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Biomedical subjects

A West

Publications and source records attributed to A West.

At least 19 recordsLinked to original sources
PubMed2005

A human ImmunoChip cDNA microarray provides a comprehensive tool to study immune responses.

DNA microarray technology has developed rapidly in recent years and has become an essential tool, providing novel approaches to biomedical research. In this paper, we describe a self-designed ImmunoChip cDNA array for immunological research. With a comprehensive selection of genes of interest, we can focus on key signalling pathways and molecular mechanisms at relatively low cost compared to commercial platforms which are usually targeted at global screening of gene expression. To validate the efficiency of the ImmunoChip, we studied T helper cell polarization to functionally distinct subsets (Th1 and Th2). We also developed a tool for quality control of cDNA microarrays that assesses the technical quality of an ImmunoChip. The information produced with the quality control tool is shown to be valuable for extracting correct information from cDNA microarrays. Gene expression measurements with ImmunoChip are in agreement with the results obtained using oligonucleotide microarrays and with published quantitative RT-PCR data. The ImmunoChip provides reliable measurements and gives new insights into various aspects of human immune responses.

DNA, Complementary↗
PubMed2003

Parkin-proven disease: common founders but divergent phenotypes.

OBJECTIVE: To compare and contrast clinical and genetic findings in six probands with parkinsonism with a parkin exon 3 438- to 477-bp deletion (Ex3Delta40) to search for evidence of a common founder. METHOD: Clinical review, parkin gene sequencing, dosage studies, and high-resolution genotype/haplotype analysis were performed. RESULTS: All subjects had two or more signs consistent with a diagnosis of possible or probable PD with age at onset younger than 45 years (mean +/- SD 29.3 +/- 10.2 years, range 16 to 42 years). Affected individuals were either homozygotes, compound heterozygotes, or Ex3Delta40 carriers with one normal parkin allele. Haplotype analysis revealed both Ex3Delta40 and Ex7 924 C-->T (R275W) mutations originated from common founders, the former most probably of Irish descent. Although three cases had Ex7 924 C-->T (R275W) and Ex3Delta40 mutations, their clinical presentation and mode of inheritance were variable. CONCLUSION: Parkin mutations on common parkin haplotypes provide testable hypotheses of parkin function in genetically defined parkinsonism.

Adult↗
PubMed2002

Evidence to suggest biased phenotypes in children with Attention Deficit Hyperactivity Disorder from completely ascertained trios.

The transmission disequilibrium test (TDT) is widely used as a robust statistical method to test for genetic association due to linkage based upon analysis of parent-proband trios. The TDT and other family-based tests (eg haplotype relative risk method) are commonly used in association studies including those of ADHD because of concerns that the case-control design has a strong tendency for false positives due to poor matching between cases and controls. Unfortunately, it is not always possible to obtain DNA from both parents in studies of this design, even where the onset of disorder is in childhood, and usually the missing parent is the father. Despite the fact that methods exist for analysis where one parent is missing, many family-based studies are based on the collection or analysis of complete trios only. However this selection process might potentially introduce bias, particularly for studies of behavioural phenotypes like ADHD because the phenotype of proband or parents might influence family stability and therefore complete parental ascertainment. We set out to examine whether children with ADHD and for whom DNA samples from fathers were missing ('duos') differed phenotypically from children for whom genotype information was available from both parents ('trios'). Children from duos showed a significantly higher frequency of DMS-IV ADHD-combined type, significantly more co-morbid conduct disorder and conduct disorder symptoms, and a trend for higher total ADHD symptom scores. Excluding duos from sample collection and analysis may result in systematic bias. If comorbid conduct disorder and ADHD-combined type index increased genetic liability, exclusion of duos could further reduce the power of the TDT (and similar tests) to detect susceptibility genes for ADHD, or replicate effects detected by case-control analysis.

Adolescent↗
PubMed2002

Stage-specific expression of Gadd45 induced by X-irradiation in rat spermatogenesis.

