Expression of the p80 region of bovine viral diarrhea virus and identification of specific antibodies to this recombinant protein in bovine sera.

A 917-base pair segment of the bovine viral diarrhea virus (BVDV) genome encoding part of the p80 region was cloned into plasmid Gex-2T expression vector for expression as a fusion protein with glutathione-S-transferase (GST). When the p80 and GST sequences were in the same reading frames, the resulting GST-p80 fusion protein had a molecular mass of 58 kilodaltons (kDa) in SDS-PAGE. Extracts of control E. coli carrying only the vector plasmid (Gex-2T) did not contain this new 58-kDa protein band. Mouse monoclonal antibody specific to BVDV-p80 recognized this recombinant protein. Seventy cattle sera that had an SN titer (to TGAC isolate of cytopathic BVDV) greater than 1:8 reacted with this recombinant protein in Western blots. Of 28 cattle sera that had SN titers less than or equal to 1:8, only one serum tested positive on Western blots.     

On feeding and nutrition in Fungiacava eilatensis (Bivalvia, Mytilidae), a commensal living in fungiid corals

The enlarged inhalant siphon of Fungiacava eilatensis opens into the coelenteron of species of fungiid corals with which it lives in commensal association. Material consisting of mucus, zooxanthellae, nematocysts, plankton and inorganic matter, is taken exclusively from the coelenteron. The very mobile foot possibly assists in food collection and in the removal of pseudofaeces; but, with large ctenidia, the bivalve is a typical ciliary feeder. Experiments with labelled zooxanthellae reveal that these are taken into the gut of Fungiacava with subsequent metabolic incorporation of products derived from them. The other prime source of food must be phytoplankton carried in with the feeding currents of the coral, itself carnivorous so that there is no competition for food between commensal and host. The Fungia zooxanthella– Fungiacava association operates as a "Troika" the productivity of which is autoregulated in proportion to the number of bivalves present. The inorganic wastes of the bivalve (as well as those of the coral) are utilized by the zooxanthellae, resultant increase in the algal component becoming available as food to the bivalve. Losses in the cycle are balanced by intake of exogenous food.     

Platelet-activating factor secreted by DDAVP-treated monocytes mediates von willebrand factor release from endothelial cells

We have previously shown that although DDAVP (1-deamino-8-D-arginine vasopressin), a synthetic analogue of the natural hormone arginine vasopressin, does not directly promote release of vWf from human umbilical vein endothelial cells (ECs), enhanced release does occur when ECs were exposed to either monocytes or to supernatants recovered from DDAVP-treated monocytes. In the present study, we have found that exposure of monocytes to DDAVP did not increase secretion of interleukins (IL)-1β, IL-6, IL-8, tumor necrosis factor (TNF-α), growth factors G-CSF (granulocyte-), GM-CSF (granulocyte, monocyte-colony stimulating factor), prostaglandins (PG) E₂, PGF_2α, or PGI₂ or purine nucleotides such as ATP and ADP. However, increased levels of platelet-activating factor (PAF) were secreted by DDAVP-treated monocytes in a time- and dose-dependent manner that positively correlated with the enhancement in vWf release from ECs. Moreover, this effect could also be elicited when lipid extracts of these supernatants or purified PAF were added directly to ECs. This response could be inhibited with (±)-trans-2,5-Bis(3,4,5-trimethoxyphenyl)-1,3-dioxolane, a specific PAF receptor antagonist, when the ECs were exposed to supernatants from DDAVP-treated monocytes or to pure PAF. The present data indicate that enhanced secretion of PAF from monocytes is one mechanism whereby DDAVP can provoke release of vWf from ECs. © 1993 Wiley-Liss, Inc.     

Sulfated oviductal glycoproteins in the rabbit: quantitation by competitive enzyme-linked immunosorbent assay

The rabbit oviductal epithelium synthesizes and secretes a family of antigenically related, sulfated oviductal glycoproteins (SOG). Anti-SOG monoclonal antibodies (Mabs) were produced and two (Mab 1 and Mab 2) were selected for further characterization. Periodate oxidation of Western blots of oviductal fluid did not affect the binding of Mab 1 or Mab 2, thus suggesting that these antibodies recognized protein rather than carbohydrate epitopes on SOG. The specificity of Mab 1 was determined by Western blot analysis of tissues obtained from estrous rabbits and from the male rabbit reproductive tract. SOG was identified in tissue extracts of both the oviductal ampulla and isthmus. Cervix was the only non-oviductal tissue with which Mab 1 cross-reacted. Mab 1 was used to isolated SOG from whole oviductal fluid by immuno-affinity chromatography. Affinity-purified SOG and Mab 1 were used to develop a quantitative, SOG-specific, competitive enzyme-linked immunosorbent assay. This assay was used to quantify SOG in rabbit oviductal fluid collected during estrus and pseudopregnancy. SOG secretion during pseudopregnancy was resolved into two transient episodes of increased secretion. Maximum SOG secretion (X = 1039 +/- 199 micrograms/day) occurred within 48 h of the induction of pseudopregnancy. A second period of enhanced SOG secretion (X = 308 +/- 46 micrograms/day) occurred during the fifth and sixth days of pseudopregnancy. Baseline SOG secretion occurred during estrus at approximately 60% of maximum postovulatory secretion.     

Effectiveness of chemical mutagenesis in multiple damage to one or both strands of plasmid DNA

Methods for site-directed multiple modification of DNA have been developed and used for modification of either one or two strands of plasmid DNA. Plasmid DNAs modified in the region of the tet gene were transformed into Escherichia coli cells and Tet colonies were screened. It was shown that multiple lesions in one DNA strand performed using either N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or sodium bisulfite were effectively repaired in the cell by error-free mechanism. In contrast, modification of two DNA strands led to induction of mutations. The efficiency of mutagenesis in the case of modification of a local region of one DNA strand with sodium bisulfite and modification of the other strand with MNNG was 1.1-7.9%. Mutations were analysed by restriction mapping and sequencing. All of them were G----A transitions.     

Interleukin-1 beta induces the production of an L-arginine-derived relaxing factor from cultured smooth muscle cells from rat aorta.

The effect of interleukin-1 beta on the production of non-prostanoid vasoactive factors by cultured rat aortic smooth muscle cells was investigated. Under bioassay conditions, the perfusate from a column of confluent cells grown on beads and treated with interleukin-1 beta (1 ng/ml for 18 to 24 hr) abolished the contraction of a canine coronary ring without endothelium contracted by phenylephrine (1 microM), while the perfusate from control cells had no effect. The relaxing activity of the perfusate was observed when transit times were increased from 1 sec to 5 min. Nitro L-arginine (100 microM) reversed the relaxations and L-arginine stereoselectively restored the relaxations. Interleukin-1 beta (1 ng/ml) evoked a time-dependent accumulation of cyclic GMP but not cyclic AMP in cultured smooth muscle cells. The transfer of fresh or stored (-70 degrees C) conditioned culture medium from interleukin-1 beta-treated cells but not from control cells, to cultured smooth muscle cells stimulated the production of cyclic GMP. These observations demonstrate that interleukin-1 beta induces the production of transferable factor which relaxes vascular smooth muscle and stimulates the production of cyclic GMP.