Mutations in the non-lysosomal cysteine protease calpain-3 cause autosomal recessive limb girdle muscular dystrophy. Pathological mechanisms occurring in this disease have not yet been elucidated. Here, we report both morphological and biochemical evidence of mitochondrial abnormalities in calpain-3 knockout (C3KO) muscles, including irregular ultrastructure and distribution of mitochondria. The morphological abnormalities in C3KO muscles are associated with reduced in vivo mitochondrial ATP production as measured by 31 P magnetic resonance spectroscopy. Mitochondrial abnormalities in C3KO muscles also correlate with the presence of oxidative stress; increased protein modification by oxygen free radicals and an elevated concentration of the anti-oxidative enzyme Mn-superoxide dismutase were observed in C3KO muscles. Previously we identified a number of mitochondrial proteins involved in β-oxidation of fatty acids as potential substrates for calpain-3. In order to determine if the mitochondrial abnormalities resulted from the loss of direct regulation of mitochondrial proteins by calpain-3, we validated the potential substrates that were identified in previous proteomic studies. This analysis showed that the β-oxidation enzyme, VLCAD, is cleaved by calpain-3 in vitro , but we were not able to confirm that VLCAD is an in vivo substrate for calpain-3. However, the activity of VLCAD was decreased in C3KO mitochondrial fractions compared with wild type, a finding that likely reflects a general mitochondrial dysfunction. Taken together, these data suggest that mitochondrial abnormalities leading to oxidative stress and energy deficit are important pathological features of calpainopathy and possibly represent secondary effects of the absence of calpain-3.
Abstract We report two female patients with a history of alcohol abuse presenting with proximal painful muscle weakness following aversion therapy with emetine hydrochloride. Muscle biopsy of Case 1 showed a reversible floccular‐shaped loss of myosin ATPase and dehydrogenase, and accumulation of PAS positive material, and a normal lipid content. Repeat biopsy showed core change with no focal loss of myosin ATPase. In Case 2, muscle biopsy was taken 1 month after commencement of emetine therapy and revealed similar but milder changes to Case 1. Electron microscopy revealed Z‐band streaming with a decrease or loss of mitochondria. Sarcotubular systems appeared normal in shape and size. Anaerobic glycolysis on homogenate from the initial biopsy of Case 1 showed generalized reduction of lactate formation, which returned to normal in the repeat biopsy.
A microscopic, three-dimensional study of the aortico-pulmonary glomus tissue in the adult cat indicates a random, continuous aggregation of glomus tissue extending from the left coronary artery to the ligamentum arteriosum. It was not possible to demonstrate unequivocally specific glomera. Serial section and the injection of the pulmonary artery with India ink failed to show a vessel of supply to glomeric tissue.
The effect of propylthiouracil-induced neonatal hypothyroidism on the development of synaptosome fraction mitochondrial membrane enzymes has been determined. Total and specific activities of two mitochondrial inner membrane (cytochrome c oxidase and succinate cytochrome c reductase) and two outer membrane enzymes (monoamine oxidase and rotenone-insensitive NADH cytochrome c reductase) were determined. The inner mitochondrial membrane enzymes developed in synchrony in euthyroid animals, but a decrease in fractional and specific activities of both inner membrane enzymes occurred in hypothyroidism. The outer membrane enzymes developed asynchronously; monoamine oxidase revealed a 9-fold increase while NADH cytochrome c reductase increased 4-fold. Hypothyroidism caused no change in total fractional activity or Km and an increased specific activity of outer membrane enzymes.
Abstract —A reproducible model of subacute methyl mercury (MeHg) intoxication was developed in the adult rat following the daily intragastric administration of 10 mg methyl mercury/kg body wt. Synaptosomes isolated from animals during the latent phase of mercury neurotoxicity (6‐10 days) demonstrated no significant change in respiratory control, State 3, State 4, or 2,4‐dinitrophenol stimulated respiration with succinate, glutamate or pyruvate plus malate. During the neurotoxic phase, a significant decline in respiratory control was evident with all substrates. Cerebellar synaptosomes revealed qualitatively similar but quantitatively greater inhibition of 2,4‐dinitrophenol stimulated respiration during the latent and neurotoxic phases with glutamate. In vitro studies of synaptosome respiration, oxidative phosphorylation and respiratory control with 5‐15 μ m ‐methyl mercury revealed a stimulation of initial State 4 respiration, loss of RCI, inhibition of State 3 but no change in the gramicidin or 2,4‐dinitrophenol uncoupled rate supported by pyruvate‐malate. Phosphate did not relieve the State 3 inhibition. At 25 μ m ‐methyl mercury and above, considerable inhibition of electron transfer occurred. At this concentration, cytochrome c oxidase was inhibited 50%. Isosmotic replacement of medium KC1 by mannitol reduced the MeHg stimulation of State 4 respiration but had no effect on MeHg inhibition of ADP stimulated respiration. Half‐maximal stimulation of State 4 respiration by MeHg occurred at [K] + ⋍ 6 m m . These findings are compatible with an energy‐linked methyl mercury induced cation translocation across the synaptosome (mitochondrial) membrane.
Previous work has suggested involvement of the muscle microvasculature in the pathogenesis of dermatomyositis (DM). Our study evaluates whether standard histochemical reactions can identify microvascular changes in muscle biopsies from patients with DM compared to myopathic and nonmyopathic controls.Muscle biopsies were obtained from 111 patients, including 45 patients with DM. Microvascular quantitation was performed on transversely oriented 1-μm toluidine blue-stained plastic sections. Histoenzymatic procedures included alkaline phosphatase (AP), nicotinamide adenine dinucleotide tetrazolium reductase (NADH-TR), succinate dehydrogenase (SDH), cytochrome C oxidase (COX), and myosin ATPase reactions.Capillary density was significantly lower in DM muscle biopsies compared to biopsies from patients with noninflammatory myopathies (NIM; n = 26) and healthy control muscle (n = 27; mean ± SD: 252 ± 114 vs 402 ± 56 and 325 ± 109 capillaries/mm(2), respectively; p values < 0.05). In contrast, a marked increase in the number of capillaries staining with NADH-TR was noted in DM compared to other idiopathic inflammatory myopathies (IIM; n = 13), NIM, and controls (49.8 ± 50.7 vs 8.0 ± 7.1, 6.7 ± 7.2, and 3.6 ± 2.8 capillaries/mm(2); p < 0.05 compared to DM). DM capillaries also demonstrated mildly increased staining with AP compared to controls; however, no increased SDH or COX reactivity was observed.DM muscle capillaries are highly reactive with NADH-TR compared to myopathic and nonmyopathic controls. The lack of staining of DM capillaries with mitochondrial SDH and COX reactions suggests that NADH-TR reactivity may be secondary to activation of the microvascular endoplasmic reticulum, rather than mitochondrial hyperplasia.
