Characterization and chromosomal location of two repeated DNAs in three Gerbillus species

Two tandemly repeated DNA sequences of Gerbillus nigeriae (Rodentia) (GN1 and GN2) were isolated and characterized. Both share a 36bp repeated unit, which includes a 20bp motif also found in primate alphoid and other repeated DNAs. The localization of GN1 and GN2 sequences on metaphase chromosomes of three Gerbillus species, G. nigeriae, G. aureus and G. nanus, was studied by fluorescence in situ hybridization (FISH). In the G. nigeriae and G. aureus karyotypes, which were shown to possess large amounts of heterochromatin and to have undergone multiple rearrangements during evolution, both GN1 and GN2 sequences were observed at various chromosomal sites: centromeric, telomeric and intercalary. In contrast, the karyotypically stable G. nanus, which does not possess large amounts of heterochromatin and seems to be a more ancestral species, possesses only GN1 sequences, localized in the juxtacentromeric regions.     

Rapid genotyping of mice with hemoglobinopathies and globin transgenes

Because of its high sensitivity, bioluminescence (BL) is an excellent alternative to radioactive quantitation of cytokine RT-PCR-derived products. BL also allows detection of amplicons at cycle numbers not normally detectable using radioactivity. No direct comparisons between these two methods have been made. In this study, the sensitivities of BL using recombinant aequorin, a flash-type luminescent tag capable of detecting signal to attomolar (10(-18) M) levels and radio imaging (RI) were directly compared. In addition, the application of BL for detecting cytokine message from biologic samples was examined. BL was 30- to 60-fold more sensitive than RI in detecting human IL-2 and CD3delta amplicons. This difference was particularly found during low cycle PCR, but was less at higher cycle numbers. The ability of BL to detect differences in cytokine message in stimulated and unstimulated human peripheral blood mononuclear cells was also evaluated. Using linear regression analysis, we observed up to 5,000-fold increases in RT-PCR amplified-mRNA in stimulated cells for IL-1alpha, IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-10 and GM-CSF compared to unstimulated cells. Changes in CD3delta, TNF alpha or IL-12 were not observed or quantitated. We present a novel aequorin-based application of bioluminescent technology to directly quantitate RT-PCR amplicons and to investigate the induction of human cytokine expression. Significant advantages of this sensitive bioluminescent method compared with radioactive methods are its abilities to quantitate amplicons in a PCR cycle range where linear detection is most robust and to analyze products in an automated, open-architecture microtiter plate format.     

The brain renin-angiotensin system contributes to the hypertension in mice containing both the human renin and human angiotensinogen transgenes

We have previously shown that mice transgenic for both the human renin and human angiotensinogen genes (RA+) exhibit appropriate tissue- and cell-specific expression of both transgenes, have 4-fold higher plasma angiotensin II (AII) levels, and are chronically hypertensive. However, the relative contribution of circulating and tissue-derived AII in causing hypertension in these animals is not known. We hypothesized that the brain renin-angiotensin system contributes to the elevated blood pressure in this model. To address this hypothesis, mean arterial pressure (MAP) and heart rate were measured in conscious, unrestrained mice after they were instrumented with intracerebroventricular cannulae and carotid arterial and jugular vein catheters. Intracerebroventricular administration of the selective AII type 1 (AT-1) receptor antagonist losartan (10 microgram, 1 microL) caused a significantly greater peak fall in MAP in RA+ mice than in nontransgenic RA- controls (-29+/-4 versus -4+/-2 mm Hg, P<0.01). To explore the mechanism of a central renin-angiotensin system-dependent hypertension in RA+ mice, we determined the relative depressor responses to intravenous administration of the ganglionic blocking agent hexamethonium (5 mg/kg) or an arginine vasopressin (AVP) V1 receptor antagonist (AVPX, 10 microgram/kg). Hexamethonium caused equal lowering of MAP in RA+ mice and controls (-46+/-3 versus -52+/-3, P>0.05), whereas AVPX caused a significantly greater fall in MAP in RA+ compared with RA- mice (-24+/-2 versus -6+/-1, P<0.01). Consistent with this was the observation that circulating AVP was 3-fold higher in RA+ mice than in control mice. These results suggest that increased activation of central AT-1 receptors, perhaps those located at sites involved in AVP release from the posterior pituitary gland, plays a role in the hypertension in RA+ mice. Furthermore, our finding that both human transgenes are expressed in brain regions of RA+ mice known to be involved in cardiovascular regulation raises the possibility that augmented local production of AII and increased activation of AT-1 receptors at these sites is involved.     

