Julius Gordon

Delayed Hypersensitivity: Indicator of Acquired Failure of Host Defenses in Sepsis and Trauma

Primary failure of host defense mechanisms has been associated with increased infection and mortality. Anergy, the failure of delayed hypersensitivity response, has been shown to identify surgical patients at increased risk for sepsis and related mortality. The anergic and relatively anergic patients whose skin tests failed to improve had a mortality rate of 74.4%, whereas those who improved their responses had a mortality rate of 5.1% (P < 0.001). This study documents abnormalities of neutrophil chemotaxis, T-lymphocyte resetting in anergic patients and the effect of autologous serum. These abnormalities may account for the increased infection and mortality rates in anergic patients. Skin testing with five standard antigens has identified 110 anergic (A) or relatively anergic (RA) patients in whom neutrophil chemotaxis (CTX) and bactericidal function (NBF), T-lymphocyte rosettes, mixed lymphocyte culture (MLC), cell-mediated lympholysis (CML), and blastogenic factor (BF) were studied. The MLC, CML and BF were normal in the patients studied, and were not clinically helpful. Neutrophil CTX in 19 controls was 117.5 ± 1.6 u whereas in 40 A patients, neutrophils migrated 81.7 ± 2.3 u and in 15 RA patients 97.2 ± 3.8 u (P < 0.01). In 14 patients whose skin tests converted to normal, neutrophil migration improved from 78.2 ± 5.4 u to 107.2 ± 4.0 u (P < 0.01). Incubation of A or control neutrophils in A serum reduced migration in A patients from 93 ± 3.7 u to 86.2 ± 3.5 u (P < 0.01) and in normals from 121.2 ± 1.6 u to 103.6 ± 2.6 u (P < 0.001). The per cent rosette forming cells in 66 A patients was 42.5 ± 3.1 compared to 53.6 ± 2.8 in normal responders (P < 0.02). Incubation of normal lymphocytes in anergic serum further reduced rosetting by 30%. Restoration of delayed hypersensitivity responses and concurrent improvement in cellular and serum components of host defense were correlated with maintenance of adequate nutrition and aggressive surgical drainage.

Host resistance in sepsis and trauma.

Host resistance to infection was measured by the in vivo response to 5 delayed hypersensitivity antigens and to sensitivity and challenge by dinitrochlorobenzene (DNCB) in 55 seriously ill or injured patients and in 50 preoperative patients. A close correlation between infections, septicemia, death related to infection and anergy was found in the postoperative and post injury patients and was predictive of these complications in the patients studied preoperatively. Decreased body cell mass was noted in both the anergic and non-anergic patients which was consistent with protein-calorie malnutrition but the two groups were not significantly different. A serum factor which inhibited cellular immunity in vitro was found in 4 patients. This factor disappeared in the two patients who recovered. The study suggests the therapeutic value of the in vivo measurement of delayed hypersensitivity in seriously ill and especially preoperative patients in whom specific or non-specific stimulation of cell mediated immunity might alter the risk of infection.

Estimating mortality risk in preoperative patients using immunologic, nutritional, and acute-phase response variables.

We measured the delayed type hypersensitivity (DTH) skin test response, along with additional variables of host immunocompetence in 245 preoperative patients to determine which variables are associated with septic-related deaths following operation. Of the 14 deaths (5.7%), 12 were related to sepsis and in 2 sepsis was contributory. The DTH response (p less than 0.00001), age (p less than 0.0002), serum albumin (p less than 0.003), hemoglobin (p less than 0.02), and total hemolytic complement (p less than 0.03), were significantly different between those who died and those who lived. By logistic regression analysis, only the DTH skin test response (log likelihood = 41.7, improvement X2 = 6.24, p less than 0.012) and the serum albumin (log likelihood = 44.8, improvement X2 = 17.7, p less than 0.001) were significantly and independently associated with the deaths. The resultant probability of mortality calculation equation was tested in a separate validation group of 519 patients (mortality = 5%) and yielded a good predictive capability as assessed by (1) X2 = 0.08 between observed and expected deaths, NS; (2) Goodman-Kruskall G statistic = 0.673) Receiver-Operating-Characteristic (ROC) curve analysis with an area under the ROC curve, Az = 0.79 +/- 0.05. We conclude that a reduced immune response (DTH skin test anergy) plus a nutritional deficit and/or acute-phase response change are both associated with increased septic-related deaths in elective surgical patients.

Macrophages in the mixed leukocyte culture reaction (MLC)

Abstract Two aspects of the mixed leukocyte culture reaction have been investigated. In the first study the capacity of two highly purified adherent cell populations, one consisting of neutrophils the other of macrophages, to support the reactivtiy of purified lymphocytes was compared. Activity was entirely confined to the latter preparation. In the second study, the ability of platelets, neutrophils, lymphocytes and macrophages to provoke a MLC reaction was studied. Macrophages were found to be 10 times as efficient in eliciting a reaction as an equal number of lymphocytes. Neutrophils and platelets appeared to be devoid of stimulating capacity. This lack of stimulation was shown to be due to a true deficiency in stimulating capacity rather than to an inhibition obtained by high concentrations of neutrophils and platelets. The two functions of the macrophage in MLC, its auxiliary helper role, and the elicitation of the reaction, may be dependent on one and the same mechanism.

