Edward Byrne

Primary LAMP-2 deficiency causes X-linked vacuolar cardiomyopathy and myopathy (Danon disease).

“Lysosomal glycogen storage disease with normal acid maltase”, which was originally described by Danon et al., is characterized clinically by cardiomyopathy, myopathy and variable mental retardation. The pathological hallmark of the disease is intracytoplasmic vacuoles containing autophagic material and glycogen in skeletal and cardiac muscle cells. Sarcolemmal proteins and basal lamina are associated with the vacuolar membranes. Here we report ten unrelated patients, including one of the patients from the original case report, who have primary deficiencies of LAMP-2, a principal lysosomal membrane protein. From these results and the finding that LAMP-2-deficient mice manifest a similar vacuolar cardioskeletal myopathy, we conclude that primary LAMP-2 deficiency is the cause of Danon disease. To our knowledge this is the first example of human cardiopathy–myopathyxa0that is caused by mutations in a lysosomal structural protein rather than an enzymatic protein.

Clinicopathological features of genetically confirmed Danon disease

Background: Danon disease is due to primary deficiency of lysosome-associated membrane protein-2. Objective: To define the clinicopathologic features of Danon disease. Methods: The features of 20 affected men and 18 affected women in 13 families with genetically confirmed Danon disease were reviewed. Results: All patients had cardiomyopathy, 18 of 20 male patients (90%) and 6 of 18 female patients (33%) had skeletal myopathy, and 14 of 20 male patients (70%) and one of 18 female patients (6%) had mental retardation. Men were affected before age 20 years whereas most affected women developed cardiomyopathy in adulthood. Muscle histology revealed basophilic vacuoles that contain acid phosphatase–positive material within membranes that lack lysosome-associated membrane protein-2. Heart transplantation is the most effective treatment for the otherwise lethal cardiomyopathy. Conclusions: Danon disease is an X-linked dominant multisystem disorder affecting predominantly cardiac and skeletal muscles.

In Vivo and in Vitro Correction of the mdx Dystrophin Gene Nonsense Mutation by Short-Fragment Homologous Replacement

Targeted genetic correction of mutations in cells is a potential strategy for treating human conditions that involve nonsense, missense, and transcriptional splice junction mutations. One method of targeted gene repair, single-stranded short-fragment homologous replacement (ssSFHR), has been successful in repairing the common deltaF508 3-bp microdeletion at the cystic fibrosis transmembrane conductance regulator (CFTR) locus in 1% of airway epithelial cells in culture. This study investigates in vitro and in vivo application of a double-stranded method variant of SFHR gene repair to the mdx mouse model of Duchenne muscular dystrophy (DMD). A 603-bp wild-type PCR product was used to repair the exon 23 C-to-T mdx nonsense transition at the Xp21.1 dys locus in cultured myoblasts and in tibialis anterior (TA) from male mdx mice. Multiple transfection and variation of lipofection reagent both improved in vitro SFHR efficiency, with successful conversion of mdx to wild-type nucleotide at the dys locus achieved in 15 to 20% of cultured loci and in 0.0005 to 0.1% of TA. The genetic correction of mdx myoblasts was shown to persist for up to 28 days in culture and for at least 3 weeks in TA. While a high frequency of in vitro gene repair was observed, the lipofection used here appeared to have adverse effects on subsequent cell viability and corrected cells did not express dystrophin transcript. With further improvements to in vitro and in vivo gene repair efficiencies, SFHR may find some application in DMD and other genetic neuromuscular disorders in humans.

