Biomedical subjects

C G Prosser

Publications and source records attributed to C G Prosser.

At least 37 records · Page 2Linked to original sources

PubMed1996

Influence of insulin-like growth factor-binding protein-2 on plasma clearance and transfer of insulin-like growth factors-I and -II from plasma into mammary-derived lymph and milk of goats.

Plasma clearance of insulin-like growth factors-I and -II (IGF-I and -II) and insulin-like growth factor-binding protein-2 (IGFBP-2) from lactating goats (n = 4) was determined following a single intravenous injection of the corresponding 125I-labelled human protein. Transfer of these proteins out of the vascular space was monitored by their subsequent appearance in mammary-derived lymph and milk. Clearance of 125I-IGFBP-2 from circulation was 0.37 +/- 0.06 ml/min/kg, which is markedly greater than that of 125I-IGF-I or -II (0.11 +/- 0.01 and 0.12 +/- 0.01 ml/min/kg respectively). This was also reflected in longer elimination half-lives for IGF-I (353 +/- 6 min) and -II (254 +/- 8 min) compared with IGFBP-2 (110 +/- 9 min). Three hours after injection of the 125I-labelled protein, the plasma:lymph ratio of trichloroacetic acid-precipitable radioactivity was 1.54 +/- 0.04, 3.3 +/- 0.6 and 4.1 +/- 0.4 for IGFBP-2, IGF-I and -II respectively. The form of 125I-IGFBP-2 in lymph was not different from that of plasma. Elevation of plasma concentrations of IGFBP-2 by its intravenous infusion significantly decreased plasma half-life of both IGF-I and -II (251 +/- 8 and 198 +/- 7 min respectively). Although the amount and rate of transfer of IGF into mammary-derived lymph was decreased slightly by IGFBP-2, concentrations eventually obtained were not different from control. However, secretion of IGFs into milk was significantly reduced by IGFBP-2, particularly in the case of IGF-I. These results are consistent with the ability of all three compounds to cross the vascular endothelium intact and of IGFBP-2 to decrease the uptake of IGF by mammary epithelium and subsequent secretion into milk. IGFBP-2 may well have acted to target plasma IGF towards non-mammary tissues, thus explaining the more rapid plasma clearance of IGFs in the presence of elevated IGFBP-2.

Animals↗

PubMed1996

Regulation of blood flow in the mammary microvasculature.

Although milk yield of cows and goats is known to be closely related to the total flow of blood through the udder, a number of studies suggest that milk yield can vary independently. No studies have attempted to measure the proportion of total flow that is nutritive. Within the mammary gland, capillary networks form a basket-like architecture surrounding each alveolus. Notably, flow in individual capillaries is not constant and varies among capillaries. Capillary flow (measured by intravital microscopy) was decreased by oxytocin, which generally increased total flow in the mammary artery, suggesting that the proportion of total flow that is nutritive can vary. In addition to classic metabolic regulators (e.g., carbon dioxide and oxygen) of tissue blood flow, the mammary gland produces a number of vasodilatory compounds, including parathyroid hormone-related protein, insulin-like growth factor-I, prostacyclin, nitric oxide, and endothelin. All of these compounds have been shown to alter mammary blood flow. Mammary tissue also contains kallikrein and angiotensin-converting enzyme, which convert circulating kinins and angiotensin, respectively, into potent vasoactive compounds. A number of these compounds are produced by epithelial cells themselves, providing a mechanism for the functioning epithelium to control its own blood supply and, hence, nutrient flow for milk synthesis. In this review, we examine the nature of the mammary microcirculation, its behavior under different conditions, and some of the regulatory features of the mammary microvasculature.

Animals↗

PubMed1996

Local secretion of nitric oxide and the control of mammary blood flow.

