Developmental regulation of SMN expression: pathophysiological implications and perspectives for therapy development in spinal muscular atrophy.

Spinal muscular atrophy (SMA), the predominant form of motoneuron disease in children and young adults is caused by loss of function of the SMN protein. On the basis of a disrupted splice acceptor site in exon 7, transcripts from a second SMN gene in humans called SMN2 cannot give rise to SMN protein at sufficient levels for maintaining function of motoneurons and motor circuits. First clinical trials with Spinraza/Nusinersen, a drug that counteracts disrupted splicing of SMN2 transcripts, have shown that elevating SMN levels can successfully interfere with motoneuron dysfunction. This review summarizes current knowledge about the pathophysiological alterations in Smn-deficient motoneurons, which lead to defective neuromuscular transmission and altered spinal circuit formation. Both pathological mechanisms are important targets for therapeutic intervention. However, the developmental time window when therapeutic interventions ideally should start is not known. Endogenous SMN expression both from SMN1 and SMN2 genes is high at early developmental stages and declines progressively in humans and mice. Thus, therapeutic SMN upregulation should start just before SMN declines below a critical threshold, and before irreversible defects occur at neuromuscular junctions and in spinal circuits. Previous results indicate that loss of Smn function leads to synaptic dysfunction during a stage of neuromuscular development when synaptic strength determines which synapses are maintained or not. This time window appears as an important target for therapy, which possibly could be supported by additional strategies that strengthen synaptic transmission.

Stiff person syndrome associated anti-amphiphysin antibodies reduce GABA associated [Ca(2+)]i rise in embryonic motoneurons.

Autoantibodies to the synaptic protein amphiphysin play a crucial pathogenic role in paraneoplastic stiff-person syndrome. Impairment of GABAergic inhibition is the presumed pathophysiological mechanism by which these autoantibodies become pathogenic. Here we used calcium imaging on rat embryonic motor neurons to investigate whether antibodies to amphiphysin directly hinder GABAergic signaling. We found that the immunoglobulin G fraction from a patient with stiff-person syndrome, containing high titer antibodies to amphiphysin and inducing stiffness in rats upon passive transfer, reduced GABA-induced calcium influx in embryonic motor neurons. Depletion of the anti-amphiphysin fraction from the patient's IgG by selective affinity chromatography abolished this effect, showing its specificity for amphiphysin. Quantification of the surface expression of the Na(+)/K(+)/2Cl(2-) cotransporter revealed a reduction after incubation with anti-amphiphysin IgG, which is concordant with a lower intracellular chloride concentration and thus impairment of GABA mediated calcium influx. Thus, anti-amphiphysin antibodies exert a direct effect on GABA signaling, which is likely to contribute to the pathogenesis of SPS.

Principles of induction chemotherapy for non-small cell lung cancer.

The results of lung cancer treatment have not significantly improved for many years. About 35% of patients with non-small cell lung cancer (NSCLC) are in clinical stage IIIA. Clinically asymptomatic distant metastases occur in the majority of these patients. In such cases only combined treatment offers a chance of cure. In the Chest Surgery Center in Lublin a clinical trial was carried out aimed to assess late results of combined treatment in patients with IIIA NSCLC. Over 700 patients were enrolled in the study. The results of the trial disclosed, that neoadjuvant chemotherapy prolonged life of the operated patients and improved their life quality. However, a question of qualification for this complex treatment and complexity of assessment criteria, still remain to be answered.

Co-regulation of survival of motor neuron (SMN) protein and its interactor SIP1 during development and in spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disease characterized by the degeneration of motor neurons in the spinal cord. The disease is caused by mutations of the survival of motor neuron 1 gene (SMN1), resulting in a reduced production of functional SMN protein. A major question unanswered thus far is why reduced amounts of ubiquitously expressed SMN protein specifically cause the degeneration of motor neurons without affecting other somatic cell types. In a first attempt to address this issue we have investigated the Smn interacting protein 1 (Sip1), with an emphasis on its developmental expression and subcellular distribution in spinal motor neurons in relation to Smn. By confocal immunofluorescence studies we provide evidence that a significant amount of Smn does not co-localize with Sip1 in neurites of motor neurons, indicating that Smn may exert motor neuron-specific functions that are not dependent on Sip1. Sip1 is highly expressed in the spinal cord during early development and expression decreases in parallel with Smn during postnatal development. Strikingly, reduced production of Smn as observed in cell lines derived from SMA patients or in a mouse model for SMA coincides with a simultaneous reduction of Sip1. The finding that expression of Sip1 and Smn is tightly co-regulated, together with the unique localization of Smn in neurites, may help in understanding the motor neuron-specific defects observed in SMA patients.

