Embryonic development in superovulated dairy cattle exposed to elevated ambient temperatures between Days 1 to 7 post insemination

Holstein heifers (n = 29) were used to determine whether thermal stress during the first 7 d of embryonic development may increase the incidence of embryonic abnormalities in dairy cattle. Heifers were acclimated to environmental chambers at 20 degrees C for 9 d and superovulated with follicle stimulating hormone-pituitary (FSH-P; 40 mg total), beginning on Days 9 to 11 of the estrous cycle. Prostaglandin F(2)alpha (Lutalyse; 50 mg total) was administered on Day 3 of FSH-P. Heifers were inseminated artificially at estrus and then maintained at either thermal neutrality (20 degrees C) or under hyperthermic conditions (daily exposure up to 16 h at 30 degrees C and 8 h at 42 degrees C) for 7 d beginning at 30 h after the onset of estrus. On Day 7 post estrus, embryos were recovered nonsurgically and evaluated morphologically for stage of development and quality. The distribution of embryos classified as normal, abnormal, retarded or as unfertilized ova, differed (P<0.001) between heat stress and thermoneutral treatments. Only 20.7% of 82 embryos recovered from stressed heifers were normal compared with 51.5% of 68 embryos from thermoneutral animals. Stressed heifers had a higher incidence of abnormal and retarded embryos with degenerate nonviable blastomeres. Responses indicated that thermal stress from 30 h after the onset of estrus to Day 7 post estrus increases the incidence of abnormal and retarded embryos in superovulated heifers.     

Conceptus development in large white and prolific Chinese Meishan pigs

Large White (LW) and Meishan (MS) gilts were killed on Days 8, 10, 11, 12, 14 and 30 of gestation. Mean diameters (mm) for MS and LW conceptuses, respectively, were: Day 8, 0.45 and 0.69; Day 10, 2.7 and 1.9; Day 11, 5.3 and 2.7, with the differences among days being affected by breed (P less than 0.01). Variation in diameter among conceptuses from LW gilts was greater (P less than 0.01) than that for MS gilts on Days 8-11, respectively: Day 8, 20 and 46%; Day 10, 29 and 38%; and Day 11, 22 and 44%. Conceptuses had elongated in 3 of 5 MS and 1 of 4 LW gilts on Day 11, 6 and 6 MS and 2 of 4 LW gilts on Day 12 and all gilts of both breeds on Day 14. These results indicate that conceptuses of MS gilts develop more rapidly and more uniformly between Days 8 and 14 of gestation. Overall, embryonic survival for Days 8-12 for gilts not having elongated conceptuses was 90.2% for MS and 73.2% for LW gilts (P less than 0.01). On Day 30 of gestation, embryonic survival was also higher (P less than 0.01) for MS (89%) than LW (55%) gilts. However, embryonic weight, crown-rump length, placental length, allantoic fluid volume, amniotic fluid volume, as well as total glucose, fructose and protein in allantoic fluid were not affected by breed. Placental weight was greater (P less than 0.01) for LW gilts. Uterine development at Day 30 of gestation, based on total length and weight of uterine horns, width of uterine horns, total endometrial surface area and total endometrial weight was greater (P less than 0.01) for LW gilts.(ABSTRACT TRUNCATED AT 250 WORDS)     

Secretory proteins of the bovine conceptus alter endometrial prostaglandin and protein secretion in vitro

The conceptus is believed to produce factors that regulate endometrial function and prevent luteolysis during early pregnancy. Endometrial tissues were collected from cyclic (n = 8) and pregnant (n = 2) cows at Day 17 post-estrus and cultured for 24 and 48 h with bovine conceptus secretory proteins (bCSP) (0%, 10%, 100%), where the amount of protein produced by a bovine conceptus during 24 h of culture is 100%. Incorporation of [3H]leucine into secreted proteins was determined and examined qualitatively by trichloroacetic acid precipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography. Levels of an intracellular endometrial inhibitor of prostaglandin synthesis were determined with a cotyledonary microsomal test system. Treatment with 10% and 100% bCSP reduced incorporation of [3H]leucine into secreted proteins. However, bCSP selectively induced two secreted proteins (13 and 10 kDa) from endometrium of cyclic cows. Prostaglandin F (PGF) secretion was decreased by bCSP treatment while prostaglandin E2 secretion was unaltered. An intracellular endometrial inhibitor of prostaglandin synthesis was induced by bCSP; synthesis of PGF by the cotyledonary prostaglandin-generating system was decreased when incubated with cytosol of endometrium treated with bCSP, but unaltered by cytosol from control tissues. In conclusion, products produced by the bovine conceptus are capable of regulating endometrial protein and prostaglandin biosynthesis in a fashion that could act to prevent luteolysis in vivo and provide endometrial secretory products for embryonic development.     

Proteins secreted by day-16 to -18 bovine conceptuses extend corpus luteum function in cows

Corpus luteum function, interoestrous interval and spontaneous uterine PGF-2 alpha (PGF) production were evaluated in 9 cyclic Holstein cows (3/group) after intrauterine injections of pooled conceptus secretory proteins, 5 beta-pregnan-3 alpha-ol-20-one, or homologous serum proteins on Days 15.5 through 21 after oestrus. A significant extension of corpus luteum lifespan and interoestrous interval were detected in cows treated with conceptus secretory proteins compared to the other 2 groups. CL lifespan and interoestrous interval were not different (P greater than 0.25) between 5 beta-pregnan-3 alpha-ol-20-one and control groups. Evaluation of spontaneous PGF responses suggested that proteins synthesized and secreted by the bovine conceptus accommodate luteal maintenance during early gestation via an attenuation of endometrial PGF production.     

