Analysis of Italian regulations on pathways of care for patients in a vegetative or minimally conscious state.

Different rehabilitation models for persons diagnosed with disorders of consciousness have been proposed in Europe during the last decade. In Italy, the Ministry of Health has defined a national healthcare model, although, to date, there is a lack of information on how this has been implemented at regional level. The INCARICO project collected information on different regional regulations, analysing ethical aspects and mapping care facilities (numbers of beds and medical units) in eleven regional territories. The researchers found a total of 106 laws; differences emerged both between regions and versus the national model, showing that patients with the same diagnosis may follow different pathways of care. An ongoing cultural shift from a treatment-oriented medical approach towards a care-oriented integrated biopsychosocial approach was found in all the welfare and healthcare systems analysed. Future studies are needed to explore the relationship between healthcare systems and the quality of services provided.

Switching from constant voltage to constant current in deep brain stimulation: a multicenter experience of mixed implants for movement disorders.

BACKGROUND AND PURPOSE: For many years deep brain stimulation (DBS) devices relied only on voltage-controlled stimulation (CV), but recently current-controlled devices have been developed and approved for new implants as well as for replacement of CV devices after battery drain. Constant-current (CC) stimulation has been demonstrated to be effective in new implanted parkinsonian and dystonic patients, but the effect of switching to CC therapy in patients chronically stimulated with CV implantable pulse generators (IPGs) has not been assessed. This report shows the results of a consecutive retrospective data collection performed at five Italian centers before and after replacement of constant-voltage with constant-current DBS devices, in order to verify the clinical efficacy and safety of this procedure. METHODS: Nineteen patients with Parkinson's disease or dystonic syndrome underwent DBS IPG CV/CC replacement. Clinical features and therapy satisfaction were assessed before surgery, 1 week after and 3 and 6 months after replacement. Programming settings and impedances were recorded before removing the CV device and when the CC IPGs were switched on. RESULTS: The clinical outcome of CC stimulation was similar to that obtained with CV devices and remained stable at 3 and 6 months of follow-up. Impedance values recorded for CV and CC IPGs were similar. Ninety-five percent of patients and physicians were satisfied with mixed implants. No adverse events occurred after IPG replacement. CONCLUSION: Replacing CV with CC IPGs is a safe and effective procedure. Longer follow-up is necessary to better clarify the impact of CC stimulation on clinical outcome after chronic stimulation in CV mode.

Coiling and neuroendoscopy: a new perspective in the treatment of intraventricular haemorrhages due to bleeding aneurysms.

BACKGROUND: Intraventricular haemorrhages (IVHs) caused by bleeding aneurysms are critical conditions that often carry a severe prognosis. Two main problems must be urgently dealt with: the secondary damage caused by intraventricular clotting and the risk of early rebleeding. A protocol of ultra-early endoscopic ventricular evacuation, after securing the aneurysm with coils, is proposed to solve this challenge in the acute phase and within a few hours of onset. METHODS: Ten consecutive patients presenting with haematocephalus from aneurysm rupture were treated in our institute with coiling and endoscopic clot aspiration extended to the whole ventricular system. The only inclusion criteria were the presence of a massive IVH and an aneurysm appropriate for coiling. Computed tomography scans obtained before (within 4 h of symptom onset in all patients) and immediately after surgery were compared for Graeb score and ventriculocranial ratio (VCR); the Glasgow Outcome Scale (GOS) was assessed at 1 year. RESULT: All patients were treated within 2 days of onset. The procedure resulted in a mean 58% removal of ventricular blood and decrease of hydrocephalus; the mean (standard deviation (SD)) Graeb score reduced from 11.5 (0.7) to 4.7 (2.2) (p<0.001) and mean ventriculocranial ratio from 0.26 (0.06) to 0.17 (0.05) (p<0.001). No rebleeding or delayed hydrocephalus needing shunt was observed. Mortality at 1 year was 30%; marked disability (GOS = 3) and good recovery (GOS = 5) were observed in 40% and 30% of patients, respectively. CONCLUSIONS: Early neuroendoscopic removal of blood casting from the lateral to the fourth ventricle after coiling of bleeding aneurysms is a feasible approach, allowing in most instances the rapid improvement of the IVH.

Caspase-independent death of Leber's hereditary optic neuropathy cybrids is driven by energetic failure and mediated by AIF and Endonuclease G.

