Autoantibody to vasoactive intestinal peptide in human circulation

In a radioassay for Vasoactive Intestinal Peptide (VIP)-binding, eight out of 33 plasma samples from healthy human subjects exhibited specific binding ranging from 2.6% to 46.7% of total [125 I]VIP. This binding was competitively displaced by unlabeled VIP. The structurally homologous peptides, Peptide Histidine Isoleucine (PHI) and secretin, were, respectively, 72-fold and 413-fold less potent than VIP in displacing bound [125 I]VIP, whereas the unrelated peptides, neurotensin, eledoisin, bombesin and metenkephalin, were without effect on the binding. The antibody nature of the VIP-binding factor was suggested by its precipitation with ammonium sulfate, attenuation after absorption with Staphylococcus aureus preparations, precipitation with antisera against human IgG and IgM, and coelution with standard IgG and IgM on anion-exchange and high-performance gel-filtration columns. Pepsin treatment of purified IgG fraction yielded a VIP-binding species with apparent molecular weight of 108 +/- 13 kDa that was precipitated by antiserum against the F(ab)2 fragment of the IgG molecule. These results demonstrate the existence in some human plasmas of an autoantibody that binds VIP.     

Some characteristics of pollen wall cytochemistry and ultrastructure in Japanese larch (Larix leptolepis Gord.)

Summary The mature pollen of Larix leptolepis Gord. (Conifer) contains five different cell types, and the plasma membrane of the vegetative cell is continuous and organized. The pollen wall is composed of two morphologically and cytochemically distinct domains: the exine and the intine. In the multilayered exine, the ektexine appears granular and the endexine, lamellar. The intine is thick and bilayered with a microfibrillar structure occupying its inner portion. Cytochemical reactions of the exine and the intine are similar to those found in angiosperms. Pollen wall involvement in the male female recognition system is discussed with respecl to the angiosperms.     

Interaction of alpha-N-Acetyl-beta-endorphin and calmodulin

Acetylation at the -amino terminal is a common post-translational modification of many peptides and proteins. In the case of the potent opiate peptide -endorphin, -N-acetylation is a known physiological modification that abolishes opiate activity. Since there are no known receptors for -N-acetyl--endorphin, we have studied the association of this peptide with calmodulin, a calcium-dependent protein that binds a variety of peptides, phenothiazines, and enzymes, as a model system for studying acetylated endorphin-protein interactions. Association of the acetylated peptide with calmodulin was demonstrated by cross-linking with bis(sulfosuccinimidyl)suberate; like -endorphin, adducts containing 1 mol and 2 mol of acetylated peptide per mole calmodulin were formed. Some of the bound peptides are evidently in relatively close proximity to each other since, in the presence of amidated (i.e., lysine-blocked) calmodulin, cross-linking yielded peptide dimers. The acetylated peptide exhibited no appreciable helicity in aqueous solution, but in trifluoroethanol (TFE) considerable helicity was formed. Also, a mixture of acetylated peptide and calmodulin was characterized by a circular dichroic spectrum indicative of induced helicity. Empirical prediction rules, applied earlier to -endorphin, suggest that residues 14–24 exhibit -helix potential. This segment has the potential of forming an amphipathic helix; this structural unit is believed to be important in calmodulin binding. The acetylated peptide was capable of inhibiting the calmodulin-mediated stimulation of cyclic nucleotide phosphodiesterase (EC 3.1.4.17) activity with an effective dose for 50% inhibition of about 3 µM; this inhibitory effect was demonstrated using both an enzyme-enriched preparation as well as highly purified enzyme. Thus, acetylation at the -amino terminal of -endorphin, although abolishing opiate activity, does not interfere with the binding to calmodulin. Indeed, -endorphin and the -N-acetylated peptide behave very similarly with respect to calmodulin association.Portions of this work are in partial fulfillment of the requirements for the Ph.D. degree from Vanderbilt University.     

Effect of nutrition and endoparasitic treatment on growth, onset of puberty and reproductive activity in Menz ewe lambs

Ewe lambs of the Ethiopian Menz breed were assigned at weaning (90+/-3 days) to four levels of nutrition (poor, low, medium and high) to achieve different premating growth rates with or without treatment for endoparasites. A concentrate mixture providing 2.5 Mcal/kg dry matter (DM) metabolizable energy and 15.2 g/kg DM digestible protein was used. Improved nutrition increased lamb postweaning average daily weight gain up to puberty by 6 to 26 g/day and the conception rate to first estrus by 9 to 16% while it reduced the mortality rate by 24 to 31% and age at first lambing by 2 to 5 months. Lambs reached puberty (age at first estrus) at 16.9+/-0.1 kg (+/-SEM) or 60% of mature body weight and 350+/-12 days of age. The onset of puberty was advanced by weaning weight (P<0.05), itself being well correlated with birth weight (r = 0.51, P<0.001), and by level of nutrition (high=299+/-19, medium=301+/-18, low=383+/-23 and poor=454+/-31 days, P<0.001) through enhanced growth rate (r = -0.82, P<0.001). No independent effect of drenching for endoparasites on pubertal development was observed (P>0.05), but its interaction with season-of-birth improved the growth of lambs born during the period of short rains (P<0.05). Overall mean litter size at first lambing was 1.07; the twinning rate was 6.5% and the birth weight was 1.9+/-0.1 kg. Up to 13.4% of newborn lambs, averaging 1.3+/-0.6 kg, died on the day of parturition. The results indicate that improved growth rate and body weight, resulting from better postweaning nutrition, affects the attainment of puberty in Menz ewe lambs. Mitigation of nutrition stress and endoparasitic infection depending on season-of-birth would thus increase the annual reproductive rate of breeding ewes and flock productivity.     

