The role of the fetal gonads and placenta in steroid production, maintenance of pregnancy and parturition in the mare.

The effects of fetal gonadectomy on steroid production and the maintenance of pregnancy in the mare were studied. Removal of the fetal gonads resulted in an immediate fall in maternal plasma concentrations of conjugated and unconjugated oestrogens whereas progestagen levels remained unchanged. Hormone profiles in mares carrying sham-operated fetuses remained similar to those in unoperated control mares. Plasma levels of 13,14-dihydro-15-oxo-PGF-2 alpha (PGFM) were much lower, and uterine contractions weaker, during labour in mares carrying gonadectomized foals than in control mares. Pregnancy was maintained until parturition at term in the mares carrying gonadectomized fetuses. However, 3 of the 4 gonadectomized foals were dysmature and died during or soon after birth. The biosynthetic pathways involved in the production of oestrogens by the feto-placental unit and the possible role of oestrogens in fetal development in the pregnant mare are discussed.     

Transdetermination of Drosophila imaginal discs cultured in vitro

Imaginal discs of Drosophila melanogaster undergo transdetermination when cultured in vivo in the abdominal cavity of adult female hosts. We report here that leg discs cultured in vitro, in a recently developed system, also undergo transdetermination. Whether cultured in vivo or in vitro, leg discs produce a similar range of specific transdetermined structures. Moreover, in comparison to discs cultured in vivo, the discs cultured in vitro exhibit a similar correlation between the amount of growth and the total frequency of transdetermination.     

Purification of the glycoprotein lectin from the broad bean (Vicia faba) and a comparison of its properties with lectins of similar specificity.

1. The lectin from the broad bean (Vicia faba) was purified by affinity chromatography by using 3-O-methylglucosamine covalently attached through the amino group to CH-Sepharose (an omega-hexanoic acid derivative of agarose). Its composition and the nature of its subunits were compared with concanavalin A and the lectins from pea and lentil. 2. Unlike the other three lectins, broad-bean lectin is a glycoprotein; a glycopeptide containing glucosamine and mannose was isolated from a proteolytic digest. 3. The mol.wt. is about 47500; the glycoprotein consists of two apprently identical subunits, held together by non-covalent forces. Fragments of the subunits, similar to those found in concanavalin A and soya-bean agglutinin, were found in active preparations. 4. Broad-bean lectin was compared with concanavalin A and the lectins from pea and lentil in an investigation of the inhibition of their action by a number of monosaccharides, methyl ethers of monosaccharides, disaccharides and glycopeptides. The most striking differences concern 3-O-substituted monosaccharides, which are strong inhibitors of the action of broad-bean, pea and lentil lectins but not of the action of concanavalin A. There is, however, no strong inhibition of the action of these lectins by 3-Olinked disaccharides.     

Properties of microsomal phospholipases in rat liver and hepatoma.

Phospholipase A1, A2 and lysophospholipase activities in microsomes of Novikoff hepatoma host rat liver and regenerating rat liver were compared using 1-[9', 10'-3H2]palmitoyl-2-[1'-14C] linoleoyl-sn-glycero-3-phosphoethanolamine, 1-[1' -3H-]hexadecyl-2-acyl-sn-glycero-3-phosphoethanolamine, and 1-[9', 10'-3H2]palmitoyl-sn-glycero-3-phosphoethanolamine as substrates. 1. Microsomes of all three tissues showed two pH dependent peaks of hydrolytic activity, one at pH 7.5 and another at pH 9.5. 2. Phospholipid hydrolytic activity in microsomes from host liver and regenerating liver require Ca2+ for hydrolysis at pH 9.5, but not at pH 7.5. Hepatoma microsomes require Ca2+ for activity at both pH values. 3. Phospholipase A1 activity, stimulated by addition of Triton X-100 to the incubation mixtures, was detected in both host liver and regenerating liver microsomes. There was no evidence of phospholipase A1 activity in hepatoma microsomes. 4. Phospholipase A2 was detected in microsomes of all three tissues using 1-[1'-3H] hexadecyl-2-acyl-sn-glycero-3-phosphoethanolamine as a substrate. The activity required calcium and was inhibited by Triton X-100. 5. Lysophospholipase activity was evident in the microsomes from all three tissues. The activity was inhibited by both Ca2+ and Triton X-100. 6. Differences were also detected between host liver and hepatoma microsomal phospholipid hydrolase activities with respect to the effect of increasing protein concentration, apparent Michaelis-Menten constants, and time course of the reaction.     

The surface morphology of normal and malignant rat liver epithelial cells in culture

Summary A direct comparison has been made between normal parenchymal cells cultured from rat liver and malignant cells from both a rat liver tumor and a spontaneously transformed line derived from the parent rat culture. In all nondividing cells there was a 3- to 5-fold increase in the population of surface microvilli on the malignant cells compared to the normal cells. Notable variations in overall morphology were also observed when cells were incubated in arginine-deficient medium. During the course of this work T. D. Allen and P. T. Iype were supported by grants from the Medical Research Council and Cancer Research Campaign, and M. J. Murphy, Jr. was supported by a Special Fellowship of the Leukemia Society of America, Inc. and the Jean Shaland Fund.     

