Genotype to phenotype correlations in mitochondrial encephalomyopathies associated with the A3243G mutation of mitochondrial DNA.

We studied 22 subjects carrying the A3243G point mutation of human mitochondrial DNA (mtDNA). In 14 cases the clinical phenotype was characterized by mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), while 8 patients had chronic progressive external ophthalmoplegia (CPEO). The proportion of A3243G heteroplasmy in muscle was determined by two methods; densitometry on a diagnostic restriction-fragment length polymorphism and solid-phase mini-sequencing. We found a highly significant inverse correlation between the percentage of A3243G mutation and the specific activity of complex I, the respiratory complex with the highest number of mtDNA-encoded subunits, suggesting a direct effect of the mutation on mtDNA translation. No correlation was observed between the percentage of mutated mtDNA and the presence or absence of specific clinical features, such as stroke, ophthalmoplegia and diabetes mellitus. However, in the MELAS group the percentage of mutated mtDNA molecules was strongly correlated with the age of onset, while no such correlation was found in the CPEO group, suggesting a different time-dependent evolution of the mutation in the two groups. Finally, in contrast with other mtDNA mutations associated with ragged-red fibres (RRF), in both MELAS3243 and CPEO3243 we observed a high proportion of RRF that were positive to the histochemical reaction to cytochrome c oxidase, a morphological feature that seems to be specific for the neuromuscular phenotypes associated with mutations affecting the tRNA(Leu(UUR)) gene.

Respiratory chain and mitochondrial DNA in muscle and brain in Parkinson's disease patients.

There are several reports of a defect of complex I in the substantia nigra (SN) of Parkinson's disease (PD) patients. To evaluate whether this is specific to dopaminergic neurons or the phenotypically relevant consequence of a widespread failure of the mitochondrial oxidative phosphorylation (OXPHOS) system, we measured respiratory enzyme activities in muscle homogenates from 16 PD patients and eight age-matched controls, and in muscle isolated mitochondria of six PD patients and six age-matched controls. We found no difference between the PD and control groups. In addition, we detected, by polymerase chain reaction, the mitochondrial DNA (mtDNA) "common deletion" (CD) in muscle specimens of 14 of 17 PD patients, but we obtained similar results in age-matched controls. In both groups, the amount of CD-specific deleted (delta) mtDNA ranged from 0.0% to 0.1%. Our data suggest that PD cannot be attributed to a multisystem decline of mitochondrial OXPHOS, and that lesions of muscle mtDNA in PD are likely due to normal aging. However, there was a remarkable accumulation of delta mtDNA in the SN of a PD patient and an age-matched control, suggesting that the SN is exquisitely sensitive to age-dependent damage of the mitochondrial genome.

A MERRF/MELAS overlap syndrome associated with a new point mutation in the mitochondrial DNA tRNA(Lys) gene.

Several members of a three-generation kindred from Sardinia were affected by a maternally inherited syndrome characterized by features of both myoclonus epilepsy with ragged-red fibers (MERRF) and mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS). Clinically, symptoms such as myoclonus epilepsy, neural deafness and ataxia were variably associated with stroke-like episodes and/or migrainous attacks. Morphologically, numerous MELAS-associated SDH-stained vessels were observed in muscle biopsies, either alone or in combination with ragged-red fibers, the morphological hallmark of MERRF. Sequence analysis of the mtDNA tRNA genes revealed the presence of a single, heteroplasmic T-->C transition at nt 8356, in the region of the tRNA(Lys) gene corresponding to the T-psi-C stem. The T-->C(8356) transition was exclusively found in the maternal lineage of our family, and the relative amount of the mutant mtDNA species in muscle was correlated with the severity of the clinical presentation. Therefore, we propose that the T-->C(8356) transition is responsible for the mitochondrial encephalomyopathy found in our family, and must be added to the expanding list of the pathogenetically relevant mutations of human mtDNA.

Innervation is required to stabilize and amplify creatine kinase activity in regenerated extensor digitorum longus muscles of rats.

