A late onset variant of ataxia-telangiectasia with a compound heterozygous genotype, A8030G/7481insA.
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Adenine↗
Biomedical subjects
A Brusco
Publications and source records attributed to A Brusco.
Increased nitric oxide synthase activity in a model of serotonin depletion.
Serotonin (5HT) containing cell bodies are localized in mesencephalic and rhombencephalic raphe nuclei. It has been proposed that 5HT could be involved in neuronal development and plasticity. In the central nervous system, nitric oxide (NO) has been postulated as a neurotransmitter and neuromodulator, and has been implicated in neurotoxicity as well as in neuroprotection. Using the nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) technique, NO synthesizing neurons were described in raphe nuclei. By immunohistochemistry, nitric oxide synthase (NOS) was found colocalized with 5HT in some dorsal raphe nucleus (DRN) neurons. In a model of inhibition of 5HT synthesis produced by daily administration of parachlorophenilalanine during 14 days, we have studied the relationship between 5HT and NO systems after 5HT depletion by histochemical and immunocytochemical methods. After the treatment, we observed an important reduction of 5HT immunostaining in the DRN and enhanced NOS activity demonstrated by NADPH-d technique, especially in the dorsomedial and ventromedial subgroups. In spite of the increased NOS activity, we could not observe significant changes in the NOS-immunoreactivity in the DRN after 5HT depletion. These results could indicate that 5HT depletion is concomitant with changes in NOS activity without affecting NOS expression in the DRN.
Animals↗
TSC1 and TSC2 deletions differ in size, preference for recombinatorial sequences, and location within the gene.
Large TSC gene rearrangements are not rare findings in tuberous sclerosis. Interestingly, all deletions, duplications and inversions so far described involve TSC2, none being associated with TSC1. In order to shed light on the structural basis of the preferential DNA rearrangements in TSC2 over TSC1 and to assess, in an unselected patient population, the prevalence of large re-arrangements in both TSC loci, we screened 202 tuberous sclerosis patients consecutively referred at our center. Southern blot analysis on EcoRI+HindIII double-digested DNA identified 19 partial or full-length gene deletions: three involved TSC1 and sixteen TSC2. The breakpoint sequence of seven internal deletions, three in TSC1 and four in TSC2, allowed us to speculate on the mechanism favoring TSC2 unequal recombinations and to identify a deletion hot spot that lies in TSC1 and that may be relevant in the routine genetic testing of tuberous sclerosis. Briefly, three major features appear to distinguish TSC1 from TSC2 deletions: (1) deletion size: all TSC1 deletions are within the transcriptional unit, whereas 12 of the 16 TSC2 deletions have at least one external breakpoint; (2) location within the gene: all TSC1 deletions are confined to the 3'end of the gene (all three 5' breakpoints being located in intron 20) thus resulting in the same frameshift mutation following amino acid K875, whereas the TSC2 internal breakpoints appear to be scattered along the gene; (3) preference for recombinatorial sequences: six out of eight internal TSC2 breakpoints map within Alu repeats, whereas none of the three TSC1 deletions appear to be Alu-mediated. Indeed, in the latter gene, unique structural features (a purine-rich tract flanked by pyrimidine-rich segments) surrounding one of the two identified breakpoint cluster regions might play a role in promoting inappropriate recombinations.
Base Sequence↗
Morphological study of 5-HT neurons and astroglial cells on brain of adult rats perinatal or chronically exposed to 2,4-dichlorophenoxyacetic acid.
2,4-D is a chlorophenoxyherbicide used worldwide. We have studied the morphological alterations of 5-HT neurons and glial cells in the mesencephalic nuclei of adult rats exposed to 2,4-D both perinatally (during pregnancy and lactation) and chronically (during pregnancy, lactation and after weaning) with quantitative methods. Pregnant rats were daily exposed to 70 mg/kg of 2,4-D from gestation day (GD) 16 to post-natal day (PND) 23 through diet. After weaning, pups were assigned to one of two sub-groups: T1 (fed with untreated diet until PND 90) and T2 (maintained with 2,4-D diet until PND 90). Brain sections were immunocytochemically stained using polyclonal anti-5-HT, anti-GFAP and anti-S-100 protein antibodies as cells markers. 2,4-D exposure during pregnancy and lactancy (T1 group) produced an increase in 5-HT neuronal area and immunoreactivity (IR) in the mesencephalic nuclei studied. However, with the chronical 2,4-D exposure (T2 group) only the 5-HT neuronal area from the dorsal raphe nucleus (DRN) was increased, suggesting an adaptable response of 5-HT neurons in median raphe nucleus (MRN). The presence of reactive astrocytes in mesencephalic nuclei and in hippocampus were also different for the two 2,4-D exposure designs, showing the existence of a correspondence between neuronal changes and astrogliosis. Results support evidences that 2,4-D alters the serotoninergic system and that 5-HT neurons of each mesencephalic nuclei show different responses to the 2,4-D exposure designs which are parallel to astrogliosis.
