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Protein antigens eliciting delayed type hypersensitivity (DTH) were analyzed and purified from the supernatants of protein-free cultures in which Salmonella typhimurium TV148 organisms had grown. DTH activity was measured by the footpad swelling test in mice immunized with living organisms of S. typhimurium TV148 or Escherichia coli K-12. DTH activity in the culture supernatant was specific to TV148-immunized mice. This activity was destroyed by pronase. DTH activity was unable to pass through an ultrafilter with an exclusion limit of 10 kD. After condensation of the supernatant and following centrifugation (100,000 g for 1 h), the DTH activities of the sediment and the supernatant were examined, and both showed DTH activity. Further analyses of DTH antigens in the supernatant by HPLC gel filtration separated the activity into three portions. The most active portion was further fractionated by hydroxyapatite HPLC, revealing the presence of two DTH antigens, with molecular weights of 65 and less than 10 kD. These results indicate that the culture supernatant of S. typhimurium TV148 organisms contains a variety of macromolecular protein DTH-eliciting antigens, and one of the antigens is 65 kD, which is dissociated partly by organic solvents into a low molecular weight (less than 10 kD) antigen.

Modification of carcinoembryonic antigen (CEA) with various chemicals was investigated. Modification of CEA with dimethylsulfate or acetic anhydride resulted in derivatives which preferentially induced delayed-type hypersensitivity (DTH) against native CEA in mice. The strength of the DTH reaction was dependent on the number and chemical nature of modifying groups as well as on the immunizing dose. The strongest DTH reaction without detectable formation of antibodies was achieved by low dose immunization using heavily methylated CEA.

The effect of the IgE-mediated reaction on the passive cutaneous anaphylaxis (PCA) reaction was studied at local skin sites of monkeys, and we found that the IgE-mediated reaction appeared to enhance the PCA reaction. Interactions among IgE-mediated reaction, PCA reaction and delayed hypersensitivity reaction were also determined. Contact dermatitis induced with DNCB was utilized as the delayed hypersensitivity reaction. The IgE-mediated reaction or PCA reaction, as well as simple serum irritation, enhanced the delayed hypersensitivity reaction. It is thus assumed that the IgE-mediated reaction enhances the PCA reaction and that this in turn accelerates the delayed hypersensitivity reaction.

The effect of Newcastle disease virus (NDV) on delayed hypersensitivity to oxazolone in CBA mice was studied. There was a significant impairment of the ability of mice to develop cutaneous hypersensitivity shortly after injection of the virus. The effect was evident when NDV was administered up to 2 days before or within 24 h after sensitization, suggesting that NDV interferes with the process of sensitization. The degree of depression was related to the dose of virus inoculated. NDV inactivated by UV irradiation or heat did not depress contact sensitivity to oxazolone. These data are considered to support the hypothesis that the depression is mediated by a direct interaction between lymphocytes and NDV.

Tartrazine is occasionally associated with some clinical changes which have been attributed to allergy. In tests on laboratory animals with tartrazine and its metabolites by methods which should have detected potential to induce antibody formation, no antibodies were detected except by methods which are artificial in terms of human exposure. Similarly, laboratory methods have shown that the metabolites of tartrazine, and in some cases tartrazine itself, can induce contact sensitization in guinea pigs, although there is little evidence that tartrazine can induce similar changes in man.

The effect of immunosuppressive reagents cyclosporin A (CsA) and rabbit anti-mouse antilymphocyte serum (ALS) on the response to alloantigens was studied in inbred mouse strains. Alloantigen was given either as a cell suspension which induced a delayed-type hypersensitivity reaction (DTH), or as a full-thickness skin graft. Dose-response studies showed that DTH reactions in CBA mice sensitised to BALB/c cells were reduced to background levels when recipient mice were treated with 100 mg/kg CsA on days 0, 4 and 6 after primary alloantigenic challenge. The response to a second challenge was significantly decreased by CsA treatment during primary or secondary exposure to alloantigen and CsA was as effective as ALS in abrogating both primary and secondary DTH reactions. Survival of full-thickness grafts of BALB/c skin on CBA mice was increased from 9 to 23 days by ALS treatment on days ––1 and +2, with grafts given on day 0. Long-term treatment with CsA, from day ––14 to +12, also prolonged graft survival from 9 to 18 days but donor-specific transfusions, with or without concomitant ALS or CsA treatment, decreased graft survival and often sensitised the recipients. This occurred with transfusions administered from ––63 to ––7 days and on the day of grafting. Thus, in H-2 mismatched mice, both CsA and ALS treatments produced a state of tolerance when administered during short-term exposure to alloantigen. With a stronger antigenic stimulus, in the form of a skin graft, ALS was only partially effective in overcoming the sensitising effects of donor-specific transfusions given prior to grafting and CsA could not prevent sensitisation of the recipient, but the continued presence of the drug delayed the onset of rejection in unsensitised recipients.

