Costimulator deficient antigen presentation by an endothelial cell line induces a nonproliferative T cell activation response without anergy.

The ability of endothelial cells to activate helper T (Th) cells by antigen presentation was studied using the murine endothelial cell line SVEC4-10 and antigen-specific murine T cell clones. SEVEC4-10 cells constitutively express vascular cell adhesion molecule 1 but not intercellular adhesion molecule 1. Interferon gamma (IFN-gamma) treatment of these cells induced class II major histocompatibility complex (MHC) expression and antigen-presenting capabilities, but did not alter surface integrin expression. IFN-gamma-treated SVEC4-10 cells were competent at mediating antigen-dependent cytokine production and proliferation of a Th2 clone. In contrast, endothelial antigen presentation to Th1 cells did not stimulate T cell proliferation. The addition of MHC mismatched spleen cells as a source of costimulatory molecules resulted in the ability of the endothelial cells to stimulate Th1 cell proliferation in an antigen-specific manner. The failure of the endothelial cell line alone to support Th1 cell proliferation correlated with the failure to stimulate interleukin 2 (IL-2) gene expression. T cell exposure to the endothelial cells plus antigen resulted in upregulation of IL-2 receptors and an enhanced response to subsequent antigen presentation by splenic antigen-presenting cells. Despite the lack of functional costimulators for IL-2 expression, antigen presentation by the endothelial cell line did not induce Th1 cell anergy, indicating that costimulator deficiency for IL-2 expression is not obligatorily linked to anergy induction. Thus, endothelial cells are capable of presenting antigens to helper T lymphocytes, but stimulate only partial T cell responses. These partial responses may serve to selectively stimulate transmigration of antigen-specific T cells and may enhance functional responses upon subsequent, extravascular antigen exposure.

Cytokine transcriptional events during helper T cell subset differentiation.

The molecular basis for changes in cytokine expression during T helper (Th) cell subset differentiation is not well understood. We have characterized transcriptional events related to cytokine gene expression in populations of naive T cell receptor-transgenic T cells as they are driven in vitro toward Th1 or Th2 phenotypes by interleukin (IL)-12 or IL-4 treatment, respectively. Quantitative reverse transcriptase-polymerase chain reaction analysis of cytokine transcripts indicates that interferon (IFN) gamma, IL-4, and IL-2 mRNA are expressed with distinct kinetics after naive T cells are stimulated with antigen and either IL-4 or IL-12. IFN-gamma mRNA appears as early as 6 h in IL-12-treated cultures, IL-4 appears only after 48 h in IL-4-treated cultures, and IL-2 is equivalently expressed in both types of cultures. Analyses were performed to determine if there were any differences in activation of IL-2 or IL-4 transcription factors that accompanied Th1 versus Th2 differentiation. These studies demonstrated that signal transducer and activator of transcription 6 (STAT6) binds to a sequence in the IL-4 promoter and that this STAT6-binding site can support IL-4-dependent transcription of a linked heterologous promoter. Prolonged activation of STAT6 is characteristic of populations undergoing Th2 differentiation. Furthermore, STAT6 is activated in an autocrine manner when differentiated Th2 populations are stimulated by antigen receptor ligation. Th1 populations derived from IL-12 plus antigen treatment of naive T cells remain responsive to IL-4 as indicated by induction of STAT6 and IL-4 mRNA. These data indicate that Th1 and Th2 differentiation represents the combination of different, apparently independently regulated transcriptional events. Furthermore, among transcription factors that bind to the IL-4 or IL-2 promoters, STAT6 is the one whose activation distinguishes Th2 versus Th1 development.

Transcription of the interleukin 4 gene is regulated by multiple promoter elements.