Ultrastructure of the salivary bladder of the nine-banded armadillo

Summary The nine-banded armadillo possesses a salivary bladder which is a dilated portion of the main duct of the submandibular gland at its origin. The wall of the bladder is composed of an epithelium, a submucosa and a thick coat of skeletal muscle. The ultrastructure of the epithelium reveals that it is complex and consists of three cell types: 1) principal cells, 2) light cells, and 3) basal cells. The general organization of the epithelium suggests that it is a transporting type of epithelium such as that found in the amphibian and reptilian urinary bladders and the mammalian gall bladder. The submucosa is composed primarily of densely-packed collagen fibers. The skeletal muscle is very vascular and richly innervated.This study was supported in part by a research grant from U.S.P.H.S. (GRS 5-S01-RR-05705)The authors wish to acknowledge the technical assistance of Elizabeth Underwood     

Human plasma R-type vitamin B12-binding proteins. II. The role of transcobalamin I, transcobalamin III, and the normal granulocyte vitamin B12-binding protein in the plasma transport of vitamin B12.

The normal human granulocyte vitamin B12-binding protein, transcobalamin I, and transcobalamin III, have been labeled with 125I-labeled N-succinimidyl 3-(4-hydroxyphenyl)propionate and utilized for plasma clearance studies performed with rabbits. Both moieties of 125I-labeled granulocyte vitamin B12-binding protein-[57Co]vitamin B12 were cleared rapidly from the plasma (is less than 90% by 5 min) by the liver. After 30 min, the bulk of the 125I reappeared in the plasma in small molecular weight (less than 1000) form and was rapidly excreted in the urine. After 60 min the bulk of the [57Co]vitamin B12 reappeared in the plasma bound to rabbit transcobalamin II and was subsequently taken up by a variety of tissues. Approximately 15% of the 125I-labeled granulocyte vitamin B12-binding protein-[57Co-a1vitamin B12 was excreted intact into the bile during the period from 10 to 80 min after injection. The hepatic uptake of the protein-vitamin B12 complex was blocked by the prior injection of desialyzed fetuin but not by native fetuin. Similar results were obtained with 125I-labeled transcobalamin III-[57Co]vitamin B12. Approximately 90% of both moieties of 125I-labeled transcobalamin I-[57Co]vitamin B12 had prolonged plasma survivals similar to that of 125I-labeled bovine serum albumin. After treatment with neuraminadase, both moieties of the 125I-labeled transcobalamin I-[57Co]vitamin B12 complex were cleared rapidly from the plasma by the liver in a manner that was indistinguishable from that observed in the case of untreated granulocyte vitamin B12-binding protein and transcobalamin III. These observations indicate that desialyzed transcobalamin I and the native forms of the granulocyte vitamin B12-binding protein and transcobalamin III are cleared from plasma by the mechanism elucidated by Ashwell and Morell (Ashwell, G., and Morell A. G. (1974) Adv. Enzymol. 41, 99-128) that is capable of clearing a wide variety of asialoglycoproteins. These observations have implications concerning the function of the human R-type vitamin B12-binding proteins, the nature of the enterohepatic circulation of vitamin B12, the biological significance of the mechanism described by Ashwell and Morell, and the etiology of the increased plasma concentration of human R-type protein that occurs frequently in chronic myelogenous leukemia and occasionally in hepatocellular carcinoma and other solid tumors.     

Iodination of glyceraldehyde 3-phosphate dehydrogenase from Bacillus stearothermophilus.

The reaction of iodine with glyceraldehyde 3-phosphate dehydrogenase from Bacillus stearothermophilus was investigated. The active-site thiol group of the cysteine residue homologous with cysteine-149 in the pig muscle enzyme was protected by reaction with tetrathionate. The apoenzyme was readily inhibited by KI3 solution at pH8, but the coenzyme, NAD+, protected the enzyme against inhibition and decreased the extent of iodination. At pH 9.5, ready inhibition of both apo- and holo-enzyme was observed. Tryptic peptides containing residues iodinated at pH 8 were isolated and characterized. One of the most reactive residues in both holo- and apo-enzymes was a tyrosine homologous with tyrosine-46 in the pig muscle enzyme, and this residue was iodinated without loss of enzymic activity. Other reactive tyrosine residues in the apoenzyme were in positions homologous with residues 178, 273, 283 and 311 in the pig muscle enzyme, but they were not readily iodinated in the holoenzyme. Histidine residues in both holo- and apo-enzymes were iodinated at pH 8 in sequence positions homologous with residues 50, 162 and 190 in the pig muscle enzyme. The inhibition of the enzyme was not correlated with the iodination of a particular residue. The results are discussed in relation to a three-dimensional model based on the structure of the lobster muscle enzyme and demonstrate that conformational changes affecting the reactivity of several tyrosine residues most probably occur on binding of the coenzyme.     

Pregnant mare serum gonadotrophin: rate of clearance from the circulation of sheep.

The process involved in the disappearance of PMSG from the blood of sheep, following a single intravenous injection, has been separated into two exponential components. Values (mean plus or minus S.E.) calculated from experiments on five animals were: metabolic clearance rate (37.8 plus or minus 1.6 ml hr-minus 1); rate constant of disposal (0.0315 plus or minus 0.0016 hr-minus 1); half-time of disposal (21.2 plus or minus 1.1 hr). The stage of the oestrous cycle, ovariectomy and the dose of PMSG used had no apparent effect on these values.