concanavalin-a and Burns

Immature myeloid cells have been implicated as a source of postburn inflammation, and the appearance of these cells correlates with enhanced upregulation of hematopoiesis. The role of proliferative cells in postburn immune changes has not been directly tested. Gemcitabine, a ribonucleotide reductase inhibitor, has been shown to deplete proliferative immature myeloid cells in tumor models while sparing mature cells, leading to restored lymphocyte function and tumor regression. We treated burn mice at postburn day 6 (PBD6) with 120 mg/kg gemcitabine. On PBD8, splenocytes were taken and stimulated with LPS, peptidoglycan, or concanavalin A. The blood and spleen cell populations were enumerated by flow cytometry or automated cell counter. In addition, mice treated with gemcitabine were given LPS or infected with Pseudomonas aeruginosa at PBD8, and mortality was monitored. Gemcitabine depleted burn-induced polymorphonuclear leukocytes and inflammatory monocytes without affecting mature F4/80 macrophages. This was accompanied by reduced TNFα, IL-6, and IL-10 production by burn splenocytes. Burn splenocytes stimulated with mitogens exhibited increased nitric oxide production relative to sham mice. In vivo treatment of burn mice with gemcitabine blocked these burn-induced changes without damaging lymphocyte function. Treatment of burn mice with gemcitabine ameliorated burn-induced susceptibility to LPS and infiltration of polymorphonuclear leukocytes into the liver and lung. Finally, gemcitabine treatment blocked the protective effect of burn injury upon P. aeruginosa infection. Our report shows that proliferative cells are major drivers of postburn immune changes and provides evidence that implicates immature myeloid cells in these processes.

Topics: Animals; Burns; Concanavalin A; Deoxycytidine; Drug Evaluation, Preclinical; Gemcitabine; Interleukin-10; Interleukin-6; Leukocyte Count; Lipopolysaccharides; Lymphocyte Activation; Macrophages; Male; Mice; Mice, Inbred C57BL; Monocytes; Myeloid Cells; Nitric Oxide; Peptidoglycan; Pseudomonas Infections; Ribonucleotide Reductases; Spleen; T-Lymphocyte Subsets; Tumor Necrosis Factor-alpha

Major physical traumas provoke a systemic inflammatory response and immune dysfunction. In a model of thermal injury in rats, we previously showed that an overproduction of nitric oxide (NO) was responsible for the collapse of lymphoproliferative responses. In the present work, we performed a time-course analysis of cell proliferation and cell death parameters in order to establish the sequence of events triggered by the high NO output in Wistar/Han rat splenocytes activated with Con A, 10 days after burn injury. We demonstrate that activated T cells from burned rats never divided whereas normal T cells underwent four division cycles. However, T cells from both burned and normal rat entered the G1 phase as shown by increase of cell size, mitochondria hyperpolarization, and expression of cyclin D1. Burned rat T cells progressed to the late G1 phase as shown by expression of the nuclear Ki-67 antigen, but they never entered the S phase. They underwent apoptosis as shown by morphological parameters, disruption of transmembrane mitochondrial potential, and DNA fragmentation. Persistent accumulation of the p53 protein accompanied these phenomena. NO synthase inhibitors antagonize alterations of cell proliferation and cell death parameters in burned rat T cells and accelerated p53 turnover.

Topics: Animals; Apoptosis; Burns; Cell Division; Cell Proliferation; Concanavalin A; Enzyme Inhibitors; Flow Cytometry; G1 Phase; Interleukin-2; Isothiuronium; Lymphocyte Activation; Male; Models, Biological; Necrosis; Nitric Oxide; Rats; Rats, Wistar; Spleen; T-Lymphocytes; Thiazines; Tumor Suppressor Protein p53

Cutaneous burn injury-induced T lymphocyte suppression is a well-known phenomenon. In this study, we evaluated the effect of treatment of burn rats with pentoxifylline (PTX) on the burn-induced suppression of T lymphocytes. Anesthetized rats were subjected to 30% total body surface area burn by exposing skin to 95 degrees C water for 10 s. T lymphocytes were isolated from sham and burn rats with or without PTX treatment (120 mg/kg, ip). T cell proliferation and interleukin (IL)-2 production in response to T cell mitogen concanavalin A was measured using 3 H-thymidine uptake and enzyme-linked immunosorbent assay, respectively. P59 fyn autophosphorylation and its kinase activity was determined using in vitro kinase assay. In addition, T lymphocyte Ca2+ signaling was assessed using Ca2+ imaging technique. Two days after injury, there was a significant decrease in mesenteric lymph node T cell proliferation and IL-2 production in burn injured rats compared with those obtained from sham-injured rats. This decrease in T cell proliferation and IL-2 production in burn-injured rats was accompanied by a significant suppression in both P59 autophophorylation and kinase activity as well as Ca2+ signaling. Treatment of burn-injured rats with PTX produced a near complete recovery of T cell proliferation and IL-2 production. Furthermore, PTX treatment also prevented the burn-mediated suppression in P59fyn and kinase activity as well as restored Ca2+ signaling similar to those observed in sham injured rats. These findings altogether suggested that PTX treatment attenuate T cell suppression in burn-injured rats and that the effects of PTX are mediated via modulating P59 fyn and Ca2+ signaling.

Topics: Animals; Burns; Calcium; Cell Division; Concanavalin A; Hematologic Agents; Immunoblotting; Interleukin-2; Lymph Nodes; Lymphocyte Activation; Male; Pentoxifylline; Phosphorylation; Precipitin Tests; Rats; Rats, Sprague-Dawley; Signal Transduction; T-Lymphocytes

The laying down of collagen and fibrous tissue is a key process in wound healing, however excessive collagen (and glycoprotein) deposition causes hypertrophic and keloid scars, eg after burns. Collagen synthesis is increased in these scars compared with normal healing, as is collagenase activity, which controls the degradation pathway of collagen. The processes of wound healing are inextricably linked to those of the acute-phase response (APR): alpha-1-acid glycoprotein (AGP), a plasma glycoprotein that undergoes both an increase in concentration and an alteration in its glycosylation pattern during the APR. This study determined that AGP isolated from the plasma of burns patients was of an increased concentration and altered glycosylation pattern compared with normal plasma and was capable of directly interacting with type I collagen. It also had a profound effect on both collagen fibril formation and collagenase activity, to a degree dependent upon the percentage body surface area burned. Additionally, the results obtained provided the basis for predicting the formation of hypertrophic scars.

Topics: Burns; Chromatography, Ion Exchange; Collagen; Concanavalin A; Enzyme-Linked Immunosorbent Assay; Glycosylation; Humans; Hydrogen-Ion Concentration; Kinetics; N-Acetylneuraminic Acid; Orosomucoid; Wound Healing

Previous studies from our laboratory established that low-fat diets prevent immunosuppression and reduce oxidative stress after a thermal injury. The purpose of the present study was to test the hypothesis that the type of dietary fatty acid influences splenocyte proliferation and oxidative stress following a burn injury. Female C3H/HeN mice were fed ad libitum six experimental diets (5% w/w lipids) differing in fatty acid composition for 10 days following a burn injury. Compared to the controls, burned mice fed whichever diet showed lower lymphoproliferative responses to concanavalin-A (Con-A) and lipopolysaccharide (LPS) (p<0.01), but not to an anti-T cell receptor monoclonal antibody (H-57). In burned animals, nitric oxide (NO) concentration was negatively correlated to the proliferation induced by Con-A (p<0.01) or LPS (p<0.05). These results suggest that: (1) dietary fatty acid type does not influence the splenocyte proliferation or oxidative stress and (2) NO production is involved in the immunosuppression following burn injury.