Neural control of creatine kinase (CK, adenosine 5'-triphosphate creatine phosphotransferase: EC 2.7.3.2) was investigated by measuring enzymatic activity and isoenzymatic representation of CK in extensor digitorum longus (EDL) muscles of adult rats during and after regeneration. Experimental models were ischemized EDLs, reversibly or permanently denervated. Results showed that, during regeneration, CK in muscle fibers was likewise modified in both reversibly and permanently denervated EDLs. After regeneration a clear dichotomy was observed between the regenerated EDLs which were innervated and recovering CK activity, and those in which innervation was prevented and were rapidly losing activity. Further to investigate the neural influence on CK turnover, merely denervated age-matched EDLs were analysed and found to lose CK activity rapidly. The major conclusions are that during regeneration muscle CK is autonomously expressed, but following regeneration neural influence becomes an absolute requirement for stabilization and amplification of CK.

[Effects of dithiocarbamate: "in vitro" and "in vivo" changes in gamma-glutamyltransferase induced by sodium diethyldithiocarbamate]. / Azione dei ditiocarbamati (DTC): alterazioni della gamma-glutamiltransferasi (GGT) "in vitro" ed "in vivo" indotte da sodiodietilditiocarbamato (NaDDTC).

Experiments of NaDDTC effect on kidney and 10,000 X g liver supernatant fraction have shown no modification of normal GGT activity, also after dialysis. A decrease of about 75% of the GGT activity was observed in rabbit 10,000 X g liver supernatant fraction after i.p. treatment with 100 mg/kg of NaDDTC for three days. It is suggested that liver MFO produces SH groups derived from NaDDTC metabolism able to interact with GGT and consequential inactivation of the enzyme.

[The biological action of dithiocarbamates. Changes in serum leucine aminopeptidase induced in vitro by sodium diethyldithiocarbamate]. / Azione biologica dei ditiocarbamati. Variazioni di attività di leucinoaminopeptidasi sierica (LAP) indotte "in vitro" da sodiodietilditiocarbamato (NaDDTC).

The Authors demonstrate that serum LAP activity in vitro is inhibited by NaDDTC. This may be related to the NaDDTC chelating-like action on the metallo-enzyme. The trend of the phenomenon follows an exponential pattern. The enzyme activity was completely restored after removal of NaDDTC from the medium by dialysis. The NaDDTC concentration able to inhibit the enzymatic activity in vitro was much higher than in vivo, in experimental animals. The Authors conclude pointing out the problem of a probable effect caused by repeated doses of dithiocarbamates more on enzyme synthesis than on metallic apoenzyme.

[Biological effects of dithiocarbamates: changes in serum alkaline phosphatase activity "in vitro" by sodium diethyldithiocarbamate (NADDTC) (author's transl)]. / Azione biologica dei ditiocarbamati: variazioni di attività di fosfatasi alcalina sierica indotte "in vitro" da sodiodietilditiocarbama to (NaDDTC).

The Authors demonstrate that serum alkaline phosphatase (s-ALP) activity in vitro is inhibited by NaDDTC. This may be related to the NaDDTC chelating action on zinc enzyme. The trend of the phenomenon follows an exponential pattern. The activity of the isozyme pool was restored after removal from medium of NaDDTC by dialysis. The NaDDTC concentration able to inhibit in vitro enzymatic activity was nearly one thousand times higher than that found in vivo, in experimental animals (rabbit). The Authors conclude that the in vivo activity of NaDDTC in different enzymatic systems, such as ALP and other metallo-dependent enzymes, cannot be explained by its chelating action alone, but also by its influence on other systems. At present investigations in this field are in progress in our laboratory.

[Rubella seroepidemiology. proposal of a predictive mathmatical model (author's transl)]. / Sieroepidemiologia della rosolia. Proposta di un modello matematico predittivo.

This paper deals with time dependent epidermic behaviour of rubella. The hypothesis that ten classes of immuno-response (Stewart test) can be reduced to three is assumed (= low, intermediate and high level). Upon these conditions a mathematical model is developed and tested using experimental data measured during six years over 1288 subjects. Initial conditions, time-constants and transition probability for the model are also estimated from experimental data.

[A mathematical model for the study of the kinetics of serum alkaline phosphatase inhibition by sodium diethyldithiocarbamate (NaDDTC)]. / Modello matematico per lo studio della cinetica di inibizione della fosfatasi alcalina sierica ad opera del dietilditiocarbammato di sodio (NaDDTC).

A mathematical model for the kinetic of the serum alkaline phosphatase inhibition by NaDDTC is described in this paper. The model is tested over an experimental data set consisting on measured % residual activity of enzyme in human serum, after inhibition. The Bessey method is used. The solution of the differential equations for the model is found to be the best fitting for the observed time-dependent phenomena. The rate constant, K, for the inhibition reaction is then computed.