Hartmut Lins

Application of inhibitor titrations for the detection of oxidative phosphorylation defects in saponin-skinned muscle fibers of patients with mitochondrial diseases.

Inhibitor titrations were applied to characterize functional changes in mitochondrial energy metabolism in the skeletal muscle of patients with mitochondrial diseases. For this we titrated the maximal mitochondrial respiration rate of saponin-skinned muscle fibers isolated from the skeletal muscle biopsy with the specific inhibitors of mitochondrial oxidative phosphorylation complexes I, IV and V-rotenone, azide and oligomycin. For three patients with deletions of mitochondrial DNA and one patient with a complex I deficiency the titrations revealed at rather normal respiration activities of saponin-skinned fibers significant differences to healthy controls: (i) The inhibitor titration curves of the affected enzyme were much steeper and (ii) for almost complete inhibition of respiration a smaller amount of the inhibitor is necessary. The detailed analysis of the titration curves within the framework of metabolic control theory indicated elevated flux control coefficients of the respective complex of respiratory chain. On the other hand, for one patient with a mitochondrial DNA depletion syndrome, decreased respiration activities of skinned fibers but no redistribution of flux control was observed. We conclude, therefore, that application of inhibitor titrations and the quantitative description of the titration curve can be a valuable approach to elucidate functional defects of mitochondrial oxidative phosphorylation.

Re-evaluation of the dysfunction of mitochondrial respiratory chain in skeletal muscle of patients with Parkinson’s disease

Summary.The origin and tissue distribution of the mitochondrial dysfunction in Parkinson’s disease (PD) remains still a matter of controversy. To re-evaluate a probably free radical-born, generalized mitochondrial impairment in PD, we applied optimized enzymatic assays, high resolution oxygraphic measurements of permeabilized muscle fibers, and application of metabolic control analysis to skeletal muscle samples of 19 PD patients and 36 age-matched controls. We detected decreased activities of respiratory chain complexes I and IV being accompanied by increased flux control coefficients of complexes I and IV on oxygen consumption of muscle fibers. We further investigated if randomly distributed point mutations in two discrete regions of the mitochondrial DNA are increased in PD muscle, and if they could contribute to the mitochondrial impairment. Our data confirm the previously debated presence of a mild mitochondrial defect in skeletal muscle of patients with PD which is accompanied with an about 1.5 to 2-fold increase of point mutated mtDNA.

Detection of Respiratory Chain Defects in Cultivated Skin Fibroblasts and Skeletal Muscle of Patients with Parkinson's Disease

Parkinson’s disease (PD) is associated with a degeneration of dopaminergic neurons. The cause of this neuronal death is still unknown. Since 1989,1,2 the hypothesis of mitochondrial respiratory chain dysfunction as a possible basis for the neuronal death has been intensively debated. It is expected that the proposed mitochondrial defect as a possible pathogenic factor for PD should include mitochondria in other cell populations than neurons. Reported data obtained from different extracerebral tissues are controversial.3,4 Therefore, the direct proof of a suspected generalized mitochondrial dysfunction in PD is still missing. However, in muscle homogenates of patients with mitochondrial myopathies the mitochondrial defect is not always detectable with conventional enzyme assays for the determination of respiratory chain enzyme complexes I + III (NADH:cytochrome c reductase), II + III (succinate:cytochrome c reductase), and IV (cytochrome c oxidase).5 Therefore, we assessed the applicability of different in situ methods for the detection of putative mitochondrial respiratory chain defects in extracerebral tissues of PD patients. The functional behavior of mitochondria in digitonin-permeabilized skin fibroblasts and in saponinpermeabilized skeletal muscle fibers of PD patients was studied applying respiration experiments with different substrates using metabolic control analysis. Furthermore, we determined the redox state of the mitochondrial NAD system measuring the laser-excited autofluorescence of NAD(P)H and fluorescent flavoproteins in permeabilized skeletal muscle fibers.6 Fifteen patients with PD (4 female and 11 male, age range 37 to 78 years, mean 58 years) had a skeletal muscle and skin biopsy. The patients had had symptomatic PD for 2 to 13 years. The median UPDRS was 20 (range 4 to 56). All patients had the akinetic-rigid type of the disease. Skeletal muscle samples from diagnostic biopsies of 32 patients with discrete myopathic EMG abnormalities but no biopsy evidence for a manifest myopathy (17 male, 15 female, age range 29 to 72 years) were used as control, and all patients gave written informed consent. The study was approved by the Ethics Committee of Magdeburg University. Bundles of fibers from fresh M. vastus lateralis were used for preparation of saponin-permeabilized fibers.

Sequential analyses of neurobiochemical markers of cerebral damage in cerebrospinal fluid and serum in CNS infections

Objective –  To elucidate the relation between release patterns and cerebrospinal fluid/serum concentrations of neurobiochemical markers of cerebral damage and their potential value as monitoring parameters in central nervous system infections.

