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The reactivity of 38 murine strains to a synthetic analogue of bacterial lipoprotein, tripalmitoyl-pentapeptide (TPP), was tested and compared with the reactivity to lipopolysaccharide (LPS). These strains include common laboratory mice and H-2 recombinant inbred lines, as well as some newly bred lines originating from animals recently captured in different regions of Europe. All animals analysed were reactive to TPP and polyclonally activated to proliferation and immunoglobulin synthesis. Large differences in mitogen reactivities of various H-2 recombinant inbred strains suggest that MHC or closely linked gene products influence the reactivity to the LPS and TPP mitogens. By analyzing the frequencies of precursor cells reactive to TPP or LPS and the isotype patterns obtained after stimulation, we demonstrated that both mitogens activate individual B cells in different ways. [ABSTRACT FROM AUTHOR]

The in vitro anti-trinitrophenyl (TNP) response induced by TNP-lipopolysaceharide (TNP-LPS) in mouse spleen cells was eliminated by passage through Sephadex G10 columns. Conditioned medium obtained from peritoneal macrophages restored the response, indicating the supportive role of such cells in the response. Conversely, polyclonal B-cell activation mediated by LPS was not affected by Sephadex G10 treatment, as judged by incorporation of 3H-thymidine, generation of specific anti-TNP plaque-forming cells, and induction of polyclonal IgM-secreting cells. The failure of the LPS moiety of TNP-LPS to induce B-cell activation without macrophage help suggests a restriction in the number of available anti-LPS receptor molecules when the Ig anti-TNP receptor interacts with the haptenic (TNP) groups. This restriction can be due to the attachment of the TNP molecules to residues located in the vicinity of the lipid A mitogenic region of the carrier. [ABSTRACT FROM AUTHOR]

Our aim was to investigate the subset distribution and function of circulating monocytes, proinflammatory cytokine levels, gut barrier damage, and bacterial translocation in chronic spinal cord injury (SCI) patients. Thus, 56 SCI patients and 28 healthy donors were studied. The levels of circulating CD14 +high CD16 - , CD14 +high CD16 + , and CD14 +low CD16 + monocytes, membrane TLR2, TLR4, and TLR9, phagocytic activity, ROS generation, and intracytoplasmic TNF-α, IL-1, IL-6, and IL-10 after lipopolysaccharide (LPS) stimulation were analyzed by polychromatic flow cytometry. Serum TNF-α, IL-1, IL-6 and IL-10 levels were measured by Luminex and LPS-binding protein (LBP), intestinal fatty acid-binding protein (I-FABP) and zonulin by ELISA. SCI patients had normal monocyte counts and subset distribution. CD14 +high CD16 - and CD14 +high CD16 + monocytes exhibited decreased TLR4, normal TLR2 and increased TLR9 expression. CD14 +high CD16 - monocytes had increased LPS-induced TNF-α but normal IL-1, IL-6, and IL-10 production. Monocytes exhibited defective phagocytosis but normal ROS production. Patients had enhanced serum TNF-α and IL-6 levels, normal IL-1 and IL-10 levels, and increased circulating LBP, I-FABP, and zonulin levels. Chronic SCI patients displayed impaired circulating monocyte function. These patients exhibited a systemic proinflammatory state characterized by enhanced serum TNF-α and IL-6 levels. These patients also had increased bacterial translocation and gut barrier damage.

We have derived and characterized a set of monoclonal antibodies (mAb) specific for the different human GH (hGH) isoforms. The binding characteristics of each antibody to the hGH isoforms (22K and 20K) were analyzed in direct and competitive immunoassays as well as by Western blot. We studied the effects of these mAb on the biological activity of hGH and showed that they specifically block their respective activities. Using these mAb, we developed several immunoassays that have been applied for the quantitation of the different hGH isoforms in body fluids. Therefore, these mAb may help to unravel the biological function of these variants.

Injection of swine peripheral blood mononuclear cells into mice with severe combined immunodeficiency (SCID), resulted in the stable long-term establishment of a functional swine immune system (SCID-sw). Swine immunoglobulins were present in the serum of SCID-sw mice and swine cells were detected in the blood as well as in lymph nodes and spleen using monoclonal antibodies raised against cell subpopulations. Swine lymphocytes from reconstituted SCID mice responded in vitro to specific antigens or mitogens. When SCID-sw mice were challenged with African swine fever (ASF) virus, ASF virus-infected cells were detected in blood and spleen, and antiviral antibodies and virus-specific T cells were generated.