Activation of T helper cell 1 (Th1) and Th2 results in transcription of the interleukin 2 (IL-2) and IL-4 cytokine genes, respectively. Whereas many of the regulatory elements and factors responsible for IL-2 transcription in T cells are well defined, little is known about parallel mechanisms that drive transcription of the IL-4 gene. Here we have analyzed the murine IL-4 promoter, both in vivo and in a Th2 clone. 3 kb of IL-4 upstream sequence is shown to be sufficient to achieve tissue-specific and inducible expression of a thymidine kinase reporter gene in vivo in a manner that mirrors the expression of endogenous IL-4. Tissue-specific and inducible expression is also demonstrated in a Th2 clone, but not in a B cell line. Deletional and mutational analysis of the IL-4 promoter demonstrated that sequences from -100 to -28 were necessary for a transcriptional response to Concanavalin A or anti-CD3 monoclonal antibody. An overlapping, yet smaller region, spanning the sequences from -60 to -28 bp was shown to be required for the response to ionomycin. Mutation of an 8-bp region from -43 to -35 of the IL-4 promoter completely abrogated IL-4 gene transcription in response to all stimuli tested. In addition, our results show that the effects of the immunosuppressive agent Cyclosporin A map to the same DNA sequences as the positive control elements. These results identify DNA sequences that are functionally important for the control of IL-4 gene transcription both in vivo and in vitro. Although these sequences are highly conserved in the human and murine IL-4 genes, they are largely not present in the IL-2 enhancer complex. Thus, cytokine-specific cis-acting elements may be one mechanism by which these two cytokine genes are differentially regulated.

Characterization of pulmonary immune responses to hyperoxia by high-dimensional mass cytometry analyses.

Prolonged exposure to hyperoxia has deleterious effects on the lung, provoking both inflammation and alveolar injury. The elements of hyperoxic injury, which result in high rates of lethality in experimental models, are thought to include multicellular immune responses. To characterize these alterations in immune cell populations, we performed time-of-flight mass cytometry (CyTOF) analysis of CD45-expressing immune cells in whole lung parenchyma and the bronchoalveolar space of mice, exposed to 48 hours of hyperoxia together with normoxic controls. At the tested time point, hyperoxia exposure resulted in decreased abundance of immunoregulatory populations (regulatory B cells, myeloid regulatory cells) in lung parenchyma and markedly decreased proliferation rates of myeloid regulatory cells, monocytes and alveolar macrophages. Additionally, hyperoxia caused a shift in the phenotype of alveolar macrophages, increasing proportion of cells with elevated CD68, CD44, CD11c, PD-L1, and CD205 expression levels. These changes occurred in the absence of histologically evident alveolar damage and abundance of neutrophils in the parenchyma or alveolar space did not change at these time points. Collectively, these findings demonstrate that pulmonary response to hyperoxia involves marked changes in specific subsets of myeloid and lymphoid populations. These findings have important implications for therapeutic targeting in acute lung injury.

Characterization and identification of interleukin 1 receptors on bovine neutrophils.

Interleukin 1 (IL-1) has been shown to modulate various functional activities of neutrophils, presumably by first binding to cell surface IL-1 receptors. In this study, we characterized and identified IL-1 receptors on bovine neutrophils using direct and competitive receptor binding studies and affinity cross-linking analysis. The results of direct binding studies demonstrated that bovine neutrophils bound 125I-labeled bovine IL-1 by a single affinity binding site (Kd = 4.6 nM, 5600 binding sites per cell). Competitive receptor binding studies demonstrated that unlabeled recombinant bovine IL-1 beta, murine IL-1 alpha, and human IL-1 beta competitively blocked neutrophil binding of 125I-labeled bovine IL-1 beta. In contrast, the IL-1 receptor antagonist (IL-1ra) demonstrated no detectable binding to bovine neutrophils as judged by direct and competitive receptor binding assays. Affinity cross-linking of 125I-labeled bovine IL-1 beta to neutrophils was used to identify cell surface IL-1 receptors. Two specific cross-linked products were observed with molecular sizes of 89 kd (a deduced receptor size of 71.5 kd) and more than 200 kd. The latter may indicate a complex of IL-1 receptor-associated signal-transducing components, IL-1 receptors, and IL-1. The presence of 100 nM unlabeled bovine, murine, or human IL-1 during the receptor binding and cross-linking reactions prevented the formation of cross-linked complexes of 125I-labeled bovine IL-1 beta and its receptor. In contrast, the IL-1ra failed to inhibit the formation of cross-linked complexes of 125I-labeled bovine IL-1 beta and its receptor.

Characterization and identification of interleukin-1 receptors on bovine fibroblasts.

