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K E Chapman

Publications and source records attributed to K E Chapman.

At least 19 recordsLinked to original sources
PubMed2006

Prenatal programming of hepatocyte nuclear factor 4alpha in the rat: A key mechanism in the 'foetal origins of hyperglycaemia'?

AIMS/HYPOTHESIS: Prenatal glucocorticoid exposure causes lifelong hyperglycaemia in rat offspring, associated with permanently increased hepatic phosphoenolpyruvate carboxykinase 2 (PCK2), the rate-controlling enzyme of gluconeogenesis. To elucidate the mechanisms underlying the 'programming' of PCK2, this study examined the effect of prenatal dexamethasone treatment on expression of transcription factors that regulate Pck2. MATERIALS AND METHODS: Real-time RT-PCR and in situ hybridisation were used to measure and localise hepatic mRNA transcribed from the genes for PCK2, hepatocyte nuclear factor 4, alpha (HNF4A), transcription factor 1 (TCF1), CCAAT/enhancer binding protein, alpha (CEBPA), CEBPB, the glucocorticoid receptor (NR3C1) and peroxisome proliferative activated receptor, gamma, coactivator 1 alpha (PPARGC1A) in foetal and adult offspring of dams treated with dexamethasone or vehicle during the last week of gestation. RESULTS: Prenatal dexamethasone exposure significantly elevated Hnf4a mRNA expression in foetal and adult liver. This resulted from increased expression of isoforms derived from the 'adult' (P1) Hnf4a promoter. In contrast, isoforms from the 'foetal' (P2) promoter were markedly suppressed by dexamethasone. Like Pck2, the increase in hepatic Hnf4a mRNA occurred exclusively in the periportal zone. Foetal Tcf1 expression was also increased by dexamethasone treatment, but this did not persist into adulthood. Prenatal dexamethasone did not affect the amounts of foetal and/or adult Cebpa, Cebpb, Nr3c1 or Ppargc1a mRNA. CONCLUSIONS/INTERPRETATION: Prenatal dexamethasone exposure caused a permanent increase in hepatic Hnf4a mRNA. This increase, which was associated with a premature switch from foetal to adult promoter predominance, was congruent with changes in Pck2 expression. These data suggest that HNF4A might mediate Pck2 overexpression and subsequent hyperglycaemia.

Animals↗
PubMed2003

Differential regulation of corticosteroid receptors by monoamine neurotransmitters and antidepressant drugs in primary hippocampal culture.

Hyperactivity of the hypothalamic-pituitary-adrenal axis is a characteristic feature of depressive illness. The centrally located corticosteroid receptors, the glucocorticoid and mineralocorticoid receptors, are thought to be important modulators of this axis and changes in the levels of these receptors, particularly in the hippocampus, may underlie the hyperactivity observed. Various antidepressant drugs increase hippocampal mineralocorticoid and glucocorticoid receptor levels in vivo. These effects are thought to be mediated via alterations in monoaminergic neurotransmission. We examined whether serotonin (5HT) and noradrenaline (NA) have direct effects on glucocorticoid receptor and mineralocorticoid receptor expression in primary hippocampal neurones, and whether antidepressants also exert direct effects on target neurones. Exposure of hippocampal cells to 5HT for 4 days increased both glucocorticoid and mineralocorticoid receptor mRNA and protein expression. The induction of mineralocorticoid receptor mRNA was completely blocked by the 5HT(7) receptor antagonist SB 269970. In contrast glucocorticoid receptor induction was insensitive to the 5HT(7) receptor, whilst studies with the 5HT(1A) receptor agonist 8-hydroxy-2-(di-n-proplamino) tetralin hydrochloride and the 5HT(1A) receptor antagonist N-[2-[4-2-[O-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexane carboxamide trihydrochloride (WAY 100635) suggest a partial role for 5HT(1A) receptors in hippocampal glucocorticoid receptor regulation. Treatment with NA for 4 days also increased glucocorticoid receptor expression but had no effect on mineralocorticoid receptor expression. This was blocked by propanolol suggesting action via beta-adrenergic receptors. Similarly to NA, fluoxetine and amitriptyline also selectively increased glucocorticoid receptor mRNA and protein levels over this time course. However, glucocorticoid receptor induction by fluoxetine or amitriptyline was not blocked by WAY 100635 or propanolol. These results show that 5HT, NA and antidepressants act directly but via distinct mechanisms on hippocampal neurones to regulate mineralocorticoid and glucocorticoid receptor expression. Thusly, manipulation of neurotransmitter or antidepressant levels in the brain may aid in reversing hypothalamic-pituitary-adrenal axis hyperactivity by restoring hippocampal corticosteroid receptor balance.

