Response of mature unprimed CD8+ T cells to Mlsa determinants.

Contrary to existing dogma, evidence is presented that proliferative responses of mature unprimed T cells to Mlsa antigens involve CD8+ cells as well as CD4+ cells. The response of CD8+ cells to Mlsa antigens proved to be heavily dependent on help from CD4+ cells, and responses were stronger in three I-E+ strain combinations than in an I-E- combination. In I-E+ combinations, CD8+ blast cells accounted for 20-25% of the blasts generated from unseparated T cells responding to Mlsa-bearing stimulator cells in vitro; similar findings applied to blast cells generated in vivo. The observation that the majority (greater than or equal to 50%) of Mlsa-stimulated CD8+ cells (and CD4+ cells) were V beta 6+ indicated that CD8+ cells respond to Mlsa antigens, per se, rather than to nonspecific stimuli. Whether CD4+ and CD8+ cells use the same or different H-2-restricting elements to respond to Mlsa antigens has yet to be resolved.

Exclusion of circulating T cells from the thymus does not apply in the neonatal period.

Although T cells arise in the thymus, migration of mature postthymic T cells back to the thymus is very limited in adult mice and is restricted to activated cells. In neonates, by contrast, we present evidence that circulating CD4+ and CD8+ T cells with a naive/resting phenotype readily enter the thymus after intravenous injection and remain there for prolonged periods. The migration of resting T cells to the neonatal thymus is largely limited to an unusual subset of cells which lacks expression of the lymph node homing receptor, leukocyte-endothelial cell adhesion molecule 1 (LECAM-1) (MEL-14). Migration of mature T cells to the thymus in neonates may be important for self-tolerance induction.

Restricted tissue distribution of Mlsa determinants. Stimulation of Mlsa-reactive T cells by B cells but not by dendritic cells or macrophages.

Evidence was sought on the tissue distribution of Mlsa determinants, a class of cell-associated non-H-2 alloantigens that is highly immunogenic for unprimed T cells. Whereas normal CD4+ T cells and an Mlsa-reactive T hybridoma gave strong responses to Mlsa-positive stimulator populations containing Ig+ B cells, anti-Mlsa responses to B-depleted stimulators were almost undetectable. The B-depleted stimulators tested included Thy-1- spleen cells from mu-suppressed mice (mice treated with anti-mu antibody from birth) and J11d- preparations of spleen dendritic cells (DC) and peritoneal macrophages (M phi) from normal mice. Each of these populations was strongly immunogenic for allo-H-2-reactive T cells. The failure to detect Mlsa determinants on Ig- APC, i.e., M phi and DC, suggests that Mlsa determinants are not typical H-2-associated peptides. The data are more compatible with a model in which Mlsa determinants represent (or form part of) an integral cell membrane molecule expressed largely, and perhaps exclusively, on B cells. T cells might recognize these molecules only in native form, "processed" Mlsa determinants being nonimmunogenic. Consistent with this possibility, no evidence was found that Mlsa-negative B cells could absorb Mlsa determinants from Mlsa-positive B cells in a chimeric environment.

Susceptibility of mice to group B coxsackie virus is influenced by the diabetic gene.

A positive correlation was found between genetic predisposition to diabetes in the mouse and susceptibility to group B Coxsackie virus in this host. Male mice of the inbred strain C57BL/Ks and the following genetic variants were used; mice homozygous for the autosomal recessive gene for diabetes (db/db), the phenotypically normal heterozygous (db/+), and the normal mice which lacked the diabetic gene (+/+). The mortality response of the +/+ mice to intraperitoneal inoculation with Coxsackie virus B4 differed from the response of the two genetic variants (db/db and db/+) derived from this strain. The db/+ variant was more susceptible to Coxsackie virus B4 than the parental background strain (+/+). The db/db variant was more susceptible than either of the other genotypes. Pathological findings of the pancreas of the three genotypes during the acute stage of infection closely paralleled the genotypically dependent susceptibility of the host.

Negative selection in vivo reveals expression of strong Mls determinants in mice with X-linked immunodeficiency.