Interleukin-1 (IL-1), a pro-inflammatory cytokine, initiates its many biological effects by first binding to cell-surface receptors. Prior to this study, there were no published reports addressing the nature of the bovine IL-1 receptor. In this study, we characterized and identified cell-surface IL-1 receptors on bovine fibroblasts. Direct binding studies using [125I]-labeled bovine IL-1 beta demonstrated that bovine fibroblasts had approximately 130 high affinity and 2,500 low affinity binding sites (Kd = 4.9 x 10(-11) M and 3.7 x 10(-9) M, respectively). Competitive binding studies using unlabeled recombinant bovine IL-1 beta, IL-2, IFN-alpha, and bovine insulin demonstrated that only unlabeled bovine IL-1 beta competitively blocked fibroblast binding of [125I]-labeled bovine IL-1 beta. Affinity cross-linking of [125I]-labeled IL-1 beta to fibroblasts demonstrated that IL-1 receptors on bovine fibroblasts have an apparent M(r) of 71.5 kD. This report provides the first characterization and identification of IL-1 receptors on bovine fibroblasts.

Evaluation of resource recovery from waste incineration residues--the case of zinc.

Solid residues generated at European Waste to Energy plants contain altogether about 69,000 t/a of Zn, of which more than 50% accumulates in air pollution control residues, mainly boiler and filter ashes. Intensive research activities aiming at Zn recovery from such residues recently resulted in a technical scale Zn recovery plant at a Swiss waste incinerator. By acidic leaching and subsequent electrolysis this technology (FLUREC) allows generating metallic Zn of purity>99.9%. In the present paper the economic viability of the FLUREC technology with respect to Zn recovery from different solid residues of waste incineration has been investigated and subsequently been categorised according to the mineral resource classification scheme of McKelvey. The results of the analysis demonstrate that recovery costs for Zn are highly dependent on the costs for current fly ash disposal (e.g. cost for subsurface landfilling). Assuming current disposal practice costs of 220€/ton fly ash, resulting recovery costs for Zn are generally higher than its current market price of 1.6€/kg Zn. With respect to the resource classification this outcome indicates that none of the identified Zn resources present in incineration residues can be economically extracted and thus cannot be classified as a reserve. Only for about 4800 t/a of Zn an extraction would be marginally economic, meaning that recovery costs are only slightly (less than 20%) higher than the current market price for Zn. For the remaining Zn resources production costs are between 1.5 and 4 times (7900 t/a Zn) and 10-80 times (55,300 t/a Zn) higher than the current market value. The economic potential for Zn recovery from waste incineration residues is highest for filter ashes generated at grate incinerators equipped with wet air pollution control.

The effects of injury on the adaptive immune response.

For more than thirty years it has been apparent that serious injury in humans and experimental animals is associated with a decrease in immune functions dependent upon T cells, the principal cells involved in initiating adaptive immune responses. This review focuses on more recent evidence that T helper cell function is altered after serious injury with loss of T helper 1 function and cytokine production and with preservation of T helper 2 function and an increased production of T helper 2 cytokines. Emphasis is placed on the importance of interactions between the innate and adaptive immune systems in the perturbed immune responses seen following injury. Immunomodulatory strategies are mentioned that have had success in animal models in ameliorating the diminished resistance to infection commonly seen after major traumatic or thermal injury. Finally, it is emphasized that immunomodulatory treatments that are successful in preventing infection may be contraindicated once infection is manifest.

Severe injury triggers antigen-specific T-helper cell dysfunction.

Although it is established that post-injury immune dysfunction involves alterations in T-cell function, the effects of injury on T-cell function in vivo are poorly understood. This study uses a mouse injury model and an antigen immunization approach to investigate the influence of injury on antigen-specific T-helper cell function. We report here that injury triggered a significant reduction in antigen-specific T-helper-1 (Th1)-dependent IgG2a antibody formation, while IgM, IgG1, and IgE production was unchanged. In addition, injury caused a reduction in cytokine production (IL-2, IFNgamma and IL-10) by antigen-stimulated T-cells. We also demonstrate that interleukin 12 (IL-12), a cytokine that promotes Th1 cell differentiation, restored IgG2a antibody formation and corrected the injury-induced reduction in antigen-stimulated cytokine production. Taken together, these findings indicate that severe injury induces a dramatic reduction in Th1 cell function in vivo and suggest that therapies designed to restore Th1 cell function may be beneficial to the injured host.