Our objective was to test the hypothesis that local production of the vasorelaxant nitric oxide could regulate mammary blood flow. In four lactating Saanen goats, the response of mammary blood flow to intraarterial infusion of the nitric oxide donor diethylamine NONOate and the inhibitor of nitric oxide synthesis N omega-nitro-arginine was measured. Diethylamine NONOate induced a rapid and sustained increase of mammary blood flow in the infused gland only, suggesting a direct effect on vasculature of the mammary gland. In contrast, infusion of N omega-nitro-arginine decreased mammary blood flow by up to 35%, and the coinfusion of arginine, the nitric oxide precursor, with N omega-nitro-arginine markedly reduced its ability to decrease mammary blood flow. The distribution of nitric oxide synthase was investigated in cryosections of caprine and bovine mammary tissue by histochemical staining for NADPH-diaphorase activity and by immunocytochemistry using specific antibodies against two nitric oxide synthase isoforms. Both techniques revealed nitric oxide synthase in the vascular endothelium and secretory epithelium of the two species. Only antibodies against nitric oxide synthase-III showed specific staining. These results suggest that the mammary gland produces and responds to nitric oxide and, further, raise the possibility that the epithelium may control its own blood supply by secreting nitric oxide.

Animals↗

PubMed1995

Metabolic response of sheep skin to a chronic infusion of a variant of insulin-like growth factor I.

The effects of a chronic (21-day) skin infusion of a variant of insulin-like growth factor I (IGF-I) (long-Arg3-IGF-I; LR3IGF-I) on short-term (48 h) responses of skin metabolism and 21-day plasma hormone concentration, wool-follicle characteristics and wool production were investigated in well-fed castrated Romney sheep. A bilateral arteriovenous preparation was used to infuse LR3IGF-I continuously into the skin on one abdominal flank and saline into the other abdominal flank of six sheep; a further six sheep had one flank infused with saline (controls). LR3IGF-I caused an initial (4-24 h) reduction in the plasma concentrations of amino acids, especially tyrosine, valine and lysine, and, after 24 h, significant (P < 0.05) reductions in blood oxygen and plasma glucose concentrations. After 4 h of LR3IGF-I infusion, there was a significant increase in blood flow (P < 0.05) and oxygen uptake (P < 0.05), and net uptake of amino acids [which was significant (P < 0.05) for valine and phenylalanine] by the LR3IGF-I-infused skin was increased. Total uptake of phenylalanine for skin protein synthesis, measured using [3H]phenylalanine uptake, was also significantly increased after 4 and 24 h of infusion. After 48 h of infusion all LR3IGF-I-dependent measurements of metabolic parameters had fallen to preinfusion values. By day 7 of the 21-day infusion there was a significant (P < 0.05) decrease in circulating endogenous IGF-I in plasma of treated sheep compared with that of control sheep, followed by a significant (P < 0.05) increase between day 7 and 21. Plasma insulin levels followed a similar pattern. There was no change at any stage of infusion in IGF-binding proteins in the plasma of the two LR3IGF-I-infused sheep tested, and it is concluded that LR3IGF-I caused a down-regulation of the type-I IGF-I receptors followed by a rise in endogenous IGF-I concentration consequent on lack of feedback regulation. After 21 days of infusion there was no effect of LR3IGF-I on wool-follicle-bulb-cell mitotic rate, bulb diameter or wool production.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acids↗

PubMed1995

Milk protein synthesis, gene expression, and hormonal responsiveness in primary cultures of mammary cells from lactating sheep.

A ruminant mammary cell culture that accurately reproduces mammary function in vitro would be a valuable tool in studies of ruminant lactation. With this in mind, we have examined milk protein synthesis and secretion, milk protein mRNA abundance, and hormonal responsiveness in primary cultures of mammary acini from lactating sheep. alpha- and beta-casein protein synthesis, beta-lactoglobulin synthesis, and alpha-casein, beta-casein, and beta-lactoglobulin secretion are maintained at high levels for 8 h in culture, but then decline to approximately 25% of maximal rates between 8 and 24 h in culture, whereas synthesis of other proteins remains unaltered. The relative abundance of alpha-S1-casein, beta-lactoglobulin, and alpha-lactalbumin mRNAs similarly decline between 8 and 24 h in culture. Extracellular labeled alpha-casein is increased fourfold in the presence of fetal calf serum (FCS). In total, FCS alters the abundance of 47 of 68 secreted proteins detected by two-dimensional electrophoresis. However, FCS and lactogenic/galactopoietic hormones had no effect on the rate of decline of mammary function and did not promote any regaining of function when present for up to 9 days in culture. These results suggest that providing its limitations are recognized, this primary cell culture system may be useful in studying some aspects of ruminant mammary function in vitro.