2,4-Dichlorophenoxyacetic Acid↗
Myopathy in a patient with chromosome 22q11 deletion.
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Abnormalities, Multiple↗
Defective function of Fas in patients with type 1 diabetes associated with other autoimmune diseases.
Fas (CD95) triggers programmed cell death and is involved in cell-mediated cytotoxicity and in shutting off the immune response. Inherited loss-of-function mutations hitting the Fas system cause the autoimmune/lymphoproliferative syndrome (ALPS). We have recently shown that ALPS patients' families display increased frequency of common autoimmune diseases, including type 1 diabetes. This work evaluates Fas function in type 1 diabetic patients without typical ALPS. Cell death induced by anti-Fas monoclonal antibody was investigated in T-cells from 13 patients with type 1 diabetes alone and 19 patients with type 1 diabetes plus other autoimmune diseases (IDDM-P). Moreover, we analyzed 19 patients with thyroiditis alone (TYR), because most IDDM-P patients displayed thyroiditis. Frequency of resistance to Fas-induced cell death was significantly higher in patients with IDDM-P (73%) than in type 1 diabetic (23%) or TYR (16%) patients or in normal control subjects (3%). The defect was specific because resistance to methyl-prednisolone-induced cell death was not significantly increased in any group. Fas was always expressed at normal levels, and no Fas mutations were detected in four Fas-resistant IDDM-P patients. Analysis of the families of two Fas-resistant patients showing that several members were Fas-resistant suggests that the defect has a genetic component. Moreover, somatic fusion of T-cells from Fas-resistant subjects and the Fas-sensitive HUT78 cell line generates Fas-resistant hybrid cells, which suggests that the Fas resistance is due to molecules exerting a dominant-negative effect on a normal Fas system. These data suggest that Fas defects may be a genetic factor involved in the development of polyreactive type 1 diabetes.
Adolescent↗
Neuroglial interactions in a model of para-chlorophenylalanine-induced serotonin depletion.
Serotonin (5HT) is involved in the development and plasticity of the CNS through the release of S-100beta, a glial trophic factor which stabilizes synapses and neuronal cytoskeleton and promotes neuronal development. S-100beta is released from glial cells after activation of glial 5HT(1A) receptors. We present in this paper the effects upon neurons and glia of a 5HT depletion induced by 14 days of treatment with para-chlorophenylalanine (PCPA) in adult rats. S-100beta, 5HT, 5HT-transporter (5HT-T) and neurofilaments (Nf-200 and Nf-68) expressions were studied by immunohistochemistry and image analysis in striatum, hippocampus, parietal and frontal cortex. Immediately after ending PCPA treatment we found increased intracellular S-100beta immunoreactivity in glial cells, reduced 5HT immunolabelling, reduced density of 5HT-T, Nf-200 and Nf-68 fibers and morphological alterations in neuronal cytoskeleton. One week after PCPA treatment S-100beta immunoreactivity decreased towards control levels, 5HT was normalized in dorsal raphe nucleus, but not in innervation areas; 5HT-T, Nf-200 and Nf-68 fiber densities increased but some neuronal cytoskeletal alterations were still present in striatum. Two weeks after PCPA treatment S-100beta had returned to control levels in most studied regions; 5HT immunoreactivity was normalized, meanwhile 5HT-T, Nf-200 and Nf-68 fiber densities increased reaching values over the control level. We propose that S-100beta could be accumulated in glial cells during the 5HT depletion period, to be released once 5HT levels have recovered. Neuronal cytoskeletal alterations and reduced fiber density may be the expression of decreased extracellular availability of S-100beta. Conversely, increased 5HT-T, Nf-200 and Nf-68 expressions, once S-100beta is normalized, may be the biological response to the growth factor release.
