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G J Macdonald

Publications and source records attributed to G J Macdonald.

At least 19 recordsLinked to original sources
PubMed1998

Epitope-specific focusing of the immune response to a minimized human chorionic gonadotropin analog.

Minimized proteins have long been used to elicit an immune response to particular regions of a protein antigen. Most efforts to derive minimized proteins have employed synthetic peptide fragments. This approach works well for linear epitopes but poorly for conformational epitopes. Here we describe a homodimeric human chorionic gonadotropin (hCG) analog that retains the conformation of related parts of hCG and elicits high affinity specific antibodies. This novel immunogen displays the tertiary structure of selected loops of the protein but lacks structures that could elicit potentially undesirable antibodies.

Amino Acid Sequence↗
PubMed1998

Positive association between expression of follicle-stimulating hormone beta and activin betaB-subunit genes in boars.

This study tested our hypothesis that inhibin/activin (I/A) betaB subunit and not follistatin (FS) gene expression relates positively to plasma FSH concentrations in the anterior pituitary gland of boars. Mature crossbred boars (n = 12) were selected for divergence in plasma FSH concentrations, and their anterior pituitary glands were evaluated for expression of the FSHbeta, I/A ssB, FS, calmodulin, and GnRH receptor (GnRH-R) genes by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and/or RNase protection assays (RPAs). Expression of I/A ssB was greater (p < 0. 01) in the six boars with high FSH than in the six with low FSH; expression of the I/A betaB-subunit gene was positively correlated to that of the FSHbeta gene (RT-PCR: r = 0.96; p < 0.01; RPA: r = 0.68; p < 0.05). In contrast, expression of the FS (p > 0.10), GnRH-R (p > 0. 08), and calmodulin (p > 0.10) genes was similar in the two groups of boars. Additionally, expression of the FSHbeta gene was correlated positively with pituitary and plasma FSH concentrations (r = 0.69 and 0.88, respectively; p < 0.05). These results support the hypothesis that activin B is partially responsible for elevated FSH concentrations in boars. Furthermore, the expression difference of the calmodulin gene observed previously between Meishan and White Composite boars represents a breed difference unrelated to FSH.

Activins↗
PubMed1997

Nitric oxide synthase inhibition in a spontaneously hypertensive rat model of diabetic nephropathy.

1. To investigate the role of nitric oxide (NO) in diabetic nephropathy the effect of nitric oxide synthase (NOS) inhibition by NG-nitro-L-arginine methyl ester (L-NAME) was observed in a streptozotocin diabetic spontaneously hypertensive rat (SHR) model. 2. Two groups of SHR (n = 8) with streptozotocin-induced diabetes were studied. One group was given L-NAME 5 mg/kg bodyweight per day in the drinking water for 8 weeks while both groups received daily subcutaneous injections of Ultratard insulin. Creatinine clearance, urinary protein excretion, urinary nitrate concentration and systolic blood pressure were measured at fortnightly intervals. Rats were killed at 8 weeks and plasma angiotensin II (AngII) was measured by radioimmunoassay. 3. Renal function (endogenous creatinine clearance) remained stable in both groups. In the L-NAME group, however, there was a progressive increase in proteinuria that was highly significant at 6 weeks (22.1 +/- 2.9 compared with 6.5 +/- 0.7 mg/ 24 h per 100 g in control SHR diabetic rats P < 0.001). 4. Systolic blood pressure was significantly elevated in the L-NAME group throughout the study compared with the control group. 5. Plasma AngII was significantly elevated in the L-NAME group compared with controls (42.8 +/- 10.3 vs 15.1 +/- 1.9 pmol/L, respectively; P < 0.05). 6. Activation of the renin-angiotensin system may account, at least in part, for the resulting vasoconstrictor activity with chronic nitric oxide depletion.

Animals↗
PubMed1996

Apoptotic cell death in the rat adrenal gland: an in vivo and in vitro investigation.

Adrenocortical cell apoptosis was studied by using an established in vivo model, the hypophysectomized rat, and an in vitro model, viz., rat adrenal glands in short-term organ culture. In vivo, apoptosis (biochemical autoradiographic analysis of internucleosomal DNA cleavage) was weak and not apparent until 12-24 h after hypophysectomy. In situ histochemical localization of 3'-end DNA strand breaks revealed that apoptosis in vivo occurred nearly exclusively in subpopulations of zona reticularis cells. Adrenocorticotropic hormone (ACTH) maintenance completely blocked these indices of apoptosis. By contrast, apoptosis (DNA fragmentation) in cultured rat adrenal glands without ACTH was extensive and relatively rapid, being apparent after 1 h and increasing with the duration of incubation. ACTH attenuated (by 44%) but did not completely block apoptosis in vitro. Thus, ACTH appears to be the sole pituitary hormone that forestalls apoptosis of terminally differentiated adrenocortical (zona reticularis) cells. However, the discrepancy between in vitro and in vivo models in terms of the magnitude and rate of DNA fragmentation suggests that, in vivo, other factors finely regulate the magnitude of adrenocortical apoptotic cell death.