Animals↗

PubMed1995

EGTA-induced disruption of epithelial cell tight junctions in the lactating caprine mammary gland.

The suitability of the Ca2+ chelator ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) to induce disruption of mammary tight junctions (TJ) and its effect on milk secretion were investigated in six goats. EGTA was administered via the teat of one gland as an isosmotic (300 mosmol/l) K-EGTA solution (68 mM EGTA), whereas the control gland received an isosmotic sucrose solution. Lactose, Na, K, and Cl in milk, blood lactose, and the presence of Evans blue (EB) in mammary lymph were used as indicators of TJ disruption. EGTA caused transient (approximately 60 h) changes (P < 0.05) in the concentration of lactose, K, Na, and Cl in milk, consistent with loss of TJ integrity. This was confirmed by a rapid (< 1 h) increase (P < 0.05) in blood lactose levels. Moreover, EB appeared in lymph < 1 h after EGTA+EB treatment. Milk secretion declined unilaterally by 15% (P < 0.05) after EGTA and did not return to baseline until approximately 60 h after EGTA. EGTA caused a unilateral, temporary (first 7 h) increase in mammary blood flow. This study shows that a rapid temporary disruption of mammary TJ can be successfully induced in vivo and that such disruption compromises milk secretion.

Animals↗

PubMed1995

Pharmacokinetics and bioactivity of intact versus truncated IGF-I during a 24-h infusion into lactating goats.

The aim of this study was to compare the plasma concentration profile, mammary blood flow response and transfer into milk of intact IGF-I with that of its truncated analogue, des(1-3)IGF-I (des-IGF-I). Each peptide was infused for 24 h into the pudic artery supplying one mammary gland of lactating goats (n = 5). Concentrations of IGF-I in plasma (from the jugular vein) rose rapidly during infusion of IGF-I or des-IGF-I to reach 510 +/- 62 and 640 +/- 32 ng/ml (mean +/- S.E.M.) respectively, compared with 262 +/- 35 ng/ml after a similar infusion of saline. Ligand blotting analysis indicated a significant increase in the intensity of [125I]IGF-I binding to the 40-43 kDa doublet (binding protein-3 (BP-3), P < 0.01) and the band at 31 kDa (P < 0.05) during infusion of either IGF-I or des-IGF-I, as compared with saline. Furthermore des-IGF-I induced a significant increase in intensity of binding to the 35 and 24 kDa bands, but IGF-I did not. Whereas [125I]IGF-I was distributed between BP-3 and the other binding proteins, [125I]des-IGF-I bound exclusively to BP-3. Mammary blood flow (MBF) increased 48 +/- 6% after 12 h of infusion of des-IGF-I, compared with an increase of 22 +/- 6% during IGF-I. The difference in response was significant at P < 0.05. In addition, more IGF-I was secreted into the milk of the infused than the non-infused gland during either infusion of IGF-I or des-IGF-I.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Isolation of cDNAs and tissue specific expression of ovine glucose transporters.

The facilitative glucose transporters are a family of proteins responsible for the transmembrane transport of glucose and other hexose sugars (1,2). In mammals, the seven glucose transporter isoforms display a characteristic tissue distribution reflecting the physiological requirement and metabolism of glucose. This report describes the isolation and sequencing of the full length ovine GLUT-3 cDNA and the tissue distribution of ovine GLUT-1 and GLUT-3 mRNA. The ovine GLUT-3 cDNA is 3854 base pairs and the coding nucleotides show 82% and 79% homology with the human and mouse GLUT-3 sequences respectively. In addition, a reverse transcriptase-polymerase chain reaction strategy is described for the rapid isolation of mammalian cDNA subclones for GLUT-1, GLUT-2 and GLUT-4. This method has been used to isolate the corresponding ovine subclones.

Amino Acid Sequence↗

PubMed1994

The use of caprylic acid for the extraction of the immunoglobulin fraction from egg yolk of chickens immunised with ovine alpha-lactalbumin.