Evidence is presented that mice with X-linked immunodeficiency (xid) express strong Mlsa,d determinants, a putative marker of the mature subset of B cells. Although young (3-5 wk) (CBA/N X DBA/2)F1 male (xid+) mice stimulated only very weak mixed lymphocyte reactions (MLR) to Mlsa,d determinants, older mice (greater than 7 wk) regularly elicited conspicuous responses, despite being totally unresponsive to TNP-Ficoll. Expression of Mlsa,d determinants by xid+ mice was also detected by the procedure of negative selection in vivo. Thus BALB/c T cells were totally depleted of Mlsa,d reactivity after blood to lymph recirculation through 10-wk old irradiated xid+ (CBA/N X DBA/2)1 male mice. Significantly, a marked (90%) reduction in the anti-Mlsa,d response also occurred with T cell filtration through 3-wk xid+ mice, i.e., mice that elicit only minimal primary MLR; filtration through 3-wk xid- normal female mice led to near-complete (99%) negative selection. Collectively these data indicate either, (a) that xid+ mice contain appreciable numbers of cells with at least some of the properties of mature B cells, or (b) that the expression of Mlsa,d determinants is not restricted to mature B cells. In either case, B cells from xid mice cannot be viewed as a simple model for immature normal B cells.

Functional analysis of two human T-cell subpopulations: help and suppression of B-cell responses by T cells bearing receptors for IgM or IgG.

Subpopulations of thymus-derived T lymphocytes bearing receptors for either IgM or IgG molecules were isolated from human peripheral blood. Those with receptors for IgM (T.M) provided help in a cell dose-dependent fashion for the pokeweed mitogen-induced differentiation of B lymphocytes in vitro, whereas cells with receptors for IgG (T.G) did not. T.G cells, on the hand, efficiently suppressed the differentiation and proliferation of B cells in the pokeweed system in the presence of helper T.M cells. This suppressive activity of T.G cells required prior interaction of the T.G cells with immune complexes. The helper activity of T.M cells was relatively radioresistant while the suppressor activity of T.G cells was radiosensitive. The results indicate that helper and suppressor functions of human T lymphocytes in this model system are mediated by different subpopulations of T cells which can be distinguished by their ability to bind IgM or IgG immune complexes, respectively.

T cell receptors for responses to Mls determinants and allo-H-2 determinants appear to be encoded on different chromosomes.

Previous studies have shown that T cell clones specific for strong Mlsa,d determinants concomitantly display apparently random reactivity to allo-H-2 determinants. One explanation for this finding is that T cell recognition of Mlsa,d and allo-H-2 determinants is controlled by separate sets of receptors. If these receptors were chromosomally unlinked, karyotypically unstable T cell hybrids with dual reactivity for Mlsa,d and particular allo-H-2 determinants would be expected, occasionally, to lose reactivity for one set of determinants, but not the other. The results presented here provide direct support for this prediction.

T cell clones with dual specificity for M1s and various major histocompatibility complex determinants.

A high proportion of T cell clones derived from bulk cultures selected to M1s a,d determinants were found to have joint specificity for allo-H-2 determinants, and vice versa. Significantly, the patterns of H-2 alloreactivity shown by clones selected to M1sa,b determinants appeared to be random. The possible implications of these findings are discussed.

Morphological and histochemical analyses of two human T-cell subpopulations bearing receptors for IgM or IgG.

Two subpopulation of circulating human T cells forming rosettes with neuraminidase-treated sheep erythrocytes were purified on the basis of the presence of receptors for IgG (TG cells) or for IgM (TM cells), and were shown to have distinguishing morphological and histochemical characteristics. TM cells had the general features of typical small- or medium-sized lymphocytes; most were easily identifiable by distinctive cytoplasmic accumulations, usually one and sometimes two large spots, of nonspecific acid esterase activity. The release of the vesicular contents on short-term culture of TG cells was inhibited by cytochalasin B. Definition of these distinguishing characteristics of TM and TG cells provides a basis for practical enumeration of these functionally distinct subpopulations of human T cells. Some of the TG cells were capable of endocytosis of IgG antibody-coated erythrocytes.