8-Hydroxy-2-(di-n-propylamino)tetralin↗
PubMed2001

Early environmental regulation of hippocampal glucocorticoid receptor gene expression: characterization of intracellular mediators and potential genomic target sites.

Environmental conditions in early life permanently alter the development of glucocorticoid receptor gene expression in the hippocampus and hypothalamic-pituitary-adrenal responses to acute or chronic stress. In part, these effects can involve an activation of ascending serotonergic pathways and subsequent changes in the expression of transcription factors that might drive glucocorticoid receptor expression in the hippocampus. This paper summarizes the evidence in favor of these pathways as well as recent studies describing regulatory targets on the promoter region of the rat hippocampal glucocorticoid receptor gene.

Animals↗
PubMed2000

C/EBP regulates hepatic transcription of 11beta -hydroxysteroid dehydrogenase type 1. A novel mechanism for cross-talk between the C/EBP and glucocorticoid signaling pathways.

Glucocorticoid action within individual cells is potently modulated by 11beta-hydroxysteroid dehydrogenase (11beta-HSD), which, by interconverting active and inert glucocorticoids, determines steroid access to receptors. Type 1 11beta-HSD (11beta-HSD1) is highly expressed in liver where it regenerates glucocorticoids, thus amplifying their action and contributing to induction of glucocorticoid-responsive genes, most of which are also regulated by members of the C/EBP (CAAT/enhancer-binding protein) family of transcription factors. Here we demonstrate that C/EBPalpha is a potent activator of the 11beta-HSD1 gene in hepatoma cells and that mice deficient in C/EBPalpha have reduced hepatic 11beta-HSD1 expression. In contrast, C/EBPbeta is a relatively weak activator of 11beta-HSD1 transcription in hepatoma cells and attenuates C/EBPalpha induction, and mice that lack C/EBPbeta have increased hepatic 11beta-HSD1 mRNA. The 11beta-HSD1 promoter (between -812 and +76) contains 10 C/EBP binding sites, and mutation of the promoter proximal sites decreases the C/EBP inducibility of the promoter. One site encompasses the transcription start, and both C/EBPalpha and C/EBPbeta are present in complexes formed by liver nuclear proteins at this site. The regulation of 11beta-HSD1 expression, and hence intracellular glucocorticoid levels, by members of the C/EBP family provides a novel mechanism for cross-talk between the C/EBP family of transcription factors and the glucocorticoid signaling pathway.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed2000

Linkage analysis of chromosome 2q in osteoarthritis.

BACKGROUND: In independent linkage studies chromosome 2q11-q24 and chromosome 2q23-35 have previously been implicated as regions potentially harbouring susceptibility loci for osteoarthritis (OA). OBJECTIVE: To test chromosome 2q for linkage to idiopathic osteoarthritis. METHODS: Using a cohort of 481 OA families that each contained at least one affected sibling pair with severe end-stage disease (ascertained by hip or knee joint replacement surgery), we conducted a linkage analysis of chromosome 2q using 16 polymorphic microsatellite markers at an average spacing of one marker every 8.5 cM. RESULTS: Our results provide suggestive evidence for a locus at 2q31 with a maximum multipoint logarithm of the odds score (MLS) of 1.22 which increased to 2.19 in those families concordant for hip-only disease (n = 311). This suggestive linkage was greater in male-hip families (MLS = 1.57, n = 71) than in female-hip families (MLS = 0.71, n = 132). CONCLUSIONS: Chromosome 2q is likely to contain at least one susceptibility locus for OA.

Adult↗
PubMed2000

5'-heterogeneity of glucocorticoid receptor messenger RNA is tissue specific: differential regulation of variant transcripts by early-life events.