The role of SMN in spinal muscular atrophy.

Childhood spinal muscular atrophy (SMA) is a common autosomal recessive disorder which is characterized by muscle weakness due to degeneration of motoneurons in the spinal cord and brainstem nuclei. Positional cloning strategies have revealed several gene candidates including the genes for the survival motoneuron (SMN) and the neuronal apoptosis inhibitory protein (NAIP). Both genes are duplicated on chromosome 5. Homozygous deletions/mutations of the telomeric SMN gene, which is expressed from both copies on human chromosome 5, are associated with the disease. Recent reports suggest involvement of the SMN protein in the formation of spliceosomal particles in the cytoplasm and in the regeneration of spliceosomes in the nucleus. These data put spinal muscular atrophy into a growing group of disorders of RNA metabolism which also include fragile-X syndrome and myotonic dystrophy. Relevance of these previous data for the pathogenesis of the disease are discussed in this review.

The human centromeric survival motor neuron gene (SMN2) rescues embryonic lethality in Smn(-/-) mice and results in a mouse with spinal muscular atrophy.

Proximal spinal muscular atrophy (SMA) is a common motor neuron disease in humans and in its most severe form causes death by the age of 2 years. It is caused by defects in the telomeric survival motor neuron gene ( SMN1 ), but patients retain at least one copy of a highly homologous gene, centromeric SMN ( SMN2 ). Mice possess only one survival motor neuron gene ( Smn ) whose loss is embryonic lethal. Therefore, to obtain a mouse model of SMA we created transgenic mice that express human SMN2 and mated these onto the null Smn (-/-)background. We show that Smn (-/-); SMN2 mice carrying one or two copies of the transgene have normal numbers of motor neurons at birth, but vastly reduced numbers by postnatal day 5, and subsequently die. This closely resembles a severe type I SMA phenotype in humans and is the first report of an animal model of the disease. Eight copies of the transgene rescues this phenotype in the mice indicating that phenotypic severity can be modulated by SMN2 copy number. These results show that SMA is caused by insufficient SMN production by the SMN2 gene and that increased expression of the SMN2 gene may provide a strategy for treating SMA patients.

Reduced survival motor neuron (Smn) gene dose in mice leads to motor neuron degeneration: an animal model for spinal muscular atrophy type III.

Spinal muscular atrophy (SMA) is caused by deletion or specific mutations of the telomeric survival motor neuron ( SMN ) gene on human chromosome 5. The human SMN gene, in contrast to the Smn gene in mouse, is duplicated and the centromeric copy on chromosome 5 codes for transcripts which preferentially lead to C-terminally truncated SMN protein. Here we show that a 46% reduction of Smn protein levels in the spinal cord of Smn heterozygous mice leads to a marked loss of the cytoplasmic Smn pool and motor neuron degeneration resembling spinal muscular atrophy type 3. Smn heterozygous mice described here thus represent a model for the human disease. These mice could allow screening for SMA therapies and help in gaining further understanding of the pathophysiological events leading to motor neuron degeneration in SMA.

[Surgical treatment of advanced lung emphysema. Does it really improve treatment results?]. / Chirurgiczne leczenie zaawansowanej rozedmy pluc. Czyzby nowa nadzieja na poprawe wyników leczenia?