The extraction and purification of serum-derived immunoglobulin fraction in the egg yolk of hens by the combined treatment of the raw egg yolk with caprylic (octanoic) acid and ammonium sulphate is described. This simple two-step method proved to be both rapid, reproducible and suitable for batch processing of pooled egg yolk. The method recovered in excess of 130 mg of immunoglobulin per egg yolk. Two chickens were inoculated at two weekly intervals with 100 micrograms each of ovine alpha-lactalbumin over a ten week period. The alpha-lactalbumin antigen was purified by a hydrophobic-interaction chromatographic procedure and further purified by a gel excision-elution process. No precipitating antibodies could be demonstrated in gel diffusion techniques with this antibody. The specificity and specific activity of the antibody were monitored by western blotting and demonstrated the presence of highly specific antibodies to ovine alpha-lactalbumin in the treated egg yolk. The extraction procedure had no adverse effects on antibody titre. We concluded, and confirmed previous reports, that the use of chickens for the production of highly specific antibodies to mammalian proteins with particular reference to milk proteins presented numerous advantages over conventional procedures.

Animals↗

PubMed1994

Increased mammary blood flow in the lactating goat induced by parathyroid hormone-related protein.

Human synthetic parathyroid hormone-related protein (PTHrP) increased mammary blood flow (MBF) following close-arterial infusion via the external pudic artery in goats during mid-lactation. MBF increased 74 +/- 8% within 30 min of the start of continuous infusion of PTHrP compared with 10 +/- 3% in controls. MBF decreased by 90 min, however, and was not different from control values for the remainder of the infusion. The increase in plasma concentrations of calcium and decrease in phosphate during PTHrP suggests that this was not due to altered activity of PTHrP, but may relate to downregulation of response or production of counter-regulatory vasoconstrictive agents within the gland. This problem was alleviated when PTHrP was infused in a pulsatile fashion. An average 14-40% increase in MBF was achieved over 6 h, but this did not alter the rate of milk secretion, suggesting that mammary hyperaemia is not sufficient by itself to increase milk yield in the normally lactating goat. MBF increased in a dose-dependent fashion, although the lowest dose used to give a detectable response was approximately 40-fold higher than the concentration normally present in the mammary venous circulation. Thus, endogenous PTHrP may not be an important regulator of MBF during lactation in the goat.

Animals↗

PubMed1994

Effects of close-arterial (external pudic) infusion of insulin-like growth factor-II on milk yield and mammary blood flow in lactating goats.

Five lactating goats were infused, via an external pudic arterial catheter, directly into the mammary gland with 0.9% (w/v) NaCl (20 ml/h), recombinant human insulin-like growth factor-I (IGF-I; 80 nmol/h), recombinant human IGF-II (133 nmol/h) or IGF-I and IGF-II combined. The infusion was for 6 h and milk yield was determined every 2 h. The ratio of milk yield in the infused relative to the non-infused gland was changed only slightly by saline (2%), but increased to 9% (P < 0.05) in response to IGF-I and 8% (P < 0.05) in response to IGF-II. When combined, both peptides increased this ratio by 6%. These effects were elicited within 2-4 h of the beginning of infusion. Mammary blood flow increased 50-80% (P < 0.05) during all IGF infusions, but only 28% during saline treatment. Plasma insulin decreased 50% (P < 0.01) during the infusion of IGF-I alone or in combination with IGF-II and 25% in response to IGF-II alone. Whereas plasma glucose increased by approximately 10% during infusion of IGF-I alone or with IGF-II, it was not altered by infusion of IGF-II only. The rapidity and unilateral nature of the milk-yield response to IGF-I and IGF-II is consistent with their acting directly on mammary tissue itself. Thus, the present results demonstrate similar local and systemic actions induced by intramammary infusion of IGF-II and IGF-I, although the magnitude of the response to IGF-II tends to be less than that to IGF-I.

Animals↗

PubMed1994

Mammary epithelial cell tight junction integrity and mammary blood flow during an extended milking interval in goats.