Glucocorticoid receptor (GR) gene expression is regulated in a complex tissue-specific manner, notably by early-life environmental events that program tissue GR levels. We have identified and characterized several new rat GR mRNAs. All encode a common protein, but differ in their 5'-leader sequences as a consequence of alternate splicing of, potentially, 11 different exon 1 sequences. Most are located in a 3-kb CpG island, upstream of exon 2, that exhibits substantial promoter activity in transfected cells. Ribonuclease (RNase) protection analysis demonstrated significant levels of six alternate exons 1 in vivo in rat, with differences between liver, hippocampus, and thymus reflecting tissue-specific differences in promoter activity. Two of the alternate exons 1 (exons 1(6) and 1(10)) were expressed in all tissues examined, together present in 77-87% of total GR mRNA. The remaining GR transcripts contained tissue-specific alternate first exons. Importantly, tissue-specific first exon usage was altered by perinatal environmental manipulations. Postnatal handling, which permanently increases GR in the hippocampus, causing attenuation of stress responses, selectively elevated GR mRNA containing the hippocampus-specific exon 1(7). Prenatal glucocorticoid exposure, which increases hepatic GR expression and produces adult hyperglycemia, decreased the proportion of hepatic GR mRNA containing the predominant exon 1(10), suggesting an increase in a minor exon 1 variant. Such tissue specificity of promoter usage allows differential GR regulation and programming.

Alternative Splicing↗
PubMed2000

11 beta-hydroxysteroid dehydrogenase type 1 is a predominant 11 beta-reductase in the intact perfused rat liver.

11 beta-Hydroxysteroid dehydrogenase type 1 (11 beta-HSD-1), a regulator of intrahepatocellular glucocorticoid activity, is bidirectional in homogenates but catalyses 11 beta-reduction (regenerating glucocorticoid) in intact primary hepatocytes in culture. To examine this discrepancy at the whole-organ level, we examined 11 beta-HSD-1 activity in the intact bivascularly perfused rat liver. On a single pass through male rat liver, 44+/-5% of 11-dehydrocorticosterone (11-DHC) recovered was 11 beta-reduced to corticosterone, whereas 10+/-1% of corticosterone was 11 beta-dehydrogenated to 11-DHC. 11 beta-Reduction was less in female liver (21+/-2%, P<0.01) and was significantly greater with perfusion of all substrate via the portal vein (50+/-3%) than via the hepatic artery (30+/-2%, P<0.05). 11 beta-Reductase activity was not saturated by 11-DHC (10(-)(9)-10(-)(6) M). Perfusion with carbenoxolone (CBX, 10(-)(6)-10(-)(3 )M) did not alter 11 beta-reduction of 11-DHC. In contrast, pretreatment with CBX in vivo (10 mg/day) for 7 days inhibited 11 beta-reductase (19+/-4% conversion, P<0.01). Concentrations of 11-DHC in male rat plasma were 44+/-6 nM. Thus 11 beta-HSD-1 is predominantly an 11 beta-reductase in the intact rat liver and is only inhibited by chronic administration of CBX. The substantial concentrations of plasma 11-DHC as substrate suggest that 11 beta-HSD-1 activity and its potential selective inhibition could modify glucocorticoid action in vivo.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1999

Changes in the expression of myometrial ryanodine receptor mRNAs during human pregnancy.

Uterine contraction is triggered by a rise in intracellular free Ca(2+) concentration ([Ca2+]i), and although ryanodine-sensitive Ca(2+) release channels (RyRs) play a key role in the regulation of [Ca(2+)](i) in skeletal and cardiac muscle, much less is known about their role in smooth muscle. In this study, we investigated the expression of RyR mRNAs (ryr1-3) during human pregnancy by examining myometrial samples (n=18) taken, with informed consent and ethical approval, from non-pregnant patients undergoing hysterectomy, and patients undergoing elective caesarean section (at term, prior to or following the onset of labour). Ca(2+) release channel expression was determined both qualitatively and quantitatively, using reverse transcription-polymerase chain reaction (RT-PCR) analysis, RNase protection assays, and in situ mRNA hybridisation. RT-PCR analysis demonstrated that all three ryr genes, as well as the gene encoding the type I inositol 1,4,5-trisphosphate receptor (InsP(3)RI), are expressed in human myometrium. Quantitation by RNase protection assays showed that ryr3 and InsP(3)RI mRNAs are the most abundant, while ryr2 mRNA is barely detectable. In situ mRNA hybridisation confirmed that ryr3 and InsP(3)RI mRNAs are both localised to myometrial smooth muscle cells. The expression of ryr2 and ryr3 mRNA is down-regulated at the end of pregnancy compared to non-pregnant myometrium, indicating that ryanodine-sensitive Ca(2+) release channels are differentially expressed. The relative conservation of ryr1 expression is consistent with a role for Ca(2+) release from ryanodine-sensitive stores in the mechanism of uterine contractility during labour.

