CASP-9: A susceptibility locus for multiple sclerosis in Italy.

Caspase-9 is a primary effector CASP that executes programmed cell death, which plays an important role in the development of multiple sclerosis (MS). Polymorphisms in the CASP-9 gene may influence its activity, thereby modulating the susceptibility to MS. To test this hypothesis, we evaluated a SNP in the CASP-9 gene in a set of Italian patients from Southern Italy and healthy control subjects. Our results suggest that the presence of the G/G genotype represents a higher risk factor in our MS population and a differential production of CASP-9 might be a contributory factor in determining the severity of MS.

The role of VLA4 polymorphisms in multiple sclerosis: an association study.

Lymphocyte and monocyte brain infiltration determines inflammation in multiple sclerosis. The trafficking of these cells into the CNS results from the VLA-4 binding with its ligand on brain endothelial cells. MS patients treated with an antibody against the alpha-4 subunit, which inhibits this interaction, prevents brain lesion development. We investigated the association between VLA-4 gene polymorphisms and MS in a study on 275 patients and 255 controls. No differences were detected, thus suggesting that these polymorphisms are not a significant genetic risk factor for susceptibility to MS in Italy.

Genetic variation in the myeloperoxidase gene and cognitive impairment in multiple sclerosis.

There is evidence that multiple sclerosis (MS) may associated with cognitive impairment in 25 to 40% of cases. The gene encoding myeloperoxidase (MPO) is involved in molecular pathways leading to beta-amyloid deposition. We investigated a functional biallelic (G/A) polymorphism in the promoter region (-463) of the MPO gene in 465 patients affected by MS, divided into 204 cognitively normal and 261 impaired. We did not find significant differences in allele or genotype distributions between impaired and preserved MS patients. Our findings suggest that MPO polymorphism is not a risk factor for cognitive impairment in MS.

GABA(B) receptor 1 polymorphism (G1465A) is associated with temporal lobe epilepsy.

BACKGROUND: Dysfunction of gamma-aminobutyric acid (GABA) (B) receptors has been implicated in the pathogenesis of temporal lobe epilepsy (TLE). OBJECTIVE: To evaluate the genetic contribution of cloned human GABA(B) receptors to TLE. METHODS: The authors genotyped 141 patients (78 women and 63 men; mean age = 49.1 +/- 18.0 years) with nonlesional TLE and 372 age- and sex-matched normal individuals for the known polymorphism G1465A in the human GABA(B) receptor 1 [GABA(B[1])] gene. RESULTS: There was a highly significant overrepresentation of the G1465A heterozygote in patients with TLE compared with controls. The A/G genotype was found in 17% of the 141 patients with TLE and in only 0.5% of the 372 controls (p < 0.0001). The authors also found that patients carrying the A allele had a significantly higher risk (p = 0.003, OR = 6.47, 95% CI = 2.02 to 20.76) of developing drug-resistant TLE. Furthermore, the age at onset of seizures tended to be lower in patients with A/G genotype, but the difference was not significant. CONCLUSIONS: The results of this study indicate that the GABA(B[1]) polymorphism (G1465A) confers a highly increased susceptibility to TLE. Moreover, it seems to influence the severity of this common epileptic disorder.

Genetic association of alpha2-macroglobulin polymorphisms with AD in southern Italy.

The authors investigated the segregation of two polymorphisms of the alpha2-macroglobulin gene (A2M-I/D and A2M-Ile1000Val) in patients with sporadic AD from southern Italy. The A2M-I and A2M-Val1000 alleles were more frequent in cases than in controls, and this effect was independent from the APOE-epsilon4 status as well as from the age at onset of AD. Moreover, subjects carrying the A2M genotype I/I-Val/Val had a threefold increase of risk for AD. These data support a population-based susceptibility for AD linked to A2M polymorphisms.

Serine base exchange enzyme in porcine lyophilised platelets: enzyme properties and modulation by AlF4- and different types of heparin.