Topics: Animals; Burns; Concanavalin A; Dietary Fats; Fatty Acids; Female; Lipopolysaccharides; Lymphocyte Activation; Mice; Mice, Inbred C3H; Nitric Oxide; Oxidative Stress; Random Allocation; Spleen

Burn injury induces immune dysfunction and alters numerous physiological parameters. While clinical studies indicate that burn injury size profoundly impacts patient immune status, only limited experimental studies have systematically addressed its impact on immune functional parameters. In the present study, mice were subjected to burn injuries of varying sizes and splenic immune cells (splenocytes and macrophages) were isolated 7 days thereafter. Burn injury suppressed splenic T-cell proliferation in an injury size-dependent manner that correlated with the release of the immunosuppressive mediators PGE(2) and nitric oxide. In addition, a shift towards an immunosuppressive Th-2 cytokine profile and a hyperactive macrophage phenotype (increased release of inflammatory mediators) was observed post-injury, however, this effect was in part independent of burn size. Thus, unlike patient survival data, burn injury-induced changes in immune function do not necessarily correlate with the size of the injury.

Topics: Animals; Burns; Concanavalin A; Dinoprostone; Female; Immunologic Deficiency Syndromes; Lipopolysaccharides; Lymphocyte Activation; Lymphokines; Macrophage Activation; Mice; Mice, Inbred C57BL; Models, Animal; Nitric Oxide; Spleen; Th1 Cells; Th2 Cells

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.

Topics: Animals; Burns; CD4-Positive T-Lymphocytes; Cells, Cultured; Concanavalin A; Gene Expression Regulation; Immunity, Cellular; Interleukin-2; Lymphocyte Activation; Male; Mice; Mice, Inbred A; NF-kappa B; Reverse Transcriptase Polymerase Chain Reaction; Spleen; T-Lymphocytes; Time Factors; Transcription Factor AP-1; Transcription, Genetic

To evaluate the effect of burn injury with and without an Escherichia coliseptic complication on T-cell proliferation, interleukin-2 production, and Ca(2+) signaling responses in intestinal Peyer's patch and splenic T cells.. Prospective, randomized, sham-controlled animal study.. University medical center research laboratory.. Adult male Sprague-Dawley rats.. Rats were subjected to a 30% total body surface area, full skin thickness burn. Infection in rats was induced via intraperitoneal inoculation of E. coli, 10(9) colony forming units/kg, with or without a prior burn.. Rat Peyer's patch and splenic T lymphocytes were isolated by using a nylon wool cell purification protocol. T-cell proliferation, interleukin-2 production, and Ca(2+) signaling responses were measured after stimulation of cells with the mitogen, concanavalin A. T-cell proliferation was determined by measuring incorporation of (3)H-thymidine into T-cell cultures. Interleukin-2 production by T-cell cultures was measured by using enzyme-linked immunosorbent assay. Intracellular T-cell Ca2(+ )concentration, [Ca(2+)](i), was measured by the use of Ca(2+)-specific fluorescent label, fura-2, and its fluorometric quantification. [Ca(2+)](i) was also evaluated by the use of digital video imaging of fura-2 loaded individual T cells. T-cell proliferation and interleukin-2 production were suppressed substantially in both Peyer's patch and splenic T cells 3 days after either the initial burn alone or burn followed by the E. coli inoculation at 24 hrs after the initial burn. There seemed to be no demonstrable additive effects of E. coli infection on the effects produced by burn injury alone. The T-cell proliferation and interleukin-2 production suppressions with burn or burn-plus-infection insults were correlated with attenuated Ca(2+) signaling. E. coli infection alone suppressed T-cell proliferation in Peyer's patch but not in splenic T cells at 2 days postbacterial inoculation; E. coli infection had no effect on Peyer's patch or splenic T cells at 1 day postinjury. On the other hand, burn injury alone caused a substantial T-cell proliferative suppression at 2 days postburn in both Peyer's patch and splenic cells and a significant suppression in T-cell proliferation on day 1 postburn in Peyer's patch but not in the spleen.. An initial burn injury suppressed T-cell proliferation at a level that it would not be further affected by a subsequent infection even if the infection by itself has the potential of suppressing T-cell proliferation. An earlier onset of T-cell suppression in Peyer's patch cells than in the spleen with burn could be attributable to an initial hypoperfusion-related intestinal mucosal tissue injury. Overall, our study supports the concept that burn injury per se can significantly suppress T-cell mediated immunity and that the intestine is an early tissue site of such suppression.

Topics: Analysis of Variance; Animals; Burns; Calcium Signaling; Concanavalin A; Escherichia coli Infections; Immune Tolerance; Interleukin-2; Male; Peyer's Patches; Random Allocation; Rats; Rats, Sprague-Dawley; Sepsis; Spleen; Statistics, Nonparametric; T-Lymphocytes

The gender difference in normal immune function has been well documented, however, there is only limited information regarding whether such a difference occurs after injury. To investigate this, we examined cell-mediated immune responses in male and female mice given a 15% total body surface area dorsal scald or sham injury. Both delayed-type hypersensitivity (DTH) and splenocyte proliferative responses were significantly suppressed in males at 1 day and in females at 7 and 10 days post burn (P < 0.01). The decreased splenocyte proliferation was found to be macrophage-dependent and suppression of both immune parameters corresponded with elevated interleukin-6 (IL-6) levels. Furthermore, post-burn treatment with an anti-IL-6 antibody partially restored the DTH response in males at 1 day and females at 10 days post injury and completely restored splenocyte proliferation. These data demonstrate a possible mechanism for the gender difference in cell-mediated immune responses after thermal injury.

Topics: Animals; Antibodies; Burns; Cell Division; Concanavalin A; Female; Hypersensitivity, Delayed; Immunity, Cellular; Interleukin-6; Lymphocyte Activation; Macrophages; Male; Mice; Mice, Inbred BALB C; Sex Characteristics; Spleen; Temperature; Time Factors

The purpose of this study was to characterize the impact of a low-fat (LF; 1% fat) diet, a high-fat (HF; 25% fat) diet, and a standard (SD; 5% fat) diet on immune and oxidative parameters in a 20% body surface area burn animal model fed ad libitum for 10 days postinjury. Although the mechanisms are poorly understood, the amount of dietary lipid in nutritional support has been shown to have immunomodulatory effects after burn injury. Burned mice fed the LF diet showed a normal response in activated splenocyte proliferation compared to burned animals that received the SD or HF diet. Animals fed the SD and HF diets presented increased production of nitric oxide and prostaglandin E2 response after burn injury, which is associated with inhibited splenocyte proliferation. The total thiol concentration in spleen cells from burned animals kept on the HF diet was significantly higher than that in unburned animals, while no increase in these oxidative parameters was observed in LF-fed burned animals. Moreover, the LF diet significantly reduced hepatic lipid peroxidation, as measured by malonaldehyde concentration, compared to the other two diets. These results suggest that the administration of a LF diet in mice after a burn injury prevents inhibition of in vitro splenocyte proliferation and reduces the intensity of oxidative stress.

