Differential effects of polyadenylic: polyuridylic acid and lipopolysaccharide on the generation of cytotoxic T lymphocytes.

In a mixed leukocyte culture (MLC) reaction of allogenic mouse spleen cells differing for H-2K or H-2D, only a weak cytotoxic response is generated. This cytotoxic response is augmented significantly if bacterial lipopolysaccharide (LPS), 5 microgram/ml, or polyadenylic acid (poly A):polyuridylic acid (poly U), 20 microgram/ml, is present in the culture. The cytotoxic cells generated in the presence of these two agents are specific for sensitizing H-2K or H-2D antigen. Two lines of evidence suggest that these two agents exert their effect at different steps in the development of cytotoxic lymphocytes: (a) the effect of poly A:U depends on the presence of adherent cells, whereas the effect of LPS is independent of the presence of adherent cells and (b) LPS promotes the development of cytotoxic cells when ultraviolet light-treated stimulating cells are used in the MLC whereas poly A:U does not.

Chloral hydrate: a solvent for biological membranes.

A buffer system conitainin chloral hydrate, taurine, and bromopyridinium lactate was used to dissolve several biological membranes and separate their protein components by polyacrylamide gel electrophoresis. This solvent system was capable of separating molecules of similar size on the hasis of their charge and allows easy recovery of the proteins Thus. aqueous chloral hydrate is an effective solvent for biological membranes.

Effect of liposomes on lymphocytes: induction of proliferation of B lymphocytes and potentiation of the cytotoxic response of T lymphocytes to alloantigens.

Liposomes of certain lipid composition prepared by the detergent removal method (Brunner, J. et al., Biochim. Biophys. Acta 1976. 455: 322) induced the proliferation of spleen cells from different mouse strains. Spleen cell populations enriched in B lymphocytes and those obtained from nude mice were induced to proliferate, whereas spleen cell fractions enriched in T lymphocytes and thymocytes were not. The mitogenic effect of liposomes resembled that of lipopolysaccharide (LPS) and it depended upon their lipid composition. Liposomes prepared from dimyristoyl lecithin (DML), 2:1 dimyristoyl lecithin:cholesterol (DML:C), 2:1 dioleoyl lecithin: cholesterol (DOL:C), and 2:1 egg yolk lecithin:cholesterol (EYL:C) were mitogenic, whereas liposomes prepared from egg yolk lecithin (EYL) alone were not mitogenic for spleen cells. The mitogenic effects of these liposome preparations were in the decreasing order DML greater than DOL:C greater than or equal to EYL:C greater than DML:C greater than EYL. The results suggest a correlation between the membrane fluidity of liposomes and their mitogenic effect. Although no proliferative response was induced on T lymphocytes, two of these liposomes, DML and EYL:C, had the ability to potentiate the cytotoxic response of T lymphocytes to alloantigens in mixed leukocyte culture. In responder-stimulator combinations which differed for the H-2K, H-2D or the entire H-2 region, these liposomes potentiated the cytotoxic response significantly. The results suggest that liposomes have an ability to modulate T lymphocyte response.

Production of chemotactic activity in mixed leukocyte cultures: maximum effect caused by H-2I region disparity.

Primary mixed mouse leukocyte culture supernatants contain an activity chemotactic for mouse peritoneal exudate cells and it can be detected within 72 h after initiation of the culture. Disparity for H-21 region leads to maximum production of chemotactic activity whereas H-2K or H-2D region differences result in the production of significantly less activity. The rate of production of chemotactic activity follows closely the rate of incorporation of 3H-thymidine, and both attain the peak on day 4 after initiation of the culture. But whereas proliferation is sensitive to gamma-irradiation, chemotactic activity production is not. It is our hypothesis that proliferating cells are primarily responsible for the production of chemotactic activity. The possible relevance of chemotactic activity production to graft rejection is discussed.

Enhancement of movement of mouse peritoneal macrophages caused by colchicine treatment.