Phosphatidylserine is one of the PKC modulators and thus it may play an important role in signal transduction. Regulation of the synthesis of this phospholipid is not yet clarified. The contrasting reports are possibly related to the existence of different enzymes which, in mammalian tissues, catalyse the exchange between free serine and the nitrogen base of a membrane phospholipid. This study demonstrates that serine base exchange reactions of commercially available lyophilised porcine platelets exhibit similar pH optima, temperature and Ca2+ dependence as observed in fresh tissues. Analysis of fatty acids composition of the three phospholipid classes involved in base exchange reactions also demonstrated a similarity with fresh platelets. Serine and ethanolamine base exchange enzyme activities were assayed in parallel in platelet lysate subjected to preincubation at various temperatures (30-60 degrees C). When dithioerithrol was omitted from the incubation medium, the two base exchange reactions were inhibited with a similar temperature-dependent pattern. Addition of the reducing agent enhanced the sensitivity to preincubation only for the serine base exchange reaction which was inhibited by 80% after preincubation at 45 degrees C. With respect to its regulation, porcine platelet serine base exchange enzyme(s) was inhibited by fluoroalluminate, a widely used G-protein activator, and stimulated by unfractionated heparin. Low mol. wt. heparin did not influence enzyme activity. Unfractionated heparin greatly stimulated SBEE activity assayed at pH 7.4, a pH value far from the optimal pH.

Dopamine D2 receptor gene polymorphism and the risk of levodopa-induced dyskinesias in PD.

OBJECTIVE: To investigate whether polymorphisms in the genes for dopamine receptors D1 and D2 are associated with the risk of developing peak-dose dyskinesias in PD. BACKGROUND: Peak-dose dyskinesias are the most common side effects of levodopa therapy for PD. The identified predictors may only partially account for the risk of developing peak-dose dyskinesias because a substantial proportion of patients never develop peak-dose dyskinesias. Genetic factors could play a role in determining the occurrence of peak-dose dyskinesias. METHODS: A case-control study of 136 subjects with sporadic PD and 224 population control subjects. We studied three polymorphisms involving the dopamine receptor D1 gene and one intronic short tandem repeat polymorphism of the dopamine receptor D2 gene. RESULTS: The polymorphisms of the dopamine receptor D1 gene were not associated with the risk of developing PD or peak-dose dyskinesias. The 15 allele of the polymorphism of the dopamine receptor D2 gene was more frequent in parkinsonian subjects than in control subjects. More important, the frequency of both the 13 allele and the 14 allele of the dopamine receptor D2 gene polymorphism was higher in nondyskinetic than in the dyskinetic PD subjects. The risk reduction of developing peak-dose dyskinesias for PD subjects carrying at least 1 of the 13 or 14 alleles was 72% with respect to the PD subjects who did not carry these alleles. CONCLUSIONS: Certain alleles of the short tandem repeat polymorphism of the dopamine receptor D2 gene reduce the risk of developing peak-dose dyskinesias and could contribute to varying susceptibility to develop peak-dose dyskinesias during levodopa therapy.

APOE and risk of cognitive impairment in multiple sclerosis.

OBJECTIVES: The APOE gene polymorphism and the -491 A/T polymorphism in its regulatory region have been associated with an increased risk for developing Alzheimer's disease. We examined these polymorphisms in multiple sclerosis (MS) patients, to determine if a genetic predisposition may explain the risk for developing cognitive decline in MS. MATERIAL AND METHODS: Eighty-nine relapsing-remitting and secondary progressive MS patients underwent to a full neuropsychological battery as well as to determination of APOE and -491 A/T polymorphisms. Genetic analysis was also performed in 107 population controls. RESULTS: The APOE polymorphism was not associated with the risk of cognitive impairment in MS patients. The AA genotype of the -491 A/T polymorphism in the APOE regulatory region was more frequent in cognitively impaired than in cognitively preserved MS subjects. CONCLUSION: The AA homozygous state of the -491 A/T polymorphism of the APOE regulatory region is associated with cognitive impairment in patients with MS.

Reboxetine in the treatment of depression in the elderly: pilot study.

Elderly patients are particularly susceptible to the potential side effects of current antidepressants due to agerelated physiologic changes. We report a pilot study to examine the tolerability of increasing doses of reboxetine, a selective noradrenaline reuptake inhibitor (selective NRI), in elderly depressed patients. Twelve elderly female patients (75-87 years) with either major depression or dysthymia received reboxetine titrated to 8 mg/day over a 4-week period. Tolerability was assessed and included the measurement of vital signs. Electrocardiograms were recorded at baseline and on days 14 and 28. Newly emergent signs and symptoms were recorded throughout the study. Efficacy was assessed using four rating scales, including the Clinical Global Impression (CGI) scale and Hamilton Depression Rating Scale (HAM-D). Eleven patients completed the study, nine received the maximal dose of reboxetine 8 mg/day, and two received maximum doses of reboxetine 6 mg/day due to cardiac rhythm changes in week 3. A total of five patients experienced tachycardia (including two with cardiac rhythm changes in week 3). At the end of the study, seven patients were "much" to "very much" improved on the CGI scale with a concomitant decrease in HAM-D total score of 22% to 41%. Reboxetine was well tolerated by the majority of patients and efficacy outweighed side effects in 75% of patients. Reboxetine 4 mg/day, increasing to 6 mg/day on the basis of individual patient tolerability, may be considered as a safe dose range for testing the efficacy and tolerability of reboxetine in long-term controlled clinical trials in elderly patients with depression.

