Endotoxemia after burn injury: effect of early excision on circulating endotoxin levels.

Serial circulating endotoxin measurements (quantitative chromogenic limulus assay) were performed in sera from 19 burned patients to determine the profile of circulating serial circulating endotoxin after burn and the effect of early wound excision on serial circulating endotoxin level. Results indicate an early endotoxemia with the peak serial circulating endotoxin levels 7 to 12 hours and 4 days after burn. More importantly, the level of circulating serial circulating endotoxin can be diminished by early excision; late wound excision was associated with a transient increase of serial circulating endotoxin level. Early excision, therefore, may play a critical role in limiting endotoxemia after burn injury.

Immunology of intermingled skin grafts in rats: preliminary results.

Successful treatment of severely burned patients with the Chinese method of intermingled skin grafting has raised questions concerning the pathophysiological and immunological mechanisms which allow the survival of mixed allogeneic and autologous tissue. In creating a rat model and measuring systemic immunological status by means of mixed lymphocyte reactions (MLR) and histology, we found comparable levels of sensitization in intermingled and allografted rats; however, allografts were rejected and intermingled graft survival was significantly prolonged. We interpret our results to indicate that the survival of intermingled grafts is dependent upon an as yet undefined local protective effect exerted by the presence of auto-skin islands in the allotypic portion of intermingled grafts.

Trauma peptide induction of lymphocyte changes predictive of sepsis.

Post-trauma immunosuppression is characterized by T-cell subpopulation changes and the presence of a low molecular weight suppressive active peptide (SAP), which suppresses T-cell blastogenesis and neutrophil chemotaxis. This study evaluated post-trauma T-cell antigens and suppressive active peptide/T-cell interactions to determine if the suppressive active peptide concentrations predictive of sepsis can cause changes in antigen expression predictive of sepsis. Human lymphocyte markers and differentiation antigens were analyzed post-trauma using flow cytometry for markers predictive of sepsis. Changes induced by purified suppressive active peptide incubated with normal human lymphocytes were similarly analyzed by flow cytometry. SAP concentrations for incubation were chosen which correlated with concentrations in patients developing clinical sepsis. Significant T-cell changes in patients who developed sepsis include: decreased total T-cells, decreased helper cells, decreased natural killer cells, increased Ia expressing mononuclear cells, increased activated T-cells, (L22) and increased IL-2 expressing cells (TAC). Suppressive active peptide can activate T-cells and cause significant increased expression of IL-2 receptors and natural killer cells. Other T-cell changes following trauma predictive of sepsis seem to occur independent of in vitro incubation with suppressive active peptides. IL-2 expressing cells are known to be more readily suppressed by the suppressive peptide. Suppressive peptide activation and subsequent inhibition of T-cells suggests a potential way to explain suppressive peptide-induced immunosuppression following trauma.

Immunosuppressive effects of a trauma-induced suppressor active peptide.

The isolation and partial characterization of an immunosuppressive glycopeptide from sera of severely burned patients has previously been reported. Recently, a monoclonal antibody to this factor and an enzyme linked immunosorbent assay for detection of the peptide have been developed. The presence of the peptide in elevated quantity has been demonstrated in serum of patients with multiple blunt trauma as well as thermally injured patients. It was determined that the peptide is capable of suppressing neutrophil chemotaxis and T-cell blastogenesis as measured by MLR. Inhibition of B-cell blastogenesis induced by the peptide as measured by LPS mitogen-induced proliferation was demonstrated to be less sensitive to suppression. Further, it appears that activated T lymphocytes, those expressing increased IL-2 receptors, are more sensitive to suppression by the peptide at lower concentrations than are nonactivated T lymphocytes.

Trauma peptide-mediated prostaglandin E2 biosynthesis: a potential mechanism for trauma-induced immunosuppression.