PURPOSE: Gadd45 is involved in the response to DNA damage in somatic cells. The effect of X-irradiation and chemical treatments on expression of Gadd45 and two other 53-regulated genes, p21 and cyclin-G, was studied in rat testis. MATERIALS AND METHODS: The reverse-transcriptase polymerase chain reaction (RT-PCR) was performed on testis extracts of control, X-irradiated (6Gy), etoposide (10 mg kg(-1)) and adriamycin (5mg kg(-1))-treated rats. For stage-specific analysis, seminiferous tubules were isolated and segments representing the 14 epithelial stages were obtained. RESULTS: In whole testis extracts, increases in Gadd45, p21 and cyclin-G expression were detectable after irradiation, but not after etoposide or adriamycin treatments. Analysis of fractions consisting of defined epithelial stages showed a high expression of Gadd45 in stages VII-XII and of p21 in stages VII-VIII. Irradiation significantly increased the level of Gadd45 mRNA in stages VI-VIII and of p21 mRNA in stages VI-I. Although no overall increase could be observed in whole testis samples of the etoposide-treated rat, stage-specific analysis revealed an induction of p21 expression in stages XIII-I. Gadd45 and cyclin-G mRNA were localized to spermatocytes and round spermatids known to express p21. CONCLUSIONS: Although X-irradiation, etoposide and adriamycinare known spermatogenic mutagens and activators of apoptosis, only X-rays induce slightly Gadd45 expression in testis. This small induction was very stage specific.

Animals↗
PubMed2001

Development and application of a sensitive high performance ion-exchange chromatography method for the simultaneous measurement of dopamine, 5-hydroxytryptamine and norepinephrine in microdialysates from the rat brain.

A high performance liquid chromatography (HPLC) method based on cation exchange separation has been developed for the measurement of dopamine (DA), 5-hydroxytryptamine (5-HT) and norepinephrine (NE) in microdialysates. The separation conditions have been optimised for using electrochemical detection. All three bioamines were resolved in less than 22 min using isocratic conditions. The optimum oxidation potential for the three bioamines was found to be +0.4 V vs. in situ Ag/AgCl reference electrode. Linear regression analysis of HPLC-peak area as a function of concentrations in the range 1-50 ng x ml(-1) gave coefficients of correlation between 0.998 and 0.999. The limit of detection for DA, 5-HT and NE was found to be between 50 and 100 pg x ml(-1) with a signal to noise ratio of 3:1. The method has been applied to the simultaneous measurement of the three monoamines in microdialysates from the medial prefrontal cortex under basal conditions and following the administration of the antipsychotic drug clozapine (10 mg x kg(-1) s.c.).

Animals↗
PubMed2001

Neurokinin-1 receptor immunoreactivity in hypotension sensitive sympathetic preganglionic neurons.

Substance P activation of neurokinin-1 (NK1) receptors on spinal sympathetic preganglionic neurons (SPN) influences blood pressure. We identified SPN likely to subserve the baroreceptor reflex and established if these neurons showed NK1 receptor-immunoreactivity. Nitroprusside (NP) infusion or inferior vena cava (IVC) constriction activated similar numbers of SPN. Of these, about 40% were NK1 receptor-immunoreactive after NP infusion, but only about 20% were NK1 receptor-immunoreactive after IVC constriction. The distribution of Fos/NK1 receptor SPN suggested that substance P may preferentially target sympathoadrenal SPN.

Animals↗
PubMed2001

alpha-Synuclein gene haplotypes are associated with Parkinson's disease.

We report haplotype analysis of the alpha-synuclein gene in Parkinson's disease (PD), extending earlier reports of an association with a polymorphism within the gene promoter. This analysis showed significant differences in haplotypes between PD cases and controls. Our analyses demonstrate that genetic variability in the alpha-synuclein gene is a risk factor for the development of PD. These genetic findings are analogous to the tau haplotype over-represented in progressive supranuclear palsy and further extend the similarity in the etiologies and pathogeneses of the synucleinopathies and tauopathies.

Adult↗
PubMed2001

Tamoxifen for the prevention of breast cancer: psychosocial impact on women participating in two randomized controlled trials.