Topics: Animal Feed; Animals; Burns; CD4-CD8 Ratio; Concanavalin A; Corn Oil; Diet, Fat-Restricted; Dietary Fats; Dinoprostone; Disease Models, Animal; Eating; Female; Immune Sera; Immunophenotyping; Lipopolysaccharides; Lymphocyte Activation; Mice; Mice, Inbred C3H; Nitric Oxide; Oxidative Stress; Receptors, Antigen, T-Cell; Spleen; Weight Gain

Major burn injury induces T-lymphocyte dysfunction. Previous studies suggest that prostaglandin (PG) E2, which is elevated post-burn, is the causative factor via a cyclic AMP-dependent process. The present study was conducted to elucidate the mechanism by which cAMP induces T-lymphocyte dysfunction following burn injury. Splenocytes were isolated from mice 7 days after receiving a scald burn covering 25% of their total body surface or sham procedure. ConA-induced proliferation by splenocytes from burned mice was significantly suppressed. Macrophage depletion of the splenocyte cultures abrogated the suppression. Concanavalin A-stimulated proliferation by macrophage-depleted splenocytes was suppressed by PGE2 and dibutyryl cAMP in both groups. The IC50 of these cAMP-elevating agents, however, was approximately 100-fold lower for cells from burned mice, indicating an increased sensitivity to cAMP. PGE2 did not suppress PMA/Ca2+ ionophore-induced T-lymphocyte activation. Addition of PMA to ConA-stimulated cultures prevented the suppression of proliferative responses by PGE2, whereas Ca2+ ionophore had no effect. Thus, the suppression of T-lymphocyte activation following burn injury is macrophage-dependent, related to an increased sensitivity to cAMP and due to an uncoupling of cell surface receptors from protein kinase C activation.

Topics: Animals; Burns; Concanavalin A; Cyclic AMP; Dinoprostone; Female; Immunosuppression Therapy; Interleukin-2; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred C57BL; Protein Kinase C; T-Lymphocytes

Macrophages (Mphi) have been implicated in the suppression of lymphocyte function following thermal injury. Splenocytes isolated from C57BL/ 6NCR female mice 4-7 days after thermal injury displayed suppressed proliferative responses to Concanavalin A (ConA) and lipopolysaccharide (LPS) and high levels of reactive nitrogen intermediate (RNI) production. Inhibition of nitric oxide synthase activity with NG-monomethyl-L-arginine restored ConA responses but not LPS responses. Surprisingly, ConA-stimulated interferon-gamma (IFN-gamma) production was increased in splenocytes from injured mice. IFN-gamma contributed to the RNI-mediated immunosuppression as antibodies against IFN-gamma reduced RNI production and immunosuppression. ConA-stimulated co-cultures of splenic Mphi from injured mice and normal splenocytes produced high levels of RNI only under conditions of cellular contact and splenic Mphi from injured mice were capable of suppressing normal splenocytes responses in co-culture. These results indicate that Mphi activity and specifically RNI production contribute to the suppression of T lymphocyte function after thermal injury.

Topics: Animals; Burns; Concanavalin A; Cytokines; Enzyme Inhibitors; Female; Immunosuppression Therapy; Interferon-gamma; Lipopolysaccharides; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred C57BL; Nitric Oxide Synthase; Nitrogen Oxides; omega-N-Methylarginine; Spleen

Immunodeficiency follows extensive burns. We investigated some underlying mechanisms in rats, 10 days after a full-thickness skin burn affecting 20% of total body surface area. In both normal and burned rats the splenocyte proliferative response to Con A was linearly and negatively correlated with nitric oxide (NO) production. In all burned rats, the proliferative response was depressed by more than 80% and NO production corresponded to a nitrite concentration above 20 microM. Proliferative responses in burned rats were fully restored in the presence of 250 microM NG-monomethyl-L-arginine (NMMA). A time course study of NO production in response to Con A, LPS, anti-CD3, and IFN-gamma showed that splenic macrophages from burned rats responded to direct and indirect stimuli more rapidly and more intensively than normal macrophages. In the second part of this work, the effect of the overproduction of NO on the synthesis of immunoregulatory and proinflammatory cytokines was investigated. Although it was inhibited, IFN-gamma production by splenocytes from burned rats remained sufficient for NO synthase induction and was restored by NMMA. Concomitantly, IL-2 concentration was enhanced but returned to normal in the presence of NMMA. TNF production was halved after burn injury and NMMA partially restored it. In contrast, IL-6 production was enhanced and increased further in the presence of NMMA. Therefore, cytokines were differently affected by burn injury and variously regulated by NO.

Topics: Animals; Burns; Cell Division; Cells, Cultured; Concanavalin A; Cytokines; Dexamethasone; Dinoprostone; Glucocorticoids; Immune Tolerance; Interferon-gamma; Interleukin-2; Interleukin-6; Lipopolysaccharides; Macrophages; Male; Nitric Oxide; Rats; Rats, Wistar; Spleen; Time Factors; Tumor Necrosis Factor-alpha

Patients with severe trauma or polytrauma frequently acquire alterations in immune functions which are correlated to dysbalanced cytokine synthesis. In these settings the role of polymorphonuclear neutrophil granulocytes (PMN) as cytokine-producing cells is less well characterized. The immunosuppressive role of interleukin (IL)-10 is well known, and increased systemic IL-10 levels are related to the severity of injury and to posttraumatic complications. We determined concentrations of IL-10 in culture supernatants of 30 individual PMN fractions isolated from 18 severely traumatized patients (15 polytraumata, Injury Severity Score: 18-41, 3 severely burned patients) admitted to intensive care units. IL-10 was analyzed by ELISA (R&D Systems, Wiesbaden, Germany). PMN were isolated from EDTA-anticoagulated peripheral blood employing a one-step procedure based on a discontinuous double Ficoll gradient. The cells [1 x 10(6)/ml RPMI 1640 supplemented with 10% fetal calf serum and 25 mM N-(2-hydroxyethyl)-piperazine-N'-(2-ethanesulfonic acid] were stimulated with 0.05% heat-killed Staphylococcus aureus (Pansorbin, Calbiochem-Novabiochem, Bad Soden, Germany) for 24 h using cell culture conditions. Our results show that PMN fractions of traumatized patients produce significantly (P<0.008) higher amounts of IL-10 (354+/-95 pg/ml, n = 30) than normal healthy donor cells (125+/-95 pg/ml, n = 7). IL-10 release from PMN fractions exceeded the release from isolated patients' peripheral blood mononuclear cells induced by similar stimulation or by stimulation with toxic shock syndrome toxin-1 (10 ng) and concanavalin A (2 microg). Our results provide evidence that PMN fractions play an active role in the development of posttraumatic immunosuppression by autocrine or paracrine mechanisms, for example, by suppressing one's own antimicrobial activities or determining the development of T-cell responses via their ability to release IL-10.

