Effect of administration of a novel recombinant bovine interferon on length of oestrous cycle in cattle.

In ruminants, extensive reproductive loss occurs during the process of maternal recognition of pregnancy and it has been suggested that trophoblast interferons may be potential therapeutic agents. This paper reports results from a trial using eight first lactation Holstein-Friesian heifers to test the efficacy of a novel recombinant bovine interferon produced in bacteria in extending the life of the corpus luteum. Oestrus was synchronised in these animals and 0.1 mg of this non-glycosylated interferon was infused into the uterus twice daily for 13 days starting approximately 12 days after oestrus. This treatment resulted in an extension of the lifespan of the corpus luteum by 5.5 days (P=0.028) compared with untreated controls. In these animals the interovulatory period was extended by 6.4 days (P=0.009). Administration of this protein did not have any adverse effects either on body temperature or on daily milk yields. The results indicate that this novel interferon may have potential therapeutic application for reducing embryo mortality.

Effect of a novel recombinant bovine interferon and trophoblast secretory products on protein metabolism by endometrial explants from cattle and sheep.

The effect of a novel recombinant bovine interferon on the rate of protein synthesis, as measured by the rate of leucine incorporation, in endometrial explants from cows and ewes was investigated and compared with the effect of trophoblast secretory proteins and of a recombinant bovine interferon-alpha1. Exposure of cultured bovine endometrial tissue to the novel recombinant bovine interferon mimicked the effect of pregnancy in reducing the rate of protein synthesis in both caruncular and intercaruncular endometrium. This effect was similar to that seen with naturally produced trophoblast secretory proteins and it was more pronounced than that seen when endometrial tissue was treated with a related recombinant bovine interferon-alpha1. Ovine endometrial cultures were unaffected by exposure to this novel recombinant bovine interferon. This novel recombinant interferon exhibited potent antiviral properties (an activity of 1.6 x 10(10) international reference units/mg protein) and may prove to be a valuable reagent in studying the maternal recognition of pregnancy in the cow and reducing embryo mortality.

Effect of fetal thyroidectomy on brown adipose tissue and thermoregulation in newborn lambs.

The effect of fetal thyroidectomy on thermoregulation in newborn lambs was investigated. Seven of 14 lambs born normally at term were thyroidectomized at Day 127 of gestation. Colonic temperature and oxygen consumption were measured during non-rapid eye movement sleep 6-45 h after birth. All lambs were then killed and perirenal brown adipose tissue was sampled for measurement of thermogenic activity (guanosine diphosphate binding), uncoupling protein and lipid contents. Thyroidectomized lambs tended to have a mean colonic temperature 2.35 degrees C lower (P = 0.067) than controls and two became hypothermic (i.e. colonic temperature < 35 degrees C). Thyroidectomized lambs exhibited lower rates of oxygen consumption (P = 0.05) and an increased incidence of shivering thermogenesis. The perirenal adipose tissue of these lambs had a lower thermogenic activity (P < 0.01), less uncoupling protein (P < 0.01) and higher lipid content (P = 0.072) compared with intact controls. It is concluded that fetal thyroidectomy results in a decreased ability of newborn lambs to utilize nonshivering thermogenesis in brown adipose tissue as well as increasing the incidence of hypothermia. These changes are associated with decreased synthesis of uncoupling protein and functional development of brown adipose tissue in the late gestation fetus.

Elevated levels of N-terminal pro-opiomelanocortin peptides in fetal sheep plasma may contribute to fetal adrenal gland development and the pre-parturient cortisol surge.

Parturition in sheep is initiated by a rapid rise in fetal plasma cortisol. There is some controversy as to the exact nature of the drive for this pre-partum cortisol surge and it is thought that factors other than ACTH may act in concert to stimulate the development of the fetal adrenal gland. We have investigated the concentrations of ACTH and other peptides derived from pro-opiomelanocortin (POMC) in the circulation of fetal sheep during the final part of gestation, using specific 2-site immunoradiometric assays. The expected rise in fetal cortisol was seen with an 880% (p < 0.01) increase in concentration of this hormone between the initial measurement period (110-119 days gestation) and the final period (139-147 days). Fetal plasma ACTH increased less dramatically (137%; p < 0.03) during this time. The most surprising finding was the presence of very high relative concentrations of the N-terminal POMC peptide N-POMC(1-77) in the fetal circulation. Initially the concentration was 289 +/- 66 pmol/l compared to ACTH concentrations of 6.4 +/- 0.8 pmol/l. In the final week of gestation N-POMC(1-77) levels, although still high, had declined to 188 +/- 35 pmol/l (ACTH having increased to 13.7 +/- 2.2 pmol/l). Fetal plasma 3 yen-MSH was found to increase towards the end of gestation when the concentration of N-POMC(1-77) was declining, suggesting some cleavage of the latter. We postulate that the N-POMC(1-77) and its fragments, acting in concert with ACTH, play a vital role in stimulating the development of the fetal adrenal cortex and provide the additional drive to the adrenal gland required to stimulate parturition.

