Characterization of chicken CD4-expressing cells.
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Animals↗
Biomedical subjects
O Lassila
Publications and source records attributed to O Lassila.
Can B cells turn on virgin T cells?
The first event in the initiation of an immune response is the capture and presentation of antigen to T cells. Such presentation involves two distinct steps: (1) display of the antigen, which requires uptake, processing and re-expression of the antigen in association with MHC molecules on the presenting cell surface; and (2) triggering, in which the presenting cell provides signals leading to the activation of the responding T cell. Two sorts of cells can capture antigens, the 'professional' antigen-presenting cells (APCs) such as dendritic cells and macrophages, and the B cells. Both types of cells can display antigens and the APCs are known to be able to trigger resting T cells. But despite in vitro evidence that certain B-cell types can reactivate previously-activated T cells, it is not yet clear whether a B cell can initiate an immune response by providing the signals necessary to activate a resting T cell. We reasoned that resting B cells should not have this capacity because of the problems this would present with tolerance to self idiotypes. By exploiting the unique properties of the avian haematopoietic system, we have examined the presenting capacity of B cells in vivo and found that resting B cells are indeed unable to activate resting T cells.
Animals↗
Immunoglobulin diversification in embryonic chicken bursae and in individual bursal follicles.
Previous studies have shown that the same immunoglobulin (Ig) V lambda gene (V lambda 1) is rearranged in all chicken B cells, and that extensive sequence diversification of this gene occurs during B cell development in the bursa of Fabricius. We used two-dimensional gel electrophoresis to compare the heterogeneity of Ig lambda light chains produced by B cells at different stages of bursal development. Somatically diversified light chains were observed in Ig molecules produced by bursal cells as early as 15 days of embryonic incubation. The two principal species of light chain observed probably represent glycosylated and nonglycosylated forms of lambda chain encoded by alleles of a single lambda gene. Extensive diversification was observed during late embryogenesis. We also studied lambda light chain diversity in cyclophosphamide-treated birds repopulated with normal bursal cells. In these birds, individual bursal follicles are repopulated by single B cell precursors. Follicular cells derived from single B cell precursors were able to produce a spectrum of light chains almost as diverse as that of the total bursal cell population. We used two monoclonal anti-idiotype antibodies to study idiotype expression in individual normal or reconstituted follicles. About 30% of follicles contained 0.1% to 5% of lymphocytes which reacted with one or both of the antibodies. The results indicate that within individual bursal follicles bursa stem cells undergo Ig hyperdiversification.
Animals↗
B cell precursors in chick embryos surgically bursectomized at 72 h of incubation.
To determine the presence of precursor B cells in chick embryos surgically bursectomized at 72 h of incubation (E-Bx) we studied chick chimeras that were produced by establishing parabiotic connections between blood vessels of chorioallantoic membranes of normal and surgically bursectomized chick embryos. Using sex chromosomes and a B cell alloantigen (Bu-1a) as markers we showed that chick embryos bursectomized at 72 h of incubation contain B cell precursors capable of colonizing the bursa of Fabricius and developing into B lymphocytes. The repopulation capacity of 14-day-old embryonic spleen cells from E-Bx recipients was tested by transferring them into age-matched X-irradiated Bu-1-disparate embryos. The results show that B cell precursors are present in 14-day spleen of chick embryos bursectomized at 72 h of incubation. These precursors carry the Bu-1 B cell alloantigen, suggesting that commitment to the B cell lineage can take place in the absence of bursa.
Age Factors↗
Attempt to detect recombination between B-F and B-L genes within the chicken B complex by serological typing, in vitro MLR, and RFLP analyses.
In search for recombinants within the chicken major histocompatibility B complex, 1155 animals from crosses between the congenic lines CB (B12) and CC (B4) were tested with alloantibodies and monoclonal antibodies for the B-F (class I), B-L (class II), and B-G (class IV) antigens and by mixed lymphocyte reaction. The absence of detectable recombination was confirmed by restriction fragment length polymorphism analysis with B-L beta and B-F probes. Together with previous reports, this indicates that the distance between the B-F and B-L loci is below 0.01 centimorgan.
Animals↗
B-cell commitment and diversification in the bursa of Fabricius.
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Animals↗
A re-evaluation of the function of the bursa of Fabricius.