The influence of positive selection on RAG expression in thymocytes

Many human cancers present deletions of the short arm of chromosome 17, which includes the TP53 locus. We detected a new polymorphism in intron 2 of the TP53 gene using PCR-SSCP and used this polymorphic site as a marker to detect loss of heterozygosity in 135 human tumors (73 soft tissue sarcomas, and 48 colorectal and 14 bladder carcinomas). Heterozygosity for this site was 41.5% in this study group and tumor-specific loss of alleles occurred in 43% of informative cases. Allelic losses were more frequently detected at this site than at that in which restriction fragment length polymorphism (RFLP) is located, as detected by the pHp53B probe. It is concluded that this novel approach has several advantages, including detection of a high incidence of informative cases and minimal tissue requirements.     

Paraoxonase (PON1) gene in mice: sequencing, chromosomal localization and developmental expression

PURPOSE: To retrospectively examine the optic disc photographs of a glaucoma population for optic disc haemorrhages, vascular occlusions and vascular abnormalities. METHODS: The optic disc photographs of 906 eyes of glaucoma and suspect glaucoma patients were examined. Optic disc photographs were taken annually, where possible, with the follow-up period varying between 1 and 14 years duration (mean, 2.89). Glaucoma patients are regularly reviewed every 4-6 months and glaucoma suspects every 1-2 years, depending on the ophthalmologist. Low-tension glaucoma patients were reviewed more frequently (mean, every 2.6 months). The results of the findings were compared to a control group of 39 subjects with a mean follow-up period of 7 years, using Fisher's exact test. RESULTS: It was found that during the period under review, 7.4% (n = 67) of eyes had optic disc haemorrhages. The highest frequency of optic disc haemorrhages (37.5%) was found in the low tension glaucoma group (P = 0.0001) followed by 11% of primary open-angle glaucoma eyes (P = 0.03). In the normal group there were three eyes with optic disc haemorrhages and one with a disc collateral, which constitutes 5.1% vascular changes in this sub-group. Of the study eyes 2.8% had central retinal vein occlusions, 1.3% branch vein occlusion, 1.2% disc vessel abnormalities (loops) and 1.1% disc collaterals. Discrete nerve fibre layer haemorrhages and microaneurysms were found in 0.8% and 1.8% of eyes, respectively. CONCLUSIONS: A total of 16.8% of the eyes observed in this study had either disc haemorrhages or vascular changes. The underlying trend of vascular and haemorrhagic changes in glaucoma are demonstrated in this sample, which is in general agreement with previous studies. The high percentage of optic disc haemorrhages in low tension glaucoma is highlighted. The presence of microaneurysms and nerve fibre layer haemorrhages is interesting but of unknown significance.     

Mammary gland expression of transgenes and the potential for altering the properties of milk

Transgenic animals are a useful in vivo experimental model for assessing the ability and impact of foreign gene expression in a biological system. Transgenic mice are most commonly used, while transgenic sheep, goats, pigs and cows have also been developed for specific, "applied" purposes. Most of the work directed at targeting expression of transgenes to the mammary gland of an animal, by using a milk gene promoter, has been with the intent of either studying promoter function or recovering the desired protein from the milk. Transgenic technology can also be used to alter the functional and physical properties of milk resulting in novel manufacturing properties. The properties of milk have been altered by adding a new protein with the aim of improving the milk, not of recovering the protein for other uses.     

The influence of filter top caging on the transmission of pinworm infections in mice

The effectiveness of filter top cages in preventing infection of mice with pinworms, Aspiculuris tetraptera and Syphacia obvelata, was investigated by introducing mice from a pathogen-free colony into a colony in which both types of pinworms were enzootic. Half of the pathogen-free mice were maintained in filter top cages, while the remainder were held in similar cages without filters. All mice thus introduced were killed and examined after 3-8 week exposure periods. No pinworm were recovered from mice in 17 of 18 cages with filter tops while mice in 12 of 16 cages without filter tops were infected.     

The influence of an irrelevant stimulus on two discriminations.

In 3 experiments, the effect of adding an irrelevant stimulus to a discrimination was examined. In Exp 1, a group of pigeons received autoshaping with an A+Bo discrimination in which 1 stimulus signaled food, A+, and a simultaneous compound of A with another stimulus, B, signaled the absence of food, ABo. A 2nd group received similiar training, except that a 3rd stimulus, C, was present in both types of trials, AC+BCo. The A+Bo discrimination was acquired more readily than the AC+BCo discrimination. Exps 2 and 3 used a negative-patterning design, A+Bo+. In both experiments, this problem was mastered more readily than when an irrelevant stimulus was used to create an AC+BCoC+ discrimination. The results fail to confirm predictions derived from elemental theories of conditioning. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The influence of cue-task association and location on switch cost and alternating-switch cost.