Lung emphysema may be one of the components of chronic obstructive pulmonary disease (COPD). Nearly 60% of patients with advanced chronic respiratory insufficiency with FEV1 < 30% expected, aged > 65 years die within the first 2 years of the disease. The treatment of COPD includes oxygen therapy, lung transplantation and/or lung volume reduction (LVR). The study presents qualification criteria for LVR in reference to oxygen therapy and lung transplantation. Crucial diagnostic features include: CT of the chest with densitometric analysis and lung perfusion scans. Patients with severe but limited dyspnea, disseminated emphysema lesions with fairly untouched areas of normal lung tissue, FEV1% < 30-35%, DLCO < 25% expected, PaCO2 < or = 50 mmHg and PaO2 < or = 50-55 mmHg, appear to be the best candidates for LVR. The results of the study indicate, that in Poland, patients with advanced emphysema, who are treated with oxygen therapy or soon will be qualified this therapy or await lung transplantation might be candidates for LVR. The early results of lung volume reduction are promising and include: decrease in total lung capacity, residual volume and breathing frequency. LVR has been in use for about 3 years and majority of authors still do not possess clinical observations long enough to obtain statistically significant comparison with other methods of treatment. Limited morbidity and low perioperative mortality enable application of LVR even in patients with severe respiratory insufficiency and/or requiring constant oxygen therapy. In conclusion considering all positive aspects of LVR it is necessary to emphasise that for overall assessment of this method a broader clinical material is required including data on duration of clinical improvement.

[Lung neoplasms in patients under 40 years of age. Treatment results in patients qualifying for surgical treatment]. / Rak pluca u chorych przed 40 rokiem zycia. Wyniki leczenia chorych kwalifikowanych do leczenia operacyjnego.

In last decades an increase of new cases of lung cancer has been observed. The global prognosis based on the analysis of the increase rate of new cases envisages that total number of lung cancer patients will reach 2 millions at the end of the XXth century. In Poland alone, the number of new cases can come up to 50,000 a year in the second decade of the XXIst century. Respectively, the number of surgically treated patients with lung cancer increases. In literature, controversial opinions on the results of surgical therapy of patients with lung cancer younger than 40 years can be found. Some authors report unsatisfactory results. The others do not confirm differences due to the age of operated patients. We present our observations based on the clinical analysis of 46 patients below 40 years of age treated surgically because of lung cancer. The aim of this analysis is the evaluation of the long-term results of surgical therapy in this group of patients. This is the first Polish report on this problem.

Relationships between divisions of the lingular bronchus and vascularization patterns in the lingula.

In 100 left human lungs the main bronchus, the pulmonary artery and the pulmonary veins were injected with 65% methyl methacrylate (Duracryl) and then digested in sulphuric acid. The resulting specimens were studied concerning the divisions of the lingular bronchus and the types of arterial and venous vascularization of the lingula. As a rule the lingular bronchus divided into two segmental bronchi. A single lingular artery was found in 80% of the cases and a single lingular vein in 58%. Atypical bronchial divisions were almost always associated with unusual types of vascularization. Patterns of bronchial division showed complete concordance with those of arterial vascularization of the lingula in 64% of the lungs and consistency with venous drainage patterns in 54%.

Oguchi disease: suggestion of linkage to markers on chromosome 2q.

Oguchi disease is a rare autosomal recessive form of congenital stationary night blindness. The condition is associated with fundus discolouration and abnormally slow dark adaptation. Earlier studies suggested that the 48 kD protein S antigen may be involved in the recovery phase of light transduction. Previous cytogenetic and linkage studies have localised the S antigen gene (SAG) to chromosome 2q37.1. In the present study markers which map to distal chromosome 2q were typed in an inbred Oguchi pedigree. The segregation data obtained suggested that the affected subjects are homozygous by descent for a region between D2S172 and D2S345. An intragenic SAG polymorphism was homozygous in all affected people and a recombination event suggested that SAG maps proximal to D2S345. Collectively, these findings support the hypothesis that a defect in S antigen may be responsible for Oguchi disease.

The estimation of factor VII in livestock plasma of domestic animals by the use of tripeptide chromogenic substrate.

The levels of factor VII in horse, cattle, sheep and pig plasmas were estimated by the use of the synthetic chromogenic substrate, benzoyl-isoleucyl-glutamyl-glycyl-arginyl-p-nitroanilide (S-2222). The highest level of factor VII was observed in the cattle plasma, the lowest one in the plasma of the pig.

