Differential expression of human COMT alleles in brain and lymphoblasts detected by RT-coupled 5′ nuclease assay

Rationale A common polymorphism, Val158Met, alters catechol-O-methyltransferase (COMT) enzyme activity and has been linked to psychiatric phenotypes. Bray et al. (2003) reported that COMT is subject to differential allele expression in brain, finding modest (13–22%) underexpression of a haplotype containing Val158. However, disparate findings by another group who used the same method, but in lymphoblasts, raise the issues of tissue specificity, magnitude of differential expression, and identity of loci altering expression.Objectives We measured COMT allele expression ratios in heterozygous human lymphoblast cell lines and brains.Methods Using transcribed single nucleotide polymorphisms as endogenous reporters, we developed an RT-coupled 5 nuclease assay for allele expression ratios and applied it to 63 COMT rs4818(C>G) heterozygotes and 68 Val158Met [rs4680(G>A)] heterozygotes.Results For rs4818(C>G), the C allele was overexpressed relative to the G allele in 18 of 27 lymphoblast lines and 23 of 36 brains. For Val158Met, Met158 was overexpressed relative to Val158 in all (29 of 29) lymphoblast lines and all (39 of 39) brains. Each of the 22 rs4818 heterozygotes without differential allele expression was a Val158/Val158 homozygote. The Met158 allele was overexpressed by 65–77% when compared with Val158 in lymphoblasts and brain. Haplotype augmented ability to predict expression in brain only. However, the expression of the Val158 allele on the high-expressing haplotype was only 19% higher than Val158 alleles on the other haplotype background.Conclusions COMT alleles are differentially expressed. The Met158 allele predicts higher mRNA expression in both brain and lymphoblasts. As exemplified here, the RT-coupled 5 nuclease assay is a reliable method for the quantitative evaluation of cis-acting regulatory effects.     

Single mothers and the use of professionals for mental health care reasons

In the present study, we examine whether higher rates of mental health service use observed among single-parent mothers is due to greater need (psychopathology) or other factors (predisposing and enabling characteristics) using a socio-behavioural model of health care use. We use data from two large surveys in Canada (the 1994-95 National Population Health Survey and the 1990 Ontario Mental Health Supplement). The bivariate results from both surveys revealed that single-parent mothers were two to three times more likely than married mothers to have sought professional help for mental health reasons over a 12-month period. Multivariate analyses showed that differences in predisposing and enabling characteristics between single and married mothers accounted for very little of the relationship between family structure and service use. Rather, differences in the prevalence of psychiatric disorders accounted for the higher use of services among single mothers. Single mothers are more likely than married mothers to seek professional help for mental health concerns. The use of services appears equitable in that need (higher rates of psychopathology) is the major factor differentiating use between married and single mothers. Further work should examine differences in pathways into formal care between single and married mothers.     

巯嘌呤甲基转移酶基因多态性位点与白血病巯嘌呤耐受性的关系

目的 分析巯嘌呤甲基转移酶(thiopurine methyltransferase,TPMT)基因型对急性白血病(AL)患儿巯嘌呤(6-mercaptopurime,6-MP)耐受性的影响,提高患儿对巯嘌呤类药物治疗的有效性和安全性。方法 应用以聚合酶链反应(PCR)为基础的2种方法并结合DNA直接测序,检测250例健康成人和280例AL患儿TPMT基因第5外显子G238C、第7外显子G460A和第10外显子A719G的3个多态性位点。详细记录160例患儿6-MP全量治疗时间、减少剂量时间和未治疗时间。结果 280例AL患儿中,10例为TPMT第10外显子A719G杂合变异,变异率为3．6％,未发现纯合变异,变异的等位基因均为TPMT*3C。AL患儿TPMT基闪变异的频率和类型与健康成人差异无显著性。在观察的160例患儿中,有28%的患儿未接受6-MP标准剂量、全疗程治疗。其中TPMT野生型者39例,占野生型患儿的26%,杂合型者6例,占杂合型患儿的60％(P=0．03)。而且,6／10例TPMT杂合型者和30／150例野生型者减少6-MP的剂量(P=0．009)。结论 TPMT基因的多态性位点与AL患儿6-MP的耐受性有关。TPMT杂合型患儿中不耐受6-MP的比例明显高于TPMT野生型者,必须中断治疗或减少剂量以避免较大毒性反应的发生：提示检测TPMT基因型有利于提高巯嘌呤类药物的有效性和安全性。     