Examined whether task-relevant information provided by task cues influences the magnitude of switch cost in a parallel manner in 74 Ss (aged 18-37 yrs). Cues presented prior to a trivalent stimulus indicated which of three tasks to perform. These cues either had a preexisting association with the to-be-performed task, or a recently learned association with the task. The results paralleled the effects of stimulus bivalence: substantial switch cost with recently learned cue-task associations and greatly reduced switch cost with preexisting cue-task associations. This suggests that both stimulus-based and cue-based information can activate the relevant task set. Alternating-switch cost was also observed with both preexisting and recently learned cue-task associations, only when all tasks were presented in a consistent spatial location. When spatial location was used to cue the to-be-performed tasks, no alternating-switch cost was observed, suggesting that different processes may be involved when tasks are uniquely located in space. Specification of the nature of these processes may prove to be complex, as post-hoc inspection of the data suggested that for the spatial cue condition, the alternating-switch cost may oscillate between cost and benefit, depending on the relevant task. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Sucrose consumption in mice: major influence of two genetic loci affecting peripheral sensory responses

Individual variability in sucrose consumption is prominent in humans and other species. To investigate the genetic contribution to this complex behavior, we conducted behavioral, electrophysiological, and genetic studies, using male progeny of two inbred mouse strains (C57BL/6ByJ [B6] and 129/J [129]) and their F2 hybrids. Two loci on Chromosome (Chr) 4 were responsible for over 50% of the genetic variability in sucrose intake. These loci apparently modulated intake by altering peripheral neural responses to sucrose. One locus affected the response threshold, whereas the other affected the response magnitude. These findings suggest that the majority of difference in sucrose intake between male B6 and 129 mice is due to polymorphisms of two genes that influence receptor or peripheral nervous system activity.     

Efficient peripheral clonal elimination of B lymphocytes in MRL/lpr mice bearing autoantibody transgenes

Peripheral B cell tolerance was studied in mice of the autoimmune-prone, Fas-deficient MRL/ lpr.H-2(d) genetic background by introducing a transgene that directs expression of membrane-bound H-2Kb antigen to liver and kidney (MT-Kb) and a second transgene encoding antibody reactive with this antigen (3-83mu delta, anti-Kk,b). Control immunoglobulin transgenic (Ig-Tg) MRL/lpr.H-2(d) mice lacking the Kb antigen had large numbers of splenic and lymph node B cells bearing the transgene-encoded specificity, whereas B cells of the double transgenic (Dbl-Tg) MRL/lpr.H-2(d) mice were deleted as efficiently as in Dbl-Tg mice of a nonautoimmune B10.D2 genetic background. In spite of the severely restricted peripheral B cell repertoire of the Ig-Tg MRL/lpr.H-2(d) mice, and notwithstanding deletion of the autospecific B cell population in the Dbl-Tg MRL/lpr.H-2(d) mice, both types of mice developed lymphoproliferation and exhibited elevated levels of IgG anti-chromatin autoantibodies. Interestingly, Dbl-Tg MRL/lpr.H-2(d) mice had a shorter lifespan than Ig-Tg MRL/lpr.H-2(d) mice, apparently as an indirect result of their relative B cell lymphopenia. These data suggest that in MRL/lpr mice peripheral B cell tolerance is not globally defective, but that certain B cells with receptors specific for nuclear antigens are regulated differently than are cells reactive to membrane autoantigens.     

Genetic influence on neurogenesis in the dentate gyrus of adult mice

To address genetic influences on hippocampal neurogenesis in adult mice, we compared C57BL/6, BALB/c, CD1(ICR), and 129Sv/J mice to examine proliferation, survival, and differentiation of newborn cells in the dentate gyrus. Proliferation was highest in C57BL/6; the survival rate of newborn cells was highest in CD1. In all strains approximately 60% of surviving newborn cells had a neuronal phenotype, but 129/SvJ produced more astrocytes. Over 6 days C57BL/6 produced 0.36% of their total granule cell number of 239,000 as new neurons, BALB/c 0.30% of 242,000, CD1 (ICR) 0.32% of 351,000, and 129/SvJ 0.16% of 280,000. These results show that different aspects of adult hippocampal neurogenesis are differentially influenced by the genetic background.     