Adolescent↗
PubMed1999

Tissue- and temporal-specific regulation of 11beta-hydroxysteroid dehydrogenase type 1 by glucocorticoids in vivo.

11Beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) catalyses the interconversion of active corticosterone and inert 11-dehydrocorticosterone. Short-term glucocorticoid excess upregulates 11beta-HSD-1 in liver and hippocampus leading to suggestions that 11beta-HSD-1 ameliorates the deleterious effects of glucocorticoid excess by its 11beta-dehydrogenase activity. However the predominant activity of 11beta-HSD-1 in vivo is 11beta-reduction, thus generating active glucocorticoid. We have re-examined the time-course of glucocorticoid regulation of 11beta-HSD-1 in the liver, hippocampus and kidney of adult male rats in vivo. Sham operation markedly reduced 11beta-HSD-1 mRNA expression in all tissues, and reduced 11beta-HSD bioactivity in liver and hippocampus when compared to untouched controls. Adrenalectomy reduced 11beta-HSD-1 expression in all tissues in the short-term (7 days), followed by subsequent recovery of enzyme activity by 21 days in liver and hippocampus. Dexamethasone replacement of adrenalectomised rats attenuated the initial decrease in hepatic 11beta-HSD-1 activity, but by 21 days dexamethasone reduced activity compared to control levels. Thus glucocorticoids regulate 11beta-HSD-1 in a complex tissue- and temporal-specific manner. This pattern of regulation suggests glucocorticoids repress 11beta-HSD-1 at least in the liver, a pattern of regulation more consistent with the evidence that 11beta-HSD-1 is an 11beta-reductase in vivo. Operational stress per se down-regulates 11beta-HSD-1 which has implications for interpretation and design of in vivo studies of 11beta-HSD-1.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1999

Brachydactyly type B: linkage to chromosome 9q22 and evidence for genetic heterogeneity.

Brachydactyly type B (BDB), an autosomal dominant disorder, is the most severe of the brachydactylies and is characterized by hypoplasia or absence of the terminal portions of the index to little fingers, usually with absence of the nails. The thumbs may be of normal length but are often flattened and occasionally are bifid. The feet are similarly but less severely affected. We have performed a genomewide linkage analysis of three families with BDB, two English and one Portugese. The two English families show linkage to the same region on chromosome 9 (combined multipoint maximum LOD score 8.69 with marker D9S257). The 16-cM disease interval is defined by recombinations with markers D9S1680 and D9S1786. These two families share an identical disease haplotype over 18 markers, inclusive of D9S278-D9S280. This provides strong evidence that the English families have the same ancestral mutation, which reduces the disease interval to <12.7 cM between markers D9S257 and D9S1851 in chromosome band 9q22. In the Portuguese family, we excluded linkage to this region, a result indicating that BDB is genetically heterogeneous. Reflecting this, there were atypical clinical features in this family, with shortening of the thumbs and absence or hypoplasia of the nails of the thumb and hallux. These results enable a refined classification of BDB and identify a novel locus for digit morphogenesis in 9q22.

Chromosomes, Human, Pair 9↗
PubMed1999

Pregnant rat myometrial cells show heterogeneous ryanodine- and caffeine-sensitive calcium stores.