Leber's hereditary optic neuropathy (LHON) is associated with mitochondrial DNA point mutations affecting different subunits of complex I. By replacing glucose with galactose in the medium, cybrids harboring each of the three LHON pathogenic mutations (11778/ND4, 3460/ND1, 14484/ND6) suffered a profound ATP depletion over a few hours and underwent apoptotic cell death, which was caspase-independent. Control cybrids were unaffected. In addition to cytochrome c, apoptosis inducing factor (AIF) and endonuclease G (EndoG) were also released from the mitochondria into the cytosol in LHON cybrids, but not in control cells. Exposure of isolated nuclei to cytosolic fractions from LHON cybrids maintained in galactose medium caused nuclear fragmentation, which was strongly reduced by immuno-depletion with anti-AIF and anti-EndoG antibodies. In conclusion, the caspase-independent death of LHON cybrids incubated in galactose medium is triggered by rapid ATP depletion and mediated by AIF and EndoG.

Neuroendoscopic management of intraventricular hemorrhage.

BACKGROUND AND PURPOSE: We reviewed our 7-year experience in neuroendoscopic management of severe intraventricular hemorrhage (IVH) to evaluate its safety, efficiency, and efficacy. METHODS: Thirteen patients with spontaneous primary or secondary tetraventricular IVH underwent neuroendoscopy. In all procedures, we used a flexible instrument. CT scans obtained before and after surgery were compared for Graeb score and ventriculocranial ratio. Glasgow Outcome Scale was assessed at 12 months. RESULTS: In all patients, the procedure resulted in a substantial removal of ventricular blood. Graeb score was reduced by 65%, and ventriculocranial ratio was reduced by 30% (P<0.002). The procedure was carried out safely even in the presence of a vascular malformation, and no rebleeding or delayed hydrocephalus was observed in any case. Mortality at 12 months was 30.7%. Favorable outcome (Glasgow Outcome Scale, 3 to 5) was observed in 61.5% of cases. CONCLUSIONS: Neuroendoscopic management of severe IVH in this cohort of patients was safe, efficiently reduced the amount of ventricular blood and ventricular dilatation, and effectively produced an outcome profile that compares very favorably with other more conventional treatments.

Endoscopic treatment of colloid cysts of the third ventricle: 9 consecutive cases.

The purpose of this study is to evaluate the efficacy of the endoscopic technique for the treatment of the colloid cysts of the third ventricle. Between August 1995 and October 1997 a series of nine patients with colloid cyst of the third ventricle (6 males and 3 females) were treated with this method. The technique, consisting of cyst fenestration, aspiration of the colloid, and coagulation of the internal layer of the wall, was always effective in restoring CSF circulation. Operating time was 54-120 min (median 67 min). We recorded only one post-operative septic complication but no signs of direct surgical morbidity. Post-operation hospital stay was 2-30 days (median 5 days). Follow up was 14-40 months (mean 27 months). We did not observe any clinical or radiological recurrence. Endoscopic treatment of colloid cysts of the third ventricle is a safe and effective alternative to the well-established approaches of microsurgical removal and stereotactic aspiration. Only a very long follow-up will answer the question of the long-term effectiveness of this method.

A stop-codon mutation in the human mtDNA cytochrome c oxidase I gene disrupts the functional structure of complex IV.

We have identified a novel stop-codon mutation in the mtDNA of a young woman with a multisystem mitochondrial disorder. Histochemical analysis of a muscle-biopsy sample showed virtually absent cytochrome c oxidase (COX) stain, and biochemical studies confirmed an isolated reduction of COX activity. Sequence analysis of the mitochondrial-encoded COX-subunit genes identified a heteroplasmic G-->A transition at nucleotide position 6930 in the gene for subunit I (COX I). The mutation changes a glycine codon to a stop codon, resulting in a predicted loss of the last 170 amino acids (33%) of the polypeptide. The mutation was present in the patient's muscle, myoblasts, and blood and was not detected in normal or disease controls. It was not detected in mtDNA from leukocytes of the patient's mother, sister, and four maternal aunts. We studied the genetic, biochemical, and morphological characteristics of transmitochondrial cybrid cell lines, obtained by fusing of platelets from the patient with human cells lacking endogenous mtDNA (rho0 cells). There was a direct relationship between the proportion of mutant mtDNA and the biochemical defect. We also observed that the threshold for the phenotypic expression of this mutation was lower than that reported in mutations involving tRNA genes. We suggest that the G6930A mutation causes a disruption in the assembly of the respiratory-chain complex IV.