Topics: Adolescent; Adult; Aged; Bacterial Toxins; Burns; Concanavalin A; Enterotoxins; Enzyme-Linked Immunosorbent Assay; Female; Humans; Interleukin-10; Interleukin-6; Leukocytes, Mononuclear; Lymphocyte Activation; Male; Middle Aged; Multiple Trauma; Neutrophils; Staphylococcus aureus; Superantigens

The purpose of this study was to try to elucidate a possible biobehavioral mechanism associated with decreased immune function in trauma patients by determining whether there is an interaction between the effects of ACTH, a stress hormone, and TGF beta, a cytokine, on peripheral blood lymphocyte proliferation. Peripheral mononuclear lymphocytes (PMLs) from healthy donors were preincubated with varying concentrations of ACTH for 24 hr, stimulated with concanavalin A and increasing concentrations of TGF beta, and incubated for 72 hr. Proliferation was assayed by tritiated thymidine incorporation. A parallel aliquot of PMLs were incubated in the presence of ACTH to determine the direct effect of ACTH on mononuclear cell TGF beta production. While harvested supernatant from cells incubated in the presence of ACTH did not contain any detectable TGF beta, ACTH as well as TGF beta were found to significantly decrease cellular proliferation independent of one another. An even greater decrease in cellular proliferation was found when both ACTH and TGF beta were used, compared to either ACTH or TGF beta alone. These results suggest a biobehavioral interaction between ACTH and TGF beta at the cellular level and that interactions to relieve stress may assist in improving function and recovery from trauma.

Topics: Adrenocorticotropic Hormone; Analysis of Variance; Burns; Cell Division; Cells, Cultured; Concanavalin A; Dose-Response Relationship, Drug; Drug Interactions; Humans; Monocytes; Psychoneuroimmunology; Reference Values; Transforming Growth Factor beta; Wounds and Injuries

Serious traumatic or thermal injury is associated with depression of cellular immunity, including the failure of T-lymphocyte proliferation in response to stimulation that depends both on production of interleukin-2 (IL-2) and on expression of functional IL-2 receptors (IL-2R). While decreased IL-2 production following thermal injury is undisputed, the status of IL-2R expression and function in this setting is controversial; therefore, we sought to investigate this issue.. A total of 220 male A/J mice (n = 22 per group) were subjected to a 20% scald burn injury or sham burn, killed 4, 7, 10, 14, or 21 days later, and splenocytes harvested. In vitro parameters of both IL-2R expression and function were measured.. On day 7, splenic lymphocyte proliferation and IL-2 production in response to mitogenic stimulation were both suppressed following burn injury to 50% and 60% of controls, respectively. Northern blot analysis revealed normal IL-2R p55 messenger RNA expression in response to mitogenic stimulation on days 7, 10, and 14 in thermally injured animals. Phenotypic IL-2R p55 expression in concanavalin A-stimulated CD3+ cells was unchanged following burn injury. Binding of fluorescein-labeled IL-2 to cell membranes was increased in burned animals at days 10 and 14. The addition of IL-2 to cultures of spleen cells from burned mice consistently restored the mitogenic response to that of the controls.. Thermal injury in this model does not result in either quantitative or functional suppression of IL-2R. Suppression of T-cell activation and proliferation, seen following thermal injury, appears primarily related to abnormal IL-2 production.

Topics: Animals; Burns; Cell Division; Cell Membrane; Concanavalin A; Gene Expression Regulation; Interleukin-2; Lymphocyte Activation; Lymphocytes; Male; Mice; Mice, Inbred A; Mice, Inbred Strains; Receptors, Interleukin-2; RNA, Messenger; Spleen; T-Lymphocytes; T-Lymphocytes, Helper-Inducer

Cyclic adenosine monophosphate (cAMP) is an intracellular second messenger that is known to convey inhibitory signals for T-cell proliferation and function. We investigated the association between this molecule and the profound immunosuppression that accompanies thermal injury.. Mice were randomized into two groups: one group was subjected to a 20% full-thickness scald burn; the second to a sham burn (control). The mice were killed on days 4, 7, or 10 after the burn injury and splenocytes were pooled and cultured for 15 minutes in the presence or absence of prostaglandin E2 (PGE2).. Levels of cAMP in splenocytes were significantly elevated on day 7 after burn in the burn group compared with the sham controls (P < .05, Wilcoxon Rank Sum Test). Incubation of splenocytes with PGE2 resulted in significantly greater levels of intracellular cAMP in cells from the burn group compared with controls on days 4, 7, and 10. Incubation of normal splenocytes with dibutyryl cAMP in the presence of concanavalin A significantly decreased cell proliferation and the production of interleukin-2. The decrease in interleukin-2 production was evident at the level of messenger RNA expression. Stimulation of splenocytes with a combination of phorbol ester and calcium ionophore, bypassing all membrane-associated events prior to protein kinase C activation, reversed the inhibitory effects of dibutyryl cAMP. Incubation of splenocytes from burned animals with H-8, a selective inhibitor of cAMP-dependent protein kinases, restored the proliferative response to that of sham controls on days 4, 7, and 10 after thermal injury.. These data indicate that elevated levels of intracellular cAMP, combined with an increased production of cAMP in response to circulating PGE2, may play a fundamental role in suppression of the immune response following thermal injury and that cAMP exerts its immunomodulatory effects prior to protein kinase C activation.

Topics: Animals; Burns; Concanavalin A; Cyclic AMP; Dinoprostone; Disease Models, Animal; Gene Expression Regulation; Immune Tolerance; Immunity, Cellular; Interleukin-2; Lymphocyte Activation; Male; Mice; Mice, Inbred Strains; Phorbol Esters; Random Allocation; RNA, Messenger; Spleen

It is well accepted that nutritional support improves the immunologic functions in burned patients. Arginine has been demonstrated to have tissue-specific properties which induce beneficial effects upon the immune system. A series of experiments are carried out, in order to evaluate the immune effect of arginine on burned mice. 1. Lymphocytes from both burned and unburned mice are harvested and incubated in various concentrations of arginine solution for 72 hours, and ConA-stimulating lymphocyte transformation is determined after incubation. 2. Burned animals are divided into four groups. Standard diets for nutritional support are formulated. These formulas contain an identical carbohydrate and lipid (61% and 15% of total energy respectively) intake with varied proportions of protein for each group (24%, 23%, 22%, and 20% of total energy, respectively), in addition. 0%, 1%, 2% and 4% of protein energy is supplied by arginine for different groups. Lymphocyte proliferations in responses to ConA stimulation are determined on the 7th postburn day. The data shows that, in vitro study, increasing arginine concentrations enhanced lymphocyte responses to ConA. The arginine needed level for optimal lymphocyte responses is 1.8 mmol/L in burned mice and 0.9 mmol/L in unburned controls. This difference indicates that in burned mice, a higher arginine concentration is required for better lymphocyte responses. On the contrary, further increase of arginine concentration do not give better response. When varied amounts of arginine are given in the diet to the burned mice, the degree of lymphocyte response to ConA is different. The amount of arginine which supplies 2% of total energy is found to be optimal.

