The effect of nitric oxide donors on nitric oxide synthase-expressing myenteric neurones in culture.

Previously, we demonstrated that intestinal inflammation leads to a postinflammatory loss of nitric oxide synthase (NOS)-expressing myenteric neurones and motility disturbances. Here, we investigated whether high NO concentrations could be responsible for the decrease in NOS neurones. Myenteric neurone cultures, prepared from guinea-pig small intestine, were incubated with NO donors [sodium nitroprusside (SNP) and 3-morpholinosydnonimine (SIN-1)]. After fixation, NOS neurones were identified by NADPH diaphorase staining and neurone-specific enolase (NSE)-positive neuronal content was assessed with an enzyme-linked immunosorbent assay (ELISA)-based method. Twenty-four hours incubation with SIN-1 (10(-3) mol L(-1)) or SNP (10(-4) mol L(-1) or higher) reduced the number of NADPH diaphorase-positive neurones. SNP incubation did not affect the NSE-positive neuronal content. Shorter incubations (SNP: 4 and 12 h) had no significant effect. The SNP-induced reduction was reversed by glutathione (GSH), but not by NO- or O-scavengers, whereas GSH depletion enhanced the decrease. The NO-dependent guanylate cyclase-blocker 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) did not affect the SNP effect. This reduction can be explained by either specific apoptosis of NOS neurones or downregulation of NOS activity. However, TdT-mediated X-dUTP nick end labelling (TUNEL stainings argue in favour of the latter. In conclusion, the NO donor SNP decreases the number of NOS-expressing myenteric neurones time and concentration dependently, without affecting the amount of neuronal material. Glutathione plays an important protective role.

A physical map of the chromosome 12 centromere.

While current sequencing efforts consider the detection of alpha satellite repeats as logical end points for map construction, detailed maps of most pericentromeric regions are lacking to confirm this hypothesis. Here we identify the different alpha satellite families present at the pericentromeric region of chromosome 12. The order, size and location of these repeats is established using radiation hybrid analysis, pulsed field gel analysis and FISH and the maps are integrated with current sequence information. For the different classes of alpha satellites present at the chromosome 12 centromere the paralogs in the human genome were mapped by FISH. Unique sequences flanking the alpha satellite repeats were identified, some of which are not represented in the current draft sequence. This mapping effort localises the different alpha satellite repeats within the pericentromeric region and anchors them in the current maps. The novel sequences identified may serve as the end point for the ongoing sequencing efforts.

APOE genotype: no influence on galantamine treatment efficacy nor on rate of decline in Alzheimer's disease.

Apolipoprotein E (APOE) has been extensively demonstrated to be a genetic risk factor for Alzheimer's disease (AD). Associations of APOE genotype have been reported with age at AD onset, rate of decline, and responsiveness to therapy. This study aimed to test these hypotheses in a large study population of AD patients. APOE genotype was determined from 1,528 Caucasian subjects, diagnosed by NINCDS/ADRDA criteria as probable AD patients, enrolled in four international placebo-controlled clinical trials of 3--12 months duration, designed to evaluate efficacy of treatment with galantamine or sabeluzole. In addition to patient demographics and baseline scores for Mini Mental State Examination, scores on the Disability Assessment for Dementia (DAD) and the cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-cog) were recorded at the start, during, and at the end of the study. APOE epsilon 4 homozygotes had a significantly lower age at disease onset compared to patients with other APOE genotypes. The epsilon 4 allele was significantly over-represented in females compared to males, and in the group of subjects with an AD family history. Based on longitudinal data of 504 placebo-treated AD patients, the linear annual rate of change in score was 5 points on the ADAS-cog scale and 11 on the DAD scale. The epsilon 4 allele copy number did not influence these rates of decline. Sabeluzole treatment was not effective in the overall group compared to the placebo-treated group, nor in any subgroup stratified by epsilon 4 allele count. Galantamine produced cognitive and functional improvement that were not affected by epsilon 4 allele count. In conclusion, our data confirm a strong association between epsilon 4 homozygotes and age at onset of AD but do not support an effect of epsilon 4 allele copy number on rate of cognitive and functional decline nor on the efficacy of galantamine in patients with AD.

Novel syndromic form of X-linked complicated spastic paraplegia.