Phenotypic expression of lethal chromosomal anomalies in human abortuses

From anatomical and cytogenetic studies of 1500 spontaneous abortuses attempts were made to establish correlations between the phenotype and karyotype of the specimens. The criteria used to determine the phenotype were based on estimated developmental age and macroscopic and microscopic examinations of embryo and placenta. Main features were described permitting, on anatomic examination of the conceptus, diagnosis of genetic anomalies, and in some instances possibly assigning the type of chromosomal aberration: viz., in monosomy X, triploidies, tetraploidies, and some trisomies. Phenotypic expression at the cellular level was studied by morphometry of the placenta and by study of in-vitro growth charcteristics of cells lines initiated from aneuploid embryos. Study of growth rate characteristics may reveal the basis for the pathogenesis of developmental arrests.     

Age-dependent silencing of globin transgenes in the mouse

Variegation of transgene expression, a heterocellular or mosaic pattern of expression seen in all mice in a given transgenic line, is a frequently observed but unexplained phenomenon. We have encountered variegation with globin transgenes; when lacZ expression is driven by globin control elements a proportion of erythrocytes express beta-galactosidase (beta-gal), while the remaining erythrocytes express none. The percentage of expressing cells is constant within each line (at any particular developmental stage), but varies between lines. Such variation may account for much of the line-to-line variability which has been reported in the expression of a transgene construct. We have now extended these observations by studying expression of several globin/lacZ transgenes with increasing age. Expression of beta-gal is variegated in all lines in adult mice, including those made with a beta-globin promoter and locus control region driving lacZ. The extent of variegation differs widely between lines, but in all lines there is a marked decline in the number of erythrocytes expressing beta-gal with increasing age. Progression of silencing continues long past the point at which globin switching is complete, suggesting that it is not related to this process. We observe that age-dependent silencing is most severe in high copy number animals. Increasing variegation of transgene expression with ageing of mice is likely to complicate interpretation of the developmental regulation of transgenes. We speculate that it reflects a general mechanism of epigenetic regulation.     

应用染色体G显带技术评估辐射剂量和染色体畸变的量效关系

目的:应用染色体G显带技术观察4MV的X线辐射后的染色体畸变情况,分析不同类型的染色体畸变和辐射剂量的关系.方法:以4MV的X线0.5、1、2、4 Gy对离体人外周血进行辐照,其淋巴细胞培养后,采用染色体G显带分析不同类型的染色体畸变率.结果:染色体双着丝粒、缺失、易位以及片段畸变和辐射剂量呈线性正相关,其线性趋势线和R<'2>值依次分别为:Y=24.1X-13.3(R<'2>=0.975);Y=10.5X-2.7(R<'2>=0.887);Y=30.2X-17.8(R<'2>=0.913);Y=53.3X-38.7(R<'2>=0.976).结论:染色体G显带技术可以较非显带技术提供更多更准确的畸变信息,为进一步研究提供借鉴.     

The influence of xenotransplant immunogenicity and immunosuppression on host MHC expression in the rat CNS

During the early stages following neural transplantation, host immune responses are initiated that are not normally found in the CNS including the induction of major histocompatibility antigens (MHC I and II). Previous laboratory findings have demonstrated prolonged survival of bovine chromaffin cells (BCC) in the rat CNS following transient immunosuppression with cyclosporin A (CSA) providing chromaffin cells are isolated from highly immunogenic passenger cells. To assess the influence of passenger and chromaffin cells on host MHC I and II expression, either BCC, nonchromaffin cell adrenal constituents (NCC), or adrenal medullary endothelial cells (EC) were implanted into the host. At 2 weeks postimplantation, robust BCC survival was obtained in CSA-treated animals. This correlated with low expression of MHC I at the host-graft border and the virtual absence of MHC II. Good BCC survival with reduced MHC I expression only was seen at 6 weeks postimplantation in animals transiently immunosuppressed (4 weeks). In contrast, poor survival was seen in the EC group (even with CSA treatment). In addition, marked MHC I and II expression was found in and around these grafts at 2 weeks, and was particularly intense in EC implanted animals. The results of this study suggest that nonchromaffin passenger cells in BCC preparations, most notably endothelial cells, can induce strong immune responses even in the presence of immunosuppression. Based on MHC staining, removal of these passenger cells can reduce host responses and improve long term survival of xenogeneic chromaffin cells in the CNS.