The timing and relation of changes in mammary epithelial cell tight junction integrity and mammary blood flow during a 36-h milking interval were studied in six lactating Saanen goats. An increase in lactose concentration in plasma, a decrease in transepithelial potential difference, and changes in ionic milk composition were used to indicate tight junction patency. After 36 h of milk accumulation, mammary tight junctions had become disrupted. Further analyses indicated that this disruption began after 21 h of milk accumulation and that mammary blood flow also started to decline after 21 h. The time when both events occurred was not significantly different from the time when milk secretion began to decline (19 h). Moreover, positive but nonsignificant correlations existed between these events. Mammary tight junctions became disrupted when milk secretion declined, suggesting that impairment of mammary tight junction integrity is associated with decreased milk secretion during an extended milking interval. The decline in mammary blood flow may be the result of a negative feedback response to a reduced demand for metabolites, which is due to a reduced rate of milk secretion.

Animals↗

PubMed1994

Effect of milking frequency and somatotropin on the activity of plasminogen activator, plasminogen, and plasmin in bovine milk.

Six pairs of identical twin cows during late lactation (213 d) were used to study the effect of milking frequency (twice vs. once daily) and bST during once daily milking on the activity of plasminogen activator, plasminogen, and plasmin in milk. Less frequent milking increased the activity of plasminogen, plasmin, and plasminogen activator in milk. The ratio of plasminogen to plasmin, a measure that is independent of milk volume, decreased during less frequent milking, suggesting that at least part of the increase in activity of plasmin was due to the accelerated conversion of plasminogen to plasmin. Changes in the activity of plasminogen and plasmin in milk were positively correlated with increases in the concentrations of milk BSA and plasma lactose, both of which are indicators of disruption of tight junctions between mammary epithelial cells, indicating that paracellular leakage may have contributed to increased protease activity in milk during less frequent milking. No correlation existed between changes in plasminogen activator and indicators of tight junction disruption, suggesting that increased activity of plasminogen activator in milk was not due to leakage across the mammary epithelium, but rather to increased local production in the mammary gland. Administration of bST during once daily milking did not significantly affect milk protease activity.

Animals↗

PubMed1992

Insulin-like growth factor binding proteins of equine serum.

Ligand blotting analysis of serum from the horse using radiolabelled IGF-I revealed a protein at 96 kDa which was not present in serum from goat, cow, sheep, deer or donkey. These latter species all displayed five labelled bands in the range 24 to 41 kDa. Conversely, these were only weakly labelled in serum from the horse. Size exclusion chromatography of horse serum pre-incubated with radiolabelled IGF-I revealed reduced binding in the 130-kDa peak compared with goat plasma, and ligand blotting analysis indicated the 96-kDa protein was present in this peak. The 96-kDa protein from horse serum binds IGF-I and IGF-II specifically and appears to be unique to this species. The nature of this protein is at present unknown.

Animals↗

PubMed1992

Secretion of insulin-like growth factor II into milk.

125I-labeled insulin-like growth factor II (IGF-II) was infused directly into the pudic artery supplying one gland of lactating goats (n = 4). Maximum specific activity for [125I]IGF-II transferred into milk from the infused gland was reached 60 min after that in plasma and was 2.5 fold greater than in milk from the non-infused gland. Inclusion of either 67.5 nmoles unlabeled IGF-II or IGF-I had no influence on the amount or pattern of secretion of [125I]IGF-II into milk from either gland. While the temporal pattern of secretion of [125I]IGF-II into milk was consistent with a transcellular mechanism of transfer, the lack of competition by unlabeled IGF-II or IGF-I suggests a non-specific mechanism is operable, which contrasts to secretion of IGF-I.

Animals↗

PubMed1992

Effect of colostrum intake on alpha-lactalbumin concentrations in serum of calves.