This study presents a family with a syndromic form of X-linked mental retardation in which four males in two generations present severe mental retardation, slowly progressive spastic paraplegia, facial hypotonia, and maxillary hypoplasia. Multipoint linkage analysis with 24 highly polymorphic markers indicated two possible candidate regions: Xp21.1-Xq21.3 (flanking markers DXS1214 and DXS990) and Xq23-Xq27.1 (flanking markers DXS8020 and DXS984). The two known loci for X-linked mental retardation and spastic paraplegia are excluded: proteolipid protein in Xp21 and L1 cell adhesion molecule in Xq28. Therefore, the syndrome in this family appears to represent a previously undescribed X-linked spastic paraplegia-mental retardation syndrome.

A European multicenter association study of HTR2A receptor polymorphism in bipolar affective disorder.

The available data on the role of 5-HT in a variety of behaviors support the hypothesis that a dysfunction in brain serotoninergic system activity contributes to vulnerability to major depression. The diversity in the electrophysiological actions of 5-HT in the central nervous system can now be categorized according to receptor subtypes and their respective effector mechanisms. In particular, the implication of central postsynaptic 5-HT2A receptor in affective disorders has been supported by findings consistent with the hypothesis of 5-HT2A receptor up-regulation in depression. For these reasons, the 5-HT2A receptor (HTR2A) gene can be considered as a candidate gene in bipolar affective disorder (BPAD). We tested the possible genetic contribution of the polymorphic DNA variation T102C in exon 1 of HTR2A (chromosome 13q14-21) gene in a large European multicentric case-control sample. Allele and genotype frequencies, as well as homo-heterozygote distributions were compared between the two groups of 309 bipolar affective disorder patients and 309 matched controls. No significant differences were observed in the allelic and genotypic (also for homo-heterozygote) distribution between BPAD and controls. These results indicate that, in our sample, the 5-HT2A receptor polymorphism studied is unlikely to play a major role in the genetic susceptibility to BPAD. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:136-140, 2000.

Linkage of mood disorders with D2, D3 and TH genes: a multicenter study.

BACKGROUND: It has been suggested that the dopaminergic system is involved in the pathophysiology of mood disorders. We conducted a multicenter study of families with mood disorders, to investigate a possible linkage with genes coding for dopamine receptor D2, dopamine receptor D3 and tyrosine hydroxylase (TH). METHODS: Twenty three mood disorder pedigrees collected within the framework of the European Collaborative Project on Affective Disorders were analyzed with parametric and non-parametric linkage methods. Various potential phenotypes were considered, from a narrow (only bipolar as affected) to a broad (bipolar+major depressive+schizoaffective disorders) definition of affection status. RESULTS: Parametric analyses excluded linkage for all the candidate genes, even though small positive LOD (Limit of Detection) scores were observed for TH in three families. Non-parametric analyses yielded negative results for all markers. CONCLUSION: The D2 and D3 dopamine receptors were, therefore, not a major liability factor for mood disorders in our sample, whereas TH may play a role in a subgroup of patients.

A new gene (DYX3) for dyslexia is located on chromosome 2.

Developmental dyslexia is a specific reading disability affecting children and adults who otherwise possess normal intelligence, cognitive skills, and adequate schooling. Difficulties in spelling and reading may persist through adult life. Possible localisations of genes for dyslexia have been reported on chromosomes 15 (DYX1), 6p21.3-23 (DYX2), and 1p over the last 15 years. Only the localisation to 6p21.3-23 has been clearly confirmed and a genome search has not previously been carried out. We have investigated a large Norwegian family in which dyslexia is inherited as an autosomal dominant trait. A genome wide search for linkage with an average 20 cM marker density was initiated in 36 of the 80 family members. The linkage analysis was performed under three different diagnostic models. Linkage analysis in the family identified a region in 2p15-p16 which cosegregated with dyslexia. Maximum lod scores of 3.54, 2.92, and 4.32 for the three different diagnostic models were obtained. These results were confirmed by a non-parametric multipoint GENEHUNTER analysis in which the most likely placement of the gene was in a 4 cM interval between markers D2S2352 and D2S1337. Localisation of a gene for dyslexia to 2p15-16, together with the confirmed linkage to 6p21.3-23, constitute strong evidence for genetic heterogeneity in dyslexia. Since no gene for dyslexia has been isolated, little is known about the molecular processes involved. The isolation and molecular characterisation of this newly reported gene on chromosome 2 (DYX3) and DYX1 will thus provide new and exciting insights into the processes involved in reading and spelling.