Injury primes the immune system for an enhanced and lethal T-cell response against bacterial superantigen.

We previously reported the high lethality of CD4+ T-cell activation in burn-injured T-cell receptor (TCR) transgenic mice. This suggested to us that T-cells may play a role in the development of the systemic inflammatory response syndrome (SIRS) which can occur after severe injury. In this study, we sought a more clinically relevant model to test the hypothesis that naturally produced bacterial toxins that are known to act as potent polyclonal T-cell activating agents may induce a similar lethal shock-like response in injured, non-TCR transgenic mice. Accordingly, sham- or burn-injured mice were treated with various doses of staphylococcal enterotoxin A (SEA), then observed for 48-hour mortality. We observed 94% and 56% 48-h mortality when burn-injured mice were given 15 microg and 10 microg of SEA, respectively, while neither SEA dose caused mortality in sham-injured mice. The assessment of serum cytokine levels demonstrated significantly elevated interleukin 2 (IL-2) and tumor necrosis factor alpha (TNFalpha) levels when compared to sham mice (P < 0.01). In vitro studies confirmed our in vivo results and also demonstrated elevated levels of interferon gamma (IFNgamma) (P < 0.01). We also observed a novel injury-dependent switch from CD4+ to CD8+ T-cells as the dominant T-cell type producing TNFalpha and IFNgamma in response to SEA stimulation in vitro. Taken together, our findings indicate that injury primes the immune system for an augmented early T-cell response that can result in a lethal shock-like syndrome.

Injury induces alterations in T-cell NFkappaB and AP-1 activation.

The immune dysfunction that occurs after severe injury involves major changes in T-cell-mediated immunity resulting in suppressed T-helper 1 (Th1) type responses and increased or persistent T-helper 2 (Th2) type cytokine production. Since little is known about what signaling pathways are responsible for this injury-induced phenotypic shift in T-cells, we undertook this study to address the molecular basis for injury effects on T-helper cell subset cytokine expression. Experiments were designed to test whether diminished IL-2 gene expression after thermal injury coincided with changes in the induction of IL-2 gene regulatory transcription factors. Electrophoretic mobility shift assays (EMSA) were used to screen for nuclear expression of changes of the IL-2 gene transcription factors. Our findings revealed that changes in mitogen-stimulated T-cell AP-1 and NFkappaB factor activation correlated directly with defective mitogen-induced IL-2 mRNA expression. We determined that there was a loss of nuclear AP-1 activation and changes in NFkappaB factor activation at 9 days after injury. T-cell nuclear extracts prepared from sham injured mice showed induction of NFkappaB2 (p52) and RelA (p65) containing NFkappaB EMSA complexes, while we detected no RelA or NFkappaB2 in EMSA complexes using T-cell nuclear extracts prepared from burn injured mice. Instead, these NFkappaB EMSA complexes contained mostly NFkappaB1 (p50). Western immunoblot analysis confirmed defective nuclear RelA translocation. Taken together, these results indicate that T-cell NFkappaB and AP-1 activation pathways may be involved in the injury-induced changes in T-cell cytokine production and the immune deviation that occurs after injury.

Anti-interleukin-10 antibody restores burn-induced defects in T-cell function.

