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Biomedical subjects

M A Watson

Publications and source records attributed to M A Watson.

At least 55 records · Page 3Linked to original sources
PubMed1999

Specificity from the synapsis of DNA elements by the Sfi I endonuclease.

The synapsis of DNA sites is a prerequisite for the reactions of many proteins that act at specific DNA sequences. The requirement for synapsis was investigated by analysing the reactions of Sfi I, a tetrameric restriction enzyme that cleaves DNA only after interacting with two recognition sites. In the presence of Mg2+, oligonucleotide duplexes with the cognate recognition sequence were cleaved rapidly, with cooperative kinetics, while non-cognate duplexes were not cleaved. In the absence of Mg2+, the primary complex formed by Sfi I with cognate DNA contained two duplexes synapsed by the tetramer: a secondary complex containing one duplex was seen only at elevated Sfi I concentrations. In contrast, the principal complex with non-cognate DNA contained one duplex bound to Sfi I. Pairs of non-cognate duplexes, or one cognate and one non-cognate duplex, generally failed to form synaptic complexes. On adding Mg2+to complexes with cognate DNA, cleavage occurred much more rapidly in the synaptic complex than in the secondary complex. DNA synapsis thus acts to enhance the specificity of Sfi I for its recognition sequence, by demanding two cognate sites for a catalytically active complex and by excluding non-cognate sites from the synaptic complex.

Catalysis↗
PubMed1999

Maternal separation: hypothalamic-preoptic area and hippocampal calbindin-D28K and calretinin in male and female infantile rats.

Calcium-binding proteins (CaPs) potentially play important roles in neurogenesis and neuronal survival. Calbindin-D28K (CALB) and calretinin (CALRET) in the medial basal hypothalamic (and preoptic area) (MBH) and hippocampus (HIPPO) from control and maternally separated male and female infantile rats were examined by Western analysis. Significantly greater levels of the CaPs in the MBH vs. the HIPPO may suggest enhanced or decreased neuroprotection, respectively, during the stress hyporesponsive period (SHRP). Male infantile rats separated from their mother's from postnatal day 2-10 displayed significant changes in CALB and CALRET for the MBH (decrease) and HIPP (increase) brain sites suggesting possible modified (negative feedback) mechanism(s) in HPA dysfunction observed during postnatal life.

Animals↗
PubMed1998

Identification of mammaglobin B, a novel member of the uteroglobin gene family.

In this report, we have identified, sequenced, and characterized the expression pattern of a novel human gene, mammaglobin B. Mammaglobin B (MGB2) is highly homologous to mammaglobin (MGB1), a previously characterized human gene whose expression is limited to the mammary epithelium and frequently up-regulated in human breast cancer cells. Based upon amino acid sequence similarities, both mammaglobin and mammaglobin B may be considered members of a larger, mammalian multigene family that includes rabbit uteroglobin, human Clara Cell 10-kDa protein (CC10), and the multimeric rat prostatein protein. Together with the human CC10 gene, mammaglobin and mammaglobin B are closely linked on human chromosome 11q13. However, despite their primary sequence similarity and close chromosomal proximity, the expression of mammaglobin and mammaglobin B is nonconcordant in both nonmalignant and neoplastic tissue.

Amino Acid Sequence↗
PubMed1998

Long-term outcome of a prospective randomized trial of conversion from cyclosporine to azathioprine treatment one year after renal transplantation.

BACKGROUND: Since the introduction of cyclosporine (CsA), 1-year renal allograft survival has improved, but concern persists about the long-term adverse effects of CsA, especially with respect to renal function and blood pressure. This randomized controlled trial was set up to establish whether withdrawal of CsA would alter long-term outcome. METHODS: Adult patients who, at 1 year after renal transplantation, had a stable serum creatinine of less than 300 micromol/L and who had not had acute rejection within the last 6 months were eligible for entry. Patients were randomized either to continue on CsA (n=114) or to stop CsA and start azathioprine (Aza, n=102). All patients remained on prednisolone. Median follow-up was 93 months after transplantation (range: 52-133 months). RESULTS: There was no significant difference in actuarial 10-year patient or graft survival (Kaplan-Meier), despite an increased incidence of acute rejection within the first few months after conversion. Median serum creatinine was lower in the Aza group (Aza: 119 micromol/L; CsA. 153 micromol/L at 5 years after randomization, P=0.0002). The requirement for antihypertensive treatment was also reduced after conversion to Aza; 75% of patients required antihypertensive treatment at the start of the study, decreasing to 55% from 1 year after randomization in the Aza group and increasing to >80% in the CsA group (55% (Aza) and 84% (CsA) at 5 years after randomization, P<0.005). CONCLUSIONS: Conversion from CsA to Aza at 1 year after renal transplantation results in improvement in both blood pressure control and renal allograft function, and is not associated with significant adverse effects on long-term patient or graft survival.