Regulation of intracellular protein degradation in the isolated perfused liver of the chicken (Gallus domesticus).

1. The effects of insulin, glucagon and a supply of exogenous amino acids on protein degradation have been studied in isolated perfused livers from growing chickens by measuring the rate of net valine release in the presence of cycloheximide. 2. Insulin inhibited protein degradation as did a supply of exogenous amino acids. 3. Addition of glucagon increased uric acid release from the livers but had no significant effect on protein degradation. 4. When the effects of the hormones and amino acid mixture are compared with published data for the rat it is evident that the action of glucagon differs in the two species.

Ovine trophoblast protein-one: evidence for possible glycosylation.

1. The polymerase chain reaction has been used to amplify specifically the cDNA coding for the secreted form of ovine trophoblast protein-one from a preparation of total cellular RNA extracted from sheep embryos removed from ewes 16 days after mating. 2. Cloning and sequencing of the amplified cDNA revealed two new sequence variants: SPW49 having 93% similarity with deduced amino acid sequences from published cDNA data, and SPW27 a variant coding for a deleted form of ovine trophoblast protein-one. 3. The gene for ovine trophoblast protein-one is intronless. 4. This study provides further evidence for the existence of an ovine trophoblast protein-one gene family. 5. Both variants contain a potential N-glycosylation site not apparent in published sequences for ovine trophoblast protein-one.

Morphological and radiochemical evidence for the metabolism of exogenous proteins by the preimplantation sheep blastocyst.

The ability of the trophoblast of the ovine preimplantation blastocyst to take up and metabolise proteins has been investigated using two experimental approaches, microscopical and radiochemical. The ultrastructure of the expanded blastocyst obtained from 14 and 17 day pregnant ewes was examined. The morphology of tissues maintained in culture for 24 h has been compared with that of fresh tissues. After culture, the cellular morphology of the explants was well preserved. Fresh and 24 h cultured tissues were incubated with horse-radish peroxidase and ferritin and these proteins subsequently were found to be localized in coated pits, caveolae and secondary lysosomes of the trophoblast. Comparison of the uptake of [3H]dextran and of 125I-labelled bovine serum albumin indicated that proteins could be taken up by cultured tissue by mechanisms in addition to simple fluid phase endocytosis. During culture of explants of blastocyst with 125I-labelled bovine serum albumin, a large fraction of the radioactivity taken up by the tissue appeared in the TCA-soluble fraction of the culture medium indicating that cultured trophoblast hydrolysed proteins. That amino acids released from captured protein could be used for protein synthesis by the trophoblast was indicated by the labelling of tissue and medium proteins after culturing explants with beta-lactamase labelled with [14C]leucine. A major product (Mr approximately 17 x 10(3) present in the medium was likely to have been ovine trophoblast protein-1. It is concluded that, during the expansion of the ovine blastocyst, the trophoblast has the ability to take up proteins, transport them to lysosomes and degrade them to amino acids which are used for protein synthesis. Thus proteins, as well as free amino acids, present in the histotrophe may be an important source of nitrogen for the sheep conceptus in the critical period just prior to implantation.

Glucose metabolism in shorn and unshorn pregnant sheep.

1. Whole-body, hind-limb and uterine tissue metabolism of glucose was studied using a combination of isotopic and arterio-venous difference techniques in shorn and unshorn pregnant sheep over the final 4 weeks of pregnancy. This was combined with the measurement of the concentrations of oxygen and carbon dioxide in arterial blood and plasma concentrations of lactate, acetate, non-esterified fatty acids, 3-hydroxybutyrate, glycerol, growth hormone (GH), insulin, glucagon, cortisol, thyroxine and 3,5,3'-triiodothyronine (T3). 2. Glucose entry rate was 28% higher in shorn ewes compared with unshorn controls, even though there was no difference in the arterial plasma concentration of glucose. This effect may have been caused by a decrease in the molar rate, insulin: glucagon (I:G), which was 40% lower in shorn ewes as a result of a significant decrease in the plasma concentration of insulin. There was no difference in the plasma concentration of cortisol or GH. 3. Blood flow across the hind-limb or uterine tissues was not significantly different between shorn and unshorn groups, neither were the net glucose uptake, glucose oxidation rate or contribution of glucose to O2 consumption across these tissues. 4. Insulin-tolerance tests performed on a separate group of shorn and unshorn ewes showed an increased sensitivity to the hypoglycaemic effects of insulin in the shorn group. 5. There was no significant difference between shorn and unshorn animals in the contribution of glucose to CO2 output or in the proportion of glucose entry rate oxidized. CO2 entry rate was 18% higher in shorn ewes compared with unshorn controls which resulted in a 26% higher estimated value for heat production. There was a 47% increase in glucose oxidation rate in shorn ewes but there was no significant difference in the proportion of total heat production which was derived from glucose. The arterial concentrations of O2 and CO2 were significantly higher in shorn ewes, which may be an indication of the higher metabolic rate in these animals. This effect may be mediated via a significant rise in plasma T3 concentration in the shorn group. 6. It is concluded that as a result of long-term cold exposure there is a significant increase in whole-body glucose entry and oxidation rates in the shorn pregnant ewe. The increase in insulin sensitivity at the same time as a decrease in plasma insulin concentration may represent a mechanism to ensure continued glucose supply to insulin-sensitive tissues while the concomitant decrease in plasma I:G stimulates hepatic gluconeogenesis.