Animals↗
Development of nitric oxide neurons in the chick embryo retina.
Nitric oxide (NO) is a gas involved in neurotransmission in the central nervous system (CNS) and in vertebrate retinas. This paper describes five types of nitrergic neurons in developing and adult chick retina using the nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd) reaction. Three of them, nitrergic types 1, 2 and 3, were observed in the inner nuclear layer, while nitrergic type 4 was observed in the ganglion cell layer; nitrergic type 5 were the retinal photoreceptors. Cell processes formed four nitrergic networks, which could be observed in the inner plexiform layer (IPL), at sublayers 1, 3a, 3b and 4. Another nitrergic network was observed in the outer plexiform layer (OPL). From hatching, the dendritic branches were completely developed in the IPL and in the OPL, forming the mentioned networks. Current evidence suggests that NO is coexpressed with other neurotransmitters in neurons of the CNS. Double-staining procedures, using NADPHd and 5HT immunohistochemistry in chicken retina, in a sequential or in an alternative manner, did not reveal the coexistence of these two neurotransmitters in the same neurons, but their networks matched in sublayers 1 and 4 of the IPL.
Animals↗
Diamond-Blackfan anemia: report of seven further mutations in the RPS19 gene and evidence of mutation heterogeneity in the Italian population.
Diamond-Blackfan anemia (DBA) is a congenital disease characterized by defective erythroid progenitor maturation and physical malformations. Most cases are sporadic, but dominant or, more rarely, recessive inheritance is observed in 10% of patients. Mutations in the gene encoding ribosomal protein (RP) S19 have recently been found in 25% of patients with either the dominant or the sporadic form. DBA is the first human disease due to mutations in a ribosomal structural protein. Families unlinked to this locus have also been reported. In an investigation of 23 individuals, we identified eight different mutations in 9 patients. These include five missense, one frameshift, one splice site defect, and one 4-bp insertion in the regulatory sequence. Seven mutations are new; one has so far been found in 8 patients and is a relatively common de novo event. Two mutations are predicted to generate a truncated protein. We also report the prevalence of RPS 19 mutations in the Italian DBA population, as shown by an analysis of 56 patients. No genotype-phenotype correlation was found between patients with the same mutation. The main clinical applications for molecular analysis are clinical diagnosis of patients with an incomplete form of DBA and testing of siblings of a patient with a severe form so as to avoid using those who carry a mutation and a silent phenotype as allogeneic stem cell donors.
Adult↗
Clustering of distinct autoimmune diseases associated with functional abnormalities of T cell survival in children.
To ascertain whether alterations of lymphocyte switching off may be associated with clustering of autoimmune diseases in children, Fas- and C2-ceramide-induced cell death was evaluated on T cell lines derived from three patients affected by clustering of autoimmune disorders. Three patterns were found: patient 3 was resistant to Fas- and C2-ceramide, patient 1 was resistant to Fas, but sensitive to C2-ceramide, patient 2 was resistant to C2-ceramide, but sensitive to Fas. By contrast, Fas- and C2-ceramide-induced cell death was normal in five children with systemic juvenile rheumatoid arthritis, five children with insulin-dependent diabetes and 10 age-matched healthy controls. Surface expression of Fas was low in patient 1, but normal in patients 2 and 3. Together with normal Fas transcripts, patients 2 and 3 displayed a transcript 152 bp longer than the normal one retaining intron 5. Our data indicate that polyreactive autoimmune syndromes may be associated with heterogeneous alteration of the immune response switching-off system.
Adolescent↗
Mutations in ribosomal protein S19 gene and diamond blackfan anemia: wide variations in phenotypic expression.