Intracellular Ca(2+) release channels such as ryanodine receptors play crucial roles in the Ca(2+)-mediated signaling that triggers excitation-contraction coupling in muscles. Although the existence and the role of these channels are well characterized in skeletal and cardiac muscles, their existence in smooth muscles, and more particularly in the myometrium, is very controversial. We have now clearly demonstrated the expression of ryanodine receptor Ca(2+) release channels in rat myometrial smooth muscle, and for the first time, intracellular Ca(2+) concentration experiments with indo 1 on single myometrial cells have revealed the existence of a functional ryanodine- and caffeine-sensitive Ca(2+) release mechanism in 30% of rat myometrial cells. RT-PCR and RNase protection assay on whole myometrial smooth muscle demonstrate the existence of all three ryr mRNAs in the myometrium: ryr3 mRNA is the predominant subtype, with much lower levels of expression for ryr1 and ryr2 mRNAs, suggesting that the ryanodine Ca(2+) release mechanism in rat myometrium is largely encoded by ryr3. Moreover, using intracellular Ca(2+) concentration measurements and RNase protection assays, we have demonstrated that the expression, the percentage of cells responding to ryanodine, and the function of these channels are not modified during pregnancy.

Animals↗
PubMed1999

Interactions between oestradiol and glucocorticoid regulatory effects on liver-specific glucocorticoid-inducible genes: possible evidence for a role of hepatic 11beta-hydroxysteroid dehydrogenase type 1.

In vitro, 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) catalyses the interconversion of active corticosterone and inert 11-dehydrocorticosterone. 11beta-HSD-1 is highly expressed in liver, where the reaction direction is 11beta-reduction, thus potentially increasing intrahepatic active glucocorticoid levels. Inhibition of 11beta-HSD-1 increases insulin sensitivity in humans in vivo suggesting that hepatic 11beta-HSD-1 plays a role in the maintenance or control of key glucocorticoid-regulated metabolic functions. We have selectively repressed hepatic 11beta-HSD-1 in rats by oestradiol administration for 42 days. This nearly completely repressed hepatic 11beta-HSD-1 mRNA expression and enzyme activity and reduced expression of hepatic glucocorticoid-inducible genes including phosphoenolpyruvate carboxykinase (PEPCK), the rate-limiting step in gluconeogenesis. Similar effects were seen after 3 weeks of oestradiol treatment. To examine whether this was due to any direct effect of oestradiol upon PEPCK, the experiment was repeated in adrenalectomised rats+/-glucocorticoid replacement. In adrenalectomised rats, oestradiol did not attenuate hepatic PEPCK, whilst glucocorticoid replacement restored this action. Oestradiol did not alter hepatic metabolism of corticosterone by pathways other than 11beta-HSD-1. These data suggest 11beta-HSD-1 plays an important role in maintaining expression of key glucocorticoid-regulated hepatic transcripts. Enzyme inhibition may provide a useful therapeutic target for manipulating glucose homeostasis.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1999

Observations on the effects of low frequency electromagnetic fields on cellular transcription in Drosophila larvae reared in field-free conditions.

Drosophila larvae reared inside a micro-metal box with an internal field strength 0.004 microT, were treated with a magnetic field of 50 Hz, 8 microT. for 20 min. Control experienced 0.004 microT. Cellular transcript levels were assessed using slot blots and quantified using a Phosphorimager. Blots were hybridised using probes against HSP 70a, Histone 1.9, and Copia. The low frequency EMFs very significantly decreased transcript levels, indicating that experimental responses may be influenced by previous exposure or lack of previous exposure.

Analysis of Variance↗
PubMed1998

Partial cloning and differential expression of ryanodine receptor/calcium-release channel genes in human tissues including the hippocampus and cerebellum.