Adrenal Cortex↗
PubMed1996

A study of angiotensin II receptors after chronic inhibition of nitric oxide synthase in the spontaneously hypertensive rat.

1. Nitric oxide (NO) synthase inhibition, induces a sustained increase in blood pressure and amplifies the pressor response to infused angiotensin II (AngII). This study was designed to investigate the contribution of AngII receptors in the elevated blood pressure and enhanced pressor response to AngII in the spontaneously hypertensive rat (SHR) chronically treated with N(G)-nitro-L-arginine-methyl ester (L-NAME). 2. Two groups of 13 week old female SHR were housed four to a box. Group I rats received L-NAME for 7 days (2.5 mg/kg per day) in their drinking water. Group II rats received water only. Blood pressure was monitored daily by tail-cuff plethysmography. Plasma AngII was measured by radioimmunoassay. Aortic and uterine receptor binding was determined by saturation analysis using [125I]-Sar8, Ile1)AngII. Data was analysed using the computer program LIGAND. 3. Mean systolic blood pressure was significantly elevated in rats treated with L-NAME compared with the control group. Plasma AngII concentration was slightly decreased in rats treated L-NAME compared with control. Densities of both aortic and uterine AngII receptors increased significantly following NO synthase inhibition. Receptor affinity in the aorta decreased in the L-NAME group compared with control. However, uterine AngII receptor affinity was unchanged. 4. We conclude that the increased blood pressure and enhanced pressor responsiveness that occurs with chronic inhibition of NO synthesis may result partly from increased vascular AngII receptor expression.

Angiotensin II↗
PubMed1996

Angiotensin II: a humoral mediator for the gastric sodium monitor.

Natriuresis in direct response to a gastric sodium stimulus (upper-gut sodium monitor) has paradoxically only been demonstrated in humans and animals on a low-sodium diet preceding each study. It is possible that the low-sodium diet itself induces or suppresses systems that mediate or oppose the ensuing natriuresis. In this study, we sought to determine whether a system activated by this diet, the renin-angiotensin system, mediates the natriuretic response. Specifically, we sought to show whether changes in the circulating concentration of angiotensin II (ANG II) may mediate the renal response to stimulation of the gastric sodium monitor. Male New Zealand White rabbits were randomly assigned to low- (0.008%) or normal (2.2%) sodium diets. After 1 wk on the experimental diet, they received a sodium load intragastrically or intravenously, and plasma ANG II was measured at 0, 5, 10, 30, 60, and 120 min. Urine was collected for 4 h after the sodium load, and plasma sodium was measured at 0, 2, and 4 h. Urinary sodium excretion was greater in the 4 h after gastric than after intravenous sodium administration (P < 0.025) in the rabbits on the low-sodium diet. No significant difference was noted in the rabbits on the normal sodium. In rabbits on the low-sodium diet, there was an immediate and significant decline in plasma ANG II after sodium was administered both intragastrically (P < 0.025) and intravenously (P < 0.05). This decrease was greater after intragastric than intravenous sodium (P < 0.0025), and the difference was still evident at 120 min (P < 0.05). No significant difference in plasma ANG II was found in the normal diet group. We conclude, therefore, that a prolonged decrease in ANG II concentration may play a role in mediating the natriuretic response to the gastric sodium monitor.

Angiotensin II↗
PubMed1996

Overexpression of beta-subunit of thyroid-stimulating hormone in Meishan swine identified by differential display.

Anterior pituitary gland RNA isolated from Meishan (MS) and White Composite (WC) sexually mature boars was compared by a newly developed method of differential expression cloning called differential display. A number of cDNAs were identified that differed distinctly in expression between these breeds. The gene for beta-subunit of thyroid-stimulating hormone (TSH-beta) was initially identified as more highly expressed in MS than in WC boars. Subsequently, RNase protection assays and TSH RIA were used to quantify the magnitude of differences in transcription and translation in both males and females. Mature MS pigs had threefold greater expression of TSH-beta (P < .05) and greater plasma TSH concentrations than mature WC pigs (P < .01). The cDNA cloning and sequence analyses indicate identity in TSH-beta mRNA between MS and WC males. Elevated plasma TSH concentration may contribute MS pigs reaching sexual maturity earlier than WC or other European breeds. The method used in this study provides a useful molecular tool 1) to detect differentially expressed genes, 2) to study genetic variation that occurs at the transcription level between individuals, populations, or breeds; and additionally, 3) to identify candidate genes that control economically important quantitative traits in livestock.