The purpose of this report is to present results demonstrating for the first time the development of rheumatoid factors to rabbit IgG-Fc as well as antirabbit F(ab')2 antibodies in guinea pigs after chronic sensitization with purified native type II bovine collagen. The sensitized animals also developed antibodies to guinea pig Ig. Antibodies to rabbit Ig arose as early as 3 weeks after bovine type II collagen injection and persisted for as long as 80 weeks when the experiment was terminated. The anti-Ig antibodies did not cross-react with the type II bovine collagen. Despite development and persistence of higher titers of RF and anti-F(ab')2 antibodies in the immunized animals, the animals failed to show clinical evidence of inflammatory polyarthritis. These results indicate that rheumatoid factors as well as antibodies to F(ab')2 arise independently of the clinical expression of disease.

In the present study the effects of a large oral dose of the contact allergen dinitrochlorobenzene (DNCB) on the distribution of T-cell subsets in the small intestines and the emigration pattern of lymphocytes from Peyer’s patches were investigated. Apart from the inflammatory effects, DNCB administration resulted in an influx on the T-helper cells in the villi. Precise quantification of T lymphocytes showed a decrease in Peyer’s patches and an increase in mesenteric lymph node cell suspensions. However, no differences could be found in the emigration rate of T-cell subsets from Peyer’s patches into mesenteric lymph nodes, suggesting that the increase of T cells in mesenteric lymph nodes results from T-helper cells directly immigrating from the villi.

Genetic restriction on the expression of delayed-type hypersensitivity (DTH) to Salmonella typhimurium in mice transferred passively with immune spleen cells was studied. After the intravenous transfer of immune C3H/HeJ (H-2Ik) cells into A.TL (H-2Ik) or A.TH (H-2Is) mice, footpad DTH responses could be evoked in the A.TL recipients, but not in the A.TH mice. When the immune cells of BALB/c or C3H/He mice were intravenously-transferred into F1 hybrids produced by mating BALB/c and C57BL/6 or C3H/He and C57BL/6, respectively, no DTH response could be evoked in these F1 hybrids that received immune parental cells. Local transfer as well as systemic intravenous transfer of immune parental cells to F1 haplotype recipients did not cause any DTH. Previous treatment of the F1 hybrid recipients with cyclophosphamide did not result in the expression of the DTH response. Transfer of immune F1 spleen cells into parental strains also did not induce DTH. When the immune cells of parental strains were transferred into F2 mice and into back-cross mice, examination of the DTH response in these mice showed that some of them did not have any obvious footpad swelling, while others revealed various magnitudes of swelling. The resistance of F1 hybrids to transfer of DTH is discussed.

Variations in opposite directions of cellular and humoral reactions observed during desensitization and immune deviation could reflect a balance controlled by a cell which dispatches information towards cells involved in delayed hypersensitivity and those involved in antibody synthesis. This dispatcher cell could be one of the targets of antigen-antibody complexes which modulate both cellular and humoral components of the immune response.

Autoimmune MRL mice develop a poor delayed-type hypersensitivity (DTH) after BCG cell wall (BCG CW) immunization. To test the possible participation of suppressor cells on DTH induction, MRL/ MpJ-lpr/lpr (MRL-lpr) and MRL/MpJ-+/+ (MRL-+/+ ) mice were pretreated with cyclophosphamide (Cy) 4 days before the immunization. The footpad reaction to the purified protein derivative of tuberculin (PPD) was enhanced remarkably by Cy pretreatment in both MRL-lpr and MRL-+/+ mice. In addition, the lymph node cells obtained from nonpretreated MRL-lpr and MRL- +/+ mice at 7 days after BCG CW immunization suppressed the DTH induction of Cy-pretreated MRL-+/+ mice. On the other hand, a positive proliferative response to PPD in vitro was seen only in the cells from Cy-pretreated MRL-+/+ and younger MRL-lpr mice, but not in the cells of nonpretreated MRL-+/+ or both pretreated and nonpretreated older MRL-lpr mice. Removal of B220-positive cells from the lymph node cell populations of Cy-pretreated MRL-lpr mice restored PPD responsiveness, but the same treatment had no effect on the cells from nonpretreated MRL-lpr mice. The results suggest that DTH responses to PPD in MRL mice immunized with BCG CW are regulated by Cy-sensitive suppressor cells. These suppressor cells occur in MRL mice irrespective of whether they carry the lpr gene. And it is also demonstrated that MRL-lpr mice have TDTH precursor cells in a sufficient number for TDTH production even after expressing lymphadenopathy.