We will briefly outline mammalian B cell ontogeny to provide a comparison with the avian model. The earliest defined stage of mammalian B cell development is the pre-B cell (itself derived from a multipotent stem cell) which expresses heavy chains within the cytoplasm and is a large, rapidly dividing cell. This cell drops out of division, reduces in size and, over about 24 hours, rearranges Ig light chain V region gene. As a consequence of light chain rearrangement and expression, intact IgM is expressed on the cell surface and the cell leaves the bone marrow as a small virgin B lymphocyte with the capacity to respond to antigen (Opstelten and Osmond 1983). This pathway occurs throughout the life of the animal and so there is constant production of B cells from the bone marrow which are recently derived from sIg- precursors, which have, in turn, recently rearranged their Ig V region genes. Thus there is a constant influx of new V region gene combinations into the peripheral mammalian B cell pool. Furthermore, the mammalian B cell repertoire is very large with estimated germ line diversity approaching 5 X 10(7) as a consequence of various germ-line V region recombinations (Honjo 1983). The pre-bursal stem cell is present in the periphery of the embryo from before day 8 to about day 16 - 17 of embryonation. Most colonisation of bursal follicles probably occurs prior to day 13. Individual bursal follicles are populated by a low number (2-3) of precursor cells. These rapidly become committed for the expression of particular V region genes, probably as a consequence of productive V region recombination. This recombination is restricted to the embryo and may not require the bursal microenvironment for its induction. The available germ-line repertoire of heavy and light chain V region in the chicken is very limited and little diversity can be generated from this initial recombination event (see Weill et al 1986). By day 12 of embryonation, sIg+ cells are present in the bursa and from this time sIg+ cells rapidly divide within the medulla of the bursal follicle. By about day 18 of embryonation, there are no pre-bursal cells in the periphery and sIg+ cells begin to seed from the bursa. It is from about this time that the cortex appears within the bursal follicle and it is tempting to suggest that cells migrate from the medulla to the cortex where further cell division occurs prior to export into the periphery.(ABSTRACT TRUNCATED AT 400 WORDS)
Animals↗
Avian B cell precursors: surface immunoglobulin expression is an early, possibly bursa-independent event.
The avian bursa of Fabricius contains about 1 X 10(4) discrete follicles, each of which is colonized by a small number of lymphoid progenitor cells during embryonic life. We have previously shown (J.R.L. Pink et al., Eur. J. Immunol. 1985. 15:617) that all, or almost all B cell progenitors in the bursae of 4-day-old chicks express cell surface IgM. In this report, we have analyzed the distribution of cell surface (s)IgM-1 allotypes within individual follicles of (M-1a/M-1b) allotype heterozygous birds. Although the majority of follicles contained a mixture of sIgM-1a+ and sIgM-1b+ cells, a significant proportion of isolated follicles contained exclusively sIgM-1a+ or sIgM-1b+ cells. Statistical analysis of the frequency of such "M-1a" and "M-1b" follicles demonstrated that the sIg+ B cells in the bursae of 4-8-week-old birds are derived from 2-4 allotypically committed precursor cells per follicle. Since we have previously shown that each bursal follicle is colonized by 2-5 pre-bursal stem cells, these cells must be committed to the eventual expression of one or other allotypic haplotype before they have undergone extensive proliferation within the bursa. In addition, we show that almost all B progenitor cells from the bursae of chicks which had been allotype suppressed as embryos were committed to synthesis of the nonsuppressed allotype, showing that this commitment was essentially complete at the time of suppression (i.e. before 19 days of incubation). Finally the bone marrow of 16-day embryos was used to reconstitute the bursal lymphocytes of cyclophosphamide-treated host embryos. Reconstitution was inhibited by anti-Ig antiserum indicating that most 16-day embryonic BM-derived bursal cell precursors also express sIgM. These results raise the possibility that expression of sIgM may be controlled by a "biological clock" rather than by any inductive capacity of the bursal microenvironment. Furthermore, these results provide further evidence that in normal birds a self-renewing sIg+ B cell population in the hatched chicken is the sole source of B cells in the adult.
Animals↗
Rearrangement of chicken immunoglobulin genes is not an ongoing process in the embryonic bursa of Fabricius.