Animals↗
PubMed1995

The groove between the alpha- and beta-subunits of hormones with lutropin (LH) activity appears to contact the LH receptor, and its conformation is changed during hormone binding.

Gonadotropins are heterodimeric glycoprotein hormones that control vertebrate fertility through their actions on gonadal lutropin (luteinizing hormone, LH) and follitropin (follicle-stimulating hormone, FSH) receptors. The beta-subunits of these hormones control receptor binding specificity; however, the region of the beta-subunit that contacts the receptor has not been identified. By a process of elimination we show this contact to be the portions of beta-subunit loops one and three found in a hormone groove created by the juxtaposition of the alpha- and beta-subunits. Most other regions of the beta-subunit can be recognized by antibodies that bind to human chorionic hormone (hCG)-receptor complexes or replaced without disrupting hormone function. Using a series of bovine LH/hCG and human FSH/hCG beta-subunit chimeras we identified key hCG beta-subunit residues in the epitopes of two antibodies that bind to hCG-receptor complexes. These epitopes include the surfaces of beta-subunit loops one and three near residue 74 on the outside of the hormone groove and parts of the C-terminal end of the "seat belt" that holds the two subunits together. The antibody that recognized residue 74 bound to receptor complexes containing most mammalian lutropins better than to the free hormones, an indication that the outside surface of the beta-subunit groove is altered during hormone binding. This region of the beta-subunit is furthest from the alpha-subunit and is recognized equally well in the free beta-subunit and in the heterodimer. Thus, the receptor associated increase in antibody binding appears due to an interaction of this portion of the beta-subunit with the receptor and not to an effect of the receptor on the relative positions of the alpha- and beta-subunits. Unlike most previous studies designed to identify portions of the beta-subunit likely to contact the LH receptor, this indirect approach provides data that are more easily interpreted because it does not rely on the use of mutations that disrupt hormone function. The approach described here should be valuable for studying the receptor interactions of other complex ligands.

Amino Acid Sequence↗
PubMed1995

Birth dates and survival after axotomy of neurochemically defined subsets of trigeminal ganglion cells.

Trigeminal (V) ganglion cells with different neurochemical phenotypes or different birth dates are affected differently by neonatal axonal transection. The aim of the present study was to determine if V ganglion cell birth date and neurochemical phenotype were correlated and if these two variables could be related to responses to neonatal axonal transection. Immunocytochemistry, histochemistry, and [3H]thymidine labelling were used to determine the birth dates of V ganglion cells recognized by antibodies directed against neurofilament protein (NF), calcitonin gene-related peptide (CGRP), and substance P (SP) and those that bound the lectin Bandierea simplicifolia-I (BS-I). All V ganglion cells were born between embryonic days (E-) 9.5 and 14.5. All ganglion cells were born between E-9.5 and E-14.5. In a normalized population (percentages normalized to equal 100%), over 90% of NF-positive V ganglion cells were born between E-10.5 and E-12.5. The majority of CGRP-positive and SP-positive ganglion cells (> 90%) were generated from E-13.5 to E-14.5 and E-12.5 through E-14.5, respectively. Almost 85% of BS-I-positive ganglion cells were generated on E-12.5 through E-14.5. Previous results and additional data from this study indicated that NF- and BS-I-positive ganglion cells are proportionally more likely to be lost after neonatal axotomy and that SP-positive cells are more likely to remain. The percentage of CGRP-positive cells in the V ganglion was not significantly altered by neonatal infraorbital nerve transection. Overall, these findings do not indicate a strong relationship between cell birth date and the probability of survival after neonatal axonal damage for all V ganglion cell phenotypes.

Animals↗
PubMed1995

Effects of endopeptidase 24.11 inhibition on plasma and tissue concentrations of vasoactive intestinal peptide.