Most of wax D (peptidoglycolipid) used here appeared to be ineffective for production of arthritis when given in a water-in-oil emulsion, while the same wax D in squalane was very effective for production of arthritis. Arlacel A as an emulsifier appeared to suppress the arthritogenicity of wax D in squalane, probably through some interaction with the arthritogenic portion of wax D. Poly 1:C seemed to remarkably enhance the arthritogenicity of wax D, even in water-in-oil emulsion. Acetylated wax D and cord factor (trehalose-dimycolate) were much less effective than poly 1:C. Delayed skin hypersensitivity to PPD, peptidoglycan and poly 1:C was also markedly affected by oil composition. However, there was no correlation between these delayed hypersensitivities and development of arthritis.

Footpad reactions elicited in DHS mice using human γ-globulin (HGG)-coated polystyrene latex particles and soluble HGG (sHGG) were compared. Both the magnitude and the persistence of DHS lesions produced by HGG-latex were considerably greater; at 24, 48 and 72 h after challenge, the level of reactivity induced by HGG-latex was 24, 41 and 71% higher, respectively, than those elicited with sHGG. Early nonspecific swelling following injection of HGG-latex in footpads of normal mice was negligible by 24 h, whereas Arthus-responsive animals did not return to control levels until 48 h. The possible advantages and limitations of using particle-associated protein to elicit DHS reactions are discussed.

Sensitized lymphoid cells could transfer to normal non-sensitized (naive) mice by 24-48 h after antigen challenge in the ear, the capacity to incorporate, at the site of antigen deposition, 5-iodo-2'deoxyuridine-125I in amounts significantly above those obtained in control mice. This was associated with a mononuclear cell infiltration in the pinna. In contrast to lymphoid cells, serum antibodies were unable to transfer a 24- or 48 hour ear reaction. The cells responsible were T lymphocytes as demonstrated by successful transfer following enrichment for T lymphocytes, and abrogation of transfer following treatment with anti-theta serum and complement. Transfer was achieved whether the naive recipients were normal, T-cell deprived, pretreated with cyclophosphamide, or lightly irradiated but not when they were heavily irradiated. Adoptive transfer of the 24-hour ear response was demonstrated with three different antigenic systems. The time-response curves were different with each system although peak reactions were obtained 5 days after sensitization of the donors in all cases. The specificity patterns of the 24-hour ear reaction on transfer were similar to those obtained in the sensitized donors. The results of these studies indicate that the radioisotopic ear method can, under defined conditions, demonstrate the existence of a state of delayed type hypersensitivity in the donors.

Footpad assay of delayed-type hypersensitivity to bovine serum albumin (BSA) was studied. Injection of saline solution of BSA (S-BSA) into footpads of sensitized mice resulted in the development of only the local Arthus-type reaction. Injection of alumprecipitated BSA (AP-BSA), however, was effective in generating both Arthus- and delayed-type reactions. The difference between S-BSA and AP-BSA in the ability of elicitation of delayed-type reaction may be attributable to the difference in the retention of these two forms of antigen in the footpad, since S-BSA was found to be eliminated much more rapidly from footpads than AP-BSA.