Assay for endogenous heparin in plasma of livestock using a synthetic chromogenic substrate.

The levels of endogenous heparin in the plasmas of horses, cows, sheep and pigs were determined with the use of synthetic chromogenic substrate benzoyl-isoleucyl-glutamyl-glycyl-arginyl-p-nitroanilide (S-2222). The lowest heparin concentrations were stated in cattle plasma, the highest ones in the plasma of pigs.

The estimation of factor VIII levels in horse, cattle, sheep and pig plasma by the use of synthetic chromogenic substrate.

Factor VIII level in horse, cattle, sheep and pig plasma was estimated by the use of synthetic chromogenic substrate S-2222 (benzoyl-isoleucyl-glutamyl-glycyl-arginyl-p-nitronilide). The highest level of this factor was stated in pig, the lowest one in sheep plasma.

The influence of Ambroxol on peroxidative processes in lung and plasma in dogs after pulmonectomy.

The purpose of the studies was to determine the effect of Ambroxol on the activity of antioxidant enzymes and on the glutathione level as well as the intensity of the peroxidative processes in the lung tissue, alveolar macrophages and plasma in dogs after unilateral pulmonectomy. On the 2nd and 6th day after the surgery the activity of antioxidant enzymes and the glutathione level were studied in the remaining lung. The levels of the lipid peroxidation products were determined in the analogous system. In both examined groups the increase in the antioxidant enzyme activity and the lipid peroxidation product levels was observed in the remaining lung after the surgery. In Ambroxol-treated animals the statistically significant increase in the antioxidant enzyme activity was noted while the intensity of peroxidative processes was found to be lower. This fact may suggest that Ambroxol stimulates the resistance of the lung tissue to the free radical activity and inhibits the lung peroxidative processes in dogs after pulmonectomy.

[The estimation of platelet factor 3 activity in horses, cattle, sheep and pigs by the use of synthetic chromogenic tripeptide substrates]. / Oznaczanie aktywnosci czynnika plytkowego 3 u koni, bydla, owiec i swin przy uzyciu chromogennego substratu trójpeptydowego.

The aim of this study was to investigate the platelet factor 3 activity in platelet-rich plasma of horse, cattle, sheep and pig, by the use of chromogenic tripeptide substrate H-D-Phenylalanyl-Pipecolyl-Arginyl-p-nitroanilide. Among species examined the highest activity of this factor was stated in pig, the lowest one in sheep. Chromogenic substrate test was 10 times more sensitive that Stypven clotting time test. Thus, the use of chromogenic tripeptide substrate is fully valuable in platelet factor 3 activity estimation in domestic animals.

[The effect of ambroxol on peroxidative processes in dog lung mitochondria]. / Wplyw ambroksolu na procesy peroksydacyjne w mitochondriach pluc psa.

The influence of Ambroxol on the Fe3+ (ADP) dihydroxyfumarate-induced peroxidation of mitochondrial lipids in dog lung was investigated. It was shown that changes in mitochondrial lipid composition include great decrease in phosphatidylcholine and phosphatidylethanolamine content, with concomitant increase in lysophosphoglyceride content. Addition of Ambroxol (5 mM and 10 mM) to the incubation medium decreases these changes in dose-dependent manner. The malondialdehyde production was greatly reduced in the presence of Ambroxol. The mode of Ambroxol action was comparable to that of other free radical scavengers.

[Effect of ambroxol on surfactant lipids in patients after resection of pulmonary tissue]. / Wplyw ambroxolu na lipidy surfaktantu u chorych po wycieciu miazszu plucnego.

Effect of Ambroxol on phospholipids and higher fatty acids of pulmonary surfactant was studied in patients with lung cancer undergoing pulmonary resection. Intravenous administration of Ambroxol was carried out for 8 days in a dose 1000 mg per day before pulmonary resection. It produced elevation of disaturated phosphatidylcholine and the general level of phosphatidylcholine. Also increase of palmitic acid and saturated fatty acids of the phospholipid fraction was found. The results of this study show that Ambroxol on a short time elevates levels of active substances of pulmonary surfactant and decreases the destructive processes of pulmonary parenchyma.