Topics: Animals; Arginine; Burns; Concanavalin A; Food, Formulated; Lymphocyte Activation; Mice; Mice, Inbred BALB C; T-Lymphocytes

The presence of increased levels of suppressor T cells after thermal injury and their relevance remain controversial. It is unclear whether suppressor T cells are the cause or result of sepsis complicating thermal injury. Spleen cells from a standardized murine burn model and sham burn controls were studied and the relationship between the levels of suppressor cytotoxic T cells (CD8, Lyt-2+), helper T cells (CD4, L3T4+), response to concanavalin A (ConA) and to phytohemagglutinin (PHA) and interleukin-2 (IL-2) production was examined. Mortality following infection via cecal ligation and puncture (CLP) of matched controls was also studied. At day 7 postburn, mean ConA (70 +/- 12% of control) and PHA response (58% +/- 5.2% of controls) and IL-2 production (43% +/- 5.4%) were significantly less than sham burn values (100%; p less than 0.05). However, the mean percentage of cells staining with anti-Lyt-2 and anti-L3T4 (9.1 +/- 0.59 and 13.9 +/- 0.65) was similar to the mean percentage in sham burn animals (9.4 +/- 0.65 and 16.6 +/- 1.1). Furthermore, no significant differences were observed between burned mice and controls in helper (17.3% +/- 1.8% burn vs. 21.2% +/- 1.7% sham) or suppressor cell levels (7.8% +/- 1.2% burn vs. 8.6% +/- 0.7% sham) or helper-suppressor ratios on day 10 postburn. Mortality of 20 litter-matched controls subjected to CLP on day 10 postburn was 90%, which was significantly greater than the sham burn mortality of 20%.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Antibodies, Monoclonal; Biomarkers; Burns; CD4-CD8 Ratio; Concanavalin A; Disease Models, Animal; Evaluation Studies as Topic; Immunologic Deficiency Syndromes; Infections; Interleukin-2; Lymphocyte Activation; Male; Mice; Phenotype; Phytohemagglutinins; Predictive Value of Tests; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory

Postburn immunosuppression in mice with full skin thickness burns was assessed after Con A stimulation of splenic lymphocytes. The immunosuppression was detected within 24 h of injury and was maximal 3-7 days later. Such lymphocyte proliferation was enhanced by depletion of adherent cells (M phi) in burned mice and could be depressed in normal mice by injection of these adherent cells from burned mice. These results suggest the presence of M phi s with a suppressor function. No difference in spontaneous blastogenic transformation (SBT) of spleen cells between the burned and normal mice was found, but after removal of the M phi s the SBT of spleen cells of burned mice became significantly elevated. Hence both M phi and lymphocytes in mouse spleen cells were activated after severe thermal injury.

Topics: Animals; Burns; Cell Adhesion; Concanavalin A; Dinoprostone; Humans; Immune Tolerance; Macrophages; Male; Mice; Mice, Inbred C57BL; Spleen

The lipid-protein complex (LPC) formed by thermal injury to skin, which has been shown to have a toxic effect on mice, and which suppresses the immune response, was tested for its specific influence on monocytes. Growth of bacterial endotoxin-stimulated peripheral blood mononuclear cells (PBMC) was inhibited in the presence of LPC, however, the inhibition was less at the time of the optimal rate of cell proliferation. Inhibition was proportional to LPC concentration. ConA-stimulated PBMC were also inhibited by LPC in a dose-related manner. PBMC, in the presence of LPC, secreted interleukin 1 (IL1) at an increasing rate as LPC concentration rose from 5 to 40 micrograms/ml, and the levels of IL1 which could be induced by endotoxin were increasingly amplified in the presence of LPC. In comparison to LPC, the native tissue proteins which were isolated from unburned skin by the same techniques which produced LPC from burned skin, were tested for their effect on PBMC. Native proteins had no effect on IL1 secretion, whether on background or endotoxin-stimulated levels. Thus, the thermally induced change in skin proteins has a specific effect on monocyte IL1 secretion which is not matched by the native proteins, indicating that burn injury to skin specifically affects the lymphokine cascade and consequent immune function.

Topics: Animals; Burns; Cell Division; Cells, Cultured; Concanavalin A; Humans; Interleukin-1; Membrane Lipids; Membrane Proteins; Monocytes; Skin

The relation between interleukin-6 (IL-6) levels and changes in serum concentrations and glycosylation (concanavalin A affinity) of two human acute-phase glycoproteins, alpha 1-acid glycoprotein (AGP) and alpha 1-protease inhibitor (PI), was studied in sequential serum samples of burn patients. The level of IL-6 was already increased at the first day following injury, and after a dip at day 2 or 3 rapidly reached a second maximal value at day 4 or 5. The serum concentrations of AGP and PI reached their maximal values after day 5 and remained at a high level throughout the total period studied (7 weeks). The concanavalin A reactivities of both acute-phase glycoproteins were found to be elevated only during the first 2-2.5 weeks. Maximal values were observed on day 2 and from day 7 to 16, following closely the rise and fall of the IL-6 serum level. After day 16, the concanavalin A affinity rapidly declined long before a decrease was observed in the serum concentrations of AGP and PI. Our previous in vitro studies have indicated an involvement of IL-6 in the induction of both secretion and increased concanavalin A affinity. This study indicates that IL-6 could play a causal role in the induction of both phenomena in vivo.

Topics: Adult; alpha 1-Antitrypsin; Burns; Concanavalin A; Female; Humans; Immunoelectrophoresis, Two-Dimensional; Interleukin-6; Male; Orosomucoid

Because of the association between the development of an immunocompromised state and an increased risk of infection, increasing attention has been focused on describing and characterizing the immune consequences of thermal injury. Results of human studies are largely based on the in vitro responsiveness of peripheral blood leukocytes, while splenocytes are generally used in the animal studies. Because the response of lymphocytes from different lymphocyte compartments may vary, we compared the responses of murine peripheral blood, splenic, Peyer's patch, and mesenteric lymph node lymphocytes to a battery of mitogens after thermal injury. Burn-induced immunosuppression was maximal in the splenic lymphocyte compartment, where the responses to all three test mitogens were depressed throughout the 28-day postburn study period. Although the PHA-induced mitogen response of lymphocytes from the other three lymphoid compartments remained suppressed throughout the study period, the response to the mitogens Con-A and PWM generally returned to normal or supranormal levels by the seventh postburn day, Therefore it appears that the effect of a thermal injury on lymphocyte function varies according to the lymphocyte compartment examined and the mitogen tested. These results raise the question of whether animal studies using splenic lymphocytes can be correlated with human studies performed on circulating blood lymphocytes.

Topics: Animals; Burns; Concanavalin A; Female; Lymph Nodes; Lymphatic System; Lymphocyte Activation; Male; Mesentery; Mice; Mice, Inbred BALB C; Mitogens; Peyer's Patches; Phytohemagglutinins; Pokeweed Mitogens; Spleen

Chronic alcoholics constitute a small but significant subgroup of burned patients. The effects of chronic alcohol exposure on immune function in burned patients has not to our knowledge been studied. This study was designed to determine the effect of chronic alcohol exposure before burn injury on immune function after injury in rats. Immune function assessed by in vivo chemotaxis and responsiveness of non-adherent splenocytes to both a T-cell mitogen, concanavalin A, and a B-cell mitogen, lipopolysaccharide, was measured at 4 days after a 20% BSA full-thickness burn injury and/or gavage of 2.4 gm/kg/day of ethanol for 14 days. Chronic ethanol ingestion before burn injury produced significant suppression in chemotaxis and response to lipopolysaccharide but not in response to concanavalin A. These results suggest that chronic alcohol exposure before injury can contribute to further impaired immune function after injury, and may lead to increased susceptibility to infection and increased mortality.

