Augmentation of delayed-type hypersensitivity by doses of cyclophosphamide which do not affect antibody responses.

Abstract

Immunization of mice with low doses of sheep red blood cells (SRBC) which are suboptimal for antibody responses, leads to sensitization for delayed foot pad reactions within 4 days (1-3) . These delayed-type hypersensitivity (DTH) reactions cannot be elicited in mice which have been immunized with larger doses of SRBC that are more optimal for antibody responses (3, 4) . It has been suggested that B-cell responses to high dose immunization cause suppression of T-cell-mediated delayed foot pad reactions (3) . In support of this notion, animals pretreated with B-cell depleting doses of cyclophosphamide (2-300 mg/kg) have markedly suppressed antibody responses and augmented delayed reactions (4-7) . We have studied delayed foot pad reactions of mice immunized with SRBC and found that by lowering the dose of cyclophosphamide pretreatment to 20 mg/kg, delayed-type hypersensitivity can be augmented without affecting antibody responses . Thus antibody is not the sole regulatory factor in mice immunized with high doses of SRBC .Groups of five to six male mice (5 wk old) were obtained from Jackson Laboratories (Bar Harbor, Maine), and were immunized intravenously with 0.2 ml of different concentrations of washed SRBC 1 wk after arrival . Cyclophosphamide (Cytoxan, Mead Johnson & Co ., Evansville, Ind.) was dissolved in sterile saline immediately before use and injected by the intraperitoneal route as a single dose of 20 mg/kg 1 day before immunization, or 200 mg/kg 2 days before immunization . 4 days after immunization, one foot pad per mouse was measured for thickness with a micrometer (Brown & Sharps Mfg. Co ., No . Kingston, R.I .) and was then tested by injection of 0.03 ml of 20% SRBC in sterile saline . The original foot pad thickness was compared to subsequent measurements to compute the percent increase in thickness. Animals were re-tested in the contralateral foot pad 10 days after immunization . Blood samples were obtained by capillary tube puncture of the retro-orbital plexus 5 and 10 days after initial immunization . The resulting serum from individual mice was twofold diluted in 0.025-ml volumes in microtray wells for measurement of hemagglutinating antibody (HA) titers . Student's t test was used to compute statistical significance (P < 0.05) of HA titers and DTH.

Role of antigen-presenting cells in the development and persistence of contact hypersensitivity.

“…There are three major ways to favor this skewing: (a) Pretreat the animals with a low dose of CY before their initial contact with antigen. CY, in the dose used, has been shown to inhibit cells in the suppressor T-cell circuit (11,12). Our results in this case contrast with the findings of Miller et al (13).…”

“…As has been noted previously (3,11), this occurs when initial immunization with TNP-conjugated cells occurs in the spleen via intravenous injection of hapten-conjugated ceils. There is no requirement for major histocompatibility complex (MHC) identity between the haptenated cells and the host for this form of specific immunosuppression to occur, and presentation need not be on macrophages.…”

Analysis of synergy between cyclophosphamide Therapy and Immunity Against a Mouse Tumour

“…However, different timings and doses of drug reduced T-cell cytotoxicity. Askenase et al (1975) showed an increase in delayed hypersensitivity reactions to sheep red cells when mice were tested 10 days after receiving 20 mg/kg of Cyclo. These mice had been given a primary immunization the day after the drug.…”

“…There are several the combined effects of anti-tumour im-ways in which synergy between drug munity and cytotoxic drugs (Mihich, treatment Fisher et at., 1975). However, in tumour cells; (2) drugs may actually some instances it has been difficult to potentiate cytotoxic immunity by indistinguish the cytotoxic effect of the terfering with regulatory mechanisms drug on the tumour from the effect of (Askenase et al, 1975;Otterness and Chang, 1976) (Porteous and Munro, 1972;Chassoux et al, 1977 Heparinized Minimum Essential Medium (MEM) was injected i.p. The cell-containing medium was recovered through an abdominal incision with a Pasteur pipette.…”

Application of a radiometric ear assay for studies of adjuvant arthritis in rats

“…Since previous work has shown that cyclophos- phamide evokes an enhanced delayed hypersensitivity response to SRBC by selectively destroying B lymphocytes in the mouse (14,15), an experiment was designed to test this finding in rats. Pretreatment with cyclophosphamide was required to obtain a positive ear response 14 days after intraperitoneal sensitization with SRBC in Sprague-Dawley rats (Table 3).…”