Acute Disease↗
PubMed1998

The human orphan nuclear receptor PXR is activated by compounds that regulate CYP3A4 gene expression and cause drug interactions.

The cytochrome P-450 monooxygenase 3A4 (CYP3A4) is responsible for the oxidative metabolism of a wide variety of xenobiotics including an estimated 60% of all clinically used drugs. Although expression of the CYP3A4 gene is known to be induced in response to a variety of compounds, the mechanism underlying this induction, which represents a basis for drug interactions in patients, has remained unclear. We report the identification of a human (h) orphan nuclear receptor, termed the pregnane X receptor (PXR), that binds to a response element in the CYP3A4 promoter and is activated by a range of drugs known to induce CYP3A4 expression. Comparison of hPXR with the recently cloned mouse PXR reveals marked differences in their activation by certain drugs, which may account in part for the species-specific effects of compounds on CYP3A gene expression. These findings provide a molecular explanation for the ability of disparate chemicals to induce CYP3A4 levels and, furthermore, provide a basis for developing in vitro assays to aid in predicting whether drugs will interact in humans.

Amino Acid Sequence↗
PubMed1998

Mutagenesis of human Mel1a melatonin receptor expressed in yeast reveals domains important for receptor function.

A yeast functional colorimetric assay was employed to test the effects of site-directed point mutations on the function of the human Mel1a melatonin receptor. Seven mutants were created in transmembrane domains III, V, and VII of the receptor to test the rhodopsin-based model of melatonin recognition. Two mutants in transmembrane domains III and VI were created to investigate the mechanisms of G protein activation in the melatonin receptor. Mutations in transmembrane domain V either potentiated agonist efficiencies (H195A) or totally abolished all responses to tested compounds (V192T+H195A). Mutation N124A in the conserved NRY motif in the end of transmembrane domain III seriously impaired receptor activation. Several mutants were found to have decreased ability to activate functional responses, reflecting the importance of these residues for receptor function. These data also suggest that activation of the receptor involves interaction of the 5-methoxy group of melatonin with the conserved histidine H195 in transmembrane domain V.

Binding Sites↗
PubMed1998

Structure and transcriptional regulation of the human mammaglobin gene, a breast cancer associated member of the uteroglobin gene family localized to chromosome 11q13.

The mammaglobin gene encodes a novel secreted protein whose corresponding mRNA is frequently up-regulated in human breast cancer. In non-malignant tissues, expression is also strictly limited to the mammary epithelium. To better understand the mechanisms controlling these patterns of expression, we have isolated the human mammaglobin gene and performed an initial assessment of its promoter activity. Mammaglobin gene architecture is very similar to that of a family of related genes that includes uteroglobin and rat prostatein subunits C1, C2, and C3. However, the mammaglobin gene itself is not well conserved phylogenetically. The human mammaglobin gene is localized by fluorescent in situ hybridization to chromosome 11 band q13, a genomic region frequently amplified in breast neoplasia. The sequence of proximal 1 kb of mammaglobin promoter contains several potential transcriptional control elements and directs high-level expression of a transfected reporter construct in human breast tumor cell lines. However, comparable levels of reporter gene expression are also seen in non-mammary human cell lines. These data suggest that, unlike related gene family members, the striking breast-specific expression and tumor-associated overexpression of mammaglobin is mediated by complex transcriptional control at more distal sequence elements.

Base Sequence↗
PubMed1998

An orphan nuclear receptor activated by pregnanes defines a novel steroid signaling pathway.

Steroid hormones exert profound effects on differentiation, development, and homeostasis in higher eukaryotes through interactions with nuclear receptors. We describe a novel orphan nuclear receptor, termed the pregnane X receptor (PXR), that is activated by naturally occurring steroids such as pregnenolone and progesterone, and synthetic glucocorticoids and antiglucocorticoids. PXR exists as two isoforms, PXR.1 and PXR.2, that are differentially activated by steroids. Notably, PXR.1 is efficaciously activated by pregnenolone 16alpha-carbonitrile, a glucocorticoid receptor antagonist that induces the expression of the CYP3A family of steroid hydroxylases and modulates sterol and bile acid biosynthesis in vivo. Our results provide evidence for the existence of a novel steroid hormone signaling pathway with potential implications in the regulation of steroid hormone and sterol homeostasis.

Amino Acid Sequence↗
PubMed1998

Androgen-dependent modulation of calbindin-D28K in hypothalamic tissue during prenatal development.