BACKGROUND: Studies have shown that susceptibility to sepsis after severe injury correlated with reduced production of T-helper 1 (Th1) cytokines (interleukin-2 [IL-2] and interferon-gamma [IFN-gamma]) and a persistence of T-helper 2 (Th2) cytokines (IL-4 and IL-10). The mechanisms responsible for this effect are not clear. We used a T-dependent antigen to study both the effect of burn injury on antigen-specific Th functions in vivo and the effect of anti-IL-10 antibody on these functions. METHODS: Male A/J mice were anesthetized and given a 25% scald burn or a sham burn. On day 0 all mice were immunized with 100 micrograms trinitrobenzene sulfonic acid (TNP) haptenated ovalbumin (TNP-OVA) in complete Freund's adjuvant. Mice (10 per group) were given 250 micrograms monoclonal rat antimurine IL-10 antibody (anti-IL-10 MAB) or control rat immunoglobin G (IgG) on day 0 and 100 micrograms anti-IL-10 MAB or IgG on day 2. On day 10 the mice were killed to obtain serum and spleen cells. TNP-specific serum antibody isotype titers were determined by enzyme-linked immunosorbent assay (ELISA). Splenocyte proliferation and cytokine-production in response to TNP-OVA or to anti-CD3 MAB were determined by tritiated thymidine incorporation and by ELISA, respectively. RESULTS: Burn injury resulted in depressed levels of the TNP-specific IgG2a antibody isotype (Th1 dependent), whereas TNP-specific IgG1 and IgE (Th2 dependent) levels were not decreased in burn versus sham burn mice. Anti-IL-10 MAB but not IgG restored the IgG2a response. Burn injury also resulted in reduced TNP-OVA-specific proliferation of splenocytes, whereas anti-CD3 proliferation was equivalent in burn and sham mice. TNP-OVA-specific IL-2 and IFN-gamma production were significantly reduced by burn injury. Anti-IL-10 MAB restored TNP-OVA-specific proliferation and antigen-specific IL-2 and interferon-gamma production by splenocytes from burn mice. CONCLUSIONS: Burn injury induces the loss of antigen-specific Th1 cell function, and IL-10 acts as a trigger to down-regulate Th1 activity after injury.

A central role for CD95 (Fas) in T-cell reactivity after injury.

BACKGROUND: Recent findings indicate that severe injury primes the immune system for an enhanced and lethal proinflammatory cytokine response against bacterial-derived superantigens. This study asked whether this response to injury involves the CD95 (Fas) signaling pathway. METHODS: To assess superantigen-mediated mortality, wild-type (WT) C57BL/6 and Fas-deficient C57BL/6 lpr (-/-) (lpr) mice underwent burn or sham injury and were challenged 2 hours later with staphylococcal enterotoxin B (SEB). Spleen cells from sham and burn WT or lpr mice were stimulated in vitro with SEB to assess injury effects on IL-2, TNF-alpha, and IFN-gamma production. RESULTS: Lpr burn mice survived the SEB challenge (100% survival), while WT burn mice showed a high mortality (17% survival, P < 001, analysis of variance [ANOVA]). Sham lpr or WT mice suffered no mortality to the SEB challenge. In vitro studies demonstrated that burn lpr mice produced significantly less TNF-alpha, IFN-gamma, IL-2 than burn WT mice (P <.01, ANOVA). Burn injury markedly enhanced SEB-stimulated IFN-gamma production by WT spleen cells and CD8+ T cells, while this did not occur in SEB-stimulated lpr spleen cells. CONCLUSIONS: These findings support the hypothesis that the CD95 (Fas) signaling pathway plays an integral role in the injury-induced enhanced and lethal T-cell reactivity against bacterial superantigens.

Interleukin-12 treatment restores normal resistance to bacterial challenge after burn injury.

BACKGROUND: Preliminary studies in this laboratory have shown that treatment with interleukin-12 (IL-12), a cytokine that induces expression of the T-helper-1 lymphocyte phenotype, in an animal model of burn injury increased survival after a septic challenge. The purpose of this study was to define the efficacy of IL-12 therapy and to explore its mechanism of action. METHODS: Adult male A/J mice were subjected to 25% full-thickness scald or sham burn. Starting on day 3 after burn, groups of mice received five daily injections of IL-12, interferon-gamma (IFN-gamma), or saline solution. Some animals received anti-IFN-gamma monoclonal antibody. At day 10 most animals underwent cecal ligation and puncture (CLP) and were observed for survival. Some animals were killed at day 10, and CD4-enriched splenocytes were stimulated with anti-CD3 antibody or concanavalin A and were studied for cytokine production and mRNA expression. RESULTS: IL-12 treatment, 25 ng daily for 5 days, increased survival of the burn group after CLP to that of the sham burn control group. Anti-IFN-gamma antibody, 500 micrograms, administered 1 day before IL-12 treatment, reduced the efficacy of IL-12. IFN-gamma treatment, 7000 units, moderately increased survival. IL-12 had no effect on survival of the sham burn group. At the time of CLP IL-12 therapy had induced a marked decrease in CD4+ lymphocyte IL-4 and a moderate increase in IFN-gamma production and mRNA expression without affecting IL-2. CONCLUSIONS: IL-12 is the most effective therapy so far tested in this burn plus CLP model. It acts at least in part through IFN-gamma. However, IFN-gamma therapy was not as effective as IL-12.