In vivo and in vitro humoral immunity in surgical patients

In vivo and in vitro humoral immunity was studied in surgical patients. Laboratory controls (LC), delayed type hypersensitivity (DTH) skin test reactive (HR), and anergic (HA) patients were immunized with tetanus toxoid. Maximum in vivo antibody levels occurred 14 days after immunization. Eighty-six, 47, and 17% of LC, HR, and HA subjects, respectively, showed a positive response (xl22 = 21.1 with Yates, p < 0.0005). Peak in vitro antibody production in unstimulated lymphocyte cultures occurred at day 6 after immunization. Antibody responses in vitro were reduced in all surgical patients, worst in HA, and correlated quantitatively with in vivo antibody responses at day 14 (Spearman rank correlation = 0.794, p < 0.001). Total IgG production in vitro was not decreased; 595, 1080, and 1538 ng IgG/culture were produced by LC, HR, and HA, respectively. These data demonstrate decreased in vivo and in vitro humoral immunity in all surgical patients, worst in those with decreased DTH responses. There is a kinetic and quantitative correlation between in vivo and in vitro responses, the latter being a biologic reflection of the integrity and magnitude of the in vivo process. Finally, failure to produce specific antibody is not due to failure of total IgG synthesis.

Suppression and augmentation of the primary in vitro immune response by different classes of antibodies.

Abstract The capacity of purified γG 1 and γG 2 anti-sheep red blood cell (SRBC) antibodies to exert antigen-specific feedback regulations on the primary in vitro immune response to SRBC was studied. Antibodies were administered to the culture in the native form, as sheep erythrocyte-antibody complexes or as pepsin-derived F(ab′) 2 antibody fragments. Marked differences in the feedback regulatory effects of γG 1 and γG 2 antibodies were found. Antibodies of the γG 1 class suppressed the immune response to SRBC, whereas γG 2 antibodies isolated from the same serum exerted an augmenting effect on antibody synthesis. These opposing feedback effects on in vitro antibody synthesis were immunologically specific, relatively insensitive to changes in antigen concentrations, and could be elicited by either adding antibodies and antigen separately to the culture or as preformed antigen-antibody complexes. Experiments comparing the activities of the F(ab′) 2 antibody fragments with the parent γG 1 and γG 2 antibodies suggested that the Fc fragments may be involved in these regulatory effects on the immune response. It is concluded that the antigen-specific suppressive and augmenting effects on antibody synthesis shown here are determined by the antibody class. In addition, we suggest that these opposing antibody-mediated feedback effects may represent one of the important elements of the immune response.

Role of Monocytes in the Mixed Leukocyte Culture Reaction

Summary Suspensions of pure lymphocytes did not interact in mixed cultures; the reaction could be restored by the addition of purified monocytes or of unfractionated leukocyte suspensions in which lymphocytes had been rendered unreactive by X-irradiation or mitomycin. These findings suggest that the monocyte is essential to the mixed leukocyte culture reaction.

Natural resistance mechanisms may play a role in protection against chemical carcinogenesis

SummaryMice deprived of B lymphocytes by the chronic administration of anti-IgM antibodies have been shown to possess a heightened natural resistance (NR) to micro-organisms, to parental bone marrow, and to natural killer (NK)-sensitive tumors in vitro and in vivo. Experiments described in this communication indicate that the latent period of primary tumors induced by the injection of methylcholanthrene (MC) is also prolonged in these mice. This observation suggests that NR mechanisms may provide protection against primary chemically induced tumors.

Characterization of small lymphocytes in the bone marrow of mice after prolonged treatment with anti-IgM antibodies

Abstract In mice treated with anti-IgM antibodies from birth, small lymphocytes in the bone marrow and spleen have been characterized by their incidence, surface markers, and turnover. Essentially complete elimination of IgM-bearing small lymphocytes followed injections of anti-IgM but this treatment did not deplete the IgM-negative small lymphocytes in the marrow. These IgM-negative cells also lacked other markers of B lymphocytes, such as receptors for Fc and complement; they failed to bind anti-mouse T-lymphocyte serum and they formed a rapidly renewing population within the marrow. This population may represent cells whose normal differentiation has been aborted by the anti-IgM antibodies or, alternatively, they may be “null” cells, distinct from B lymphocytes.

Erythrocyte morphology and clustering of fluorescent anti-A immunoglobulin.

PLASMA membranes of many cells are fluid, with the protein and lipid components mobile in the plane of the membrane1,2, as changing distribution patterns of fluorescent labelled lectins and antibodies bound to cell surfaces have shown1–4. Such observations have not been made with intact human erythrocytes, which are thought to be an exception5–7. Ferritin-labelled concanavalin A (con A)8 and soybean agglutinin (unpublished results of C. Kuettner, A. Staehlin, and J.A.G.) have been shown to be randomly arranged on the membranes of intact erythrocytes. But some clustering of con A receptors on the membranes of intact erythrocytes was shown by freeze etching when con A-coated erythrocytes were treated with anticon A antibodies8. We now report evidence for clustering of fluorescein-labelled anti-blood group A antibodies on the surfaces of intact human A erythrocytes associated with alteration of erythrocyte morphology.