We report a molecular analysis of the chicken Ig loci in single bursal follicles from 3- to 7-week-old chickens. Each follicle contained between 10(5) and 3 X 10(5) cells. The Ig gene rearrangement patterns obtained were compared to the pattern observed with the corresponding total bursal DNA. The results obtained for the light chain locus imply that a very small number (two on average) of rearrangement events takes place in each follicle. For the heavy chain locus similar results were obtained, each follicle showing a more restricted pattern than the total bursa. These data favor a model in which each follicle is colonized by a very few prebursal stem cells that are committed to a particular Ig gene rearrangement at the very beginning of the development of the embryonic bursa. The role of the bursa as the organ in which such a committed stem cell population for the B-cell lineage arises is discussed.
Animals↗
Sensitivity of human fetal hemopoietic cells to NK cell-mediated lysis.
Hemopoietic cells from human fetal livers at 11 to 18 weeks of gestational age were studied as to whether they were sensitive to adult and autologous fetal NK cells. Cell line K562 was used as control target in the 51Cr release assay and in the cold target competition assay. Adult peripheral blood NK cells did not kill liver-derived hemopoietic cells of four fetuses tested. Neither did fetal liver cells, whether or not expressing spontaneous cytotoxicity against K562 cells, destroy autologous hemopoietic liver cells. Hemopoietic liver cells from six fetuses were studied in the cold target competition assay. Liver cells of two fetuses clearly competed with lysis of K562 cells by adult NK cells. Thymus cells from one 18 week-old fetus had a weak capacity to compete with lysis of K562 cells by adult cells, whereas cells from the spleen and liver of the same fetus did not. It is concluded that early fetal liver hemopoietic cells are not susceptible to lysis by NK cells but an interaction between fetal cells and NK effector cells results in a depressed NK cell activity against NK-sensitive target cells.
Cytotoxicity, Immunologic↗
Mott cells are plasma cells defective in immunoglobulin secretion.
Plasma cells containing intracellular inclusions of immunoglobulin (Russell bodies) are known as Mott cells, and are found in large numbers in lymphoid organs in autoimmune mice. Hybridoma technique was used to produce cell lines of this phenotype by fusing spleen cells from a NZB mouse with a nonproducing hybridoma cell line (Sp2/0-Ag14), allowing us to carry out studies of this cell type at the biochemical level. Ultrastructurally the inclusions were distended cisternae of rough endoplasmic reticulum, suggesting a block in the secretory pathway of the cells. Biosynthetic labeling studies confirmed that these cell lines have either a complete or partial block of secretion of immunoglobulin, possibly due to an abnormal light chain.
Animals↗
Micromethod for determination of antibacterial capacity in amniotic fluid.
A new method for assessing antibacterial activity in amniotic fluid (AF) is presented. Sample sizes of 100 microliter were incubated at 37 degrees C in an equal amount of growth medium with inoculated bacteria. The quantification of bacterial growth was established by measuring the change in optical density with an automated spectrophotometer. In this study, Escherichia coli, type K 12, was used for inoculation and the bacterial growth was assessed after 6, 18, 30, 42, and 54 hours' incubation. The variation coefficients in intra-assay and interassay measurements were 3.8 and 5.2%, respectively. Shaking of the incubated sample significantly changed the results by increasing the number of viable cells in the most probable number method and decreasing the optical density difference in the spectrophotometric method. Nevertheless, the correlation between the results in both methods was good (r = 0.94, p less than 0.05, and r = 0.98, p less than 0.01, respectively). By filtering the AF sample before incubation, a great deal of antibacterial activity was removed. The reproducibility, simplicity, and rapidity of this spectrophotometric method of assessing antibacterial activity in AF may make it a useful clinical tool.
Amniotic Fluid↗
Natural killer cell activity of human fetal liver cells after allogeneic stimulation.
Cells isolated from the liver of human fetuses were confronted with mitomycin C-treated adult allogeneic cells. After this mixed leukocyte culture (MLC)-type reaction, the cytotoxic activity of fetal cells was tested against K562 cell line in a 4-h 51Cr release assay. Three of the seven fetuses tested (8 to 11 weeks of gestational age) expressed marginal cytotoxic activity before cultivation. Cells from one 8-week-old and one 9-week-old fetus were slightly more cytotoxic when cultured in the presence of allogeneic cells than when cultured in the medium only. Production of gamma interferon (IFN-gamma) was not detected in these cultures. Cells of one 18-week-old fetus expressed strong cytotoxic activity against K562 cells after MLC. Thymocytes from the same fetus were not cytotoxic, either before or after MLC against K562 cells. The results indicate that the 'prethymic' human liver contains cytotoxic cells able to spontaneously kill natural killer (NK)-sensitive target cells. Generation of strong NK-like cytotoxicity from noncytotoxic precursor cells was observed only after the thymus becomes lymphoid, suggesting that thymus-processed cells may regulate the generation of NK-like cytotoxic activity. The results suggest a different ontogeny of spontaneous and MLC-induced NK-like cells in the human fetus.
