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Biomedical subjects

Peter J Fuller

Publications and source records attributed to Peter J Fuller.

29 records · Page 2Linked to original sources
PubMed2003

Interdomain interactions in the mineralocorticoid receptor.

The potential for interaction between the N-terminal domain and the C-terminal region (hinge and ligand-binding domain) of the mineralocorticoid receptor (MR) was examined using the mammalian-2-hybrid assay. The MR C-terminal region was fused to the GAL4 DNA-binding domain (GAL4-MRC). To examine if the AF-2 is involved in the interaction, as has been reported for other steroid hormone receptors, it was inactivated by point mutation (E962A). The N-terminal domain was fused to the VP16 transactivation domain (VP16-MRNT). In the mammalian-2-hybrid assay both GAL4-MRC and GAL4-MRC(E962A) interact with VP16-MRNT in an aldosterone-dependent manner. The GAL4-MRC(E962A) construct was used in subsequent experiments to examine the AF-2-independent N/C-interaction. The MR antagonist spironolactone inhibits the aldosterone-mediated association of the two domains. GAL4-MRC(E962A) interacts weakly with the GR or AR N-terminal domains in the presence of aldosterone. No dimerization between GAL4-MRC(E962A) and VP16-MRC is observed. Interestingly, cortisol produces a much weaker N/C-interaction than aldosterone, and it is possible that the N/C-interaction may contribute to observed functional differences in the MR bound to the two ligands.

Aldosterone↗
PubMed2003

Dissecting mineralocorticoid receptor structure and function.

The molecular mechanisms by which aldosterone regulates epithelial sodium transport in the distal colon and the distal nephron remain to be fully elucidated. Aldosterone acts via the mineralocorticoid receptor (MR) to induce the expression of genes whose products are involved in sodium transport. The structural basis of MR interactions with aldosterone has been examined by creating chimeras of the MR and the closely related glucocorticoid receptor; we have exploited differences in ligand-binding specificity to determine the region(s) of the MR that confer aldosterone-binding specificity. These findings have been related to a three-dimensional model of the MR based on the crystal structure of the progesterone receptor. These studies have been extended to include the characterisation of interactions between the N- and C-termini of the MR. We have characterised six genes that are regulated in vivo in the distal colon and/or kidney of the rat that are directly and acutely regulated by aldosterone administration: the three subunits of the epithelial sodium channel, serum and glucocorticoid-induced kinase, channel-inducing factor and K-ras2A. These studies provide insights into the molecular pathways that mediate aldosterone-induced amiloride-sensitive epithelial sodium transport.

Animals↗
PubMed2002

Molecular pathogenesis of granulosa cell tumours.

Granulosa cell tumours (GCT) of the ovary arise from granulosa cells of the ovary on morphological, biochemical and molecular criteria. In order to understand the molecular pathogenesis of these tumours better we have sought to define their molecular phenotype, to identify activating mutations of the FSH-signalling pathway, to characterise their estrogen receptor expression and to explore the hypothesis that GCT may be resistant to inhibin. The pattern of gene expression observed in GCT suggests a phenotype which is similar to that of late preovulatory granulosa cells which would be consistent with activation of the FSH receptor signalling pathway, however, there is no evidence for activating mutations of either the FSH receptor or the associated trimeric G-proteins. Estrogen receptor beta is abundantly expressed in GCT. The various subunits and isoforms of the activin-inhibin receptor are expressed in GCT. These observations provide a basis for future studies of GCT, including further characterisation of signalling pathways known to be important in the regulation of granulosa cell growth and differentiation.

Activin Receptors↗
PubMed2002

Influence of diet complexity on intestinal adaptation following massive small bowel resection in a preclinical model.

AIMS: To investigate the effect of dietary complexity on intestinal adaptation using a preclinical model. METHODS: Four-week-old piglets underwent a 75% proximal small bowel resection or transection operation (control). Post-operatively, animals received either pig chow (n = 15), polymeric formula (n = 9), polymeric formula plus fiber (n = 6), or elemental formula (n = 7). RESULTS: The weight gain of all groups was reduced compared with controls that were fed the same diet. Animals that had a resection, which were fed elemental formula, had significantly reduced weight gain compared with the other groups (4.7 4.2 vs 30.7 7.1 kg chow and 11.5 1.3 kg polymeric formula). Villus height was increased in the jejunum, ileum and terminal ileum of resected animals compared with controls in animals fed with pig chow, polymeric formula and elemental formula. The animals that had a resection had a significant reduction in the transepithelial conductance (10.4 5.5 vs 25.4 6.5 mS/cm2) and 51Chromium-EDTA flux (2.8 1.9 vs 4.8 4.9 microL/h per cm2) compared with the controls. CONCLUSIONS: A complex diet was found to be superior to an elemental diet in terms of the morphological and functional features of adaptation following massive small bowel resection.

Adaptation, Physiological↗
PubMed2002

Estrogen receptor-alpha and -beta expression in microvascular endothelial cells and smooth muscle cells of myometrium and leiomyoma.

