Becker (BMD) dystrophic patients, and transgenic studies in mdx mice. Recent progress in characterization of AAV serotypes from various species has demonstrated that alternative AAV serotypes are far more efficient in transducing muscle than the traditionally used AAV2. This article summarizes the current progress in the field of recombinant adeno-associated viral (rAAV) delivery for DMD, including optimization of recombinant AAV-microdystrophin vector systems/cassettes targeting the skeletal and cardiac musculature.

Animals↗
PubMed1992

Reconstruction of the 2.4 Mb human DMD-gene by homologous YAC recombination.

The human dystrophin gene, mutations of which cause Duchenne and Becker muscular dystrophy, measures 2.4 Mb. This size seriously limits its cloning as a single DNA fragment and subsequent in-vitro expression studies. We have used stepwise in-vivo recombination between overlapping yeast artificial chromosomes (YACs) to reconstruct the dystrophin gene. The recombinant YACs are mitotically stable upon propagation in haploid yeast cells. In contrast, specific combinations of YACs display a remarkable mitotic and meiotic instability in diploid cells. Non-disjunction is rare for overlapping YACs, but increases upon sporulation of diploid cells containing non-overlapping molecules. We have exploited this feature in a three-point recombination to bridge a 280 kb gap between two non-overlapping YACs for which no YAC of proper polarity existed. Our largest recombinant YAC measures 2.3 Mb and contains the entire muscle specific DMD-gene with the exception of a 100 kb region containing the in-frame exon 60. The latter segment has a high tendency to undergo deletions in multi-molecular interactions, probably due to the presence of as yet unidentified instability-enhancing sequences. Fluorescent in situ hybridizations confirmed that the 2.3 Mb DMD YAC contained Xp21-sequences only and indicated a compact tertiary structure of the DMD-gene in interphase lymphocyte nuclei. We conclude that the yeast system is a flexible, efficient and generally applicable tool to reconstruct or build genomic regions from overlapping YAC constituents. Its application to the human dystrophin gene has provided many possibilities for future studies.

Chromosomes, Fungal↗
PubMed1998

Preimplantation diagnosis of non-deletion Duchenne muscular dystrophy (DMD) by linkage polymerase chain reaction analysis.

The use of preimplantation diagnosis for sex determination and detection of exon deletion means that unaffected babies can be born to parents suffering from Duchenne muscular dystrophy (DMD). However, those who do not have exon deletion should also be considered for further investigation. A new method, known as linkage analysis, has been developed to diagnose the presence of non-deletion DMD in preimplantation embryos. Linkage analysis uses informative intragenic and flanking markers to track the chromosome bearing the mutated gene. The present study reports the analysis of two polymorphic sites, in blastomeres biopsied from embryos from a female carrier of DMD. A single male embryo was obtained who had inherited alternate maternal alleles to the woman's affected surviving son, and this embryo was transferred.

Adult↗
PubMed1987

Assignment of the locus order DXS28-DXS67-DMD as a spin-off from diagnostic DNA marker analysis in a family with Duchenne muscular dystrophy.

During diagnostic segregation analysis for seven DNA markers, linked to and flanking the locus for Duchenne muscular dystrophy (DMD), a family was identified in which a boy with a recombinant X chromosome had inherited his maternal grandmother's alleles at the loci DXS43 (D2/Pvu II) and DXS28 (C7/Eco RV), and his maternal grandfather's alleles at DXS67 (B24/Msp I) and DXS84 (754/Pst I). Combined with earlier data this finding strongly suggests the locus order DXS28-DXS67-DMD. Another recombination event, identified in the same family, supports the previously established order DMD-DXS84-OTC. The diagnostic importance of flanking markers, and the likelihood of false diagnostic conclusions due to possible double crossovers, with and without demonstrable neighbouring single crossover events, are discussed.

Genetic Linkage↗
PubMed1993

Diagnosis of DMD carrier status in a family with no known affected males.

A 30-year-old woman and her two-year-old daughter were found by chance to have moderately raised serum creatine kinase (CK) levels. Since the mother was pregnant, the authors investigated the possibility that the two females were carriers of the common Duchenne muscular dystrophy (DMD) gene. No immunohistochemical abnormality was detected in the mother, but in the daughter a clear mosaic pattern of dystrophin positive and negative fibres was found, indicating carrier status for DMD. These data indicate that a diagnosis of DMD carrier status can be made even in families without a positive history for this disorder; therefore, immunocytochemical studies, using antidystrophin antibodies, should be performed on all females with raised CK levels, including the youngest.