Colchicine in the concentration range of 10(-8) to 10(-4)M enhances the movement of mouse peritoneal macrophages while at higher concentrations it inhibits movement. Maximum movement is observed in the concentration range of 10(-6) to 10(-4)M.

Characteristics of a low-molecular-weight factor extracted from mouse tumors that affects in vitro properties of macrophages.

A dialysable low-molecular-weight factor capable of affecting in vitro properties of macrophages was extracted from four different mouse tumors. This factor not only modulates closely related properties of peritoneal macrophages such as spreading and migration but also inhibits lipopolysaccharde-induced tumoricidal activity of these cells. It can be extracted not only from tumor tissues but also from tumor cells grown in vitro. The appearance of this factor is unique to tumors and it is not present in detectable quantities in normal tissues. The factor from one of the tumors, Lewis lung carcinoma, was purified extensively and the partially purified factor retains all the above effects on macrophages. It is not sensitive to pronase or a mixture of bovine spleen phosphodiesterase II, E. coli alkaline phosphatase and pancreatic ribonuclease. The factor is lipid-like in character and it is soluble in both organic solvents and aqueous media. It has ionizable group(s) and is anionic at neutral pH but non-ionic under acidic conditions.

Effects of prostaglandins on the spreading, adhesion and migration of mouse peritoneal macrophages.

The effects of prostaglandins on the in vitro properties of mouse peritoneal macrophages namely spreading, adhesion and migration were investigated. PGE1 and PGE2 inhibit the spreading and adhesion of complete Freund's Adjuvant induced peritoneal macrophages significantly at concentrations of 1 ng per ml and above whereas they enhance the migration of these cells at concentrations of 100 ng per ml and above. PGA2 and PGB2 are less potent as they inhibit spreading and adhesion only at a concentration of 1 microgram per ml. At his concentration PGB2 enhances migration whereas PG2 has no effect. PGF 2alpha has no effect on the spreading, adhesion and migration of macrophages in the concentration range of 0.1 ng to 1,000 ng per ml.

Estrogen action in rooster liver. Modification of a tRNASer-inactivating activity.

The hepatic synthesis of the yolk protein, phosvitin, can be induced in rooster by the administration of estrogen and this provides a model for studying the regulatory mechanisms involved. Since phosvitin is unusually rich in serine the effect of estrogen on the metabolism of serine transfer RNA (tRNASer) was examined. Rooster liver contains an activity capable of inactivating rooster liver tRNASer. This activity has specificity in that it inactivates only part of the tRNASer in the unaminoacylated form. Upon estrogen administration this activity is modified with a change in specificity and the modified activity is able to inactivate only a significantly smaller amount of tRNASer than the unmodified activity. This novel mechanism may be involved in the regulation of some species of tRNASer and thus may be part of the translational control involved in estrogen-induced phosvitin synthesis.

Purification and functional characterization of a low molecular weight immune modulating factor produced by Lewis lung carcinoma.

Previously, we reported that mouse tumors contain a low molecular weight factor which is capable of affecting in vitro properties of macrophages. We now describe the purification to homogeneity of the only factor with macrophage modulating activity from one of these tumors Lewis lung carcinoma. The purified tumor factor appears to be a carboxylic acid and it retains all of the macrophage modulating properties which are characteristic of the unfractionated crude tumor extracts. This factor therefore reverses the spreading of macrophages, enhances the migration of macrophages out of capillary tubes and inhibits their chemotactic and tumoricidal activities. The purified tumor factor also inhibits immune responses of spleen cells. Specifically, proliferation of normal lymphocytes in response to mitogens and to alloantigens, as well as the generation of cytotoxic T lymphocytes in a mixed leukocyte culture, are suppressed when these cells are cultured in the presence of the purified tumor factor. The mixed leukocyte reaction of lymphoid cells obtained from mice which had been treated with the tumor factor is reduced compared to the response of lymphocytes obtained from normal mice. It is possible that there is a relation between production of this factor by tumor cells and survival of the tumor in a host.