1. In this study, we sought to determine the effect of endopeptidase 24.11 inhibition on the rate of metabolism of vasoactive intestinal peptide. The effect of such inhibition on the concentration of vasoactive intestinal peptide in two tissues was also investigated. 2. Male Sprague-Dawley rats were given the endopeptidase 24.11 blocker UK77,568 (10 mg/kg) or vehicle as a single intravenous injection or as a daily injection for 4 days. Two hours after the final or single injection, the rats were anaesthetized and blood was sampled to determine plasma concentrations of vasoactive intestinal peptide and angiotensin II. The hearts and kidneys were harvested and snap-frozen in liquid nitrogen. The plasma and tissue concentrations of vasoactive intestinal peptide and the plasma concentration of angiotensin II were determined by radioimmunoassay. In a separate group of experiments, male Sprague-Dawley rats were anaesthetized and carotid and jugular catheters were inserted. One hour after intravenous administration of UK77,568 or vehicle, an infusion of vasoactive intestinal peptide (10 pmol min-1 kg-1) was commenced via the jugular catheter. Blood was sampled to determine the vasoactive intestinal peptide concentration 1 h after commencing the vasoactive intestinal peptide infusion to calculate the metabolic clearance rate. 3. Plasma vasoactive intestinal peptide increased after acute (P < 0.05) but not chronic administration of UK77,568, while the concentration of vasoactive intestinal peptide in the heart increased after chronic administration (P < 0.0005). The concentration of vasoactive intestinal peptide in the kidney was unchanged after both acute and chronic endopeptidase 24.11 blockade. Plasma angiotensin II decreased significantly in the chronic group (P<0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin II↗
PubMed1995

Modulation of cholesteryl ester hydrolase messenger ribonucleic acid levels, protein levels, and activity in the rat corpus luteum.

Previous studies have shown that the induction of functional luteolysis (loss of progesterone production) with either prostaglandin F2 alpha (PGF2 alpha) treatment or hypophysectomy (APX) diminished neutral cholesteryl ester hydrolase (CEH) activity in the corpus luteum (CL) and that prolactin (PRL) replacement of APX animals prevented luteolysis and maintained CEH activity at control levels. More recent studies have shown that CEH is the same protein as hormone-sensitive lipase (HSL) and that CEH/HSL activity may be regulated by phosphorylation. However, the possibility that CEH/HSL activity may be under transcriptional and/or translation control has not been excluded. Therefore, in the present study we examined whether PGF2 alpha treatment, APX, or inhibition of PRL secretion by bromocryptine (BrC) treatment modulated CEH/HSL mRNA and/or protein levels in a coordinate fashion with CEH activity. Furthermore, we examined whether CEH/HSL mRNA and/or protein levels changed after luteinization of the ovary and after natural functional regression. PGF2 alpha treatment and APX significantly reduced CEH activity; and PGF2 alpha treatment, APX, and BrC treatment significantly reduced CEH/HSL protein and mRNA levels. PRL replacement after APX substantially blocked the reductions in CEH activity, CEH/HSL protein, and CEH/HSL mRNA levels. PRL replacement during BrC treatment significantly inhibited the reductions in CEH/HSL protein and mRNA levels. CEH/HSL mRNA levels increased twofold after luteinization. Whereas CEH/HSL mRNA levels remained elevated after natural luteal regression, CEH/HSL protein significantly decreased. In summary, the luteolytic actions of PGF2 alpha, APX, and BrC resulted in coordinate reductions in luteal CEH activity, protein levels, and mRNA levels; PRL replacement significantly reversed the luteolytic effects of APX and BrC; natural luteal regression resulted in a reduction in CEH/HSL protein without a concomitant reduction in CEH/HSL mRNA. These results suggest that ovarian CEH activity is controlled at the level of both transcription and translation, and that PRL is important for continued CEH/HSL mRNA transcription in the CL.

Animals↗
PubMed1995

Vasoactive intestinal peptide regulates angiotensin II catabolism in the rabbit.

Although vasoactive intestinal peptide (VIP) is natriuretic it stimulates renin and aldosterone secretion. Therefore, to effect a natriuresis, VIP may need to modulate the sodium conserving actions of the renin angiotensin system (RAS) by another means. One possibility is that it alters the rate of disappearance from the circulation of one or more components of the RAS. We sought to determine whether VIP regulates the rate of catabolism of angiotensin II (Ang II). Steady state metabolic clearance studies of Ang II were undertaken with and without simultaneous VIP infusion. These studies were performed in rabbits on low, normal and high sodium diets, as dietary sodium has been shown to affect the metabolism of both VIP and Ang II. The effects of VIP on plasma Ang II concentration and secretion were also studied. VIP decreased Ang II catabolism in rabbits on low (P < 0.05) and normal sodium diets (P < 0.05). Plasma levels of Ang II increased significantly in response to VIP in rabbits on these diets (low, P < 0.04; normal, P < 0.05). In contrast, in rabbits on a high sodium diet VIP increased the rate of catabolism of Ang II (P < 0.001). Thus we conclude that the effect of VIP on sodium excretion may be modulated by its effects on Ang II metabolism. The decrease in Ang II catabolism seen in rabbits on low and normal sodium diets may prevent or ameliorate any natriuresis while the more rapid degradation of Ang II which occurs in dietary sodium excess may enhance the natriuretic effect of VIP.