Mutations of the ribosomal protein S19 (RPS19) gene were recently identified in 10 patients with Diamond Blackfan anemia (DBA). To determine the prevalence of mutations in this gene in DBA and to begin to define the molecular basis for the observed variable clinical phenotype of this disorder, the genomic sequence of the 6 exons and the 5' untranslated region of the RPS19 gene was directly assessed in DBA index cases from 172 new families. Mutations affecting the coding sequence of RPS19 or splice sites were found in 34 cases (19.7%), whereas mutations in noncoding regions were found in 8 patients (4.6%). Mutations included nonsense, missense, splice sites, and frameshift mutations. A hot spot for missense mutations was identified between codons 52 and 62 of the RPS19 gene in a new sequence consensus motif W-[YFW]-[YF]-x-R-[AT]-A-[SA]-x-[AL]-R-[HRK]-[ILV]-Y. No correlation between the nature of mutations and the different patterns of clinical expression, including age at presentation, presence of malformations, and therapeutic outcome, could be documented. Moreover, RPS19 mutations were also found in some first-degree relatives presenting only with isolated high erythrocyte adenosine deaminase activity and/or macrocytosis. The lack of a consistent relationship between the nature of the mutations and the clinical phenotype implies that yet unidentified factors modulate the phenotypic expression of the primary genetic defect in families with RPS19 mutations.
Adolescent↗
A recurrent breakpoint in the most common deletion of the Ig heavy chain locus (del A1-GP-G2-G4-E).
Human Ig heavy chain constant regions are encoded by a cluster of genes, the IGHC locus, on 14q32.3. Several forms of IGHC deletions and duplications spanning one to five genes have been described in different populations, with frequencies of 1.5-3.5% and 4.5-44%, respectively. Despite the common occurrence of these gene rearrangements, little is known about the breakpoint sites; evidence obtained from deletions in the IGHC locus and in other regions of the human genome suggests that they preferentially occur in highly homologous regions and might be favored by a variety of recombinogenic signals. We present here a detailed study of three homozygotes for the most common type of IGHC multiple gene deletion, spanning the A1-GP-G2-G4-E genes. Using a combination of Southern blotting, long-range PCR, and automated sequencing, the unequal crossover events of all of the six studied haplotypes have been mapped to a region of approximately 2 kb with almost complete homology between EP1-A1 and E-A2, flanked by two minisatellites. These results are consistent with the hypothesis that segments of complete homology may be required for efficient homologous recombination in humans. The possible role of minisatellites as recombination signals is inferred, in agreement with current knowledge.
Blotting, Southern↗
Definition of the smallest pathological CAG expansion in SCA7.
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Adult↗
Polyvariant mutant CFTR genes in patients with chronic pancreatitis.
Several authors have reported an association between mutations of the cystic fibrosis transmembrane conductance regulator gene (CFTR) and chronic pancreatitis. CFTR gene transcription and protein efficiency are influenced by two polymorphic loci, (TG)m and M470V, other than the T5 allele, whose role is already well-established. The TG11/T5 haplotype is commonly found in healthy subjects, while the TG12/T5/V470 and TG13/T5/V470 haplotypes are present in congenital bilateral absence of the vas deferens (CBAVD) patients. While the T5 allele is a mutation that is over-represented in patients with chronic pancreatitis, no data are available concerning the possible allelic preference at the other two polymorphic loci, (TG)m and M470V, in these patients. For this reason, we screened 39 patients with chronic pancreatitis for the most common CFTR mutations found so far in the Italian population; in addition, we examined the length of the polypyrimidine (poly-T) tract in intron 8, the (TG)m length and the M or V codon at position 470. CFTR mutations were found in 3 patients. Poly-T variant typing identified genotype T5/T7 in 5 patients and T5/T9 in 1 patient. Direct sequencing of intron 8 in patients with the T5 variant revealed the TG12/T5/V470//TG11/T7/V470 genotype in 5 patients and TG10/T9//TG11,T5 genotype in 1 patient. In patients with chronic pancreatitis, the T5 allele is frequently associated with TG12 and V470, a haplotype already reported in CBAVD cases and quite uncommon in healthy subjects.
Adult↗
Tryptamine, serotonin and catecholamines: an immunocytochemical study in the central nervous system.
