Number Of Hemolytic Plaque-forming Cells Research Articles

Development of responsiveness and incidence of bispecific cells as revealed by in vitro assessment of the maturation of mouse bone marrow cells

The kinetics of the maturation of B cells were studied in adult thymectomized, lethally irradiated, and bone marrow-reconstituted mice. The spleen cells of the recipients were taken at various intervals after transfer and cultured in vitro with trinitrophenylated sheep erythrocytes (TNP-SRBC). The cultures were supplemented with either allogeneic culture supernatant or educated T-cell helper activity. Appearance of functional B cells in the bone marrow inoculum was assessed by the number of hemolytic plaque-forming cells (PFC) on the fourth day of culture. In a parallel series the frequency of surface Ig-bearing cells was determined by using fluorescent rabbit anti-mouse Ig serum. When helped by allogeneic culture supernatants, differentiating bone marrow cells showed a slower rate of maturation into functional B cells than when helped by specifically educated T cells. But in both cases the recovery of responsiveness reached the same level (number of PFC/10 6 cells) as that of normal spleen cells 55 to 60 days after transfer. During the initial periods of recovery, bispecific PFC (reacting both to TNP and to native SRBC determinants) were detected regularly in numbers far exceeding their frequency in normal spleen cell cultures; in some experiments, the number of bispecific PFC amounted to as much as 30% of the total PFC, whereas, when the bone marrow cells completely recovered (sixtieth day), the frequency of bispecific PFC was similar to that found in normal spleen cell cultures. The number of surface Ig-bearing cells also reached a normal level after the fiftieth day following transfer. In general, the degree of functional maturation was better correlated with the cells bearing surface Ig in the shape of rings or caps, whereas the predominance of spot-bearing cells indicated immaturity of the population.

Suppression of Graft-vs-Host Reaction by a Spleen Extract

Abstract In 1970, we reported the inhibitory effect of an alcohol extract of calf spleen (S4) on the in vitro proliferation of stimulated lymphocytes (1). The active molecule has been found to be a protein, with a molecular weight of 45,000 (2). Its inhibiting activity in vivo has been shown to be tissue specific and not species specific (1), fulfilling, therefore, the main criterion for a chalone (3, 4). The reduction of the number of hemolytic plaque-forming cells in the spleens of mice immunized with sheep red blood cells and the inhibition of in vitro lymphocyte transformation and proliferation induced by phytohemagglutinin (PHA) or histocompatibility antigens in the mixed lymphocyte culture (5, 6) suggested that this extract could exert an immunosuppressive action. Similar results have been obtained in our laboratory by Kiger in 1971 with a similar thymus extract (7) and later with thymus and spleen extracts by Houck et al. (8).

Cell components in the immune response: I. Gradient separation of immune cells

Cell suspensions from the spleens of normal mice were separated into four subpopulations by differential flotation on a discontinuous albumin gradient. Individual populations or combinations of populations were cultured under standard conditions in the presence of foreign erythrocytes and the number of hemolytic plaque-forming cells was measured at day 4. Marked cooperative effects were demonstrated in the size of the response when cells from the densest band were cultured with small numbers of cells from the lightest band. The dense band was shown to contain the precursor cells.

Cell components in the immune response: II. Cell attachment separation of immune cells

Cell suspensions from the spleens of normal mice were separated by differential attachment to glass beads and plastic surfaces. Individual populations or combinations of populations were cultured in the presence of foreign erythrocytes and the number of hemolytic plaque-forming cells were measured at day 4. Marked cooperative effects were demonstrated in the size of the response when nonattached cells were cultured with small numbers of attached cells. It was shown that precursor cells were present predominantly in the nonattached cell population, and the development of a population of antibody-forming cells by day 4 was increased in the presence of attached cells. Similar cooperative effects were demonstrated when attached cells were prepared from cells obtained from the peritoneal cavity. Such cells were inhibitory at higher concentrations.

An in Vitro Primary Immune Response to 2, 4, 6-Trinitrophenyl Substituted Erythrocytes: Response against Carrier and Hapten

Summary We have observed an in vitro response to the trinitrophenyl determinant (TNP) when spleen cell suspensions from unimmunized mice were cultured with the trinitrophenyl group coupled to sheep erythrocytes (TNP-SRBG) as immunogen. The response was measured by a progressive increase in the number of hemolytic plaque-forming cells assayed with TNP coupled to non-cross-reacting burro erythrocytes.

Antibody formation in mice infected with Salmonella typhimurium by primary immunization with sheep erythrocytes or bacterial cells.

Infection of mice with a strain of Salmonella typhimurium, known to multiply in macrophages, resulted in a marked alteration of their antibody response to further antigenic stimulation with sheep red blood cells (sRBC) or killed bacterial cells. The mice infected with a wild-type S. typhimurium and simultaneously immunized with sRBC, showed a significantly stimulated immune response to sRBC as revealed by the increased number of hemolytic plaque-forming cells in their spleens during the period following such immunization. Whether the animals were infected intraperitoneally or intravenously did not affect the stimulant effect of the infection on the immune response to sRBC. Injection of killed bacteria in amounts up to 106 organisms, instead of infection with live organisms, did not elicit any effective enhancement of antibody formation to sRBC. In cases of attenuated S. typhimurium infection, the stimulant effect on antibody formation to sRBC was obvious in the mice infected with the organisms simultaneously with or 1 week prior to the sRBC injection, while a slight suppression of antibody formation was observed in mice infected 3 weeks before the injection with sRBC. A similar effect of the infection was observed when the serum anti-bacterial antibody response was examined after stimulation with bacterial antigens. The altered immune responses after Salmonella infection are discussed in relation to the adjuvant effect of bacterial endotoxins and also to the acquired immunity to mouse typhoid.

Decrease in number of mitoses from precursers of cells synthesizing antibodies after immunisation of mice thymectomized at birth

It appears that in thymectomized CF 1 mice the number of hemolytic plaque-forming cells is very diminished, whereas the number of precursor cells is normal. This finding suggests that in thymectomized mice precursor cells the mitosis number is lower.