Some properties of the cell-free and cell-associated hemolysins of Escherichia coli were studied. Several strains of E. coli that were isolated from intestines of pigs with edema disease produce large quantities of cell-free hemolysin when grown in the presence of an extract of meat. The component of meat that stimulates production of cell-free hemolysin is not extracted by lipid solvents and is not dialyzable. The cell-free hemolysin is an acidic substance that occurs in two forms. It is inactivated by trypsin but not by lecithinase, lysozyme, ribonuclease, or deoxyribonuclease, shows optimum activity between pH 7 and 8, and requires calcium ion for activity. It does not appear to be an enzyme. The kinetics of the lytic reaction are most consistent with the hypothesis that one molecule of cell-free hemolysin is sufficient to lyse one erythrocyte and that it is inactivated in the lytic reaction. The cell-free hemolysin does not sufficiently damage the cell during the prelytic period to cause lysis after the hemolysin-calcium-erythrocyte complex has been disrupted. The cell-associated hemolysin was not separated from the cell by autolysis, freezing, sonic treatment, or treatment with trypsin or lysozyme. It appears to be closely associated with the metabolic status of the cell. Organisms that are highly hemolytic under usual conditions of assay immediately lose most of their hemolytic capability in the presence of sodium cyanide, streptomycin, nalidixic acid, and rifampin.

Establishment of pregnancy in cattle has been proposed to depend on production of a conceptus protein, bovine trophoblast protein-1 (bTP-1), which has a high degree of sequence homology with bovine interferon-alpha (bIFN-alpha), especially the alpha II subfamily. A preparation of bovine conceptus secretory proteins enriched for bTP-1 has antiviral and physico-chemical properties similar to other bIFN-alpha. Antiviral activity is initially detectable in uterine flushings on Day 14 of pregnancy, when the conceptus measures 4-5 mm in length, and increases as the conceptus elongates through Day 18. Day 17 conceptuses produce more than 10(6) U antiviral activity during 24 h of culture. All IFNs induce the enzyme 2',5'-oligoadenylate synthetase, which catalyzes production of 2',5'-oligo(A), which in turn is involved in antiviral and growth inhibitory effects of IFNs. This enzyme activity is induced in Madin-Darby bovine kidney cells by the partially purified bTP-1 preparation similarly to IFN-alpha, -beta, and -gamma. Likewise, the partially purified bTP-1 and bIFN-alpha 1 induce 2',5'-oligo(A) synthetase activity in monolayers of endometrial epithelial and stromal cells. Compared to epithelial cells, stromal cells have higher baseline activity of 2'-5'-oligo(A) synthetase activity (p less than 0.01) and show a greater degree of induction in the presence of either the partially purified bTP-1 or bIFN-alpha 1 (p less than 0.01). Also, 2',5'-oligo(A) synthetase of endometrial stromal cells is induced to a greater degree by our enriched bTP-1 preparation than by bIFN-alpha 1 (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

THE cell-free haemolysin produced by growing certain strains of Escherichia coli on meat-broth medium was originally described by Lovell and Rees (1960) and has been termed a-haemolysin by Smith (1963). Since there have been a number of reports describing other filterable haemolysins (Snyder and Koch, 1966;Walton and Smith, 1969;Muranyi and Juhasz, 1971;Rennie and Arbuthnott, 1971) which might be confused with the a-haemolysin studied in these experiments, the properties that distinguish a-haemolysin from the others produced by E. coli are summarised here. A second type of filterable cell-free haemolysin, first described by Snyder and Koch (1966), differs from a-haemolysin in that it is produced on chemically defined media and is thermostable at 56°C. It is produced later in the growth phase than is a-haemolysin and its appearance is accompanied by a fall in the pH of the culture. A similar type of haemolysin, described by Rennie and Arbuthnott (1971), is produced on chemically defined medium in the presence of various carbohdyrates. It is possible that the haemolytic effects described in both of these reports may be due to the lytic effect of organic acids formed from the carbohydrates in the culture medium, as reported in related studies by Muranyi and Juhasz (1971).Walton and Smith (1969) have described a third type of filterable haemolysin, which they termed y. This haemolysin is found in mutants resistant to nalidixic acid and either its production or its release appears to be affected by nalidixic acid.The fourth haemolysin produced by E. coli is the cell-associated or /3-haemolysin. 8-Haemolytic activity has not been dissociated from intact cells and appears to be dependent on cellular metabolism (Smith, 1963; Short and Kurtz, 1971).It is not possible to determine which of these four types of haemolysin are produced by various strains of E. coli on the basis of the appearance of colonies on blood agar plates, since both a-and p-haemolysin-producing strains give rise to a clear zone of haemolysis (Smith, 1963). Smith found that 53-76% of the faecal samples from healthy pigs, sheep, and oxen contained haemolytic E. coli. He found that 63-92% of the haemolytic strains