Further studies on prevalence of hydatidosis in slaughtered animals from North Jordan

Examination of 471 sheep, 118 goats, 157 cattle and 56 camels slaughtered in abattoirs in North Jordan was carried out during March-May 1984. Drought conditions that prevailed during the preceding winter led to slaughtering old female sheep (greater than or equal to 4 years) due to scarcity of food, which allowed us to analyse the prevalence of hydatidosis in various age groups of sheep. An overall infection rate of 27.8, 1.7, 5.8 and 10.7 percent was found in sheep, goats, cattle and camels, respectively. The infection rate was as low as 1.5 percent in male and 1.9 percent in female sheep under 2 years of age. However, the rate of hydatid infection increased with age and reached as high as 63.7 percent in ewes 4 years of age and older. The percentage of animals with fertile cysts was also highest in sheep (68.7 percent of infected animals) and increased with age reaching 100 percent in ewes which were 10 years of age or older. Analysis of all cysts recovered from the livers and lungs of infected ewes from various age groups revealed a sharp increase in the mean total number of cysts in age groups over 8 years of age. The fertility rate of the cysts in the liver was significantly greater in ewes 6 years old or more (64.8--78.6 percent) than in younger age groups (8.7-46.2 percent). In the lung, the fertility rate increased progressively with age reaching as high as 97.9 percent in ewes 10 years old or more. These findings of high infection and fertility rates of hydatid disease in sheep, particularly of older age groups, prompt plans for further epidemiological studies and control programmes.     

Monoclonal antibodies against human ovarian tumor associated antigen NB/70K: Preparation and use in a radioimmunoassay for measuring NB/70K in serum

Summary Murine monoclonal antibodies (MCAs) against human ovarian tumor associated antigen NB/70K have been prepared. One of these MCAs, NB12123, was chosen for the development of a radioimmunoassay for measuring serum NB/70K levels. In this assay, the average NB/70K level in 75 normal, healthy controls was 11.9 activity units (AU) with an SD of 14.9 AU. The normal cut off value for this assay was set at 45 AU (mean +2 SD). 24 of 46 (52%) ovarian cancer patients, 7 of 18 (39%) patients with benign ovarian cysts or tumors and 3 of 85 (4%) control samples had elevated serum NB/70K levels. Comparison of NB/70K levels measured in the NB12123 assay with levels measured in an assay using a polyclonal antiNB/70K previously developed in our laboratory [13] indicated that although both assays had approximately the same percentage of positive ovarian cancer patient samples, there appeared to be no correlation between the absolute NB/70K levels measured by the two assays. The rank of ovarian cancer patient samples was also different for the two assays. Also, almost 40% of patients with benign ovarian cysts and tumors had elevated serum NB/70K levels as measured by the NB12123 assay as compared to 0% for the polyclonal assay. Reciprocal cross-blocking experiments, absorption studies, and immune precipitate analysis indicated that both the monoclonal NB12123 assay and the polyclonal antiNB/70K assay measured the same population of NB/70K molecules. However, the polyclonal antibody recognizes epitopes in addition to that recognized by NB12123. Taken together, these results suggest that the epitope recognized by NB12123 is not as specific for malignant ovarian tumors as the epitope(s) recognized by polyclonal antiNB/70K and/or that more than the one epitope detected by the MCA is responsible for the specificity for ovarian cancer of the polyclonal NB/70K assay. In spite of this, the greater sensitivity and range of the monoclonal NB12123 assay make it possible to monitor serum NB/70K levels in ovarian cancer patients. In four patients examined, the fluctuating serum NB/70K levels appeared to correlate well with clinical statusSupported in part by ACS # PDT 231 and a grant from the Elsa U. Pardee Foundation     

Stereochemistry of the hydrolysis of α,α-trehalose by trehalase,determined by using a labelled substrate

(1,1′-¹³C)α,α-Trehalose was obtained in 37% yield from the Pavia condensation of 2,3,4,6-tetra-O-benzyl-d-(1-¹³C)glucopyranose, in dichloromethane in the presence of trifluoromethanesulfonic anhydride, followed by the usual deprotection techniques. The hydrolysis of this substrate by cockchafer trehalase was monitored at 37° by using ¹³C-n.m.r. spectroscopy with short recording times. Equimolecular amounts of α- and β-d-glucopyranose are released simultaneously by the action of the enzyme. This result is consistent with a bimolecular substitution mechanism, taking into account previous results involving C-2 asymmetric participation in the catalytic step of hydrolysis of α,α-trehalose. For comparative evaluation of its accuracy, the usual polarimetric technique was also used for the determination of the anomeric configuration of the d-glucose released by the action of the enzyme on α,α-trehalose.