Intravenous immunoglobulin therapy for acute cerebellar ataxia

A case of acute cerebellar ataxia without any prodromal illness showed cerebellar hypoperfusion on 123 I-iodoamphetamine single photon emission computed tomography. The symptoms did not resolve spontaneously or with methylprednisolone pulse therapy but disappeared rapidly with intravenous immunoglobulin therapy.

Conclusion: Intravenous immunoglobulin therapy is worth considering in acute cerebellar ataxia that does not respond to high-dose steroid therapy.     

IL-10 and TNF-alpha polymorphisms in a sample of Sicilian patients affected by tuberculosis: implication for ageing and life span expectancy

Human longevity seems to be directly correlated with optimal functioning of the immune system, suggesting that some genetic determinants of longevity might reside in those polymorphisms for the immune system genes that regulate immune responses, in particular cytokine gene polymorphisms. In fact, modification of cytokine network is a constant report in studies on age related modification of immune response. Moreover cytokine polymorphisms studies are indicating their involvement in the reshaping of cytokines network as an integral part of the scenario related to a successful ageing. A particular role might be attributed to the influence of cytokine polymorphisms on the efficiency of immune response against infectious diseases that have been the principal selection in oldest old. Here are reported data on the evaluation of the frequency of the functional polymorphisms at genes coding for TNF-alpha (-308G-->A) and IL-10 (-1082G-->A), analysed by ARMS-PCR, in a group of Sicilian patients affected by chronic lung tuberculosis (TBC) compared to that from a group of healthy individuals living in the same region. Data obtained demonstrated a reduction of -308GG TNF homozygous individuals in TBC affected subject group. In the same group a reduction of IL-10 -1082A/* carriers was found. Our results seem to suggest that multiple genetic traits may affect the capacity to cope with an infectious agents and this might predispose to an overt disease. Moreover these data are in agreement with previous reports suggesting that a balanced interaction among pro- and anti-inflammatory molecules it is a key point for conditioning the life span expectancy.     

A C/T single nucleotide polymorphism at the tyrosine kinase domain of the insulin receptor gene is associated with polycystic ovary syndrome

OBJECTIVE: To examine whether the insulin receptor (INSR) gene contributes to genetic susceptibility to the polycystic ovary syndrome (PCOS). DESIGN: Case-control study. SETTING: Academic endocrinology clinic. PATIENT(S): Ninety-nine women with PCOS as defined by the National Institutes of Health consensus and polycystic ovaries on ultrasonography, and 136 healthy controls. MAIN OUTCOME MEASURE: Frequency of genotypes of a single nucleotide polymorphism of the INSR gene in patients and controls. RESULT(S): After stratification of participants by body mass index, the frequency of the uncommon T allele of the INSR single nucleotide polymorphism was significantly increased in lean patients with PCOS (body mass index < or =27 kg/m2) compared with lean controls (relative risk, 2.1). CONCLUSION(S): The INSR gene is a susceptibility gene for PCOS among lean patients with PCOS. It remains to be determined whether the exon 17 C/T single nucleotide polymorphism is the susceptibility single nucleotide polymorphism for PCOS or whether it is in linkage disequilibrium with another INSR gene polymorphism.     

Identification of Mutations Associated with Attenuation of Virulence of a Field Sendai Virus Isolate by Egg Passage