Regional localization of a gene for nonspecific XLMR to Xp11.3-p11. 23 (MRX51) and tentative localization of an MRX gene to Xq23-q26.1.

Two families with nonspecific X-linked mental retardation (MRX) are presented. In the first family, MRX51, three male patients showed mild to borderline mental retardation. Multipoint linkage analysis yielded a maximal LOD score of 2.10 between markers DXS8012 and DXS1003, localizing the MRX51 gene at Xp11.3-p11.23. In the second family, XLMR7, three men showed moderate mental retardation (MR), and one possible female carrier had mild MR. Multipoint linkage analysis yielded an LOD score of 1.80 between markers DXS8063 and DXS1047, situating the disease gene at Xq23-q26.1. When the analysis was performed considering the affected female to be an expressing heterozygote carrier of the disease mutation, a maximal LOD score of 2.10 was found in the same region.

Genetic refinement and physical mapping of a chromosome 18q candidate region for bipolar disorder.

Recent genetic studies have implicated chromosome 18 in bipolar disorder (BP) with putative loci in the pericentromeric region and on 18q. We reported linkage to chromosome 18q21.33-q23 in a large family. In this study we typed additional markers in the family and were able to reduce the candidate region significantly. All affected family members are sharing alleles for markers spanning a genetic distance of maximal 8.9 cM. Haplotype analysis provided a marker order in agreement with published genetic and physical maps. Using yeast artificial chromosomes, we constructed a contig map that will help to identify positional candidate genes for bipolar disorder.

Homozygosity mapping to the USH2A locus in two isolated populations.

Usher syndrome is a group of autosomal recessive disorders characterised by progressive visual loss from retinitis pigmentosa and moderate to severe sensorineural hearing loss. Usher syndrome is estimated to account for 6-10% of all congenital sensorineural hearing loss. A gene locus in Usher type II (USH2) families has been assigned to a small region on chromosome 1q41 called the UHS2A locus. We have investigated two families with Usher syndrome from different isolated populations. One family is a Norwegian Saami family and the second family is from the Cayman Islands. They both come from relatively isolated populations and are inbred families suitable for linkage analysis. A lod score of 3.09 and 7.65 at zero recombination was reached respectively in the two families with two point linkage analysis to the USH2A locus on 1q41. Additional homozygosity mapping of the affected subjects concluded with a candidate region of 6.1 Mb. This region spans the previously published candidate region in USH2A. Our study emphasises that the mapped gene for USH2 is also involved in patients from other populations and will have implications for future mutation analysis once the USH2A gene is cloned.

Association analysis of the 5-HT2C receptor and 5-HT transporter genes in bipolar disorder.

We selected 42 patients with bipolar disorder type I (BPI) and 40 healthy controls for genetic analysis of DNA polymorphisms in the serotonin receptor 2c (5-HTR2c) and serotonin transporter (5-HTT) genes. No significant associations were found in the total patient sample. However, when the individuals were divided according to gender, trends for association with both polymorphisms (P = 0.051 for 5-HTR2c and P = 0.049 for 5-HTT) in female patients were observed. These results suggest that variations in these genes may be responsible for a minor increase in susceptibility for bipolar disorder in women.

X-linked severe mental retardation and a progressive neurological disorder in a Belgian family: clinical and genetic studies.

The combination of X-linked mental retardation (XLMR) and neurological disorders occurs in a number of syndromes. Differential diagnosis mostly depends on clinical data and mapping of responsible genes by linkage analysis. We present a Belgian family with severe XLMR and a progressive neurological disorder with ataxia, spasticity and convulsions. Biochemical investigations, neuroimaging and neuropathology were normal. Linkage analysis pointed to region Xq27-28 as the probable locus for the genetic defect. The sequence of the L1CAM cDNA, a possible candidate gene, proved to be normal in the patients. This suggests the presence of a genetic factor on Xq27-28, different from L1CAM, which can lead to severe XLMR and a progressive neurological disorder.

The X-linked infantile spasms syndrome (MIM 308350) maps to Xp11.4-Xpter in two pedigrees.

We report 2 families with X-linked infantile spasms syndrome (X-linked West syndrome). Data from clinical examination, biochemical analysis, neuroimaging, and neuropathology are discussed. In these families, genetic linkage analysis was able to locate the disease gene to the distal part of the short arm of the X chromosome, between Xpter and Xp11.4. This is the first report of linkage with genetic markers in this disorder. Although most cases are sporadic, further unraveling of the genetic background of the familial cases might greatly improve our understanding of infantile spasms.