In vitro exposure of peripheral-blood-adherent mononuclear cells or amnion cells to nanomolar quantities of a trauma-associated immunosuppressive peptide resulted in an increased biosynthesis of prostaglandin E2 (PGE2). Trauma peptide enhanced prostaglandin E2 biosynthesis by as much as 425% compared to buffer controls. The addition of trauma peptide to mixed lymphocyte cultures significantly inhibited [3H]thymidine incorporation by human peripheral blood lymphocytes. Addition of indomethacin (an inhibitor of prostaglandin biosynthesis) to mixed lymphocyte cultures did not significantly abrogate the immunosuppressive activity of the peptide. These results indicate that suppression of T lymphocyte blastogenesis by trauma peptide is probably mediated by at least two mechanisms: (1) by increased PGE2 biosynthesis, induced by trauma peptide, and (2) through a non-cyclooxygenase-mediated pathway.

Trauma, sepsis, and the immune response.

Burn patients, multiple trauma patients, and patients undergoing major surgical operations often suffer from acquired immunologic deficits that predispose them to life-threatening sepsis. This paper reviews the current research in this area, with emphasis on identifying the components of the immune response affected by injury, elucidating the mediators of immunologic change, and determining new therapeutic approaches for correcting immunologic deficits. Lessons learned from the study of immune deficiency disease are reviewed, as are basic observations of burn- and trauma-induced immune depression.

Generation and activity of suppressor peptides following traumatic injury.

Severe trauma is known to produce pathophysiologic changes leading to the generation of immunosuppressive compounds. With recent advances in biotechnology, a number of these factors have been identified and characterized. Many of these substances have been found to be degradation products of normal serum and tissue proteins. These degradation products have profound biologic activity both in vivo and in vitro. This report briefly focuses on a number of these factors and summarizes the current work involved in the determination of the identity and mechanisms of a previously reported suppressor-active peptide isolated from the serum of trauma patients.

Immunologic monitoring of infection risk in trauma patients: research questions and an approach to the problem.

Sepsis due to injury-related immunosuppression is generally accepted to be the leading cause of morbidity and mortality following trauma. A clear relationship between the amount of burn injury and immunosuppression can be demonstrated, but quantitative relationships between the actual amount of blunt or penetrating injury, a "state" of immunosuppression, and the subsequent development of sepsis has not been clearly established. We have studied and attempted to characterize multiply traumatized patients to identify which components or immune parameters suggest the opportunity to predict sepsis. This report briefly reviews the literature in this area and summarizes current work from our laboratory attempting to identify potential markers.

Toxic epidermal necrolysis complicated by severe wound sepsis: a case study.

The case of a 74-year-old female who developed toxic epidermal necrolysis (30% TBSA) is presented. Despite early, aggressive treatment, the patient developed severe wound sepsis and multiple-system organ failure. The fulminant wound sepsis was preceded by early endotoxemia. Patient serum suppressed T lymphocyte blastogenesis in the mixed lymphocyte reaction test and neutrophil chemotaxis in vitro. Endotoxin removal (affinity chromatography) resulted in a reverse of serum suppressive activity against T lymphocytes but did not reverse inhibition of PMN chemotaxis. Circulating endotoxin levels remained detectable during the entire course while blood cultures remained negative. It is our belief that early endotoxemia (detected before clinical signs of wound sepsis becomes apparent) leads to immunosuppression and other systemic complications.

Long-term depressed immune function in patients splenectomized for trauma.

Peripheral blood from asplenic trauma patients (ASP) was analyzed for immunoglobulin concentrations, complement levels, T- and B-lymphocyte populations, and mitogen response of T cells, and compared to a similar analysis performed on the blood of normal controls (C). The interval from splenectomy to testing averaged 1,471 +/- 193 days (mean +/- SEM) in the ASP. Total lymphocyte count averaged 2,941 +/- 234 in the ASP with a T-cell count of 2,030 +/- 182 and a B cell count of 351 +/- 58. The average control lymphocyte count of 1,769 +/- 147 was significantly less than ASP (p less than 0.001) as were the T-cell count of 1,328 +/- 107 (p less than 0.005) and the B-cell count of 124 +/- 18 (p less than 0.001). Responses to PHA were diminished in ASP lymphocytes by 38% at 3 days (p less than 0.01) and by 49% at 5 days (p less than 0.001) when compared to C. Levels of IgM were significantly decreased (p = 0.05) in ASP. Levels of C3, C4, and C5 were similar in ASP and C. These data demonstrate persistent abnormalities in immune function in adult ASP without underlying lymphoreticular disorders and suggest a possible explanation for the increased septic risk in this patient group.