Child, Preschool↗
PubMed1984

Duchenne muscular dystrophy involving translocation of the dmd gene next to ribosomal RNA genes.

Duchenne muscular dystrophy (DMD) is a severe X-linked disorder leading to early death of affected males. Females with the disease are rare, but seven are known to be affected because of a chromosomal rearrangement involving a site at or near the dmd gene on the X chromosome. One of the seven has a translocation between the X and chromosome 21. The translocation-derived chromosomes from this patient have been isolated, and the translocation is shown to have split the block of genes encoding ribosomal RNA on the short arm of chromosome 21. Thus ribosomal RNA gene probes may be used to identify a junction fragment from the translocation site, allowing access to cloned segments of the X at or near the dmd gene and presenting a new approach to the study of this disease.

Animals↗
PubMed1992

Different mosaicism frequencies for proximal and distal Duchenne muscular dystrophy (DMD) mutations indicate difference in etiology and recurrence risk.

In about 65% of the cases of Duchenne muscular dystrophy (DMD) a partial gene deletion or duplication in the dystrophin gene can be detected. These mutations are clustered at two hot spots: 30% at the hot spot in the proximal part of the gene and about 70% at a more distal hot spot. Unexpectedly we observed a higher frequency of proximal gene rearrangements among proved "germ line" mosaic cases. Of the 24 mosaic cases we are aware of, 19 (79%) have a proximal mutation, while only 5 (21%) have a distal mutation. This finding indicates that the mutations at the two hot spots in the dystrophin gene differ in origin. Independent support for the different mosaicism frequency was found by comparing the mutation spectra observed in isolated cases of DMD and familial cases of DMD. In a large two-center study of 473 patients from Brazil and the Netherlands, we detected a significant difference in the deletion distribution of isolated (proximal:distal ratio 1:3) and familial cases (ratio 1:1). We conclude from these data that proximal deletions most likely occur early in embryonic development, causing them to have a higher chance of becoming familial, while distal deletions occur later and have a higher chance of causing only isolated cases. Finally, our findings have important consequences for the calculation of recurrence-risk estimates according to the site of the deletion: a "proximal" new mutant has an increased recurrence risk of approximately 30%, and a "distal" new mutant has a decreased recurrence risk of approximately 4%.

Blotting, Southern↗
PubMed2006

[Identification of disease-causing point mutations in DMD patients' dystrophin gene without large deletions/duplications].

OBJECTIVE: To detect the disease-causing point mutations in the dystrophin gene of Duchenne muscular dystrophy (DMD) patients. METHODS: The approach of denaturing high performance liquid chromatography (DHPLC) coupling with sequencing was used to screen the point mutations of 79 exons and the untranslated regions of dystrophin gene without large deletions/duplications, which was in 6 unrelated DMD probands from 6 DMD families. RESULTS: Five disease-causing mutations, 697-698insGT, C616T, G1255T, C4279T, and C2302T, were ides created the new stop codons in downstream sites of mutations, respectively. In addition to the disease-causing point mutations, a point mutation T5586+61A in intron 39 was also found at patient 3, and a missense mutation A694T in exon 8 was detected at patient 5. Four point mutations, C2168+13T, 5740-13dupG, G5234A and C5280T, were also detected at patient 6 whose causative point mutation was unavailable. Seven point mutations have not been reported previously. Bi-directional PCR amplification of specific alleles (Bi-PASA) method was established to distinguish the haplotypes of heterozygote or homozygote in a single PCR reaction. CONCLUSION: Via automated DHPLC screening or detecting the subexonic mutations in dystrophin gene is feasible to clinical laboratories, and also is a superior method in terms of sensitivity and efficiency.

Base Sequence↗
PubMed1996

[The sequences of intron 50 and 51 of DMD gene and the deletion mechanism of 51st exon].

OBJECTIVE: To investigate the relationship between the structural characters of 50th and 51st introns and deletion mechanisms of 51st exon of DMD gene. METHODS: We cloned the 3.1 Kb-Hind I human genome DNA fragment containing the 50 and 51 introns of DMD gene, sequenced the fragment completely, and analysed the character of the sequences with computer. RESULTS: The length of the fragment was 3179, including the entire 51st exon and partial 50th and 51st intron sequences. There were 36 short tandom repeats, 22 direct and inverted repeats, and 8 homologous sequences in the 50th and 51st introns. A TATA box was found in the intron 50 and it may be another new promotor of DMD gene. CONCLUSION: The hypothesis, which the recombination of repeat sequences may result in the deletion of 51st exon, is put forward.

