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R D Johnsen

Publications and source records attributed to R D Johnsen.

11 recordsLinked to original sources
PubMed1999

The contribution of molecular genetics in the diagnosis and management of neuromuscular disorders.

It is true that the recent advances in molecular genetics have generated a medical revolution. This is especially true for the inherited neuromuscular disorders. There have been many spectacular recent discoveries with new genes being found and their protein products identified. One of the most remarkable aspects of this progress is the nexus which has developed between the basic discovery and its clinical application. As soon as a new genetic mutation is reported, the information may be used immediately to establish the molecular diagnosis for that disorder in any part of the world which has a DNA laboratory. This is done by using primers derived from the published DNA sequences using the polymerase chain reaction (PCR). This development is of immense value for the clinician as it provides an exact molecular diagnosis often with prognostic information and the test results can be used for genetic counselling and prenatal diagnosis. One of the unexpected outcomes of this work has been the surprising variation which has been shown to exist between genotype and phenotype. Previously, one mutation was believed to be responsible for one clinical disorder. However, it is now known that one genotype may be responsible for a variety of phenotypes and vice versa. In the field of neuromuscular disorders the most notable advances have occurred for Duchenne muscular dystrophy and the related dystrophinopathies and for the group of limb girdle muscular dystrophies, especially the subgroup of sarcoglycanopathies. Other areas are the congenital myopathies, the 'channel-opathies' and the mitochondrial cytopathies. In this review the most commonly used molecular genetic and immunocytochemical methods using antibodies to the protein product are outlined together with the principles of their application in the neuromuscular clinic. Included are the provisos and pitfalls which need to be kept in mind in the interpretation of DNA results for each patient.

Female↗
PubMed1998

Direct dystrophin and reporter gene transfer into dog muscle in vivo.

Bacterial beta-galactosidase cDNA was injected without lipofectin into 41 sites in dog muscle and expression was seen in 22 of them. The cDNA and lipofectin was injected into 35 similar sites and expression was seen in 21. Expression was seen in a maximum of 2.5% of muscle fibers and 23.21% of nonmuscle cells. A total of 106 muscle sites were injected with the minigene with and without lipofectin. In 4 of the 45 sites injected with the minigene without lipofectin human dystrophin was expressed around the periphery of 0.3% of the fibers. Bacterial beta-galactosidase cDNA was injected into the peritoneal cavity of 4 pups, 2 of which also received lipofectin. In all 4, expression was seen in liver, spleen, and mesenteric lymph node. In the 2 pups that received lipofectin, expression was also seen in the diaphragm, intercostal, and abdominal muscles of 1 and in the diagphragm and intercostal muscles of the other. These experiments show that human dystrophin transgene expression can be obtained in dog muscle. However, other methods will be required to increase the degree of expression before gene therapy trials can be undertaken.

Animals↗
PubMed1993

Two distinct mutations in a single dystrophin gene: identification of an altered splice-site as the primary Becker muscular dystrophy mutation.

A single base change in the 5' splice-site of intron 19 has been identified as the cause of the Becker muscular dystrophy in a family which had previously been deduced to carry both a major deletion and another, at that stage unidentified, mutation in the same dystrophin gene [Laing et al., 1992]. RNA from a muscle biopsy of one of the Becker muscular dystrophy patients in the family was analysed using the reverse transcriptase-polymerase chain reaction (RT-PCR) to study the mature gene transcript. Exon 19 was deleted from the dystrophin mRNA but present at the genomic level. The loss of exon 19 in the mature mRNA was found to be associated with an A to C mutation in the 5' splice site of intron 19. Deletion of exon 19 should alter the reading frame of the mRNA and be associated with a severe form of muscular dystrophy; however, low levels of normal-size dystrophin message and dystrophin were present in this patient. The distance between the splice-site mutation and the secondary deletion in the dystrophin gene is such that it would seem unlikely that the initial base change could act as a premutation for the deletion. Specific primers to detect the splice-site mutation have been designed and used to genotype all relatives.

Alleles↗
PubMed1993

Comparison of basic fibroblast growth factor in X-linked dystrophin-deficient myopathies of human, dog and mouse.