Reversal of SAP-induced immunosuppression and SAP detection by a monoclonal antibody.

A murine monoclonal IgG antibody (MAb) to column-isolated trauma-induced suppressor active peptide (SAP) was produced and utilized in these studies for the further characterization of SAP. Specificity of the antibody was confirmed by enzyme-linked immunosorbent assay (ELISA), passive immunoblotting, and reversal of SAP-induced neutrophil chemotaxis inhibition. ELISA analysis revealed binding of anti-SAP MAb to a serum protein present in both whole burn and normal serum, but only to burn serum using a less than 25,000-mw serum fraction. This suggests that SAP may be an injury-induced degradation product of a greater than 25,000-mw serum protein. Immunoblotting using a less than 25,000-mw burn serum fraction demonstrated MAb binding to a single low molecular weight protein band. Using the MAb in an ELISA immunodiagnostic procedure, it appeared that SAP levels were significantly elevated in the sera of burned patients who died from their injuries compared to levels in sera of controls or patients who survived.

The immunosuppressive activity of C1q degradation peptides.

We have previously reported the existence and activity of a collagen-like, low molecular weight, suppressive peptide complex in patients with greater than 40% BSA burns. Since C1q participates in the inflammatory response and contains a collagen-like sequence, we have tested, in vitro, the putative generation and immunologic activity of C1q peptide fragments under physical conditions present in burned patients. Nanogram quantities of heat- and enzyme-generated fragments of C1q were shown to be suppressive in vitro to neutrophil chemotaxis, and the mixed lymphocyte response (MLR). The addition of lymphocytes pretreated with C1q fragments suppressed ongoing MLR, indicating the activation of suppressor cells by the peptides. Rabbit anti-C1q globulin was found to reduce the suppressive activity of the collagen-like suppressor which was isolated from human burn sera. Our results therefore suggest that C1q may be an early source of degradation peptides which have strong nonspecific immunosuppressive activity following thermal injuries.

Immunologic parameters in burned patients: effect of therapeutic interventions.

Complex immunologic alterations occur following thermal injury. To further delineate the intricacies of the immune response, a longitudinal profile of immunologic parameters was investigated in burned patients with specific reference to clinical criteria (resuscitation, plasma exchange, surgical excisions, sepsis). During a 17-month period, 26 adult patients with a mean age of 32.6 years and a mean burn size of 45.6% TBSA were evaluated with serial (twice weekly) assays of immunocompetence. The immunologic variables monitored included complement components, fibronectin, immunoglobulins, acute-phase reactants, serum proteins, catecholamines, and the mixed lymphocyte reaction. Resuscitation from burn shock and clinical sepsis were associated with a wide array of serologic abnormalities and lymphocyte suppression. Plasma exchange and surgical excision and grafting procedures were also characterized by multiple serologic changes and improvement in lymphocyte function. No specific serologic parameter correlated well with cellular function; however, patterns of humoral alterations were consistently present and may represent a combined effect.

Suppression of in vitro lymphocyte and neutrophil responses by a low molecular weight suppressor active peptide from burn-patient sera.

Thermal injury produces profound pathophysiological changes in the severely burned patient. Primary among these is the modulation of immunity, leading to episodes of immunosuppression and thus increasing the risk of sepsis and possible death. We herein report the isolation of a low molecular weight suppressor active peptide (SAP) which appears to be responsible for many of the observed immunologic changes in burned patients. SAP suppressed T-lymphocyte blastogenesis in the mixed lymphocyte reaction (MLR) and inhibited neutrophil chemotaxis (CTX) in vitro. Characterization of SAP revealed a complex structure comprised of (1) a peptide component rich in glycine, serine, and alanine; (2) a carbohydrate component containing sialic acid; and (3) a fatty acid component, tentatively identified as prostaglandin E. The immunosuppressive activity of SAP is dependent upon the presence of all three structural components. The molecular weight of SAP was estimated to be 3654 as determined by Amicon cell ultrafiltration and amino acid analysis. The isoelectric point of SAP was estimated by chromatofocusing and ion-exchange chromatography to be between 3.2 and 3.6. We hypothesize that the suppressor active peptide may be comprised of cellular or tissue components released into the circulation at the time of injury.