Molecular characterization of myophosphorylase deficiency in a group of patients from northern Italy.

We studied a group of 14 patients from Northern Italy with myophosphorylase deficiency. The disease presented considerable clinical and biochemical heterogeneity, which was reflected at the molecular level. The clinical presentation was typical in 3 patients, mild in 7 (exercise intolerance), and severe in 4 (fixed weakness). Enzyme activity was undetectable in 10 patients, below 3% of control in 3, and 13% of control in one. Enzymatic protein was detectable immunologically only in 1 patient. Myophosphorylase mRNA was present in 8 patients, but in 7 of them it was reduced in amount. Two patients were homozygous for the common nonsense R49X mutation, 5 were heterozygous. Two missense mutations not previously observed were identified in this group of patients. The frequency of alleles with the R49X mutation was significantly lower in this group of patients than in previously reported series. Myophosphorylase deficiency is genetically heterogeneous even among patients living in a small region and with a common ethnic background.

Residual muscle cytochrome c oxidase activity accounts for submaximal exercise lactate threshold in chronic progressive external ophthalmoplegia.

The data from histological, biochemical, and mitochondrial DNA (mtDNA ) studies of muscle biopsies from 10 patients affected with chronic progressive external ophthalmoplegia (CPEO) were related to dynamic and metabolic parameters of incremental submaximal exercise. Maximum power output was reduced in all patients as compared to controls. Analysis of the venous lactate curve during exercise revealed a lactate threshold at exercise levels ranging from 40 to 50% of the predicted maximal power output. An earlier significant increase in lactate could be detected by calculating the mean delta lactate. Lactate values were inversely correlated with the cytochrome c oxidase (COX) activity of isolated muscle mitochondria. No relationship was found between lactate values and the number of ragged red fibers, or cytochrome c oxidase-negative fibers of the proportion of deleted mtDNA measured in muscle biopsy specimens. The discussion underscores the value of lactate kinetics in assessing skeletal muscle function, as well as the use of muscle COX levels to predict the effectiveness of wild-type complementation of deleted skeletal muscle mtDNA in in vivo contractile performance of CPEO subjects.

MtDNA mutations associated with Leber's hereditary optic neuropathy: studies on cytoplasmic hybrid (cybrid) cells.

Leber's hereditary optic neuropathy (LHON) has been associated with "primary" and "secondary" mtDNA missense point mutations, and a synergistic role has been proposed for secondary mutations. No previous study has investigated the effects of LHON primary or primary plus secondary mutations on the respiratory competence of cell lines. We constructed and compared cybrid cell lines obtained from two unrelated LHON patients both carrying the common 11778/ND4 primary mutation. One of the patients also carried the 13708/ND5 and 4216/ND1 secondary mutations. The cybrid clones were evaluated for growth efficiency, oxygen consumption, complexes I, III and IV enzymatic activity and mitochondrial protein synthesis. Complex activity and mitochondrial protein synthesis were not significantly changed in cybrid clones from the patients. Oxygen consumption was significantly decreased in all clones carrying the 11778/ND4 primary mutation demonstrating its pathogenic role in impairing cell respiration. Clones also carrying the secondary mutations showed an even lower oxygen consumption and a significantly higher doubling time, suggesting that the co-presence of the secondary mutations could be relevant in further reducing the cell fitness.

Myophosphorylase deficiency affects muscle mitochondrial respiration as shown by 31P-MR spectroscopy in a case with associated multifocal encephalopathy.

We report here a glycogen storage myopathy type V associated with multifocal encephalopathy. The patient, a 43-year-old male with increased serum CK, a heavy drinker and smoker, had been affected by generalized epilepsy since age 24, after a cranial injury. He had had a right hemiparesis 2 years before coming to our observation and a transient left hemiparesis the following year. CT and MRI of the brain showed multiple hemispheric lesions consistent with an ischemic process, as suggested by single photon emission tomography of the brain. Muscle biopsy showed a vacuolar myopathy, and myophosphorylase activity was 13% of the normal mean. Phosphorus magnetic resonance spectroscopy (31P-MRS) performed on resting calf muscles showed increased PCr to ATP and decreased PCr to P(i) ratios. During both aerobic and ischemic exercise 31P-MRS failed to show any cytosolic acidification and phosphomonoesters (PME) accumulation, two MRS findings in agreement with McArdle's syndrome diagnosis. Mitochondrial respiration was also affected as shown by a low PCr to P(i) ratio at rest and by a low rate of PCr re-synthesis during recovery from aerobic exercise. This latter finding in McArdle's disease can be explained by decreased mitochondrial substrate availability, which in turn can contribute to the phenotypic manifestations of the disease.