Mitochondrial respiratory chain activity in idiopathic dilated cardiomyopathy

BACKGROUNDnCardiomyopathy is well recognized in mitochondrial diseases in which it has been associated with defects of mitochondrial function, including cytochrome-c oxidase (COX) deficiencies. This study explores the respiratory chain activity, particularly of COX, in patients with cardiomyopathy to determine whether a relationship exists between respiratory enzyme activity and cardiac function.nnnMETHODS AND RESULTSnMyocardial specimens from the left ventricular wall of explanted hearts were obtained from subjects with ischemic (n = 6) or nonischemic dilated (n = 8) cardiomyopathy. Assays for citrate synthase (CS) and complexes II/III and IV activity were performed on cardiac mitochondria and homogenate. Enzyme activities were normalized to CS activity and compared with control activities (n = 10). A significant reduction in COX and/or CS activity was identified in mitochondrial preparations from the transplant group and correlated significantly with ejection fraction (P < .05), although this does not prove a causal relationship. Significantly reduced CS activity in homogenate was identified, suggesting decreased mitochondrial volume in addition to decreased COX activity. Measurements in cardiac homogenates failed to show a significant reduction in COX activity (P > .05) in the transplant group, suggesting that the use of prefrozen tissue homogenates may underestimate existing mitochondrial respiratory defects in cardiac tissue.nnnCONCLUSIONSnMitochondrial function is altered at a number of levels in end-stage cardiomyopathy. Defective COX activity resulting in deficient adenosine triphosphate generation may contribute to impaired ventricular function in heart failure. Agents capable of improving mitochondrial function may find an adjuvant role in the treatment of cardiac failure.

Immunotherapy of ocular myasthenia gravis reduces conversion to generalized myasthenia gravis

Background Several retrospective studies have suggested that immunotherapy, including prednisolone, azathioprine and thymectomy, reduces progression of ocular myasthenia gravis to generalized myasthenia gravis. This study examines the effect of immunotherapy on generalization rates in ocular myasthenia patients who are acetylcholine receptor (AChR) antibody-positive. Methods Retrospective record review of 34 patients from three university-based hospitals with neurology and neuro-ophthalmology services in Australia. In all patients, positive AChR antibodies were recorded, the initial symptoms were purely ocular, and all had at least 2 years of follow-up. The patients who developed generalized myasthenia gravis were compared with those who remained purely ocular. Results There were 21 patients who developed generalized myasthenia gravis. Of these 21, only 2 (9.5%) had received prior immunotherapy. Among the 13 patients whose symptoms remained purely ocular, 10 (76.9%) had received prior immunotherapy. Conclusions In this study, most of the patients who progressed from ocular myasthenia to generalized myasthenia had not received prior immunotherapy. This study adds weight to the call for a prospective trial of early immunotherapy in patients with ocular myasthenia.

Pigmentary retinopathy associated with the mitochondrial DNA 3243 point mutation

Fourteen patients from four unrelated families were studied to determine the prevalence of retinal pigmentary abnormalities associated with the MELAS A to G 3243 point mutation. Neurologic and ophthalmic examinations, retinal photography, pattern shift visual evoked potentials, and electroretinography were performed in all patients. Eight of the 14 patients had retinal pigmentary abnormalities characterized by symmetric areas of depigmentation involving predominantly the posterior pole and midperipheral retina. None of the patients had optic atrophy and only one patient with pigmentary retinal abnormalities had impaired visual acuity. None of the diabetic subjects (n = 6) had signs of diabetic retinopathy. Fluorescein angiography demonstrated mottled hyper- and hypofluorescent areas indicating multiple window defects in the retinal pigmentary epithelium. Visual evoked potentials showed delayed P100 responses in four of the eight patients with retinal pigmentary abnormalities. We conclude that there is a high prevalence of retinal pigmentary abnormalities in patients with MELAS A to G 3243 point mutation. These abnormalities are usually asymptomatic and best detected by retinal photography.

A new dominant distal myopathy affecting posterior leg and anterior upper limb muscles.