We examined the influence of androgens on fetal medial basal hypothalamic and preoptic area (MBH-POA) calbindin-D28K levels (via Western analysis) by treating pregnant rats with testosterone or flutamide, (an androgen receptor blocker). MBH-POA calbindin-D28K levels were significantly decreased in flutamide-treated male fetuses, whereas, MBH-POA calbindin-D28K levels were significantly increased in testosterone-treated female fetuses compared to controls. These results suggest that MBH-POA calbindin-D28K is modulated during prenatal development by androgens.

Anal Canal↗
PubMed1998

Human glutathione S-transferase P1 polymorphisms: relationship to lung tissue enzyme activity and population frequency distribution.

The association between glutathione S-transferase (GST) activity as measured by 1-chloro-2,4-dinitrobenzene (CDNB) conjugation and genotype at exon 5 and exon 6 of the human GSTP1 gene was investigated in normal lung tissue obtained from 34 surgical patients. These samples were genotyped for previously identified polymorphisms in exon 5 (Ile105Val) and exon 6 (Ala114Val) by PCR-RFLP and direct sequencing. GST enzyme activity was significantly lower among individuals with the 105 Val allele. Homozygous Ile/Ile samples (n = 18) had a mean cytosolic CDNB conjugating activity of 74.9 +/- 3.8 nmol/mg per min; heterozygotes (n = 13) had a mean specific activity of 62.1 +/- 4.2 nmol/mg per min and homozygous Val/Val (n = 3) had a mean specific activity of 52.5 +/- 4.5 nmol/mg per min. The CDNB conjugating activity measured for the Ile/Ile genotype group was significantly different from that observed in the Ile/Val group (P = 0.03), and from Ile/Val and Val/Val genotypes combined (P = 0.009). Mean GST activity values were consistently lower in individuals with genotypes containing the 105 valine allele, regardless of smoking exposure. Genotypes at codon 114 were also assessed but the mean GST activity was not significantly lower in individuals with the 114 valine allele. A new haplotype, present in two samples who were homozygous 105Ile and had a 114Val, was identified and proposed as GSTP1*D. Frequencies of the exon 5 and exon 6 polymorphisms were determined in samples obtained from European-Americans, African-Americans and Taiwanese. The differences observed were highly significant suggesting the possibility of GSTP1 genotype-associated, ethnic differences in cancer susceptibility and chemotherapeutic response.

Asian People↗
PubMed1998

Factors associated with involuntary return to a psychiatric emergency service within 12 months.

OBJECTIVE: This study examined patient characteristics and other factors that contributed to the involuntary return of patients to a psychiatric emergency service within 12 months of an initial evaluation in the service. The findings were used to consider whether the pressure to limit duration of hospital stays under managed care contributed to the patients' return to the emergency service. METHODS: Structured observations of evaluations of 417 patients admitted to the psychiatric emergency service were completed at seven county general hospitals in California. Twelve months after the initial evaluation, mental health and criminal justice records were reviewed for evidence of the patients' return for emergency psychiatric evaluation at any of the seven hospitals. Factors associated with patients' return to the psychiatric emergency service were evaluated using multivariate modeling. RESULTS: Of the 417 patients initially evaluated, 121, or 29 percent, were involuntarily returned to the psychiatric emergency service within 12 months. The likelihood of involuntary return was increased by a psychotic diagnosis and indications of dangerousness at the initial evaluation. Having insurance also increased the likelihood of involuntary return. CONCLUSIONS: The patient's initial condition in the psychiatric emergency service was found to be the best predictor of involuntary return. Brief hospitalization--an average of six days--after the evaluation did not have a significant prophylactic effect, perhaps because the reduced length of inpatient stay in the managed care environment did not allow adequate resolution of the patient's clinical condition.

Adult↗
PubMed1997

An association between the allele coding for a low activity variant of catechol-O-methyltransferase and the risk for breast cancer.

Mounting evidence suggests that catechol metabolites of estradiol may contribute to the development of estrogen-induced cancers. O-Methylation, catalyzed by catechol-O-methyltransferase (COMT), inactivates catechol estrogens. COMT is polymorphic in the human population, with 25% of Caucasians being homozygous for a low activity allele of the enzyme (COMT(LL)). We hypothesized that low activity COMT may be a risk factor for human breast cancer and designed a PCR-based RFLP assay to determine COMT genotype in a cohort of 112 matched, nested case-control samples. In the total study population, the odds ratios for the association of breast cancer risk with COMT(HL) and COMT(LL) genotypes were 1.30 [confidence interval (CI), 0.66-2.58] and 1.45 (CI, 0.69-3.07), respectively. Postmenopausal COMT(LL) women had a greater than 2-fold increased risk of developing breast cancer [odds ratio (OR), 2.18; CI, 0.93-5.11]. The association of COMT(LL) with the development of postmenopausal breast cancer was stronger and statistically significant in those women with a body mass index >24.47 kg/m2 (OR, 3.58; CI, 1.07-11.98). When COMT(LL) was combined with either glutathione S-transferase (GST) M1 null or with GSTP1 Ile-105-Val/Val-105-Val (intermediate/low activity, respectively) genotypes, the risk for developing postmenopausal breast cancer was also significantly increased. Our findings suggest that the allele encoding low activity COMT may be an important contributor to the postmenopausal development of breast cancer in certain women.