Cell Division↗
Monoclonal antibodies to Bacteroides fragilis lipopolysaccharide.
Monoclonal antibodies (MoAbs) to the lipopolysaccharide (LPS) of Bacteroides fragilis were produced by immunizing mice before hybridization with bacterial outer membranes solubilized with Triton X-100. Nineteen stabile clones were established. They all produced antibodies that reacted more strongly with purified B. fragilis LPS than with crude sonicated antigen in an enzyme immunoassay. Four MoAbs were studied by immunoblotting and enzyme immunoassay inhibition. Immunoblotting confirmed that the target of the MoAbs was LPS. Marked and homogeneous staining occurred in the immunoblotting with both purified LPS and outer membranes in the molecular weight range of 8,000 to 27,000. In enzyme immunoassay inhibition, MoAbs reacted positively with 93 to 96% of B. fragilis strains, including prototype strains ATCC 23745 and NCTC 9343. Within the B. fragilis group, the MoAbs reacted positively with two of five B. ovatus strains and two to six of nine B. thetaiotaomicron strains. No marked cross-reactivity with other bacteria was observed. These results confirm earlier findings that the B. fragilis LPS contains an immunodominant antigenic determinant common to almost all B. fragilis isolates.
Animals↗
Immune function in patients with ovarian carcinoma treated with irradiation or with cytostatics.
The immunocompetence of 25 patients with ovarian cancer was followed up during and after radiation or chemotherapy, by using the number of lymphocytes, E-rosette-forming cells, and mitogen responses as parameters. In comparison with chemotherapy, the irradiation caused a profound and permanent immunosuppression. In patients treated with chemotherapy, the number of E-rosette-forming cells and the mitogen responses were fewer in the patients who developed recurrence or dissemination.
Adult↗
Thymus-dependent immune functions in chickens bursectomized with colchicine applied to the anal lips.
Thymus-dependent immune functions were investigated in chickens bursectomized neonatally with colchicine solution given per anum. Antibody responses to thymus-dependent antigens sheep red blood cells (SRBC) and human gamma-globulin (HGG) were delayed, reaching the normal level after the third antigen stimulation. Also the mitogenic responses of peripheral blood lymphocytes were preserved, and no changes in the thymic morphology were found. In contrast, antibody responses to bursa-dependent antigen Brucella abortus were low and the switch of immunoglobulin isotypes from IgM to IgA and IgG was disturbed. It can be concluded that neonatal bursectomy with cloacal administration of colchicine does not significantly affect T cell functions, whereas B cell functions are partially deficient.
Animals↗
Characteristics of chicken intraembryonic cells that express B-L (Ia-like) antigen under the influence of cultured bursal epithelium.
In-vitro-cultured bursal epithelium (BE) and BE-conditioned medium (BECM) induce B-L antigen on chicken intraembryonic cells. Cells prepared from 9-day-old intraembryonic mesoderm were fractionated in accordance with cell size by linear albumin gradient sedimentation at 1 g. Two cell types could be distinguished on which expression of the B-L antigen was altered after a 6-h incubation with the bursal epithelial component. One fraction contained small mononuclear cells with low sedimentation velocity (less than or equal to 3 mm/h) and low spontaneous proliferation activity. These cells responded strongly to BECM and showed a slight but not significant response to BE (index after incubation with BECM 3.8, with BE 1.7, as compared with RPMI medium control). The other fraction was composed of large mononuclear cells with sedimentation velocity greater than 9 mm/h and with high spontaneous proliferation. These cells showed a response of equal magnitude to both BE and BECM (index after incubation with BE 2.0, with BECM 2.3). These results suggest that the bursa of Fabricius has influence on two different cell types: a large, probably primitive undifferentiated cell, responding equally to bursal cellular contacts and BE culture supernatant, and a small mononuclear cell type, probably more mature, responding more clearly to the bursal humoral factor.
Animals↗