Objective: To report a dominant, slowly progressive early onset distal myopathy with sparing of the tibialis anterior. Methods: Twelve affected and two possibly affected members from an Australian kindred were examined and investigated by EMG, imaging studies, histopathology, and genetic analysis. Results: Affected patients had a slowly progressive condition with symmetric, distal weakness and wasting of the anterior upper and posterior lower limbs, with sparing of tibialis anterior, even in advanced disease. All patients remained ambulant and there was no evidence of cardiac or respiratory muscle involvement. Serum creatine kinase levels were either normal or mildly elevated. Imaging studies showed widespread involvement of the posterior and lateral leg compartments. Proximal muscles were radiologically abnormal only in advanced disease. Muscles that were mildly affected clinically appeared normal on imaging. EMG in nine patients showed widespread myopathic changes. Muscle histopathology in four patients showed either end stage muscle or nonspecific myopathic findings without inflammation or vacuoles. Microsatellite markers for distal myopathy loci were analyzed and all known distal myopathy phenotype genes and linkage regions were formally excluded by multipoint analysis. Conclusions: The affected patients in this kindred display a clinically distinct myopathy, with selective involvement of posterior lower and anterior upper limb muscles. The genetic analysis suggests the existence of one more distal myopathy locus.

DNA electroporation in vivo targets mature fibres in dystrophic mdx muscle

Non-viral gene transfer into skeletal muscle is enhanced by electroporation and myotoxin preconditioning of muscle following plasmid injection. We investigated in vivo delivery of naked DNA to mdx mouse muscle, utilising enhanced green fluorescent protein reporter vector (pEGFP) and a corrective nucleic acid to promote targeted corrective gene conversion at the mutant mdx mouse dystrophin (DMDmdx) locus. Electroporation, myoablation with bupivacaine and a combined protocol, were applied to mdx muscle. We report up to 90% EGFP expression in electroporated mdx tibialis anterior muscle. Muscles preconditioned with bupivacaine showed low transgene expression with or without EP. Single EGFP+ve muscle fibre explants showed EGFP expression in mature fibres in preference to satellite cells. We observed a two-fold increase (P<0.005; t) in dystrophin protein, accompanied by wild-type (wt) DMD transcript in muscles injected with corrective nucleic acid over contralateral saline-injected TAs. By targeting the muscle fibres in preference to the satellite cells, plasmid-bourne transgenes delivered to dystrophic muscle will not penetrate the regenerative component of muscle. Whether in the context of targeted corrective gene conversion or therapeutic non-viral transgenes, under these conditions periodic re-administration will be required to promote phenotypic benefits in dystrophic muscle.

A novel clinical phenotype of myopathy, sensorimotor neuropathy, infertility, and hypogonadism with multiple mitochondrial DNA deletions

We describe a patient with myopathy, sensorimotor neuropathy, hypogonadism, and infertility with abnormal sperm mobility and morphology. Analysis of the deltoid muscle DNA revealed a G to A change at nt 1102 in the twinkle gene and multiple mitochondrial DNA deletions. Histochemistry revealed “ragged-red” fibers and many cytochrome-c oxidase negative fibers (32%) that lacked the mitochondrial encoded respiratory chain subunits I and II and the nuclear encoded subunit VIc. Respiratory chain enzyme analysis showed severe deficiency of complex I, III, and IV. This patient has no documented family history of progressive external ophthalmoplegia, which suggests either a sporadic or autosomal-recessive syndrome. This case is a novel phenotype for twinkle gene mutations and multiple mitochondrial DNA deletion syndromes, as these syndromes generally follow an autosomal-dominant inheritance pattern.

Names of infamy: Tainted eponyms

The use of eponyms is controversial. A distinction must be made between those doctors and scientists after whom disorders and syndromes are named in honour of their discoveries, and those whose discoveries were made as a result of maltreatment of defenceless prisoners, utilizing specimens from victims of Nazi extermination policies, and euthanasia victims of racial policies. The second group of scientists should have their names expunged from the historical record, and their deeds brought to the attention of their colleagues. We are not however advocating the abolition of eponyms in general, only tainted ones.