Aged↗
PubMed1997

Pilot study of free and conjugated urinary mutagenicity during consumption of pan-fried meats: possible modulation by cruciferous vegetables, glutathione S-transferase-M1, and N-acetyltransferase-2.

Epidemiological and experimental evidence indicates that consumption of fried meats in conjunction with certain genotypes of phase I and II metabolism genes poses an elevated risk for colorectal cancer. Parallel to this, the consumption of cruciferous vegetables is associated with a reduced risk of colon cancer. Therefore, we designed a 6-week pilot feeding study to evaluate the effect of these variables on urinary mutagenicity, which is a biomarker associated with fried-meat consumption. Eight subjects were fed fried meats daily for six weeks; four ate cruciferous vegetables, and four ate non-cruciferous vegetables. Urinary mutagenicity was evaluated in the presence of S9 in strain YG1024 of Salmonella, which is a frameshift strain that overproduces acetyltransferase. C18/methanol extracts of 24-h urines collected once each week were tested unhydrolyzed (free mutagenicity) and hydrolyzed (total mutagenicity); the difference between the two was the conjugated mutagenicity. Although not significant, the levels of conjugated urinary mutagenicity doubled among crucifera consumers and decreased to 30% of the initial levels among non-crucifera consumers, suggesting the possibility that crucifera may enhance the level of conjugated urinary mutagenicity resulting from consumption of fried meats. Such an effect would be consistent with the documented ability of cruciferous vegetables to induce phase II enzymes. The NAT2 rapid phenotype was significantly associated with approximately 2-fold increases in conjugated (p = 0.05) and total (p = 0.004) urinary mutagenicity relative to NAT2 slow subjects, consistent with the elevated risk confirmed by the NAT2 rapid phenotype for colorectal cancer among meat consumers. An approximately 2-fold increase in urinary mutagenicity among the GSTM1- subjects relative to the GSTM1+ subjects approached significance for free (p = 0.18) and total (p = 0.13) urinary mutagenicity. This is the first report on (a) the mutagenicity of hydrolyzed urine, which was consistently more mutagenic than unhydrolyzed urine; (b) the potential enhancement of conjugated urinary mutagenicity by crucifera; and (c) the association of the rapid NAT2 and possibly the GSTM1- phenotype with elevated levels of fried meat-associated urinary mutagenicity.

Aged↗
PubMed1997

Calbindin-D28k is regulated by adrenal steroids in hypothalamic tissue during prenatal development.

Regulation of calbindin-D28k (CALB) in the medial basal hypothalamus (MBH) from male and female fetuses was examined by Western analysis. Control fetal males displayed significantly higher MBH CALB levels compared to females at gestational day 20. Whereas, in general, the lowest CALB levels were recorded in male and female fetuses from long-term prenatally stressed or fetuses from adrenalectomized pregnant rats. These data indicate that corticosterone regulates MBH CALB expression during prenatal development and CALB may be implicated in modulating the sexual differentiation of neural structures within the MBH during perinatal development.

Adrenal Cortex Hormones↗
PubMed1997

Developmental expression of calretinin in the medial basal hypothalamus and amygdala from male and female rats.

Developmental expression of calretinin in the medial basal hypothalamic (MBH) and amygdala region was examined by Western analysis. Males displayed significantly higher calretinin levels compared to females in the MBH (but not the amygdala) on gestational day 19 and 20. These data imply that hormonal factors may regulate developmental MBH calretinin expression. In turn, sexually dimorphic brain structures might be influenced by calretinin levels that can alter sexually dimorphic patterns of steroidogenesis, cellular migration or programmed cell loss mechanism(s) during neuronal development by modulating intracellular calcium concentrations.

Amygdala↗
PubMed1997

Parent stress and coping in NICU and PICU.

The purpose of this study was to identify and compare parental perceptions of their stress and coping experiences with children in pediatric intensive care units (PICU) and the neonatal intensive care units (NICU). The sample consisted of 31 NICU and 20 PICU parents. Parents in both units experienced the most stress from alteration in their parenting role and in their infants' behavior and appearance. Parents of children in PICU found assistance with parenting role more helpful than parents of children in NICU. Parents with children in the PICU perceived problems-focused coping more helpful than parents with children in the NICU; parents of children in NICU found emotion-focused coping more helpful than parents of children in PICU. Parents in both units considered problem-focused coping more helpful than appraisal- or emotion-focused coping.

Adaptation, Psychological↗