Induction of neonatal tolerance to Mlsa antigens by CD8+ T cells.

Antigen-specific tolerance of T cells to minor lymphocyte stimulatory (Mls) antigens can be induced in mice by neonatal injection of foreign lymphohematopoietic cells. Although immune responses to Mlsa antigens are controlled by B cells, CD8+ T cells were the most effective cell type for induction of Mlsa tolerance. Tolerance was evident in both thymus and lymph nodes and could be induced by as few as 2 x 10(4) CD8+ T cells; these cells were 50 to 100 times as potent as CD4+ cells or B cells in causing functional tolerance and deletion of V beta 6+ T cells. Thus, intrathymic contact with antigens expressed on CD8+ T cells may play an important role in controlling the normal development of tolerance.

T cell reactivity to MHC molecules: immunity versus tolerance.

The specificity of mature CD8+ and CD4+ T lymphocytes is controlled by major histocompatibility complex (MHC) class I and class II molecules, respectively. The MHC class specificity of T cells is stringent in many assays, but is less evident when cells are supplemented with exogenous lymphokines. The repertoire of T cells is shaped through contact with MHC molecules in the thymus and involves a complex process of positive selection and negative selection (tolerance). Tolerance of immature T cells to MHC molecules can reflect either clonal deletion or anergy and results from intrathymic contact with several cell types, including epithelial cells and cells with antigen-presenting function. Unlike immature T cells, mature T cells are relatively resistant to tolerance induction. In certain situations partial unresponsiveness of mature T cells can be achieved by exposing T cells to foreign MHC molecules expressed on atypical antigen-presenting cells. Tolerance is rarely complete, however, and the precise requirements for tolerizing mature T cells are still unclear.

Activation of polyphosphoinositide hydrolysis in T cells by H-2 alloantigen but not MLS determinants.

Murine minor lymphocyte-stimulating (Mls) determinants are cell surface antigens that stimulate strong primary T cell responses; the responding T cells display restricted T cell receptor (TCR) V beta gene usage. Interaction of T cells with mitogens or major histocompatibility complex (MHC) antigens activated the polyphosphoinositide (PI) signaling pathway, but this pathway was not triggered by Mls recognition. However, interleukin-2 (IL-2) secretion and proliferation to all three stimuli were comparable. Thus, although recognition of both allo-H-2 and Mls determinants is thought to be mediated by the TCR, these antigens appear to elicit biochemically distinct signal transduction pathways.

Antigen presentation and T cell development in H2-M-deficient mice.

HLA-DM (DM) facilitates peptide loading of major histocompatibility complex class II molecules in human cell lines. Mice lacking functional H2-M, the mouse equivalent of DM, have normal amounts of class II molecules at the cell surface, but most of these are associated with invariant chain-derived CLIP peptides. These mice contain large numbers of CD4+ T cells, which is indicative of positive selection in the thymus. Their CD4+ cells were unresponsive to self H2-M-deficient antigen-presenting cells (APCs) but were hyperreactive to wild-type APCs. H2-M-deficient APCs failed to elicit proliferative responses from wild-type T cells.

T-cell activation by superantigens.

Superantigens stimulate powerful T-cell responses that can have marked effects in vivo, sometimes leading to shock or even death. The demonstration that strong T-cell responses to superantigens in vivo can be followed by tolerance, reflecting either clonal elimination or anergy, has provided important insights into how mature T cells can be regulated. Further progress in understanding the factors that control these responses relies heavily on defining the specific interactions between T-cell receptors, superantigens and major histocompatibility complex molecules which lead to T-cell activation as well as on the characterization of the specific signal transduction events and molecules involved in this activation. Significant progress has been made, during the past year, in the first area and these findings are summarized below; though less information is available in the latter area, recent observations relevant to this issue are discussed.

Intrathymic and extrathymic clonal deletion of T cells.

Clonal elimination accounts for self-tolerance induction in the thymus and also affects mature T cells responding to exogenous antigens in the periphery. Recent evidence on the microenvironments, cell-cell interactions and signalling requirements for clonal deletion of immature and mature T cells is discussed.