The appearance of mononuclear cells, mast cells and mucous cells in the airways was studied in Balb/c mice in relation to cell-mediated immunity reflected as delayed hypersensitivity (DH). The animals were pretreated with cyclophosphamide (Cy) to increase cell-mediated immunity and to decrease the influence of antibodies. Mice pretreated with Cy and epicutaneously sensitized with picrylchloride (PiCl) had stronger DH reactions as compared with sensitized mice not pretreated with Cy. The Cy treatment decreased the IgG antibody formation after sensitization. The Cy-pretreated, sensitized and challenged mice had increased numbers of mucus-producing cells and to some extent also mononuclear cells in their lungs compared with sensitized and challenged non-Cy-treated animals. However, the mucous cell numbers were also increased in Cy-treated, but nonsensitized mice in which most of the airway epithelium had differentiated into mucus-containing cells. The present results indicate that local cell-mediated immunity involves the appearance of mononuclear cells and mucus-producing cells in the lung. This inflammatory reaction is enhanced by Cy treatment, although mucous cell differentiation seems to be induced by exposure to Cy alone.

Delayed skin reactor (DSR), the material in Streptokinase-Streptodornase responsible for eliciting delayed hypersensitivity reactions in man, is stable as a lyophilized powder or when stored at -70°C. Storage at a pH above or below 6.0 leads to a decrease in the ability of DSR to stimulate lymphocyte proliferation in vitro. Its molecular weight is 30,000 as estimated by sodium dodecyl sulfate gel electrophoresis and it is composed of approximately 260 amino acid residues without apparent sulfhydryl-bridged subunits. The extinction coefficient of DSR (A 1% 280 nm) is 2.6.

The effect of experimental long-term azathioprine administration on pre-existing in vitro and in vivo manifestations of delayed hypersensitivity was examined. Adult ewes were administered 3 mg/kg/day azathioprine for periods of up to 28 days. Neither phytohemagglutinin (PHA)-induced nor tuberculin-induced lymphocyte transformation was affected. Similarly, pre-existing skin test positivity to tuberculin showed no observable change following drug therapy. These results agree with the majority of those involving clinical situations and indicate that immunosuppression with azathioprine cannot be effectively monitored by either the lymphocyte transformation response to PHA or by changes in the in vivo or in vitro manifestations of pre-existing delayed cutaneous hypersensitivity.

The effects have been studied of three different chemical modifications of timothy grass pollen extract on various immunological properties. The ability to induce IgG antibody with specificity for native antigen was least affected by glutaraldehyde treatment; IgE antibody production was reduced to a similar extent by all three modifications; there was no increase in IgM production; delayed reactions were reduced. New antigenic determinants were introduced by all the modifications, but the effect was minimal following glutaraldehyde treatment. The significance of these results and the potential application of modified allergen in hyposensitisation therapy are discussed.

Cremophor Elo proved as potent as Freund's complete adjuvant for eliciting skin reactivity to bovine serum albumin in guinea pigs. It enhanced delayed hypersensitivity to sheep erythrocytes in mice while hemagglutinin antibody titers were not affected. This adjuvant activity is likely to play a critical role in hypersensitivity reactions to Cremophor-containing drugs, e.g. alphadione.

Preliminary fractionation of Olea european pollen extract has been performed. At least 10 antigenic fractions have been found by crossed electrophoresis. After Sephadex gel filtration, two fractions with a molecular weight of 160,000 and 65,000 have been obtained. The fractions were evaluated for allergenic activity by two in vitro techniques: polystyrene tube radioimmunoassay (PTRIA) and basophil degranulation test (BDT), and by skin tests. All tests indicated that the most reactive fractions were those in the 65,000 molecular weight peak. BDT has been shown to be a very reliable method as compared with histamine release and other parameters. Although common antigenic fractions have been found for Lollium perenne and O. europea, no cross-allergenicity has been shown by PTRIA inhibition.

The humoral, cellular and neutrophil responses of uraemic patients maintained by haemodialysis were compared with those of renal transplant recipients. Humoral immunity was reduced only in the transplant group. Cell-mediated immunity was abnormal in both groups but more decreased in the transplant group studied particularly within 3 months of transplantation. The neutrophil bactericidal capacities were defective in both groups. Infectious complications were often associated with defective cellular immunity, but fatal infections occurred only in transplant patients over 40 years of age with combined cellular and neutrophil defects. No single test was predictive of graft survival in uraemic patients. The depressed cellular immune responses in uraemia, of the degree seen in the immunosuppressed transplant recipient, may explain the better prognosis of renal transplants compared with other organ transplants.