Burn injury primes naive CD4+ T cells for an augmented T-helper 1 response.

BACKGROUND: The mechanisms responsible for altered T-cell responses and cytokine production after injury are not well understood. We used T-cell receptor (TCR) transgenic mice to study burn injury effects on naive versus antigen-activated CD4+ T cells in vivo. METHODS: One week after sham or burn injury, lymph node cells were prepared from TCR transgenic mice and stimulated with TCR transgene-specific antigens. T-cell proliferation was measured and culture supernatants were tested for interleukin-2 (IL-2), interferon-gamma (IFN-gamma), IL-4, and IL-10 by enzyme-linked immunosorbent assay (ELISA). Burn injury effects on antigen-activated T cells were studied by immunizing TCR transgenic or wild-type mice at the time of injury. RESULTS: The antigen-stimulated proliferation of native CD4+ T cells was unaffected by burn injury and no increase in T-helper 2 (Th2)-type cytokine production was observed. Instead, burn injury augmented INF-gamma production by naive CD4+ transgenic T cells, and IL-2 production was marginally reduced. Thus, burn injury primed native T cells for an enhanced Th1-type response. In contrast, antigen-specific proliferation, IL-2, and IFN-gamma production by T cells harvested from immunized wild-type mice were suppressed. Unexpectedly, high mortality was observed when burn-injured TCR transgenic mice were immunized. CONCLUSION: Our results show that burn injury has differential effects on naive and antigen-activated CD4+ T cells and can prime naive CD4+ T cells.

Interleukin 10 is not essential for survival or for modulating T-cell function after injury.

BACKGROUND: Interleukin 10 (IL-10) is thought to be protective in injury and sepsis. However, we recently reported that IL-10 antagonism can be beneficial after burn injury. This study used IL-10-deficient (IL-10 [-/-]) mice to further define the role of IL-10 after injury. METHODS: Wild-type (WT) C57BL/6 or IL-10 (-/-) mice were anesthetized, sham or burn injured, and immunized subcutaneously with a T-cell-dependent protein antigen. Ten days later antigen-specific serum antibody isotype formation was measured by enzyme-linked immunosorbent assay. In addition, antigen-stimulated splenic T-cell proliferation and cytokine production (interleukin 2, interferon gamma, and tumor necrosis factor-alpha) were measured. RESULTS: Burn-injured IL-10 (-/-) mice survival (80%) was equivalent to that of burn-injured WT mice (74%). An injury-dependent loss of T-helper 1 (Th1)-type antibody isotype (IgG2a) formation occurred in both WT and IL-10 (-/-) mice. In vitro studies indicated that burn injury caused reduced antigen-stimulated splenic T-cell proliferation and Th1-type (interleukin 2 and interferon gamma) cytokine production in WT and IL-10 (-/-) mice, whereas burn-injured IL-10 (-/-) mice produced high levels of antigen-stimulated tumor necrosis factor-alpha. CONCLUSIONS: IL-10 is not essential for survival after burn injury or for several injury-induced changes in adaptive immune function, including Th1-type antibody isotype formation, T-cell proliferation, and Th1-type cytokine production.

Glucose homeostasis in magnesium-deficient rats.

Glucose homeostasis was studied in rats fed diets containing 750,200, or 100 mg/kg Mg for 6 to 8 weeks, from the age of 4 weeks. Weight gain of the rats receiving 200 and 100 mg/kg diets was decreased. This resulted from both a lower food intake and reduced effectiveness of the ingested food. Fed or fasting plasma glucose levels were similar in the three groups. During an intravenous glucose tolerance test, the rate of glucose disappearance was higher in Mg 100 rats than in controls. By contrast, during an oral glucose tolerance test, the rise in plasma glucose was greater and more sustained in Mg 100 rats. During both tests, the insulin response was markedly lower in Mg-deficient rats than in controls. This could be partially due to the reduced insulin content of the pancreas of these animals. The impairment of tolerance to oral glucose was corrected by 5 weeks on a high-Mg diet. After intravenous injection of insulin, the fall in plasma glucose levels was also slightly more pronounced in Mg 100 rats. During no test did we observe a significant difference between glucose or insulin responses in Mg 200 or Mg 750 rats. These results, thus, show that chronic Mg deficiency alters several parameters of glucose homeostasis in the rat.