The two estrogen receptors, ERalpha and ERbeta, are likely to have roles in the pathophysiology of fibroid development. They have been detected in myometrial and leiomyoma (fibroid) tissue, but the cell types expressing ERalpha and ERbeta have not been determined. ERs have also been detected in human endothelial cells. The aims of the present study were to determine whether pure populations of myometrial microvascular endothelial cells (MEC) express ERalpha and ERbeta, to compare MEC ERalpha/ERbeta expression with that of pure populations of myometrial smooth muscle cells (SMC) and to determine if ERalpha/ERbeta are differentially expressed in MEC and SMC of myometrium and fibroids from nine paired samples. Using RT-PCR (for ERalpha and ERbeta) and Western blotting (for ERalpha only), we demonstrated that all cultures of early passage myometrial and fibroid SMC (>99% pure) expressed ERalpha but not ERbeta, while myometrial and fibroid MEC (>99% CD31+) constitutively expressed ERbeta. However, both myometrial and fibroid MEC showed variable expression of ERalpha, with approximately 60% of MEC samples expressing ERalpha. While the majority (6/9) of MEC from myometrial and fibroid pairs demonstrated the same pattern of ERalpha expression, 3/9 pairs showed discordant ERalpha expression. These results show that ERalpha and ERbeta are differentially expressed in SMC and MEC of human myometrium and fibroids. Since ERalpha and ERbeta mediate opposing transcriptional activities, any effect of estrogen on the growth and development of fibroids is likely to be complex and may involve both SMC and MEC.

Adult↗
PubMed2002

17Beta-estradiol up-regulates vascular endothelial growth factor receptor-2 expression in human myometrial microvascular endothelial cells: role of estrogen receptor-alpha and -beta.

Estrogen has a cardiovascular protective role in women due in part to its effect on the vasculature. The roles of the two estrogen receptors (ERs), ERalpha and ERbeta, in the vascular actions of estrogen are unclear, as are effects of estrogen on microvascular endothelial cells (MEC) derived from sex steroid-responsive tissues. The present study demonstrates that 17beta-estradiol, but not progesterone, increases vascular endothelial growth factor (VEGF) receptor (VEGFR) expression on human myometrial MEC measured using biotin-recombinant human (rh) VEGF(165) and flow cytometry. This response occurred in a time- and dose-dependent manner, with significantly increased rhVEGF(165) binding at 3 h and maximal responses between 0.1 and 10 nmol/liter 17beta-estradiol, which was blocked by the antiestrogen ICI 182,780. Approximately 60% of samples demonstrated this response to 17beta-estradiol. All samples of myometrial MEC expressed both ERbeta mRNA and protein demonstrated by semiquantitative RT-PCR and Western blotting. However, ERalpha mRNA and protein were expressed in only 13 of 21 MEC samples. There was a significant association between ERalpha expression in myometrial MEC and their ability to respond to 17beta-estradiol by increasing rhVEGF(165) binding. 17beta-estradiol increased VEGFR-2 expression in ERalpha-expressing MEC isolates, which also demonstrated increased rhVEGF(165) binding, but failed to have these effects on ERalpha negative samples. Similarly, 17beta-estradiol augmented VEGF-induced MEC proliferation in ERalpha-expressing MEC samples, which was blocked by ICI 182,780. These observations suggest that 17beta-estradiol increases VEGFR-2 expression on human myometrial MEC promoting endothelial cell proliferation, an effect that varies between subjects and appears to be mediated primarily by ERalpha.

Adult↗
PubMed2002

Sertoli-Leydig cell tumor of the ovary, a rare cause of precocious puberty in a 12-month-old infant.

We report a 12-month-old infant who presented with a 4-month history of isosexual precocious puberty secondary to an estrogenizing Sertoli-Leydig cell tumor of the ovary. Total serum immunoreactive inhibin and subunits A and B were markedly elevated before surgical resection and subsequently decreased 7 wk later into the normal prepubertal range. Twenty weeks following surgical removal, the patient presented again with central precocious puberty; inhibin B levels were raised on this occasion, a luteinizing releasing hormone stimulation test confirmed central precocious puberty. This is the youngest reported occurrence of this rare sex cord stromal neoplasm. The prognosis of this extremely rare tumor presenting at this early juvenile stage is uncertain. This report illustrates the usefulness of serum inhibin as a tumor marker during therapeutic suppression with leuprorelin acetate for central precocious puberty. Analysis of genomic and tumor DNA revealed a normal nucleotide sequence for the LH receptor and the Galpha(s) gene. To understand the molecular pathogenesis of this tumor we analyzed mRNA levels for the inhibin A and B subunits, FSH receptor, LH receptor aromatase, steroidogenic factor-1 and the ER beta genes. Molecular characterization reveals the presence of genes specific for granulosa and Leydig cells; the relative expression of these genes, in addition to its histologic characteristics, suggests that this tumor may result from a dysdifferentiation of a primordial follicle.

Androstenedione↗
PubMed2002

Inhibin-activin receptor subunit gene expression in ovarian tumors.

Granulosa cell tumors of the ovary (GCT) express the inhibin subunit genes and secrete dimeric inhibin. Transgenic mice null for the alpha-inhibin gene develop GCT. It has been suggested that this apparent contradiction may be reconciled if the human GCT are resistant to the tumor-suppressive effects of inhibin. Inhibin receptors have recently been characterized as consisting of either betaglycan or p120 in association with the type II or type I activin receptor subunits (ActR), respectively. To test the hypothesis that GCT may exhibit loss of inhibin receptor expression we have examined the expression of the receptor subunits in a cohort of GCT and in mucinous and serous cystadenocarcinomas and normal ovary. Expression was determined by RT-PCR using gene-specific primers and probes combined with Southern blot analysis of the PCR products. The ActRI subunits and ActRIIA exhibited widespread albeit variable expression across tissues, with the highest levels in the serous tumors. ActRIIB expression was relatively low in the mucinous tumors and high in the GCT. Betaglycan expression was abundant, widespread, and variable across all tissues; highest mean levels occurred in the GCT and normal ovary. p120 expression was low or absent in all tissues except the GCT. Within the GCT there was parallel expression of the ActR subunits, betaglycan and p120; the levels, however, varied considerably between tumors. Expression of betaglycan and p120 in most GCT argues against the hypothesis, but does not exclude the possibility that low or absent expression of p120 might be significant in a subset of these tumors.

Activin Receptors↗