Studies on delayed hypersensitivity to protein antigen. Induction of delayed hypersensitivity by chemically modified antigen.

it was shown in our previous paper that mice primed with chemically modified bacterial alpha-amylase (BaA), which was neither cross-reactive with anti-BaA antibody nor able to induce a humoral anti-BaA response, developed enhanced responses to a subsequent challenge with native BaA and that the magnitude of the immunological memory was closely related to the priming dose of modified BaA. This paper describes the experimental conditions for induction of delayed hypersensitivity (DH) by modified BaA in relation to the development of immunological memory for antibody response to native BaA. Mice primed with either an intraperitoneal (i.p.) or subcutaneous (s.c.) injection of modified BaA in complete Freunds adjuvant (CFA) developed enhanced anti-BaA as the immunogen and modified BaA as the eliciting antigen, the relationship of anti-BaA responses to a subsequent challenge with BaA. In contrast, when mice were immunized with an s.c. injection of the modified BaA only, a significant level of DH to native BaA could be induced, as measured by the footpad reaction (FPR). The highest degree of DH was observed in mice given 50 micrograms of modified BaA. DH was detectable within 5 days and persisted for 25 days after immunization. In the reciprocal combination of native BaA as the immunogen and modified BaA as the eliciting antigen, the relationship of anti-BaA responses to DH was examined. The primary anti-BaA responses induced by an i.p. injection of large doses of BaA was markedly higher than those induced by an s.c. injection, while DH was exhibited only in mice given s.c. injection of BaA in CFA. With respect to DH to native BaA induced by the modified BaA, it was shown that C3H/He mice were high and C57BL/6 mice were low responders.     

Studies on delayed hypersensitivity to protein antigen. Antigen-specific suppression of delayed hypersensitivity by chemically modified antigen.

An extensively modified protein antigen (methylated bacterial α-amylase, M-BαA) which was neither reactive with anti-BαA antibody nor able to induce a humoral anti-BαA response, retained the ability to prime native BαA-specific T cells which were responsible for the enhanced anti-BαA response to subsequent immunization with BαA and delayed hypersensitivity (DH). The splenic T cell-rich fraction from mice primed with M-BαA collaborated with a native BαA-primed B cell-rich fraction to give a good adoptive IgG anti-BαA response in syngeneic irradiated mice, whereas M-BαA-primed B cell fractions failed to cooperate with native BαA-primed T cell fractions. Splenic T cells from mice given a subcutaneous (s.c.) injection of M-BαA in complete Freund's adjuvant (CFA) exhibited DH in syngeneic cyclophosphamide-treated mice. In the present study, native and methylated BαA were tested for their ability to generate suppressor T cells capable of inhibiting the development of DH. An intraperitoneal (i.p.) injection of either native or methylated BαA in incomplete Freund's adjuvant (IFA) interferred with the development of DH to M-BαA by an s.c. injection of the same antigen in CFA. Transfer of spleen cells from mice given an i.p. injection of either of these antigens 5 days previously, suppressed antigen-specifically induction and expression of DH in the syngeneic recipient mice. The suppressive activity was sensitive to treatment with anti-θ antiserum plus complement. These results indicate that the early phase of inhibition of DH after an i.p. injection is in part mediated by suppressor T cells and that M-BαA cross-reacts with native BαA at the suppressor T cell level as well as the level of effector T cells in DH.     

Management of advanced Hodgkin's disease in remission

Patients with Hodgkin's disease (stage IIIB and IV) after being rendered with combination chemotherapy disease-free, were alternatively allocated to either a group receiving levamisole treatment (Decaris) or to a control group receiving no further therapy until there was evidence of recurrent disease. Laboratory examinations showed that impaired cell mediated immune responses found after termination of intensive chemotherapy were favourably influenced by levamisole treatment. In addition, the duration of remission seems to be longer in the levamisole treated group, although the limited number of patients in the study do not allow a definite conclusions to be drawn.     

Immunorestorative properties of thymostimulin (TS) in patients with Hodgkin's disease in clinical remission

Summary Fifteen Hodgkin's disease patients (8 male, 7 female) aged 19–72 years, who had been in complete unmaintained remission for 1 year or more when the study was initiated, were given 50 mg thymostimulin (TS) IM daily for 60 consecutive days. When compared with 26–30 age- and sex-matched controls, as a group the patients' circulating E_NR⁺, OKT ₃ ⁺ , and OKT ₄ ⁺ cells were depressed (0.001P .06), whereas their OKT ₈ ⁺ cell population was not. Low (>1 SD or >2 SD below mean in controls) or borderline (mean value of two subsequent tests >1 SD below mean in controls) values of E_NR⁺, OKT ₃ ⁺ , and OKT ₄ ⁺ cells were seen in nine (group I) of the 15 patients tested, while the remaining six patients (group II) had normal T-cell proportions. Following TS treatment, the proportions of E_NR⁺, OKT ₃ ⁺ , and OKT ₄ ⁺ cells increased to normal in all group I patients. The T-cell levels, however, decreased to pretreatment values 60–70 days after completion of TS therapy. TS had no effect on the group II patients whose T-cell percentages had initially been normal. Spontaneous cell-mediated cytotoxicity (SCMC) was assessed in 11 patients, and irrespective of the baseline values, there was a significant enhancement (P<0.005) by day 15 of TS administration, which was maintained during treatment. SCMC, however, returned to pretreatment levels 60–70 days after TS was discontinued. The delayed skin test reactivity to DNCB was significantly depressed in all cases. Although TS restored the T-cell proportions, it failed to reverse DNCB reactivity from negative to positive in any of the patients tested. TS can thus restore defective T-cell frequencies and can enhance cytolytic functions that are potentially important in host immunosurveillance, but it apparently failed to improve the skin reactivity to neoantigen.     