Review: Assessment of completeness of reporting in intervention studies using livestock: an example from pain mitigation interventions in neonatal piglets.

Accurate and complete reporting of study methods, results and interpretation are essential components for any scientific process, allowing end-users to evaluate the internal and external validity of a study. When animals are used in research, excellence in reporting is expected as a matter of continued ethical acceptability of animal use in the sciences. Our primary objective was to assess completeness of reporting for a series of studies relevant to mitigation of pain in neonatal piglets undergoing routine management procedures. Our second objective was to illustrate how authors can report the items in the Reporting guidElines For randomized controLled trials for livEstoCk and food safety (REFLECT) statement using examples from the animal welfare science literature. A total of 52 studies from 40 articles were evaluated using a modified REFLECT statement. No single study reported all REFLECT checklist items. Seven studies reported specific objectives with testable hypotheses. Six studies identified primary or secondary outcomes. Randomization and blinding were considered to be partially reported in 21 and 18 studies, respectively. No studies reported the rationale for sample sizes. Several studies failed to report key design features such as units for measurement, means, standard deviations, standard errors for continuous outcomes or comparative characteristics for categorical outcomes expressed as either rates or proportions. In the discipline of animal welfare science, authors, reviewers and editors are encouraged to use available reporting guidelines to ensure that scientific methods and results are adequately described and free of misrepresentations and inaccuracies. Complete and accurate reporting increases the ability to apply the results of studies to the decision-making process and prevent wastage of financial and animal resources.

Inhibition of dihydrofolate synthetase by folate, homofolate, pteroate and homopteroate and their reduced forms.

Dihydrofolate (H2-folate) synthetase (EC 6.3.2.12) was isolated from Escherichia coli B. A radiochemical assay was developed to determine the activity of H2-folate synthetase in order to study the effects of folate metabolites and antimetabolites which would interfere with the microbiological assay method previously used. The effects of folate and pteroate derivatives on the activity of this enzyme were investigated to determine if inhibition of this enzyme could constitute a site of action for these compounds as chemotherapeutic agents or a site of metabolic regulation. H2-folate synthetase was inhibited by its product, H2-folate, and by the antimetabolite dihydrohomopteroate, with apparent Ki values of 23.4 and 9.2 muM, respectively.

Controlling mature CD4+ T cell responses.

Immune responses are by necessity highly regulated to achieve the appropriate balance of aggression and restraint. Among the many factors involved in maintaining this balance are the interactions between accessory molecule receptors expressed on T cells and their ligands on antigen-presenting cells. Our studies during the past several years have focused on defining how particular accessory molecule interactions influence the activation of naïve CD4+ T cells and the subsequent development of effector function. In this article, we discuss our findings on the effects of distinct accessory molecules with particular attention to the unique roles of LFA-1 and CD28 during different phases of the naïve CD4+ cell response.

Regulation of D-xylose utilization by hexoses in pentose-fermenting yeasts.

The aldopentose D-xylose is one of the most abundant sugars in plant biomass and its efficient microbial utilization is of fundamental importance in the overall bioconversion of lignocellulosic materials into liquid fuels and chemicals. The discovery of pentose-fermenting yeasts in the early 1980's led to world wide interest because of the perceived potential for improved D-xylose fermentation to enhance the prospect of biomass conversions. However, the utilization of D-xylose by pentose-fermenting yeasts can be adversely affected by the hexoses, mainly D-glucose and D-mannose, which are usually present in high amounts in lignocellulosic hydrolysates. Research in the past several years has uncovered some of the regulatory effects of D-glucose on D-xylose utilization. However, much remains unknown about the mechanisms responsible for these effects. This review summarizes the current state of knowledge on the induction, repression and inactivation of D-xylose utilization in pentose-fermenting yeasts.

Normal fetal cardiac dimensions obtained by perpendicular imaging.

This study provides normal fetal cardiac dimensional data in a large patient group over a wide range of gestational ages. Perpendicular imaging decreased lateral resolution error, resulting in normal values with narrower confidence limits than in prior studies.