Angiotensin II↗
PubMed1994

Accuracy of single concentration estimations of platelet angiotensin II receptor number. Its usefulness in screening for pregnancy-induced hypertension.

Platelet angiotensin II (AngII) receptor number has been suggested as a screening test for pregnancy-induced hypertension. However, markedly different false-positive rates have been reported, perhaps the result of differing methods used. We sought therefore to compare the two methods. Platelet AngII receptor number was determined by saturation analysis with computerized curve fitting and specific binding at a single radioligand concentration. The two methods were compared by correlation and by plotting their differences v their means, to determine their limits of agreement. There were significant correlations between the value obtained by saturation analysis and each of the three single ligand concentrations studied (1 nmol/L, P < .001; 500 pmol/L, P < .001; and 250 pmol/L, P < .01). However, for none of the three did the regression line approach the line of equality. Assessment of agreement by comparing differences and means for each subject showed increasing scatter with increasing receptor number and 95% confidence intervals too large to be clinically relevant. We conclude that the receptor number estimated from specific binding at one ligand concentration differs significantly from that obtained by saturation analysis. The limits of agreement of the two methods are wide and we urge caution in the use of single ligand concentration methods for estimating binding site densities.

Adult↗
PubMed1994

Differential regulation of uterine and glomerular angiotensin II receptors in normal and hypertensive pregnancy in the rat.

1. In the non-pregnant state uterine and glomerular angiotensin II (AII) receptors have been shown to regulate in a similar fashion. This study sought to determine whether such parallel regulation occurred during pregnancy. We also investigated the role of plasma AII in these changes. 2. These studies were performed in Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). The SHR has increased receptor number, increased sensitivity to infused AII and decreased plasma volume compared to the WKY. These features are also seen in pregnancy induced hypertension (PIH). 3. Age matched female WKY and SHR were exposed to an appropriate breeder and sacrificed on day 14 of gestation. Plasma AII was measured by radio-immunoassay. Uterine and glomerular receptor binding was determined by saturation analysis using 125I(Sar1,Ileu8)AII. Data were analysed using the program LIGAND. 4. Uterine and glomerular AII receptors showed different patterns of regulation during pregnancy. The uterine AII receptor affinity decreased significantly in both strains at 14 days of gestation and receptor number also decreased significantly in WKY. In contrast, glomerular AII receptor affinity did not change significantly in either strain. Receptor numbers in the glomeruli increased significantly compared to their respective non-pregnant controls. 5. We conclude that the uterine and glomerular AII receptors do not regulate in a parallel fashion in pregnancy. Plasma AII concentration does not appear to be involved in the regulation of either uterine and glomerular receptor expression in pregnancy.

Angiotensin II↗
PubMed1994

Effect of neutral endopeptidase on plasma and tissue concentrations of vaso-active intestinal peptide.

1. This study sought to determine if neutral endopeptidase metabolizes vaso-active intestinal peptide (VIP) and whether changes in the activity of this enzyme might explain the change in VIP metabolism which follows gastric sodium loading. The study also investigated whether prolonged inhibition of neutral endopeptidase was associated with inhibition of further secretion of VIP. 2. Male Sprague-Dawley rats were given the neutral endopeptidase inhibitor UK77,568 (10 mg/kg), or vehicle, tail vein injected, once or daily for 4 days. Two hours after injection the rats were anaesthetized, blood sampled to determine plasma concentrations of VIP and the hearts were harvested. Plasma and tissue concentrations of VIP were determined by radio-immunoassay. 3. Plasma VIP increased in response to UK77,568 at day 1 (P < 0.0005) but did not differ from control at day 4. The concentration of VIP in the heart did not increase at day 1 but had increased significantly at day 4 (P < 0.01). 4. It was concluded that neutral endopeptidase may metabolize VIP. Further, the increased concentration of VIP in the heart and the return of its plasma concentration to control levels at day 4 may be consistent with suppression of continued VIP secretion.

Animals↗