Topics: Alcoholism; Animals; Burns; Cells, Cultured; Chemotaxis, Leukocyte; Concanavalin A; Lipopolysaccharides; Male; Rats; Rats, Inbred Strains; Spleen

Human alpha 1-acid glycoprotein (AGP) has been shown to modulate various cellular and humoral immune reactions in vitro. Using glycosidase-modified derivatives of AGP, the importance of its carbohydrate moiety with regard to these effects has been noted. In normal serum, three molecular AGP forms interacting differently with concanavalin A (Con A) are present. The ratio of these forms is often changed during various physiopathological conditions. In this study, we could show that differences exist between the three AGP forms with regard to their immunomodulatory effectiveness. At physiological concentrations, the Con A-nonreactive variant AGP-A induced a stronger inhibition of the anti-CD3 stimulated lymphocyte proliferation than the other forms. Interestingly, AGP-A was also found to be responsible for the stimulation of lymphocyte proliferation induced by low AGP concentrations in vitro. Both immunomodulatory effects of AGP were abrogated by desialylation of the glycoprotein. These results support an immunomodulatory role of AGP in conditions characterized by a changed ratio of the differently glycosylated AGP forms.

Topics: Burns; Concanavalin A; Humans; Inflammation; Lymphocyte Activation; Orosomucoid

The time course of changes in the concentrations of individual acute-phase proteins (APRs) in the plasma was examined in the model of rats exposed to a single (sublethal) and repeated (fatal) scalding. These data were interrelated with the rate of survival, plasma volume, corticosterone level, and immunosuppressive potency of the serum. The infliction of a sublethal scalding composing 20% body surface area resulted in an initial 50% reduction of the plasma volume and a severalfold increase of the corticosterone level and immunosuppressive activity of the serum. A subsequent normalisation of these deviations proceeded in parallel with an increased rate of APR synthesis. An early infliction of a second scald was fatal. It led to a 60% reduction of the plasma volume, a lack of plasma APRs, and an additional enhancement of the immunosuppressive activity of serum.

Topics: Acute-Phase Proteins; Animals; Burns; Cell Division; Concanavalin A; Corticosterone; Glycoproteins; Male; Rats; Thymus Gland

The effect of in vivo administration of Isoprinosine (ISO) on, i) the proliferation of splenic lymphocytes in response to the T-cell mitogen, concanavalin-A (Con-A) and, ii) the natural killer (NK) cell cytotoxicity was studied following a full skin thickness burn injury in a rat model. Administration of ISO (100 mg/kg body wt/day) twice daily, resulted in significant augmentation of the proliferative responses of lymphocytes compared to non-treated burned animals, at 7 days post injury. However, it did not effect the lymphoproliferation at 14 days post injury, the time period at which a complete suppression of lymphocyte proliferation was observed in burned non-treated animals. Also, the proliferation of lymphocytes from normal nonburned animals was not affected by treatment with ISO. ISO treatment of the burned animals resulted in a significant increase in the NK cytotoxicity compared to non-treated burned animals. As with Con-A responses, ISO administered to control nonburned animals did not have any effect on NK cell cytotoxicity. Our studies thus indicate that ISO can be a potential immunomodulator of suppressed immune function following thermal injury, particularly in patients whose lymphocyte responses to T cell mitogen Con-A are not completely suppressed.

Topics: Adjuvants, Immunologic; Animals; Burns; Concanavalin A; Inosine; Inosine Pranobex; Killer Cells, Natural; Lymphocyte Activation; Rats; Rats, Inbred Strains; Time Factors

In serum from five patients with severe burns, alpha 1-proteinase inhibitor (alpha 1-PI) was analyzed and then isolated by immunosorption chromatography. By Con A-Sepharose chromatography alpha 1-PI was separated into two types of fractions: the first containing the Con A-non-reactive isoforms and the second containing the Con A-reactive isoforms. The increase of alpha 1-PI serum level in burn patients is associated on the fifth day after the burn with a significant shift toward species enriched in bi-antennary oligosaccharides (Con A-reactive isoforms). This latter change passed very quickly and ten days after the burn, whereas the alpha 1-PI serum level was still high, the difference in proportions of Con A-reactive and non-reactive isoforms was not statistically significant. With respect to the difference in oligosaccharide structure, it appeared that the glycan moiety was involved in the inhibitory effect on natural killer cell activity. At the same concentration, purified alpha 1-PI and retained alpha 1-PI isoforms had an equal effect, whereas the non-retained alpha 1-PI isoforms were more efficient (P less than or equal to 0.01). Purified alpha 1-PI and its isoforms inhibited the natural killer cell activity in a dose-dependent manner.

Topics: alpha 1-Antitrypsin; Blood Proteins; Burns; Concanavalin A; Dose-Response Relationship, Drug; Glycosylation; Humans; Killer Cells, Natural

We conducted studies to determine the effects of parenteral therapy with indomethacin, ibuprofen, and piroxicam on key immunologic and hematologic alterations induced by thermal injury. Drugs (10-20 mg/kg) or placebo were administered intramuscularly to thermally injured guinea pigs at 3 h postburn and then daily for nine days postburn. All three drugs inhibited production of 6-keto prostaglandin F1 alpha and thromboxane B2 in wound fluid and concomitantly restored the bactericidal activity of polymorphonuclear leukocytes (PMNLs) against Pseudomonas aeruginosa to normal. Indomethacin also increased the proliferative response of splenic lymphocytes to concanavalin A; however, ibuprofen and piroxicam had no effect on this response. None of the drugs affected the extent of systemic complement consumption, thrombocytopenia, leukocytosis, or leukopenia in the injured animals. These results suggest that the PMNL bactericidal defect induced by thermal injury is preventable or reversible and that the mechanisms responsible for this defect are inhibitable by nonsteroidal anti-inflammatory drugs.