The activation of a phosphatidylinositol (PI) 3-kinase is one of the earliest intracellular responses to T lymphocyte stimulation. We have used a human T cell line, Jurkat, and an antibody that recognizes the clonotype of its T cell receptor (TcR) to characterize the association of PI 3-kinase with the TcR and its activation in response to TcR cross-linking. We show that the TcR is constitutively associated with the alpha isoform of the PI 3-kinase p85 regulatory subunit. Stimulation of Jurkat cells through the TcR results in the rapid and transient activation of the associated PI 3-kinase. In addition, our results indicate that the tyrosine kinase pp56lck is essential for PI 3-kinase activation via the TcR.

Helper cells, with specificity for the haptens (4-hydroxy-3-nitro-phenyl)acetyl (NP) or (4-hydroxy-5-iodo-3-nitro-phenyl)acetyl (NIP), were raised in B10.BR mice by in vivo priming and in vitro long-term enrichment with hapten-derivatized syngeneic spleen cells. Upon co-culture with the homologous antigen (NP or NIP self), selected helper cells specifically responded by proliferation and by inducing large numbers of B cells to clonal expansion and immunoglobulin secretion. Criss-cross experiments demonstrated the nonheteroclitic nature of antigen recognition by helper cells, as the proliferative and helper cell activities were in every case one order of magnitude higher when confronted with the homologous hapten used for immunization.

Polyclonal activation of resting B lymphocytes by either lipopolysaccharide or specific helper cells recognizing antigens on B cell membranes results in selective patterns of IgG subclass expression among plaque-forming cells. We have studied the IgG subclasses of plaque-forming cells generated in cultures of purified B cell blasts selected for reactivity to either LPS or helper cells, and restimulated by either lipopolysaccharide, specific helper cells, or as "bystanders", by nonspecific B cell growth factors. Development of IgG1 plaque-forming cells is observed only when clonal expansion is maintained by specific helper cells, whereas IgG3 secretion specifically requires stimulation by lipopolysaccharide and the absence of helper cell activity. Furthermore, exposure of resting B lymphocytes to specific helper cell induces, in 48 h, an irreversible loss of the potential to produce IgG3. Other than showing that helper cell-dependent B cell growth and maturation is more complex than previously suspected, these results suggest that differentiation signals or factors induce specific DNA recombination and deletion events.

Mice were injected with DNP-derivatives of thymus dependent (DNP-OA) and thymus-independent (DNP-Ficoll and TNP-LPS) antigens, and the response to TNP-LPS in vitro was studied after several priming periods. DNP-OA priming decreases the amount of cells responding to TNP-LPS in vitro. In the case of DNP-Ficoll and TNP-LPS-primed cells, there is an initial burst of responsiveness to TNP-LPS, which progressively decreases until an abolishment of TNP-LPS responsiveness is found at day 40 after immunization. The sensitivity to TNP-LPS reappears as new precursor cells differentiate into mature cells. We suggest that B cells progressively gain the capacity to respond to thymus-independent and thymus-dependent antigens (B1-B2 differentiation) and that challenge with a particle antigen increases the ratio of maturation throughout the pathway.

Polyclonal stimulation of normal splenic B lymphocytes with either lipopolysaccharide (LPS) or helper T lymphocytes specific for B cell surface antigens results in the selective expression of IgG subclasses by the secretory cells: in addition to IgM-secreting plaque-forming cells (PFC), thymus-independent stimulation leads to the development of IgG2 and IgG3 PFC, while helper cell-dependent activation leads to IgG1 and IgG2 PFC. This cannot be solely explained by selective stimulation of distinct B cell subpopulations, because purified LPS-reactive blasts if restimulated by helper cells switch to IgG1 while if maintained with LPS switch to IgG3. The simultaneous stimulation of splenic B cells with LPS and helper cells results in additive IgM and IgG2 responses, but in the selective suppression of IgG3 PFC with a concomitant synergic enhancement of IgG1 responses. These results are interpreted to indicate that the expression of IgG C genes in proliferating B lymphocytes is directed by the quality of nonspecific stimuli.

Wheat germ agglutinin (WGA), added to Mishell-Dutton-type cultures of nude spleen cells challenged with erythrocyte antigens, allows a primary specific plaque-forming cell (PFC) response. WGA, however, does not display direct mitogenicity to either T or B lymphocytes. The activity of WGA in the specific PFC response is macrophage-dependent and parallels a nonspecific PFC response in the same cultures.