Expression of muscle-type phosphorylase in innervated and aneural cultured muscle of patients with myophosphorylase deficiency.

Patients with McArdle's myopathy lack muscle glycogen phosphorylase (M-GP) activity. Regenerating and cultured muscle of patients with McArdle's myopathy presents a glycogen phosphorylase (GP) activity, but it is not firmly established whether M-GP or non-M-GP isoforms are expressed. We have cultured myoblasts from biopsy specimen of five patients with McArdle's myopathy. Skeletal muscle was cultured aneurally or was innervated by coculture with fetal rat spinal cord explants. In the patients' muscle biopsies and in their cultured innervated and aneural muscle we studied total GP activity, isoenzymatic pattern, reactivity with anti-M-GP antiserum, and presence of M-GP mRNA. There was no detectable enzymatic activity, no immunoreactivity with anti-M-GP antiserum, and no M-GP mRNA in the muscle biopsy of all patients. GP activity, M-GP isozyme, and anti-M-GP antiserum reactivity were present in patients' aneural cultures, increased after innervation, and were undistinguishable from control. M-GP mRNA was demonstrated in both aneural and innervated cultures of patients and control by primer extension and PCR amplification of total RNA. Our studies indicate that the M-GP gene is normally transcribed and translated in cultured muscle of patients with myophosphorylase deficiency.

Dystrophin-positive myotubes in innervated muscle cultures from Duchenne and Becker muscular dystrophy patients.

Nerve-muscle co-cultures from five Duchenne muscular dystrophy (DMD) patients and one Becker (BMD) patient, were studied by immunocytochemistry with antibodies against different portions of dystrophin. Four DMD patients had a deletion in the dystrophin gene. Some dystrophin-positive myotubes were detected in a few samples of all DMD cases. PCR amplification of exon 8 of the dystrophin gene ruled out a contamination from rat spinal cord during innervation. Our results in three DMD cases, may be explained by a clonal selection of dystrophin-positive fibers observed in muscle biopsies, while in the other two cases, a "frame-restoring" mutation might account for the presence of dystrophin-positive myotubes. The possible expression of "dystrophin-related protein" or dystrophin immature isoform was considered. In the BMD case an abnormal truncated dystrophin was found in innervated muscle cultures, as well as in muscle biopsy.

Hypertrophic cardiomyopathy with mitochondrial myopathy. A new phenotype of complex II defect.

Two brothers, 25 and 19 years old, were affected by asymmetrical hypertrophic cardiomyopathy. The older brother had waddling gait and weakness of the proximal girdle muscles, while the younger had a broad-based gait and weakness of selected limb girdle muscles. EMG exam was myopathic. Serum enzyme, CPK and aldolase were elevated. Histochemical reactions in muscle revealed "core-like" areas, subsarcolemmal rims of mitochondria and lipid accumulation. Succinate-dehydrogenase stain showed a lack of activity in both biopsies, with the exception of intrafusal fibers. Microphotometric quantitative measurements confirmed the defect in both biopsies. Biochemical measurements of several mitochondrial enzymes in muscle showed a reduced activity of succinate-dehydrogenase (33%) and succinate-cytochrome C reductase (36-47%) which are both components of complex II. On myocardial biopsy lipid and mitochondrial abnormalities were found. This mitochondriopathy represents a new phenotype of partial complex II defect.

MELAS mutation in mtDNA binding site for transcription termination factor causes defects in protein synthesis and in respiration but no change in levels of upstream and downstream mature transcripts.