Synthesis of ethanolamine phosphoglycerides in rat cerebral cortex subjected in vitro to experimental hypoxia with and without hypocapnia.

Slices and homogenates from rat cerebral cortex were used to study the effect of hypoxia, with or without hypocapnia, on phosphatidylethanolamine synthesis. The incorporation of [1-3H]ethanolamine into the corresponding phospholipid was greatest in slices treated with pure nitrogen, intermediate when the nitrogen contained 5% CO2, and least in slices treated with 95% O2-5% CO2. The role of hypocapnia in reinforcing the effect due to hypoxia did not require the integrity of the cell because similar results were obtained by treating homogenates with pure nitrogen or nitrogen plus 5% CO2. In both cases the synthesis of phosphatidylethanolamine was abolished by the addition of EGTA and the degradation of newly synthesized phospholipid by phospholipases was similar to that obtained in controls. When the homogenate was not buffered, changes in the pH due to experimental treatment influenced the response to Ca2+ and to hypoxia plus hypocapnia. Intracellular calcium ions are thought to play a role in the response of cerebrocortical slices to N2-treatment. In fact, although the incorporation was greater in complete medium that contains 2 mM Ca2+ than in the same medium prepared without the addition of this ion, the relative increase of incorporation due to N2-treatment was greater in the medium lacking added Ca2+.

Different mechanisms regulate phosphatidylserine synthesis in rat cerebral cortex.

Transduction of extracellular signals through the membrane involves both the lipid and protein moiety. Phosphatidylserine participates to these processes as a cofactor for protein kinase C activity and thus the existence of a regulatory mechanism for its synthesis ought to be expected. In plasma membranes from rat cerebral cortex, the activity of serine base exchange enzyme, that is mainly responsible for phosphatidylserine synthesis in mammalian tissues, was reduced by the addition to the incubation mixture of AlF4- or GTP-gamma-S, known activators of G proteins, whereas ATP was almost uneffective. GTP-gamma-S inhibited the enzyme activity only at relatively high concentration (> 0.5 mM). When the synthesis of phosphatidylserine in the same cerebral area was investigated by measuring the incorporation of labelled serine into the phospholipid in the homogenate buffered at pH 7.6, ATP had an inhibitory effect as GTP-gamma-S and AlF4-. Heparin activated both serine base exchange enzyme in plasma membranes and phosphatidylserine synthesis in the homogenate. The preincubation of plasma membranes in the buffer without any other addition at 37 degrees C for 15 min reduced by 30% serine base exchange enzyme activity. The remaining activity responded to the addition of GTP-gamma-S but was insensitive to 5 mM AlF-4, a concentration that inhibited by 60% the enzyme assayed without preincubation. These results indicate the existence of different regulatory mechanisms, involving ATP and G proteins, possibly acting on different enzymes responsible for the synthesis of phosphatidylserine. Since previous studies have shown that hypoxia increases the synthesis of this phospholipid in brain slices or homogenate (Mozzi et al. Mol Cell Biochem 126: 101-107, 1993), it is possible that hypoxia may interfere with at least one of these mechanisms. This hypothesis is supported by the observation that in hypoxic homogenate 20 mM AlF-4 was not able to reduce the synthesis of phosphatidylserine as in normoxic samples. A similar difference between oxygenated and hypoxic samples, concerning their response to AlF4-, was observed when the incorporation of ethanolamine into phosphatidylethanolamine was studied. The incorporation of choline into phosphatidilcholine was, on the contrary, inhibited at a similar extent in both experimental conditions.

Severe erythrocyte adenylate kinase deficiency due to homozygous A-->G substitution at codon 164 of human AK1 gene associated with chronic haemolytic anaemia.

A child of Italian origin with a congenital haemolytic anaemia had spectrophotometrically undetectable erythrocyte adenylate kinase (AK) activity. Her parents and brother had approximately 50% normal AK activity, and AK electrophoresis of red blood cell (RBC) crude extract on cellulose acetate strips showed the presence of the normal allele AK1-1. No AK band was detected in the AK electrophoresis of the proband, in whom the erythrocyte 2,3-diphosphoglycerate (2,3DPG) and glutathione (GSH) concentrations were normal whereas adenosine triphosphate (ATP) concentration, pyruvate kinase (PK) and glucose-6P-dehydrogenase (G6PD) activities were increased, reflecting the high reticulocyte count (6.9%). No other evident enzymatic defect was detected by standard procedures. Analysis of AK gene exons, based on polymerase chain reaction-single-strand conformational polymorphism (PCR-SSCP), clearly showed an abnormality in the fragment containing exon 6. The subsequent sequence analysis of this abnormal fragment revealed homozygous and heterozygous A-->G substitutions in the proband and in the parents and brother respectively at codon 164, corresponding to a tyrosine-->cysteine substitution in the AK protein.