Topics: 6-Ketoprostaglandin F1 alpha; Animals; Anti-Inflammatory Agents, Non-Steroidal; Blood Bactericidal Activity; Blood Cell Count; Burns; Complement System Proteins; Concanavalin A; Cyclooxygenase Inhibitors; Disease Models, Animal; Guinea Pigs; Ibuprofen; Indomethacin; Kinetics; Lymphocyte Activation; Neutrophils; Phagocytosis; Piroxicam; Pseudomonas aeruginosa; Spleen; Thromboxane B2

In serum from 8 severely burned patients, haptoglobin (Hp), alpha 1-acid glycoprotein (AG) and alpha 1-antitrypsin (AT) were found to be increased by factors of 5, 6 and 2 respectively. Ceruloplasmin (Cp) was slightly decreased. In order to appreciate possible modifications to the structure of their attached N-glycans, whole sera were fractionated on concanavalin A (Con A)-Sepharose and respective glycoproteins measured by laser nephelometry using a monospecific antiserum. In the serum from normal as well as burned patients Hp was almost entirely bound to the immobilized lectin (but eluted with 300 mmol/l alpha 1-methylglucoside) and Cp was bound at about 92%. For AG, in contrast, the fraction without affinity for Con A, 25% in normal serum, decreased to 5% in patients, whereas the retained species increased in proportion. A very weakly reactive fraction (which was only retarded and eluted without alpha-methylglucoside) amounted to 72% in both types of serum. When reduced and alkylated, this intermediate fraction gave rise to both non-retained and retained species always in a proportion of about 1/3. On the whole one concludes that there is a significant shift for AG in burned patients towards species enriched in bi-antennary (Con A-reactive) glycans. For AT a minor part was not recognized by the lectin and about 27% was retarded. The latter, which increased in burned patients, gave rise mainly to retained species after reduction and alkylation. This again suggests a shift to bi-antennary glycans.

Topics: Acute-Phase Proteins; alpha 1-Antitrypsin; Blood Proteins; Burns; Ceruloplasmin; Chromatography, Affinity; Concanavalin A; Glycoproteins; Haptoglobins; Humans; Immunoglobulins; Isoelectric Focusing; Polysaccharides; Sepharose

A second suppressive state (S-SupS) in interferon (IFN) response was demonstrated when mice with third-degree burns of approximately 30% of the body surface area (TI-mice) were stimulated in vivo by Staphylococcal enterotoxin A, a gamma IFN inducer. The first suppressive state in IFN production, appearing 3 to 7 days after thermal injury, was mediated by the generation of splenic suppressor macrophages. The S-SupS was demonstrated approximately 3 weeks post thermal injury. It persisted for almost 3 weeks and gradually disappeared by 7 weeks. Spleen cells from mice during the S-SupS produced less IFN in vitro than normal mouse splenic mononuclear cells (MNC) when stimulated with concanavalin A (Con A). Splenic MNC of mice during the S-SupS inhibited IFN production when they were co-cultured with normal mouse splenic MNC in the presence of Con A. Since this suppressor cell activity could not be removed from TI-mice splenic MNC by carbonyl iron treatment, or by a technique of adherence to a plastic surface, it is suggested that two different cell populations which are capable of suppressing the IFN response of TI-mice exist at different time periods following burn injury.

Topics: Animals; Burns; Cells, Cultured; Concanavalin A; Depression, Chemical; Enterotoxins; Interferon-gamma; Iron Carbonyl Compounds; Lymphocytes; Macrophages; Mice; Mice, Inbred BALB C; Organometallic Compounds; Spleen; T-Lymphocytes, Regulatory; Time Factors

We have previously described a mouse model of postburn suppression of cell-mediated immunity (CMI). Ear swelling (ES) in response to contact antigen challenge is depressed maximally 14 days following a 25% steam burn and recovers to control levels 3 weeks postburn. Splenic lymphocyte proliferation in response to Concanavalin A (Con A) is also depressed 14 days postburn. Splenic T-lymphocyte subset analysis with monoclonal antibodies for helper cells (Lyt 1.2) and suppressor cells (Lyt 2.2) reveals that T-helper cells reach a minimal level and T suppressor cells reach a maximum level 14 days postburn. The helper: suppressor ratio (HSR) reaches its nadir at day 14. Treatment of burned mice with low-dose cimetidine (2 or 10 mg/kg/day), but not high-dose (50 mg/kg/day), for 14 days restores CMI. Low-dose cimetidine also normalizes the HSR but does not effect postburn depression of mitogen responsiveness. Low-dose cimetidine probably restores CMI by inhibiting suppressor cells, whereas high doses provide more global inhibition. Recovery of mitogen responsiveness may require continued cimetidine presence in culture.

Topics: Animals; Burns; Cimetidine; Concanavalin A; Dinitrofluorobenzene; Female; Immunity, Cellular; Lymphocyte Activation; Mice; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory

Reduced in vitro T cell mitogen-induced transformation, low proportion of T cells and increased proportion of non-T cells were found in blood mononuclear cells of patients with severe burns 3-12 days after the injury. High spontaneous proliferation of non-T cells was observed and could be related mainly to the B cell fraction. Monocytes mediated suppression of mitogen-stimulated T cell proliferation. We further studied the role of monocytes in the enhanced suppressor activity of Con A-activated T cells and found that in this assay system, the patient's T cells mediated suppression in collaboration with monocytes. In vitro, increased suppressor function was probably the result of in vivo stimulation of inhibitory activity ascribed to both monocytes and T cells of patients. Addition of indomethacin to cell cultures markedly reduced suppression of lymphocyte proliferation. Less significant reduction was noted when the patient's T cells were activated in vitro by Con A. Adjuvant treatment of burn patients with indomethacin may play a role in alleviating suppression of immune response in these patients.

Topics: Adolescent; Adult; Aged; Burns; Cell Communication; Concanavalin A; Female; Humans; Immune Tolerance; Leukocyte Count; Lymphocyte Activation; Male; Middle Aged; Monocytes; T-Lymphocytes; T-Lymphocytes, Regulatory

As reported previously, gamma interferon (IFN-gamma) production was selectively decreased in thermally injured mice (TI-Mice) and spleen cell cultures from the mice following stimulation with various IFN inducers. The decrease in IFN production was associated with splenic suppressor macrophages. The present study demonstrated that TI-Mice treated with Ge-132 (TGe-Mice) produced IFN following stimulation with IFN-gamma inducers. The level of IFN activity detected in the sera of TGe-Mice approximated that of controls. Similar results were obtained when spleen cells prepared from TGe-Mice were stimulated with IFN-gamma inducers in vitro. While transfer of spleen cells from TI-Mice resulted in the suppression of IFN production in normal mice (N-Mice) stimulated with IFN-gamma inducers, the transfer of spleen cells derived from TGe-Mice did not induce suppression of IFN production in N-Mice. Mononuclear cells (MNC) prepared from N-Mice produced IFN following concanavalin A (Con A) stimulation when they were co-cultured with macrophage-enriched populations (PAC) obtained from the spleens of TGe-Mice. In contrast, MNC stimulated with Con A did not produce IFN activity when they were co-cultured with PAC of TI-Mice. These results suggest that the generation of suppressor macrophages in TI-Mice may be altered by the administration of Ge-132.