Tryptamine, a serotonin-related indolamine, could be involved in the modulation of catecholaminergic and serotoninergic systems interaction. Despite previous reports on this topic, the morphological relationship among these systems is not well described. We studied the interaction among serotoninergic and catecholaminergic with tryptaminergic systems by double immunostaining at the level of light microscopy. Mesencephalic rat brain sections treated according to the Schiff quenching method were double immunostained using peroxidase and fluorescein labeled antibodies. Primary antibodies to anti-tryptophan hydroxylase (TrpOH), anit-tyrosine hydroxylase (TH) and anti-tryptamine (T) were used to demonstrate serotoninergic, catecholaminergic and tryptaminergic neurons respectively. A morphometric study was performed in order to analyze the different morphological characteristics of each system. The results showed that (i) T+ and TrpOH+ neurons are localized in the same areas but their morphology is significantly different. Moreover morphometric parameters of T+ neurons were significantly different from those TrpOH+ or TH+ neurons; (ii) The number of TrpOH+ neurons was larger than T+ neurons; (iii) T+ neurons were dominant in the lateral dorsal raphe nucleus. TrpOH+ neurons were more numerous in the central area of the dorsal raphe nucleus; (iv) Coexpression of TrpOH and T was demonstrated in the somata of dorsal raphe nucleus neurons; (v) TrpOH+ neurons from raphe nuclei and TH+ neurons from substantia nigra are contacted by T+ fibres. The present morphological evidence supports a functional relationship among these three aminergic systems.
Animals↗
Dihydropteridine reductase deficiency: physical structure of the QDPR gene, identification of two new mutations and genotype-phenotype correlations.
Dihydropteridine reductase (DHPR) is an enzyme involved in recycling of tetrahydrobiopterin (BH4), the cofactor of the aromatic amino acid hydroxylases. Its deficiency is characterized by hyperphenylalaninemia due to the secondary defect of phenylalanine hydroxylase and depletion of the neurotransmitters dopamine and serotonin, whose syntheses are controlled by tryptophan and tyrosine hydroxylases. The DHPR cDNA has been cloned and mapped on 4p15.3. In the present study we report the genomic structure of the DHPR gene (QDPR). This gene includes seven exons within a range of 84-564 bp; the corresponding introns are flanked by canonic splice junctions. We also present a panel of PCR primers complementary to intronic sequences that greatly facilitates amplification of the gene and provides a genomic DNA approach for mutation detection. We have used this approach to study six patients with DHPR deficiency. Four known mutations (G23D, H158Y, IVS5G+ 1A, R221X) and two new mutations (Y150C and G218ins9bp) were found. The Y150C mutation was found in compound heterozygosity with G23D, a mutation always associated with a severe phenotype in homozygous patients. This patient has an intermediate phenotype (good response to monotherapy with BH4). The mutant enzyme for Y150C was expressed in an E. coli system. Comparison of its kinetic parameters with those of the G23D mutant enzyme showed that it is not as effective as the wild-type enzyme, but is more active than the G23D mutant. This patient's intermediate phenotype is thus due to the mild DHPR mutation Y150C. Correlations between genotypes and phenotypes were also found for the other mutations.
Alleles↗
Molecular characterization of immunoglobulin G4 gene isoallotypes.
The molecular bases of classical serological immunoglobulin allotypes are progressively uncovered through detailed characterization of the relevant genes. Here we describe two isoallotypic determinants of the G4 gene. In the first, Leu 309, as in G1 and G3, is changed to Val, as in G2; studies on myeloma proteins have long assigned the immunologically defined nG4 m(a)/(b) to the same position. The two molecular variants, here called IGHG4*L309 and IGHG4*V309, are allelic in IGHC haplotypes with a single G4 gene, but can be found together in cis in G4-duplicated haplotypes. A second isoallotypic variant was found at codon 409, where either Arg, as in G1 and G3, or Lys, as in G2, can be found. Both isoallotypes are associated with several 'silent isoallotypic' substitutions dispersed through the hinge, CH2 and CH3 domains. This suggests segmental gene conversion as the common mechanism of origin.
Base Sequence↗
Congenital bilateral absence of vas deferens with a new missense mutation (P499A) in the CFTR gene.
We describe a congenital bilateral absence of the vas deferens (CBAVD) patient with a compound heterozygosity in the cystic fibrosis transmembrane regulator (CFTR) gene for a stop mutation W1282X and a new missense mutation P499A. The P499A is interpreted as a mild mutation whose phenotypic effects, in this case limited to the development of wolffian duct derivatives, are revealed only in combination with a severe CFTR mutation.
Adult↗