The pathogenetic mechanism of the mitochondrial tRNA(LeuUUR) gene mutation responsible for the MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes) syndrome was investigated in transformants obtained by transfer of mitochondria from three genetically unrelated MELAS patients into human mitochondrial DNA (mtDNA)-less (rho 0) cells. Marked defects in mitochondrial protein synthesis and respiratory activity were observed in transformants containing virtually pure mutant mtDNA, as compared to the parent of the rho 0 cells (the 143B cell line) or to transformants containing exclusively wild-type mtDNA, derived from one of the patients or a maternally related asymptomatic individual. A striking protective effect against the mutation was exerted in the transformants by levels of residual wild-type mtDNA above 6%. The MELAS mutation occurs within the mtDNA binding site for a protein factor (mTERF) that promotes termination of transcription at the 16S rRNA/tRNA(LeuUUR) gene boundary. A marked decrease in affinity of purified mTERF for the mutant target sequence was observed in in vitro assays. By contrast, RNA transfer hybridization experiments failed to show any significant change in the steady-state amounts of the two rRNA species, encoded upstream of the termination site, and of the mRNAs encoded downstream, in the transformants carrying the MELAS mutation.

Variability of the expression of muscle mitochondrial damage in ocular mitochondrial myopathy.

In this study we comparatively analysed deltoid histochemistry, biochemistry and mitochondrial DNA (mtDNA) in two groups of ten sporadic ocular mitochondrial myopathies (OMM), respectively with and without ragged red fibres (RRF). (1) All but one RRF--patients presented the mild form of OMM with blepharoptosis but without ophthalmoplegia; (2) the occurrence of cytochrome c oxidase deficient (COX-) fibres was significantly higher in the RRF+ group, but four RRF- cases also showed COX- fibres; (3) no difference was observed in biochemical findings between the groups; (4) two RRF- patients without COX- fibres showed mtDNA heteroplasmy; (5) in two RRF- patients without deltoid mtDNA deletion, biopsy of an eyelid muscle showed significant mitochondrial alterations. These results suggest that the expression of a mitochondrial defect can vary and that the absence of RRF in a skeletal muscle biopsy does not necessarily rule out the diagnosis of OMM, if other data support that.

Blood staining of the cornea following radial keratotomy.

Although topical anesthesia is commonly used for radial keratotomy, a regional intraorbital block may offer better control of the eye in anxious or uncooperative patients. Some complications of retrobulbar injections can produce severe loss of vision. Besides perforation of the globe and optic atrophy due to nerve perineural or intraneural injection of the anesthetic, posterior orbital hemorrhages have been reported, with consequent proptosis, tightness of the eyelids, and even occlusion of the central retinal artery. We report an unusual complication: blood staining of the cornea.

Liver fatty acid-binding protein in two cases of human lipid storage.

FABPs in the various tissues play an important role in the intracellular fatty acid transport and metabolism. Reye's syndrome (RS) and multisystemic lipid storage (MLS) are human disorders characterized by a disturbance of lipid metabolism of unknown etiology. We investigated for the first time L-FABP in these two conditions. Affinity purified antibodies against chicken L-FABP were raised in rabbits, and found to cross-react specifically with partially purified human L-FABP. L-FABP content in liver samples of two patients with RS and MLS was investigated by immuno-histochemistry, SDS-PAGE and ELISA. L-FABP immuno-histochemistry showed increased reactivity in the liver of RS patient and normal reactivity in MLS liver. L-FABP increase in RS liver was confirmed by densitometry of SDS-PAGE and ELISA method. By these two methods the increase amounted to 180% and 199% (p less than 0.02), respectively, as compared to controls. A possible role of L-FABP in the pathogenesis of RS is discussed.

[Ultrasonography in the diagnosis of chronic inflammatory intestinal disease]. / L'ecotomografia nella diagnosi della malattia infiammatoria cronica intestinale.

The accuracy of ultrasonography (US) in diagnosing active inflammatory bowel disease (IBD) is assessed on the basis of a randomized prospective study of 61 patients. Twenty-six of the patients were affected with Crohn's disease (CD) and 12 with ulcerative colitis, while the remaining 23 patients were control subjects with no specific chronic IBD. The US signs considered as significant for active CD and UC were: --visualization of a typical target image, that is a hyperechoic center corresponding to luminal bowel content, surrounded by a hypoechoic ring corresponding to loop walls; --at least 2 of the following: solid abdominal mass, distended loops, luminal narrowing, reduced peristalsis, stiff loops, and accumulation of fluid between the loops. US sensitivity and specificity for CD were 77% and 95.6%, respectively. As for UC, no significant results were obtained. In our experience, US is a reliable method for detecting alterations and, especially, complications typical of CD in its active phase. Considering the young age of the patients affected with CD and the number of exams they must undergo, US is considered as a useful tool in disease follow-up.