Studies of structure-activity relationships of human interleukin-2

Human interleukin-2 (IL-2) has 3 cysteine residues; cysteines 58 and 105 form an intramolecular disulfide bridge, whereas cysteine 125 has a free sulfhydryl group. In this study, site-specific mutagenesis has been used to modify the cysteine residues of recombinant Escherichia coli-derived IL-2 (rIL-2) to evaluate the functional structure of IL-2. Substitution or deletion of cysteine 105 disrupted the disulfide bridge and yielded a mutant protein which was 8-10 times less active than wild type rIL-2. A similar modification at position 58, however, reduced the activity of rIL-2 by more than 250-fold. Although substitution of serine for cysteine 125 did not affect IL-2 activity, deletion of cysteine 125 or deletion of amino acids in the vicinity of this cysteine yielded mutant proteins with little, if any, activity. These results indicate that the protein structure in the vicinity of both positions 58 and 125 is more critical than that close to position 105. These findings may provide a clue to the understanding of the functional structure of human IL-2.     

Characterization of human interleukin 2 derived from Escherichia coli.

Interleukin 2 isolated from Escherichia coli cells expressing the human interleukin gene has been characterized. The observed properties of the protein have been compared with those properties which can be deduced from the DNA sequence alone and the published properties of natural human interleukin 2. The purified E. coli-derived interleukin 2 is a monomeric protein of Mr 15 000 with a sedimentation velocity of 1.86S. The amino acid composition of the protein and isoelectric point (7.7) are consistent with that part of the translated DNA sequence of the gene corresponding to the mature protein. A single disulphide bridge was identified between Cys-58 and Cys-105. C.d. suggested that interleukin 2 is predominantly alpha-helical in secondary structure. The E. coli-derived protein differed from natural interleukin 2 in the presence of N-terminal methionine and also in the absence of a carbohydrate moiety. Removal of the coding region for the first three amino acids of the natural interleukin 2 protein sequence (Ala-Pro-Thr) by site-specific mutagenesis resulted in a protein with N-terminal serine. The possibility that the specificity of the E. coli ribosomal methionine aminopeptidase may not recognize the sequence NH2-Met-Xaa-Pro is discussed (where Xaa is any amino acid residue).     

Codon usage in the vertebrate hemoglobins and its implications

A study of codon usage in vertebrate hemoglobins revealed an evolutionary trend toward elevated numbers of CpG codon boundary pairs in mammalian hemoglobin alpha genes. Selection for CpG codon boundaries countering the generally observed CpG suppression is strongly suggested by these data. These observations parallel recently published experimental results that indicate that constitutive expression of the human alpha-globin gene appears to be determined by regulatory information encoded within the structural gene. The possibility is raised that, in the absence of selection, CpG decay can be used to date the evolutionary origin of a mammalian alpha pseudogene from its active alpha gene.      

Evolution of crystallins: expression of lens-specific proteins in the blind mammals mole (Talpa europaea) and mole rat (Spalax ehrenbergi)

The mole (Talpa europaea; Insectivora) and the mole rat (Spalax ehrenbergi; Rodentia) both have degenerated eyes as a convergent adaptation to subterranean life. The rudimentary eye lenses of these blind mammals no longer function in a visual process. The crystallin genes, which display a lens-specific expression pattern, were studied in these blind mammals and in related species with normal eyes by hybridizing their genomic DNAs with probes obtained from cDNA clones for alpha A-, alpha B-, and beta Bp-crystallins from calf and gamma 3- crystallin from the rat. For all crystallin genes examined, the hybridization signals of mole and mole rat genomic DNA were comparable, respectively, with those of shrew and of rat and mouse, normal-vision representatives of the orders Insectivora and Rodentia. The expression of the crystallins at the protein level was tested by using antiserum specific for alpha-crystallin in immunofluorescence reactions on lens sections of mole and mole rat eyes and by using antisera against the beta- and gamma-crystallins on sections of the mole eye. All antisera gave positive fluorescence reactions exclusively with lens tissue of these blind mammals, indicating that the crystallins are still normally expressed despite the fact that these lenses have had no function in a visual process in these mammals for at least many million years. These findings apparently imply that some unknown selective advantage has conserved the crystallin genes and their expression after the loss of normal function of the lenses.      

Inhibition of uterine prostaglandin-F2 alpha production by bovine conceptus secretory proteins

The effect of bovine conceptus secretory proteins (CSP) on uterine prostaglandin (PG)-F2 alpha production was evaluated in dairy cattle following injection of estradiol-17 beta. Intrauterine injections of dialyzed serum proteins (Control, n = 5) or CSP (n = 5) were administered from days 15 through 18 post-estrus. Following intrauterine treatments on day 18, all cows were injected with E2 (3 mg) to stimulate uterine PGF2 alpha production. Plasma concentrations of progesterone (P4) and 15-keto-13,14-dihydro-PGF2 alpha (PGFM) were determined by RIA. The PGFM responses following E2 challenge were decreased (p less than 0.01) for cows receiving CSP versus serum proteins into the uterine lumen. Individual PGFM, P4 and cycle length responses are discussed. Data suggest that proteins secreted by the bovine conceptus suppress uterine PGF2 alpha production during pregnancy recognition in the cow.