Protective effects of early interleukin 10 antagonism on injury-induced immune dysfunction.

HYPOTHESIS: Interleukin 10 (IL-10) plays a central role in the development of postinjury immune suppression, and early in vivo IL-10 antagonism can be protective. DESIGN: Male A/J mice underwent sham or burn injury and were treated with monoclonal anti-IL-10 antibody or control antibody at 1 day or 3 days after injury. Their ability to survive polymicrobial sepsis induced by the cecum ligation and puncture (CLP) technique was then tested. The response of sham- and burn-injured mice and burn-injured mice treated with anti-IL-10 to immunization with a T-cell-dependent antigen, trinitrophenyl (TNP)-haptenated ovalbumin (TNP-OVA) was also assessed. MAIN OUTCOME MEASURES: Mortality was monitored for a total of 7 days after CLP to assess the effect of anti-IL-10 therapy on the survival of burn-injured, immunecompromised mice. Serum antibody isotype formation was measured in sham- and burn-injured mice and burn-injured mice treated with anti-IL-10 to determine how IL-10 antagonism influenced helper T-cell responses in vivo. In vitro cytokine production by antigen-stimulated spleen cells was assessed to study the effect of blocking IL-10 activity at 1 day vs 3 days after burn injury. RESULTS: Treating mice with anti-IL-10 at 1 day after injury significantly improved CLP survival, whereas delaying treatment 3 days had no beneficial effect. The analysis of T-cell function in vivo as determined by serum antibody isotype formation indicated that anti-IL-10 treatment at 1 day or 3 days after injury increased T helper cell 1-type antibody formation to sham injury levels by day 10. Moreover, these treatments restored the injury-induced reduction of antigen-stimulated IL-2, interferon gamma, and IL-10 production. CONCLUSIONS: Interleukin 10 plays an early role in the development of burn injury-induced immune suppression. Its in vivo inhibition at 1 day after injury may be a useful approach toward preventing the development of injury-induced immune dysfunction and may do so by restoring T-cell function and cytokine production.

[Linoleic acid in nutritional balance: experimental studies]. / L'acide linoléique dans l'équilibre alimentaire: recherches expérimentales.

In the male rat a diet rich in beef fat facilitates the occurrence of hyperinsulinemia after a glucose load whereas fats rich in linoleic acid produce no such effect. The combination of saturated fat and saccharose facilitates the occurrence of hypertriglyceridemia in the male rat, no such effect is produced by the combination of fats rich in linoleic acid and saccharose. Linoleic acid prevents natrium chloride from provoking hypertriglyceridemia in male and in female rats subjected to a diet enriched in saccharose and fat. Estrogen-induced hypertriglyceridemia in castrated animals is strongly inhibited if the diet is rich in linoleic acid. Physical effort can prevent saccharose combined with saturated fats from inducing hypertriglyceridemia.

Production and purification of bovine monocyte-derived interleukin 1.

Few studies have addressed the biological and molecular nature of bovine interleukin 1 (IL-1). In an effort to increase our understanding of the role of bovine IL-1 in bovine immunology, we investigated various parameters of its production by LPS-stimulated monocytes in vitro. Bovine monocytes isolated by our methods constitutively released IL-1 activity, as measured by the murine thymocyte IL-1 assay. Monocyte release of IL-1 activity was further augmented when the cells were incubated with 0.005-10 micrograms per ml of Escherichia coli lipopolysaccharide (LPS). The presence of 1, 5, or 10 percent heat-inactivated fetal bovine serum (FBS) enhanced LPS-stimulated bovine monocyte release of IL-1 activity as compared with monocytes cultured under serum-free conditions. We used a combination of size-exclusion and reverse-phase high-performance liquid chromatography (HPLC) to purify bovine IL-1 from serum-free monocyte culture supernatants. Size-exclusion HPLC resulted in a single peak of biological activity with an approximate molecular weight of 18,000 daltons. Further purification by reverse-phase HPLC demonstrated at least three major molecular species with IL-1 activity. Besides providing information about production of IL-1 by bovine monocytes in vitro, this study also describes a protocol to purify bovine IL-1 for future studies addressing its biological functions.