Base Sequence↗
PubMed2000

Elevated p21 mRNA level in skeletal muscle of DMD patients and mdx mice indicates either an exhausted satellite cell pool or a higher p21 expression in dystrophin-deficient cells per se.

Abnormalities in proliferation and differentiation of the dystrophin-deficient muscle are a controversial aspect of the pathogenesis of Duchenne muscular dystrophy (DMD). Analyses of molecules involved in cell cycle modulation do not exist in this context. Cells withdrawn from the cell cycle permanently express p21. The fact that p2 1, in contrast to other cell cycle proteins, is not diminished when myotubes are reexposed to growth media, allocates this cyclin-dependent kinase inhibitor a special function. Here we report for the first time statistically increased p21 mRNA levels in dystrophin-deficient muscle tissue. Only 42% of conventional RT-PCRs from six muscle samples of human controls yielded positive results but almost all skeletal muscle biopsy samples (87%) from DMD patients (n=5). For p21 mRNA quantification in murine muscle samples we were able to use the exact real-time TaqMan PCR method due to generally higher p21 mRNA levels than in human muscles. In addition, contamination with fibroblasts can be excluded for the murine samples because they do not demonstrate fibrosis at the age of 350 days but start to lose their regenerative capacity. In accord with the results in humans, we observed p21 mRNA levels in mdx mice that were approx. four times as high as those in control mice. Elevated p21 mRNA level may indicate a shift in cell composition towards differentiated p21 expressing cells as a result of an exhausted pool of undifferentiated, non-p21-expressing satellite cells due to previous cycles of de- and regeneration. Alternatively, dystrophin-deficient cells per se may express higher p21 levels for unknown reasons. Although we cannot distinguish between these possibilities, the eventual transfec tion of a patient's own satellite cells with p21 antisense oligonucleotides may enable the dystrophic process to be influenced.

Actins↗
PubMed1992

Alterations of protein degradation and 2-D protein pattern in muscle cells of MDX and DMD origin.

Intracellular protein turnover of MDX, DMD and normal muscle was determined by [35S]methionine pulse-chase experiments and subsequent high resolution 2-D gel electrophoresis. In MDX myotubes intracellular degradation of short-lived and long-lived proteins was markedly increased by a factor of 1,4-2,1. In wildtype the rate of degradation of short-lived proteins was approximately 2.6%/h, whereas in MDX these proteins were degraded by 5.7%/h. Long-lived proteins were degraded in wildtype at a rate of 1.8%/h, and in MDX at a rate of 2.5%/h. Furthermore, we have described a 51.000 Da protein with an IEP of 5.1 (p51/5.1), whose net content is highly and specifically reduced in cultured MDX and DMD muscle cells as well as in isolated MDX muscle fibers. Treatment with calcium-channel blockers Dantrolene and Verapamil inhibited the degradation of p51/5.1 in MDX myotubes by more than 90% in contrast to controls.

Actins↗
PubMed1986

Calcium activated neutral proteases (milli- and micro-CANP) and endogenous CANP inhibitor of muscle in Duchenne muscular dystrophy (DMD).

Calcium activated neutral protease (milli- and micro-forms) and its endogenous inhibitor have been quantified in muscle from Duchenne muscular dystrophy (DMD) patients. The specific activities of both the enzymes are found to be significantly elevated. Some of the properties studied indicate that the enzymes from DMD and normal are not qualitatively different. The ratios of milli- to micro-enzyme in normal and disease state suggest that these enzymes have independent roles to play. The absence of a significant increase in the level of the endogenous inhibitor is probably indicative of its mode of regulation, in disease condition.

Adolescent↗
PubMed1992

A homologue of dystrophin is expressed at the blood vessel membrane of DMD and BMD patients: immunological evidence.

Muscles from Becker muscular dystrophy (BMD) and Duchenne muscular dystrophy (DMD) patients were analysed using monoclonal and polyclonal antibodies raised against different regions of the dystrophin molecule. On blot, two of the antibodies detected a protein of Mr 400K in muscle extracts from all patients, including a BMD patient with a deletion which spanned more than 40% of the central rod domain of the Xp21 encoded dystrophin. Immunocytochemical labelling of tissue sections from the same patients showed that the same two antibodies labelled a protein at the surface membrane of smooth muscle fibers in blood vessels of both BMD and DMD muscles. Thus we have demonstrated a 400K blood vessel-associated protein, which is immunologically homologous with dystrophin, for at least two epitopes from the carboxy terminal and the central rod domains must be encoded by another gene than the dystrophin gene.

Adult↗