We have reported that attenuation of the virulence of a field Sendai virus (SeV) isolated by egg passage is associated with an impediment of viral genome replication in mouse respiratory cells (Kiyotani et al., Arch Virol 146, 893–908, 2001). To determine the molecular basis for the attenuation, we sequenced entire genomes of representative SeV clones isolated during egg passages and compared those with that of the parental SeV clone E0. E15cl2, a 165-fold attenuated clone in 50% mouse lethal dose (MLD₅₀) isolated at the 15th egg passage, possessed only four mutations in the entire genome: U to A at position 20 (U20A) and U24A in the leader promoter region and A9362G and A12174U in the L gene from the 5-end of antigenome. The former mutation in the L gene was silent and the latter changed deduced amino acid Ser at position 1207 to Cys (Ser1207Cys) in the L protein, a catalytic subunit of viral polymerase. E30cl2, a further 6-fold attenuated clone isolated at the 30th egg passage, had an additional four mutations: A8074G (Glu461Gly) and A8077G (Asp462Gly) in the hemagglutinin-neuraminidase (HN) gene and A13598C (silent) and G13927A (Ser1791Asn) in the L gene. On the other hand, a virulent revertant clone, E30M15cl5, which was obtained by 15 mouse passages of E30cl2 and had 250-fold mouse virulence compared to E30cl2, possessed eight mutaions: A24U in the leader, C1325U (silent) in the nucleocapsid gene, G8074A (Gly461Glu) in the HN gene, G10433U (Lys626Asn), C13598A (silent), A13927G (Asn1791Ser), C14626U (Thr2024Ile) and A15272C in the L gene. Among these, the mutations in the leader and the HN gene and two of the mutations in the L gene (C13598A and A13927G) were true reversions to E0. The significance of the mutations detected in the leader as well as in the L and HN genes was discussed in the context of attenuation of SeV pathogenicity by egg passage.     

The nucleotide sequences of the parathyroid gene in primates (suborder Anthropoidea)

Nucleotide sequences of the parathyroid (PTH) gene of 12 species of primates belonging to suborder Anthropoidea were examined. The PTH gene contains one intron that separates two exons that code the sequence of prepro and PTH, respectively. The intron of the PTH gene in Cebus apella, Callithrix jacchus, and Saguinus oedipus was 102 bp long, whereas a 103-bp intron was observed in the remaining species. Phylogenetic analysis using the nucleotide sequences of PTH revealed that these 12 species of primates of suborder Anthropoidea could be divided into two groups of the infraorder Platyrrhini (C. apella, C. jacchus, and S. oedipus) and the infraorder Catarrhini (Macaca fascicularis, Macaca fuscata, Cercopithecus aethiops, Papio hamadryas, Presbytes obscura, Hylobates lar, Pongo pygmaeus, Pan troglodytes, and Pan paniscus). The latter infraorder could be further subdivided into two subgroups belonging to the superfamily Cercopithecoidea (M. fascicularis, M. fuscata, C. aethiops, P. hamadryas, and P. obscura) and the superfamily Hominoidea (H. lar, P. pygmaeus, P. troglodytes, and P. paniscus). The deduced amino acid sequences of PTH gene between 12 species of nonhuman primates and human revealed no amino acid substitution in mature PTH among orangutans, chimpanzees, and humans. The results indicated that the PTH gene is very conserved among primates, especially between great apes and humans. The apes are the most suitable animals to be used for studying the bone metabolism and applying the knowledge to clinical use in humans.     

Discrimination of single bars by the honeybee (Apis mellifera)

The bees learn to come for a reward to a very simple pattern, a black bar in a fixed position on a white background, in a Y-choice apparatus, with the targets presented in the vertical plane at a fixed range. They were trained on a number of different arrangements of a single bar on one or both targets. The trained bees were then given appropriate tests to discover what cues they had learned. A cue is an essential parameter that is recognized, not the whole pattern. At the choice point they learn exactly which way to look for consistent cues. After training on a single broad bar versus a blank target, they respond in tests to any area of black where they expect to see it, and are less able to detect it the more it has been displaced from the training position. They are more sensitive to vertical than to horizontal displacement of the bar. The cue is anything black of the right size. They do not recognize the shape or orientation of the bar. When trained to discriminate between two bars at right angles to each other, centred on the reward hole, the cue is the edge orientation at the expected places on the targets, and the bees are less able to discriminate the orientation cues the more they are displaced. When trained on a pair of broad black bars in different positions, the cues are the vertical positions of the centres. Division of the bar into squares, or making the edges stepped, removes the orientation cue but not the position cue. Addition of a large black spot or a checkerboard background to the original bar prevents discrimination, as if the spatial reference frame is disturbed. In training, or testing trained bees, parallax does not assist the discrimination of orientation.