Administration of estrogen to gilts on Days 9 and 10 of pregnancy results in total embryonic loss by Day 18. The present study examined changes in the uterine endometrial surface and secretion during conceptus attachment in control and estrogen-treated (Days 9 and 10) pregnant gilts. Gilts were unilaterally hysterectomized on either Days 12 and 14 or Days 16 and 18 of gestation. Uterine horns were flushed with saline and conceptuses were evaluated. Intact conceptuses were recovered from all control gilts, whereas estrogen-treated gilts contained normal intact conceptuses only on Day 12 of gestation. Antiviral activity, which reflects conceptus viability, was reduced (p less than 0.01) in uterine flushings after Day 14 in estrogen-treated gilts. Culture of endometrial explants with [3H]glucosamine revealed several glycoproteins that are synthesized during the period of conceptus attachment; however, no difference in glycoprotein synthesis between treatment groups was detected by analysis with two-dimensional PAGE and fluorography. Analyses of the uterine epithelium by scanning and transmission electron microscopy demonstrated that estrogen administration caused an alteration in the uterine surface, a thinning of the uterine epithelial glycocalyx, and a reduction of cationic ferritin binding to the microvilli of the uterine epithelium. Results indicate that conceptus mortality after early administration of estrogen is associated with alterations in the uterine endometrial surface during the period of conceptus attachment in the pig.

The signal for maternal recognition of pregnancy in cattle is bovine trophoblast protein-1 (bTP-1), a Type I trophoblast interferon. One of the many functions of interferons is the induction of the 2',5'-oligoadenylate (2-5[A]) system, which is involved in cell division and selective degradation of mRNA. The present study focused on the cellular changes of 2-5(A) synthetase in bovine endometrium during the estrous cycle and early pregnancy. Ability of recombinant bTP-1 to stimulate activity of the enzyme in uterine cells throughout the estrous cycle was also evaluated in vitro. Surface epithelium, glandular epithelium, and stroma were enzymatically separated in cows on Day 5, 10, 15, or 18 postestrus and on Day 15 or 18 of pregnancy. Cell samples were lysed and frozen for determination of the endogenous cellular content of 2-5(A) synthetase. Additional cells obtained during the estrous cycle were cultured and treated with increasing doses of recombinant bovine interferon alpha (rbIFN-alpha) or recombinant bTP-1 (rbTP-1). During the estrous cycle, 2-5(A) synthetase was greatest on Day 5 and declined approximately 10-fold by Day 15 (p < 0.01). Cellular content of 2-5(A) synthetase was similar among all three endometrial cell types. In the gravid horn of pregnant animals, presence of a conceptus significantly increased (p < 0.01) 2-5(A) synthetase in all endometrial cell types compared to levels on Days 15 and 18 of the estrous cycle. On Day 18, levels of 2-5(A) synthetase were 30-fold greater (p < 0.01) in epithelium (surface and glandular) from pregnant cows compared to that from cyclic cows.(ABSTRACT TRUNCATED AT 250 WORDS)