Phosphatidylserine synthesis in rat cerebral cortex: effects of hypoxia, hypocapnia and development.

Phosphatidylserine, which is necessary for protein kinase C activity, is synthesized in mammalian tissues by the Ca(2+)-dependent base exchange enzyme. The synthesis of phosphatidylserine is greater in slices or homogenates of rat cerebral cortex subjected to hypoxia by N2 treatment when compared with O2 plus 5% CO2. An intermediate effect was observed when the treatment was done with N2 plus 5% CO2. Incorporation rates were dependent on Ca2+ in Krebs-Henseleit Ringer bicarbonate medium, being greater with 2 mM Ca2+ than with the same medium prepared without Ca2+. The increase of phosphatidylserine synthesis, due to hypoxia, was, on the contrary, more evident in the medium lacking added Ca2+. Similar results were obtained with the homogenates. This suggests that elevation of intracellular Ca2+, caused by hypocapnia and hypoxia, may be responsible for the greater incorporation of serine into phosphatidylserine. In both cerebrocortical slices and homogenate, [14C]serine incorporation decreased with development both in O2 plus 5% CO2 and N2-treated preparations. However, in younger rats (14-18 days) hypoxia induced a lesser increase of phosphatidylserine than in 40 day old animals. We suggest that a regulatory mechanism for phosphatidylserine synthesis is established during development and that N2-treatment can increase phosphatidylserine synthesis by interfering with this regulatory mechanism.

The complexity of psychiatric nosography and the "simplicity" of molecular genetics.

Are molecular genetic approaches, related at the present time to psychiatric categories, mostly conventional? If the psychiatric field is to be related with the plasticity of the brain, and then to its ability to be reorganized relative to experiences, it seems to be more important to study the relation between genetics and brain plasticity. It therefore seems more significant today, to be raising questions rather than gathering data.

Affective disorders and S-adenosylmethionine: a new hypothesis.

S-Adenosyl-L-methionine (AdoMet) is a safe and probably effective antidepressant agent in certain forms of clinical depression. This article presents a new hypothesis to account for the mechanism of action of S-adenosylmethionine in such illnesses, based upon the known biochemistry of this compound, and upon current knowledge of clinical and genetic aspects of affective disorders. Giulio Cantoni, S. Harvey Mudd and V. Andreoli postulate that at least some major mood disorders are due to abnormalities affecting the AdoMet-dependent methylation of a substance in the CNS. For convenience and without prejudging the chemical structure of this substance, they call it 'barinine'. The model requires that barinine be subject to AdoMet-dependent methylation and that methylbarinine be subject to metabolic demethylation to regenerate the original barinine. Methylbarinine should be mood elevating, whereas barinine itself should not be. Depression is a result of abnormalities lowering the normal steady-state concentration of methylbarinine, whereas mania results from an abnormal elevation of methylbarinine.

Promazine. A major plasma metabolite of chlorpromazine in a population of chronic schizophrenics.

N-Demethylation and dehalogenation of chlorpromazine (CPZ) were compared in six psychotic inpatients and in rats orally treated for 4 weeks with a daily CPZ dose of 5.4 (mean value) and 20 mg X kg-1 body weight, respectively, by measuring drug and metabolite plasma levels by means of a gas-liquid chromatography-nitrogen/phosphorus detector method. In patients the major plasma metabolite was found to be promazine (PZ), as identified by capillary GC-MS analysis. In rats, on the contrary, PZ represented only a small proportion of the compounds detected in plasma. The mean [PZ]/[CPZ] ratio after 4 weeks of treatment was 1.64 in patients and 0.08 in rats. The relative frequency of the N-demethyl metabolites in plasma, however, was similar in the two species. The mean [N-monodemethylated CPZ]/[CPZ] and [N-didemethylated CPZ]/[CPZ] ratios after 4 weeks of treatment were 0.45 and 0.24 in patients and 0.56 and 0.25 in rats, respectively. These findings suggest that dechlorination of CPZ in psychotic patients represents an important metabolic pathway.