Studies were initiated to determine the effects of the different phases of the nematode life cycle on the delayed-type hypersensitivity (DTH) capabilities of the host. Mice were injected intraperitoneally with 4 X 10(6) viable or heat-killed BCG at 0, 3, 7, 14 and 28 days after oral infection with 200 Trichinella spiralis larvae. Groups were tested for DTH 28 days after BCG inoculation by the footpad swelling assay. Infection with T. spiralis was found to suppress the response in mice administered viable BCG 0 or 3 days later. Mice injected with viable bacteria 7 days after T. spiralis regained the ability to develop in vivo DTH reactions against old tuberculin. Groups administered viable BCG 14 and 28 days after nematode infection yielded potentiated DTH responses, when compared with control mice. Animals infected with T. spiralis and later injected with heat-killed BCG required a longer interval to develop specific immunopotentiation. Maximum immunopotentiation seemed to be related to the presence of larvae in muscle tissue (28 days). These findings suggest that the effect of T. spiralis on the immune capabilities of the host is dependent on the different anatomical locations of the parasite.

Peritoneal exudate macrophages in guinea pigs and peripheral blood monocytes in man are the most readily available cells sensitive to the migration-inhibiting factor(s) (MIF) induced by tuberculin or insoluble concanavalin A in supernatants of stimulated lymphocyte cultures. The scarcity of MIF-sensitive cells is probably the main reason for the unsatisfactory results obtained with direct and indirect MIF tests when using white blood cells as indicator cells. Isolated horse monocytes represent an alternative sensitive source of indicator cells for human MIF assays, whereas guinea pig peritoneal exudate macrophages appear to be less sensitive and to show large individual variations in sensitivity to human MIF. The species specificity of MIF from various origins shows various patterns and is briefly discussed.

Guinea pigs were immunized intracutaneously into the ears with sheep red blood cells (SRBC). Application of a sensitizing dose of the contact allergen dinitrochlorobenzene (DNCB) onto the same ears was shown to suppress or enhance the humoral response to SRBC depending on the time of application. When guinea pigs were sensitized to a contact allergen, application of a sensitizing dose of a non-related allergen on the same ears either had no effect or caused a clear enhancement of the development of delayed type hypersensitivity (DTH). Strongest enhancement was found when both sensitizations were performed on the same day. Further experiments on the effects of a concomitant DTH reaction elicited at the site of application of a contact allergen showed a strong potentiation of DTH when B-cell suppression was minimized by pretreatment with cyclophosphamide (CY). It was considered that CY-DTH-immunopotentiation might be a useful tool for achieving a higher level of sensitivity after epicutaneous sensitization.

A diet containing 20% by weight of fat rich in unsaturated fatty acids reduced the ability of guinea pigs to form antibody and the delayed hypersensitivity response in vivo against a single antigen. Reduced responses in vivo are manifested on first challenge only, subsequent antigenic challenges showed responses similar to those of animals fed a normal diet. However, cells from animals fed large amounts of unsaturated fats when cultured in vitro with antigen and sera of normal animals fed a low-fat diet did not show reduced delayed hypersensitivity (macrophage migration inhibition and lymphocyte transformation). The serum of animals fed high-fat diets greatly inhibited the response to mitogens and to antigen in vitro of lymphocytes from tuberculin-sensitized animals fed either low or high-fat diet. As serum from animals fed low-fat diets did not have this effect, it is postulated that the inhibitory activity is due to a lipid or lipoprotein derivative of unsaturated fatty acids. Animals fed high-fat diets remained healthy, and showed no disorder related to the fat and no susceptibility to infection.

Delayed hypersensitivity to a soluble protein has been analyzed in mice. Ear swelling, 24 h after injection of a test dose of antigen, has been used to monitor the response to a subcutaneous tail injection of egg albumin in complete Freund's adjuvant. Maximal 24 h ear swelling was observed 8 days after sensitization with either 50 or 100 microgram of antigen. These delayed reactions, which were characterized by mononuclear infiltrates, were relatively uncomplicated by Arthus reactions. The latter, antibody-mediated reactions were apparent at later times after immunization. More intense 24 hr reactions were elicited in C57BL/6 than BALB/c mice. Passive transfer of sensitivity has been accomplished with lymph node cells.