A chromosome 18 genetic linkage study in three large Belgian pedigrees with bipolar disorder.

The contribution of genetic factors to the susceptibility for affective disorders has been firmly established. Recent reports found evidence for a susceptibility locus for affective disorders in 2 regions on chromosome 18. We describe 3 large Belgian pedigrees with multiple patients with affective disorders. Both chromosome 18 regions were investigated in the 3 families, using parametric and nonparametric segregation methods. In the pericentromeric region, all evidence was against a disease gene in our families. Also the data obtained for the distal part of 18q, argue against a genetic susceptibility factor in our sample.

Analysis of the tyrosine hydroxylase and dopamine D4 receptor genes in a Croatian sample of bipolar I and unipolar patients.

We selected 83 patients with bipolar disorder type I or unipolar recurrent major depression and 71 healthy controls for genetic analysis of the tyrosine hydroxylase and the dopamine D4 receptor gene. No significant association was found between bipolar disorder type I and unipolar recurrent major depression and the polymorphisms located near these genes. Therefore, the hypothesis that the tyrosine hydroxylase and the dopamine D4 receptor genes may be involved in the etiology of bipolar disorder and unipolar recurrent major depression is not supported in our study.

Positive association between the GABRA5 gene and unipolar recurrent major depression.

The gamma-aminobutyric acid (GABA) neurotransmitter system has been implicated in the pathogenesis of mood disorders. To test this hypothesis we carried out an association study between a dinucleotide repeat polymorphism in the GABAA receptor alpha 5 subunit gene and bipolar and unipolar mood disorders. Our results suggest a possible involvement of this gene in unipolar but not in bipolar disorder.

Regional localization of two genes for nonspecific X-linked mental retardation to Xp22.3-p22.2 (MRX49) and Xp11.3-p11.21 (MRX50).

Two families with nonspecific X-linked mental retardation (XLMR) are presented. In the first family, MRX49, 5 male patients in 2 generations showed mild to moderate mental retardation. Two-point linkage analysis with 28 polymorphic markers, dispersed over the X-chromosome, yielded a maximal LOD score of 2.107 with markers DXS7107 and DXS8051 at theta = 0.0, localizing the MRX49 gene at Xp22.3-p22.2, between Xpter and marker DXS8022. Multipoint linkage analysis showed negative LOD values over all other regions of the chromosome. In the second family, MRX50, 4 males in 2 generations showed moderate mental retardation. Pairwise linkage analysis with 28 polymorphic markers yielded a LOD score of 2.056 with markers DXS8054, DXS1055, and DXS1204, all at theta = 0.0. Flanking markers were DXS8012 and DXS991, situating the MRX50 gene at Xp11.3-Xp11.21, in the pericentromeric part of the short arm of the X chromosome.

Linkage and association of the HLA gene complex with IDDM in 81 Danish families: strong linkage between DR beta 1Lys71+ and IDDM.

Many studies have shown an association of IDDM with polymorphisms in the HLA region on chromosome 6p21. Previously our case-control study in the Belgian population showed significant association between IDDM and certain HLA class II alleles, in particular Lys71+, encoding DRB1 alleles. In the present study, 81 Danish multiplex IDDM families and 82 healthy Danish controls were examined for polymorphisms in the HLA-DRB genes and 54 of the 81 families for polymorphisms in HLA-B, -DQA1, -DQB1, -TNFA, and -TNFB genes. The results confirm our previous studies in the Belgian population and show that DRB1Lya71+/+ homozygotes have a relative risk (RR) of 103.5. Linkage between IDDM and DRB1 alleles that encode Lys71+ was shown by affected zib pair analysis which showed strong linkage (p < 1 x 10(-6). By family based association studies, the DRB1Lys71+ was identified as the allale which increased susceptibility to develop IDDM most in the HLA region (haplotype relative risk = 8.38). Haplotype analysis confirmed the increased risk contributed by DRB1Lys71+ alleles and in addition showed that DRB1Lys71- provides protection against IDDM even in the presence of DQB1Aep47-. These results indicate that DRB1Lys71+ screening is a powerful test compared to full HLA typing to determine the risk for a random person to develop IDDM in the Danish population, with an even higher probability than shown previously for the Belgians.