Gastric adenocarcinoma with leptomeningeal carcinomatosis as the presenting manifestation : an autopsy case report

Leptomeningeal carcinomatosis is a clinically important and severe complication in patients with cancer. Leptomeningeal involvement as a secondary event in gastric carcinoma is rarely reported and usually occurs late in advanced disease. Herein, we report a case of leptomeningeal carcinomatosis as the initial manifestation of a previously asymptomatic gastric adenocarcinoma. The clinical features and the appropriate diagnostic procedures are discussed.

Laser-excited fluorescence studies of mitochondria) function in saponin-skinned skeletal muscle fibers of patients with chronic progressive external ophthalmoplegia

The functional behavior of mitochondria in skeletal muscle of patients with chronic progressive external ophthalmoplegia was studied by laser-excited fluorescence measurements of NAD(P)H and flavoproteins in saponin-skinned fibers. Variations in the mitochondrial content and the presence of partially respiratory chain-inhibited mitochondria can be detected using this novel method.

Paraneoplastic necrotizing myelopathy with hypertrophy of the cauda equina.

Sirs: Recently, Burton et al. [1] described clinical and radiological characteristics of two patients presenting with hypertrophic radiculopathy of the cauda equina. Because removal of a bronchial carcinoid in one patient led to clinical improvement, a paraneoplastic origin was suggested. We report clinical and autopsy data of a patient with non-small cell lung cancer (SCLC), who developed a paraneoplastic necrotizing myelopathy [7] together with hypertrophic radiculopathy. A 71-year-old male patient with non-SCLC had been treated with chemotherapy leading to remission. Two years later he presented with pain in his lumbar spine and both legs, difficulty in walking, constipation, and impaired micturition. Neurological examination revealed paraparesis (right 2/5, left 4/5 MRC) with abolished tendon reflexes, flexor plantar responses, and a sensory level at D10. Prior to admission, EMG had shown chronic denervation of leg muscles. Sensory and motor conduction velocities and action potentials were in the lower normal range, F-wave latencies of the posterior tibial and peroneal nerve in the upper normal range. Tibial somatosensory evoked potentials could not be elicited. While no classical onconeural antibodies were found (immunoblot against cerebellar extract, blots against recombinant Hu-, Yo-, and Ri-antigens), serum antibodies against oligodendrocytes and Purkinje cells not identical with anti-CV2 could be detected by immunohistochemistry. CSF showed normal values for total protein, IgG, and lactate, but an increased cell count (16/μl) without tumour cells. Myelography of the lumbar and thoracic spine ruled out compression of the spinal cord and nerve roots. Within five weeks, continuous deterioration resulted in complete paraplegia and subsequent ascension of the transverse syndrome to a cervical level. The patient died of bronchopneumonia and septicaemia. Postmortem examination revealed necrotic areas in the central part of the cord throughout its whole length. The leptomeninges were unremarkable. Histologically, necrosis of the white and gray matter, demyelination, and focal accumulation of macrophages were present, most pronounced in the lumbar segments (Fig. 1). Intraspinal vessels were normal. Examination of the cauda equina demonstrated thickening of several nerve roots in a randomly distributed fashion (Fig. 2A). The tumourlike lesions were of firm consistency without necroses or haemorrhages. Microscopy showed proliferation of connective tissue, resulting in irregular separation of the nerve fibres (Fig. 2B). However, the cellularity of the Schwann cells was not increased, and no schwannoma or neurofibroma was observed. Mitotic figures were absent, and no proliferation was detected by Ki67-immunostaining. Only occasionally were single T lymphoctes observed within the nerve roots. Again blood vessels were normal. Semithin sections of a sural nerve sample showed a slightly reduced density of myelinated nerve fibres, but no acute demyelination, onion bulb formation, inflammatory infiltrates, or vasculitis. The brain was unremarkable except for moderate widening of the perivascular spaces in the basal ganglia. Autopsy also disclosed radiologically undetected remnants of the non-SCLC. Although no MRI could be performed (because the patient had a cardiac pacemaker), the pathomorphological findings of the cauda hypertrophy correspond well with the radiological features described by Burton et al. [1]. In contrast to LETTER TO THE EDITORS

Sudden progression of a glioblastoma in partial remission

There are only sparse data on viral CNS infections in patients with malignant glioma. We report a case of fatal herpes encephalitis in a patient with glioblastoma in partial remission and provide a short review of the literature.

Detection of mitochondrial defects by laser fluorimetry

The mitochondrial function in skeletal muscle biopsies of three patients with chronic progressive external ophthalmoplegia, having deletions of the mitochondrial DNA, was studied by laser-excited fluorescence measurements of NAD(P)H and flavoproteins in saponin-skinned fibers. We detected substantially elevated steady state redox states of the mitochondrial NAD-system in the muscle fibers of these patients. Moreover, the respiratory chain-linked autofluorescence changes in the muscle fibers of these patients were larger in comparison to controls indicating substantial alterations of the mitochondrial content. These results are in line with the presence of elevated numbers of partially respiratory chain inhibited mitochondria in the skeletal muscle of chronic progressive external ophthalmoplegia patients. (Mol Cell Biochem 174: 97–100, 1997)