Cellular Ca2+ signalling is an important factor in the control of neuronal metabolism and electrical activity. Although the roles of Ca2+-release channels are well established for skeletal and cardiac muscle, less is known about their expression and roles in the central nervous system, especially in the human brain. We have isolated partial complementary DNAs derived from the human ryanodine receptor Ca2+-release channel genes (ryr1, ryr2 and ryr3), and examined their expression in the human hippocampus and cerebellum. For comparison, we have included in our analysis an inositol trisphosphate Ca2+-release channel type I complementary RNA probe. All four messenger RNAs show widespread distribution in the human hippocampus, where ryr2 is the most abundant isoform, and all four are expressed in the human cerebellum. However, striking differences were seen between ryr and inositol trisphosphate Ca2+-release channel type I complementary RNA expression in the cerebellum, with inositol trisphosphate Ca2+-release channel type I messenger RNA being largely restricted to, and very highly expressed, in Purkinje cells, whereas ryr1, ryr2 and ryr3 were all expressed predominantly in the granular layer. The widespread expression of ryr isoforms in the human hippocampus and cerebellum suggests that ryanodine receptor proteins may have a central role in Ca2+ signalling and Ca2+ homeostasis in the human central nervous system. These may include roles in fundamental processes like synaptic plasticity. Furthermore, these Ca2+-release channels may be involved in pathogenic processes such as excitotoxicity, where excessive rises in intracellular Ca2+ concentration mediate neuronal cell death.

Aged↗
PubMed1998

11Beta-hydroxysteroid dehydrogenase 1 in adipocytes: expression is differentiation-dependent and hormonally regulated.

11Beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) catalyses the reversible metabolism of physiological glucocorticoids (cortisol, corticosterone) to inactive metabolites (cortisone, 11-dehydrocorticosterone), thus regulating glucocorticoid access to receptors. 11Beta-HSD-1 expression is regulated during development and by hormones in a tissue specific manner. The enzyme is highly expressed in liver, where it may influence glucocorticoid action on fuel metabolism, processes also important in adipose tissue. Here we show that 11beta-HSD-1 is expressed in white adipose tissue, in both the adipocyte and stromal/vascular compartments, and in the adipocyte cell lines 3T3-F442A and 3T3-L1. In these cells, 11beta-HSD-1 expression is induced upon differentiation into adipocytes and is characteristic of a 'late differentiation' gene, with maximal expression 6-8 days after confluence is reached. In intact 3T3-F442A adipocytes the enzyme direction is predominantly 11beta-reduction, activating inert glucocorticoids. The expression of 11beta-HSD-1 mRNA is altered in fully differentiated 3T3-F442A adipocytes treated with insulin, dexamethasone or a combination of the hormones, in an identical manner to glycerol-3-phosphate dehydrogenase (GPDH) mRNA (encoding a key enzyme in triglyceride synthesis and a well-characterised marker of adipocyte differentiation). The demonstration of 11beta-HSD-1 expression in adipocytes and its predominant reductase activity in intact 3T3-F442A adipocytes suggests that 11beta-HSD-1 may play an important role in potentiating glucocorticoid action in these cells. 3T3-F442A and 3T3-L1 represent useful model systems in which to examine the factors which regulate 11beta-HSD-1 gene expression and the role of 11beta-HSD-1 in modulating glucocorticoid action in adipose tissue.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1997

Medical and physiological aspects of the 11beta-hydroxysteroid dehydrogenase system.

11Beta-hydroxysteroid dehydrogenases (11beta-HSD) catalyse the interconversion of active glucocorticoids (cortisol, corticosterone) and their inert 11-keto derivatives (cortisone, 11-dehydrocorticosterone). The type-2 isozyme (11beta-HSD-2) is a high-affinity dehydrogenase that catalyses the rapid inactivation of glucocorticoids, thus ensuring selective access of aldosterone to otherwise non-selective mineralocorticoid receptors in the distal nephron. Mutations of the gene encoding 11beta-HSD-2 are responsible for the syndrome of apparent mineralocorticoid excess, in which cortisol illicitly occupies mineralocorticoid receptors, causing hypertension and hypokalaemia. 11Beta-HSD-2 is also highly expressed in the placenta and mid-gestation fetus, where it may protect developing tissues from the often deleterious actions of glucocorticoids upon fetal growth and organ maturation. 11Beta-HSD-1 is probably an 11beta-reductase in vivo. Its function is obscure, but may amplify glucocorticoid action during the diurnal nadir, drawing upon the substantial circulating levels of 11-keto steroids. Both isozymes are regulated during ontogeny and by a series of hormonal and other factors. 11Beta-HSD provide an important control of glucocorticoid action at a cellular level, and may represent new targets for therapeutic intervention.

11-beta-Hydroxysteroid Dehydrogenases↗