Binding of polyclonal antibodies specific for bFGF was examined in tissue sections of myopathic and normal muscles from humans, dogs and mice. The proposal tested was that differences in the amount or distribution of bFGF in muscles of the 3 species, might correlate with the limited muscle regeneration seen in humans and dogs afflicted with x-linked muscular dystrophy, in contrast with the sustained new muscle formation in mdx mice with the homologous myopathy. There was a striking difference between the species in the binding of bFGF antibodies to extracellular matrix, particularly at the periphery of myofibres; binding was pronounced in mouse but weak or absent in human and dog muscle. Binding to muscle nuclei and sarcoplasm was also stronger in mice than in humans and dogs, and in all species was more pronounced in foetal than adult muscle. Increased binding of bFGF antibodies was seen in damaged and regenerating muscle cells in all myopathic specimens where these were present. This was associated with the regenerative process rather than with myopathy, as a similar pattern of bFGF expression was seen in mouse muscle regenerating after experimental crush injury. The higher extracellular staining for bFGF around the periphery of mouse myofibres correlated with the successful muscle regeneration in dystrophic mice. Results suggest that bFGF at the fibre periphery might stimulate a local increase in the numbers of muscle precursor cells which can respond to injury in the mdx mouse.

Adolescent↗
PubMed1992

Two distinct mutations in a single dystrophin gene: chance occurrence or premutation?

We report on a kindred segregating 2 distinct mutations of a dystrophin gene. DNA analysis showed that the second mutation, a deletion, arose in the same gene carrying the primary defect which produced a Becker phenotype in the affected males. The DNA data for this family are reported and the alternative explanations of chance occurrence and premutation are discussed to explain these unusual findings.

Child↗
PubMed1991

The diagnosis of Duchenne and Becker muscular dystrophies: two years' experience in a comprehensive carrier screening and prenatal diagnostic laboratory.

This article describes the diagnostic algorithm being used for the management of the 148 families affected by Duchenne or Becker muscular dystrophy who are known to the Molecular Neurogenetics Laboratory in the Department of Neuropathology, Royal Perth Hospital. In 60 families from whom DNA has been obtained, 41 mutations (39 deletions and two duplications) of the Duchenne muscular dystrophy gene (DMD) have been identified by means of complementary DNA (cDNA) probes. DNA-based screening has clarified the carrier status of 45 at-risk women, and 13 pregnancies have been monitored. In addition, cDNA screening of all relevant patients with autosomal recessive muscular dystrophy, spinal muscular atrophy or limb-girdle muscular dystrophy facilitated the correct diagnosis of Becker muscular dystrophy in three patients.

Algorithms↗
PubMed1981

Muscle lipids in Duchenne muscular dystrophy.

The lipids of muscle and adipose tissue from normal males and of muscle from males with Duchenne muscular dystrophy were investigated. Triglyceride, the major neutral lipid, showed similar fatty acid compositions in all tissues examined. When the phospholipids of dystrophic muscle and of normal adipose tissue were compared with those of normal muscle, it was found that there was an increase in the proportion of sphingomyelin in dystrophic muscle, while adipose tissue had higher proportions of sphingomyelin and lysophosphatidylcholine but lower choline phosphoglyceride. In dystrophic muscle only small alterations from normal were observed in the fatty acid compositions of the individual phospholipids, whereas the phospholipids of adipose tissue had quite distinctive fatty acid compositions. An atrophic muscle sample resulting from poliomyelitis consisted almost entirely of connective tissue and fat and had a phospholipid composition similar to that of adipose tissue. From a comparison of the results for all the types of tissue studied, it is evident that the increase in sphingomyelin in dystrophic muscle biopsies and the changes in the fatty acid compositions of individual phospholipids may be accounted for by the increased amounts of fat and connective tissue which are present in dystrophic muscle samples. In a case each of polymyositis, limb girdle muscular dystrophy and an autosomal recessive form of muscular dystrophy, the results obtained for the phospholipid composition of the muscle sample were also normal or consistent with some contamination from fat and connective tissue.

Adipose Tissue↗