Seven Friesian calves were fed colostrum for four days beginning within 24 hours of birth, and milk thereafter. The concentration of alpha-lactalbumin in serum was measured by specific radioimmunoassay and compared to IgG assayed by electroimmunodiffusion. Serum concentrations of alpha-lactalbumin peaked at 387 +/- 85 ng ml-1 within eight hours of initial intake of colostrum, declining to 12 +/- 3 ng ml-1 by day 6. IgG rose steadily to 17 mg ml-1 by 48 hours of birth and remained relatively constant thereafter. The temporal pattern of alpha-lactalbumin in serum following colostrum intake confirms previous studies suggesting reduced absorption of colostral proteins between 24 and 36 hours. The presence of variable amounts of alpha-lactalbumin in serum even after 17 days, however, indicates limited transfer of milk-derived proteins across the gut at this time. The data further show that cessation of maximal gut transfer does not relate to molecular weight of transferred protein.

Animals↗

PubMed1992

Transfer of insulin-like growth factors I and II from plasma to lymph in young goats.

The plasma clearance of intravenously injected 125I-labelled insulin-like growth factor I (IGF-I, n = 13) and IGF-II (n = 12) and their transfer into lymph draining the foreleg of 3.5- to 8-week-old British Saanen goats was studied. Both peptides were initially distributed into a volume equivalent to the plasma volume, while the final distribution spaces for IGF-I and IGF-II were 90 +/- 9.8 and 125 +/- 12 ml/kg live weight respectively. There were two phases to the plasma clearance of both IGF-I and IGF-II, with the half-lives of both phases for IGF-I (9.6 +/- 0.9 and 287 +/- 23 min, first and second phase respectively) being significantly (P less than 0.001) longer than those of IGF-II (4.2 +/- 0.6 and 172 +/- 18 min, respectively). The radioactivity transferred into lymph originated from intact IGF-I and IGF-II as well as degraded products of these compounds, as assessed by precipitation with trichloroacetic acid and gel filtration. Levels of undegraded IGF-I in lymph were 50% greater than IGF-II. While more than 90% of either peptide was bound to specific IGF-binding proteins in plasma, in lymph 34 +/- 2% of IGF-I and 23 +/- 3% of IGF-II remained in the free form 60-80 min after injection. The plasma: lymph ratio for free IGF-I was 2:1 and for bound IGF-I, 8:1. The corresponding values for IGF-II were 3:2 and 8:1 respectively. These results provide direct experimental evidence for transfer of undegraded IGF-I and IGF-II from blood into lymph of the foreleg, consistent with the ability of these factors to act in an endocrine role in growing tissues. Differences between plasma clearance and transfer of IGF-II into lymph compared with IGF-I may be due to its greater cellular uptake and/or degradation in vivo.

Animals↗

PubMed1992

Effect of growth hormone treatment on the distribution of insulin-like growth factor-I between plasma and lymph of lactating sheep.

Plasma and mammary efferent lymph concentrations of insulin-like growth factor I (IGF-I) were determined in lactating ewes before and after treatment with GH (10 mg/day) for 3 days. The lymph:plasma ratio of IGF-I increased from 0.34 to 0.47 after GH treatment when the IGF-I content of plasma increased by 19.4 nmol/l (from 32.1 nmol/l) and lymph by 13.7 nmol/l (from 10.7 nmol/l). This increase in the relative content of IGF-I in lymph was associated with increased lymph content of IGF-I in a lower molecular mass pool (nominally 50 kDa) derived by size exclusion chromatography. GH treatment increased the total binding capacity for IGF-I in both high (150 kDa) and low (50 kDa) molecular mass pools of plasma and the 150 kDa pool in lymph but there was a proportionally greater increase in 50 kDa total binding in lymph relative to plasma. Further, GH treatment increased the 'saturation' of the 50 kDa binding proteins but decreased the 'saturation' of the 150 kDa fraction, in both plasma and lymph. Ligand blot analysis of IGF-binding proteins (IGFBPs) in plasma and lymph showed that GH treatment of lactating sheep increased IGFBP-3 and decreased IGFBP-2 in plasma and lymph. Radioimmunoassay of IGFBP-2 showed that while GH treatment reduced the plasma content of IGFBP-2 by about half, the lymph:plasma ratio was increased from 0.68 to 0.87. GH treatment of lactating ewes not only increased the IGF-I content of plasma but increased the apparent efficiency of transfer of IGF-I across capillary endothelium to mammary efferent lymph.

Animals↗