The irreversible loss of alanine and of glycine in fetal and sucking lambs.

Estimates have been made of the irreversible loss of alanine and of glycine in chronically catheterized fetal lambs and in sucking lambs using [U-14C]-labelled radioisotopes. The experiments in the fetal lambs were carried out at least 5 d after implantation of catheters. The mean concentration of glycine in fetal femoral arterial blood between 102 and 129 d conceptual age was 755 mumol/l and this was not significantly different from that in maternal venous blood. The mean concentration of alanine in fetal femoral arterial blood during the same period of gestation was 229 mumol/l and this was significantly greater than that in maternal venous blood. Assuming a catenary model, the mean irreversible loss of glycine, determined using the single-injection technique, in three fetal lambs of 107, 111 and 127 d conceptual age was 17 mumol/min per kg, whereas in two fetal lambs aged 106 and 109 d into which the isotope was infused continuously the mean irreversible loss, calculated from the specific activity of glycine 5 h after the start of infusion of the tracer ('pseudo plateau'), was 12 mumol/min per kg. In a sucking lamb, 9 d after birth, the irreversible loss of glycine was 11 mumol/min per kg. The mean irreversible loss of alanine, determined by the single-injection technique assuming a catenary model in five fetuses between 112 and 121 d conceptual age was 14 mumol/min per kg, and in two sucking lambs, 9 and 11 d after birth, it was 5.1 mumol/min per kg. When a two-pool model was assumed in which entry of metabolite was not directly into the sampling pool but was by way of the second pool, then the mean irreversible loss of glycine in the three fetuses was 23 mumol/min per kg and of alanine in the five fetuses was 32 mumol/min per kg. Calculations based on the alternative two-pool model did not alter appreciably the rates of irreversible loss of either alanine or glycine in the sucking lambs. From a comparison of the specific activities of the amino acids and of carbon dioxide in blood during the course of the experiments, it was found that in the fetuses 0.96% of the CO2 present in blood was derived from alanine and only 0.12% was derived from glycine. It was calculated that not more than 1.6 mumol alanine/min per kg and 0.29 mumol glycine/min per kg could have been converted to CO2 in the fetal lambs.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of propionic acid on urea synthesis by sheep liver.

Propionate reduced substantially the rate of ureagenesis by slices of sheep liver whereas butyrate did not inhibit urea synthesis. The site of inhibition of urea synthesis by propionate occurs at some point between the fixation of ammonia and the formation of citrulline since 0.5 mM propionate inhibited by 80 per cent the synthesis of citrulline by mitochondria isolated from sheep liver. Since the apparent Ki for propionate is approximately 1.7 mM, the inhibitory effect of propionate on urea synthesis could be of physiological significance in sheep.

Phase I evaluation of vindesine in children: a Southwest Oncology Group pilot study.

Twenty-one children (19 with leukemia) were given 34 courses of vindesine on a weekly or twice-weekly schedule in escalating doses. Thirty-three courses were fully or partially evaluable for response and/or toxicity. Granulocytopenia was the dose-limiting toxicity. Transient jaw, neck, or bone pain was common after each dose. Motor weakness, paresthesias, and constipation were neither frequent nor severe. In this Phase I study, vindesine had some antileukemia activity in children previously treated with vincristine and other drugs. Phase II studies are warranted and a starting does of 1.85% mg/m2 twice weekly appears tolerable.

Childhood multifocal osteosarcoma--diagnosis by 99mtechnetium bone scan: a case report.

A 14-year-old boy with osteosarcoma presented with evidence of a solitary bone lesion by clinical and radiographic examination. A preoperative 99mtechnetium bone scan revealed multiple skeletal osteoblastic abnormalities and upon biopsy of one of these lesions, the diagnosis of multifocal osteosarcoma was established. This unusual case dramatically illustrates the value of 99mtechnetium bone scan in preoperative staging of patients with osteosarcoma.