PURPOSE: The purpose of this study was to evaluate the psychosocial implications of tamoxifen versus placebo in women who are at increased risk of breast cancer. PATIENTS AND METHODS: The 488 women in the psychosocial study were recruited from participants in two placebo-controlled, double-blind, randomized, controlled trials that investigated the efficacy of tamoxifen in the prevention of breast cancer in women who are at high familial risk. During a 5-year period, repeated assessments were made of anxiety, psychological distress, and sexual functioning using standardized questionnaires before treatment at baseline and at 6-month intervals during the trial. RESULTS: Questionnaire completion over 5 years was good, with 71.1% of women returning at least 8 of 10 follow-up assessments. Although scores from individuals showed considerable fluctuation and variation over time, changes in anxiety, mood, and sexual functioning were not associated with treatment group. The number of symptoms reported at 48 months via a self-report checklist were not associated with treatment group, but vasomotor symptoms were more frequent among tamoxifen-treated women. Symptoms of low energy, breast sensitivity, and visual blurring were reported most frequently in the placebo group. CONCLUSION: In general, these results are comparable to those from the National Surgical Adjuvant Breast and Bowel Project psychosocial study despite differences in study populations, methodology, and instruments. The long-term use of tamoxifen and other selective estrogen response modulators as preventive agents in high-risk groups has been questioned, but we found no evidence of treatment-related side effects that affect women's psychosocial and sexual functioning.

Adult↗
PubMed2001

X-irradiation--induced changes in the progression of type B spermatogonia and preleptotene spermatocytes.

In response to induced DNA damage, proliferating cells arrest in their cell cycle or go into apoptosis. Ionizing radiation is known to induce degeneration of mammalian male germ cells. The effects on cell-cycle progression, however, have not been thoroughly studied due to lack of methods for identifying effects on a particular cell-cycle phase of a specific germ cell type. In this study, we have utilized the technique for isolation of defined segments of seminiferous tubules to examine the cell-cycle progression of irradiated rat mitotic (type B spermatogonia) and meiotic (preleptotene spermatocytes) G1/S cells. Cells irradiated as type B spermatogonia in mitotic S phase showed a small delay in progression through meiosis. Thus, it seems that transient arrest in the progression can occur in the otherwise strictly regulated progression of germ cells in the seminiferous epithelium. Contrary to the arrest observed in type B spermatogonia and in previous studies on somatic cells, X-irradiation did not result in a G1 delay in meiotic cells. This lack of arrest occurred despite the presence of unrepaired DNA damage that was measured when the cells had progressed through the two meiotic divisions.

Animals↗
PubMed2001

Expression profiling using microarrays fabricated by an ink-jet oligonucleotide synthesizer.

We describe a flexible system for gene expression profiling using arrays of tens of thousands of oligonucleotides synthesized in situ by an ink-jet printing method employing standard phosphoramidite chemistry. We have characterized the dependence of hybridization specificity and sensitivity on parameters including oligonucleotide length, hybridization stringency, sequence identity, sample abundance, and sample preparation method. We find that 60-mer oligonucleotides reliably detect transcript ratios at one copy per cell in complex biological samples, and that ink-jet arrays are compatible with several different sample amplification and labeling techniques. Furthermore, results using only a single carefully selected oligonucleotide per gene correlate closely with those obtained using complementary DNA (cDNA) arrays. Most of the genes for which measurements differ are members of gene families that can only be distinguished by oligonucleotides. Because different oligonucleotide sequences can be specified for each array, we anticipate that ink-jet oligonucleotide array technology will be useful in a wide variety of DNA microarray applications.

Cells, Cultured↗
PubMed2001

Identification and characterization of the human parkin gene promoter.

Compound mutations and homozygous loss of function of the parkin gene causes juvenile and early onset, autosomal recessive parkinsonism. Pathologically, the disease is associated with loss of dopaminergic neurons in the substantia nigra pars compacta and locus ceruleus, usually without Lewy body pathology. Hemizygous families have been described that may harbor mutations outside of the open reading frame. The parkin gene promoter has yet to be characterized, and therein, mutations in hemizygous families may plausibly be identified. To identify the promoter of the parkin gene, the transcription start site was defined by a combination of primer extension and 5' RACE. Five kilobases of DNA 5' to the parkin start codon were directly sequenced from a BAC containing parkin exon 1 and evaluated for promoter motifs. The parkin promoter lacks TATA or CAAT boxes and appears to share homology to the alpha-synuclein promoter. Deletion constructs demonstrated core promoter activity and tissue specific enhancing regions in HEK-293T and SH-SY5Y cells.

5' Untranslated Regions↗
PubMed2000

Cytoplasmic catalytic subunit of protein kinase A mediates cross-repression by NF-kappa B and the glucocorticoid receptor.