Topics: Animals; Antineoplastic Agents; Burns; Concanavalin A; Enterotoxins; Female; Germanium; Interferon Inducers; Interferon-gamma; Kinetics; Macrophages; Mice; Mice, Inbred BALB C; Newcastle disease virus; Organometallic Compounds; Poly I-C; Propionates

The immunosuppressive effect of burned tissue was studied using a mouse burn model. To evaluate the immunologic status an in vivo measure of cell-mediated immunity (CMI) involving contact sensitization of mice by painting the skin with dinitrofluorobenzene was used; mice were challenged 5 days later by painting the ear with the same antigen. Ear swelling in response to antigenic challenge was used as a quantitative measure of CMI; diminution in ear swelling in treatment mice compared to sensitized, unburned control mice indicated the degree of immunosuppression. A full-thickness steam burn covering 20% body surface ares (BSA) was profoundly immunosuppressive as reflected by ear swelling of 45 to 60% of that found in normal mice; partial thickness burns and burns of 10% BSA extent were not significantly immunosuppressive. Transfer into unburned mice of burned skin equivalent in size to a 20% BSA burn eschar resulted in marked immunosuppression, but transfer of smaller amounts of burned skin, or of larger amounts of unburned skin and normal and burned liver tissue, did not produce immunosuppression. Mice receiving a very high-temperature (300 degrees C), dry burn were only slightly more suppressed than mice receiving a standard steam burn. Normal immunity was preserved in burned mice which received daily application of cerium nitrate to the wound for 7 days, but application of other topical agents commonly used in burn treatment did not preserve immunity. Postburn immunosuppression thus appears related quantitatively to toxic factors in burned skin, and these toxic factors can be abrogated in burned mice by the topical application of cerium nitrate.

Topics: Animals; Burns; Concanavalin A; Female; Immune Tolerance; Immunosuppressive Agents; Lipopolysaccharides; Lymphocyte Activation; Mice; Polymyxin B; Stress, Physiological; Tissue Extracts

The effect of thermal injury on the response of interferon (IFN) production in vivo and in vitro after stimulation with eight representative inducers was investigated in a mouse model. The response of mice to immune IFN (IFN-gamma) inducers, staphylococcal enterotoxin A, concanavalin A, and a specific antigen for BCG-sensitized lymphocytes (purified protein derivative) was impaired after a 30% total body surface area third-degree burn. Suppression of IFN-gamma production was observed at day 2 and persisted until day 7 after burn. Decreased IFN-gamma production correlated closely with the percentage of total body surface area burned. When virus type IFN (IFN-alpha/beta) inducers, Newcastle disease virus, polyriboinosinic-polyribocytidylic acid, 10-carboxymethyl-9-acridanone, and E. coli endotoxin, were administered to mice, no change in IFN response was observed after thermal injury. Similar results were obtained when spleen cells obtained from thermally injured mice were stimulated with IFN-gamma inducers in vitro. These studies suggest that although the capacity for IFN-alpha/beta production remains intact in thermally injured mice, IFN-gamma production may be selectively decreased in burned animals and in their spleen cells.

Topics: Animals; Burns; Concanavalin A; Disease Models, Animal; Immune Tolerance; Interferon Inducers; Interferons; Mice; Mice, Inbred BALB C; Newcastle disease virus; Poly I-C; Staphylococcal Protein A; Time Factors; Tuberculin

This study was performed to investigate the cell-mediated immune response in burned patients with no septic episodes. The results show that burned patients with percentage body burn higher than 20 had an impaired lymphocyte reactivity to phytohaemagglutinin and conconavalin A. This hyporesponsiveness appeared on day 3-4 and in all cases reached its maximum on day 7-8 post burn, while recovery occurred between day 11 and 29 depending on the severity of the injury. The serum from immunodepressed patients was able to inhibit the response to phytohaemagglutinin and conconavalin A of normal lymphocytes. This immunosuppressive activity was present very early after injury (on day 1-2) and before the onset of lymphocyte hyporesponsiveness to mitogens and was no longer detectable on day 7-8 post burn, when patient lymphocytes showed the greatest hyporesponsiveness to mitogens. This late depression was due to T suppressor cells.

Topics: Adolescent; Adult; Aged; Burns; Concanavalin A; Humans; Immune Sera; Immune Tolerance; Immunity, Cellular; Middle Aged; Phytohemagglutinins; T-Lymphocytes, Regulatory

Serial blast transformation in vitro was measured in peripheral lymphocytes from 38 patients with major thermal injury. Lymphocytes were tested with the antigens streptokinase-streptodornase (SKSD), mumps and purified protein derivative (PPD), the mitogens concanavalin A (Con A) and phytohemagglutinin (PHA), and in the one-way mixed lymphocyte reaction. Statistically significant suppression by the burn injury was noticed in all measurements except response to PHA. One-time measurements were not significantly different between the patients who survived and the patients who did not survive their burn injuries. However, serial determinations of responsiveness to the three natural antigens SKSD, mumps and PPD, as well as the mixed lymphocyte reaction accurately reflected prognosis.

Topics: Adolescent; Adult; Aged; Antigens; Burns; Concanavalin A; Humans; Immunocompetence; Lymphocyte Culture Test, Mixed; Lymphocytes; Middle Aged; Phytohemagglutinins; Streptodornase and Streptokinase

Topics: Adult; Burns; Concanavalin A; Humans; Immunity, Cellular; Lymphocytes; Middle Aged; Mitogens; Phytohemagglutinins

The high morbidity after severe thermal insult is believed to be related partially to a resultant decrease in immunocompetence. We tested the ability of phytohemagglutinin (PHA) and Concanavalin (Con A) to stimulate lymphocyte transformation in 17 patients with moderate to severe thermal injury (greater than 25% BSA). The patients acted as their own controls and the per cent change in their mitogen response was measured over time. Eight acutely burned patients who subsequently developed severe sepsis (Group I) had decreased ability (mean, 12% of normal) to proliferate in response to PHA, and six of these died of severe sepsis. The depressed response appeared 4 to 7 days postinjury and predated clinical evidence of sepsis by 2 to 4 days. Cells from four patients who had mild infectious complications (Group II) demonstrated greatly augmented mitogen responses (mean + 243%) approximately 7 to 10 days postinjury. Five burn patients whose clinical course was sepsis free (Group III) exhibited only minimal changes in their mitogen responses (mean +30%). Although the Con A responses of the patients' cells corresponded less to their pathology, Group I patients whose cells exhibited depressed PHA responsiveness also had diminished Con A responses. Group II patients' cells also showed increases in Con A-induced stimulation. Group III patients, who had only slightly augmented PHA responses, had minimal decreases of the Con A-induced lymphocyte transformation. Many severely burned patients develop septicemia as a result of their large wound surfaces. The appearance of decreases in mitogen-induced proliferation, however, appears to characterize those patients who will be unable to handle the septic challenge.

Topics: Adult; Aged; Bacterial Infections; Burns; Concanavalin A; Escherichia coli Infections; Female; Humans; Immunocompetence; Klebsiella Infections; Lymphocyte Activation; Male; Middle Aged; Phytohemagglutinins; Prognosis; Pseudomonas Infections; Sepsis; Staphylococcal Infections; T-Lymphocytes; Time Factors

Normal mice or mice burned 6 days previously were injected intraperitoneally with either saline or 50 mug of concanavalin A. Four days after the injection the peritoneal macrophages were collected and examined in vitro. Macrophages from concanavalin A-injected mice appeared activated because several in vitro parameters of activity were increased. Examination of macrophages from burned animals revealed a selective depression of macrophage activity.

Topics: Adhesiveness; Animals; Burns; Candida albicans; Cell Adhesion; Concanavalin A; Erythrocytes; Female; Glass; In Vitro Techniques; Injections, Intraperitoneal; Macrophages; Mice; Mice, Inbred Strains; Peritoneum; Phagocytosis; Rabbits; Sodium Chloride; Staining and Labeling