A randomized trial to evaluate the immunorestorative properties of thymostimulin in patients with Hodgkin's disease in complete remission

Summary A total of 19 Hodgkin's disease (HD) patients (12 male, 7 female) aged 26–67 years, who had been in complete unmaintained remission for 6 months or more when the study was initiated, were randomly given 50 mg thymostimulin (TS) i.m. daily (G1) or every other day (G2) for 35 days. A third group (G3) was not treated. Then TS, at the same dose was administered twice a week for the following 22 weeks in patients both initially receiving loading or intermittent TS treatment. When compared with age-and sex-matched controls, as a group, the patients' circulating OKT ₃ ⁺ , OKT ₄ ⁺ , OKT ₁₁ ⁺ and E-AETR⁺ cells were depressed (P<0.001 for both proportions and absolute numbers), whereas their OKT ₈ ⁺ cell population was not. Following 5 weeks of daily TS administration, the proportions and numbers of all T cell fractions significantly increased in G1 patients (P<0.03 for all the comparisons tested), while following intermittent TS treatment (G2) only the proportions of OKT ₃ ⁺ and OKT ₁₁ ⁺ cells (P<0.03), but not of other T cell fractions, significantly increased. In addition, no significant changes in the absolute numbers of T cell fractions were observed in this group of patients. Furthermore, no spontaneous variations in the T cell pool size occurred in untreated patients. TS maintenance therapy did not produce any further improvement in the size of overall T cells and T cell subsets but sustained percentage and absolute numbers of these cells during administration and the absolute number of T cells even after discontinuation of therapy. The TS-induced improvement in the T cell pool was not associated with any change in the size of circulating non-T lymphocytes and monocytes. In vitro phytohemagglutinin-induced interleukin-2 (IL-2) and gamma-interferon (IFN-) synthesis was assessed in 11 patients (3 G1, 4 G2, and 4 G3). Although it was not statistically significant, a rise in IL-2 and IFN- production was observed in TS-treated patients, but not in untreated controls. TS failed to exert any effect on the serum circulating levels of neopterin, type I and II IFN, beta-2 microglobulin (B₂-M) and immunoglobulins (Ig). TS can thus improve defective T cell frequences and numbers and may modulate IL-2 and IFN- production.     

Studies on the mechanism of suppression of delayed hypersensitivity by the antischistosomal compund niridazole.

Niridazole given in a single oral dose of 100 mg/kg to guinea pigs sensitized to ortho-chlorobenzoyl chloride-bovine gamma-globulin (OCB-BGG) regularly abolished delayed cutaneous reactivity. Little effect was observed, however, when cells from these animals were tested in vitro with either direct or indirect assays for migration inhibitory factor (MIF). On the other hand, sera taken from nonsensitized guinea pigs after they had received 100 mg/kg of niridazole markedly diminished antigen-induced inhibition of migration of sensitized peritoneal exudate cells in vitro. The immunosuppressive effects of such sera could not be produced by niridazole itself, thereby suggesting an effect of niridazole metabolites. This suppressive activity was readily removed from the serum by dialysis. The active serum blocked the production of MIF by sensitized lymph node cells but did not affect the action of preformed MIF on macrophages. The effect of this serum was reversible; lymph node cells incubated for 24 hr with active serum, then washed and reincubated with antigen in normal serum, produced normal amounts of MIF. These studies suggest that metabolites of niridazole, but not the parent compound itslef, suppress delayed hypersensitivity in guinea pigs and prevent MIF production by lymphocytes without affecting the macrophage response to MIF.     

The activity of neutrophil myeloperoxidase in patients with Hodgkin's disease.

Neutrophil myeloperoxidase activity has been studied in twenty one patients diagnosed with Hodgkin's disease. The presented data indicate no differences in total MPO activity, whereas we observed some differences in the percentage of granulocytes with different degree of scores. Changes in the intensity of reaction may indicate the possibility of exocytosis as a mechanism releasing MPO from the cell to the surrounding area. In peripheral, circulating neutrophils, such a phenomenon seems to be of no avail and may disorganise anti-cancer defence.