Negative transcriptional regulation or cross-coupling between NF-kappa B (RelA) and the glucocorticoid receptor (GR) is proposed to play a regulatory role in human physiology and disease. Despite previous advances, the biochemical basis of this phenomenon remains a subject of controversy. We show here that the inhibition of GR activity by RelA does not require the RelA DNA binding, transactivation, or nuclear localization domains. Surprisingly, RelA repression of GR is abolished by mutation of the conserved protein kinase A (PKA) site at amino acid residue 276 of RelA. We show that GR associates in vivo and in vitro with the catalytic subunit of PKA (PKAc) in a ligand-independent manner and that GR transcription depends on PKA signaling. Indeed, we demonstrated that GR-mediated inhibition of NF-kappa B transactivation is PKAc-dependent. In contrast to previous models, we suggest that the cross-coupling requires a cytoplasmic step and is regulated by a PKAc-associated signaling.

Catalytic Domain↗
PubMed2000

Stability of carbohydrate deficient transferrin (CDT) in stored blood samples.

The alcoholism marker CDT was determined on 257 blood samples 1-2 days after the blood samples were taken and again after storage for 7 months at +4 degrees C. The differences between the pairs of CDT values were so large that the determination of CDT after long term storage of the blood sample has no evidential value.

Alcoholism↗
PubMed2000

ROC analysis of alcoholism markers--100% specificity.

A combination of 4 so-called markers of alcoholism, i.e. methanol, acetone + 2-propanol, gamma-glutamyltransferase and carbohydrate deficient transferrin, was investigated in 341 blood samples from alcoholics and non-alcoholics. From the history of alcohol consumption, four defined subgroups were formed: non-alcoholics divided into (A) 33 persons with no ethanol consumption during the past year and (B) 60 persons with daily consumption less than 40 g ethanol. Alcoholics were divided into (C) 177 persons with no ethanol at the time of admission/first blood sampling (withdrawal therapy) and (D) 71 persons with positive ethanol levels on admission/first blood sampling. All markers showed different extents of overlap between the collectives of alcoholics and non-alcoholics. By logistic regression, a formula was developed combining these markers with different mathematical weights. Thus an "Alc-Index" could be calculated for each individual. The ROC curve connecting all individual values gives an ideal form with 100% specificity and nearly 93% sensitivity. The threshold between the collectives of alcoholics and non-alcoholics was defined by the Alc-Index value 1.7. This was associated with no false positives among the non-alcoholics while nearly 93% of the alcoholics exceeded this index. The ROC-based calculation of the Alc-Index thus seems to be the most effective method for the diagnosis of alcoholism.

1-Propanol↗
PubMed2000

Characterization of human HtrA2, a novel serine protease involved in the mammalian cellular stress response.

Human HtrA2 is a novel member of the HtrA serine protease family and shows extensive homology to the Escherichia coli HtrA genes that are essential for bacterial survival at high temperatures. HumHtrA2 is also homologous to human HtrA1, also known as L56/HtrA, which is differentially expressed in human osteoarthritic cartilage and after SV40 transformation of human fibroblasts. HumHtrA2 is upregulated in mammalian cells in response to stress induced by both heat shock and tunicamycin treatment. Biochemical characterization of humHtrA2 shows it to be predominantly a nuclear protease which undergoes autoproteolysis. This proteolysis is abolished when the predicted active site serine residue is altered to alanine by site-directed mutagenesis. In human cell lines, it is present as two polypeptides of 38 and 40 kDa. HumHtrA2 cleaves beta-casein with an inhibitor profile similar to that previously described for E. coli HtrA, in addition to an increase in beta-casein turnover when the assay temperature is raised from 37 to 45 degrees C. The biochemical and sequence similarities between humHtrA2 and its bacterial homologues, in conjunction with its nuclear location and upregulation in response to tunicamycin and heat shock suggest that it is involved in mammalian stress response pathways.

Alanine↗
PubMed2000

Language mapping in pretreatment planning of patients with cerebral arteriovenous malformation: a PET study.

PURPOSE: Three patients who had cerebral arteriovenous malformations underwent language mapping by positron emission tomography (PET) to identify the dominant hemisphere for language and the spatial relation of lesions to language areas. METHODS: Mapping was performed to assess the risk that surgery could cause neurologic deficits and to plan other therapeutic strategies. RESULTS: The information obtained by PET language mapping changed the course of management for these patients. Two patients were treated with stereotactic radiosurgery and one patient with surgical resection. CONCLUSIONS: PET imaging can be a valuable noninvasive tool for mapping the functional cortex and for preoperative planning of different treatment options in patients with arteriovenous malformations.

Adolescent↗