The influence of two lysolecithin analogues on the development of contact sensitivity (CS) to picryl chloride in mice was studied. Both substances were shown to have an adjuvant effect on the primary CS response, but depending on dosage of adjuvant and antigen, the timing of adjuvant injection, and the strain of mice employed, the relative activities of the two substances varied considerably. Large amounts of one of the adjuvants were less stimulatory than small amounts. Both substances had a suppressive effect on the development of CS following repeated administration of picryl chloride. The results are discussed in relation to hypotheses on the cellular targets for, and mode of action of, immunological adjuvants.

Various methods have been used to enhance the ability of laboratory animals to develop contact sensitivity to so-called weak sensitizers. The present studies have examined whether diets supplemented with vitamin A acetate (VAA) could enhance induction and elicitation of contact sensitivity to oxazolone in the mouse. Application of 0.3% oxazolone to VAA-fed mice resulted in increased DNA synthesis in the draining lymph nodes and increased ear thickness after challenge, in comparison with mice on a stock (standard) diet. Moreover, VAA-fed (but not control) mice were sensitized to 0.03% oxazolone as shown by DNA synthesis, increased arrival of syngeneic 51Cr-labelled lymphocytes in the draining lymph nodes, and ear swelling after challenge. These findings demonstrate that vitamin A enhances delayed hypersensitivity responses, and point to a role for vitamin A in developing an animal model capable of detecting weak contact sensitizers after topical application.

Two groups of cows were sensitized to ovalbumin (OA) during lactation by a subcutaneous injection of OA either in Freund’s complete or incomplete adjuvant; two groups of animals injected with the respective adjuvants only served as controls. Intraluminal infusion of microgram quantities of OA in uninflamed mammary glands elicited inflammation, i. e., release of leukocytes into the milk, only in the animals previously sensitized to OA. This indicates that an antigen-specific inflammatory pathway can be established in the mammary gland by parenteral sensitization. Immune-mediated release of leukocytes into the milk could not be reproduced by infusing complement fixing OA-anti-OA complexes into the lumen of uninflamed glands of unsensitized cows.

A kinetic study (0–72 h) was performed on the cellular composition of the exudate in delayed-type hypersensitivity (DTH) induced by the injection of cloned helper T cells into the footpad of C57BL/6J mice. In this model the reaction was maximal at 24 h, as assessed by local swelling. After an initial phase of inflammatory cell infiltration, we observed pronounced degranulation of mast cells, occurring at 15–19 h. Administration of antihistamines shortly before this time significantly inhibited the increase in footpad thickness at 24 h, suggesting mast cell mediators as a major cause of swelling in DTH. Furthermore, a new arrival of inflammatory cells, particularly eosinophils, was clearly correlated with mast cell degranulation. The latter, which might function to suppress local reactions, persisted as the only intact cell type after 48 h. Supplementary examination of peripheral blood smears showed successive peaks of neutrophils (at 3 h), monocytes (at 15 h) and finally eosinophils (after 20 h). These results suggest a local and systemic cascade in DTH which depends on mast cell degranulation for its full expression.

Delayed type hypersensitivity (DTH) lesions have been difficult to evaluate objectively in the mouse because they are usually assessed in terms of an increase in footpad swelling or ear thickness. We have developed a radioisotopic method which not only reduces the observer's error but also gives an objective measurement of the cellular activity in the lesions. In brief, 10 mul of the test antigen is injected intradermally into the left pinna and either nothing or the same volume of a control solution into the right. 10 h later, a 2-muCi pulse of 5-iodo-2'deoxyuridine-125I is given intravenously, the ears cut off at the hairline 16 h later and the radioactivity counted in a gamma spectrometer. The following was obtained as evidence that the increased radioactivity of the left pinna over the right was a measure of the extent of a DTH response: (1) the ear reaction was delayed in mice without serum antibodies becoming maximum at 24 h; (2) there was a mononuclear cell infiltration in the left pinna and autoradiographs revealed radioactive label bound to these cells; (3) athymic mice could not develop a 24-hour ear reaction, and (4) antigens known not to activate T cells did not elicit the ear response. Cell transfer studies will be described in a subsequent paper. Different sensitization regimes were required with different antigens in order to obtain the highest levels of DTH as tested by the ear response, and the maximal ear reaction occurred at different days. The ear reaction showed the specificity expected of a DTH response.

The recent identification of a T-cell-derived antigen-binding molecule (TABM), Trichinella spiralis factor (Tric-F), isolated from culture supernatants of lymphoid cells from mice infected with the intestinal helminth T. spiralis, has led to investigation of the ability of Tric-F to induce a T-cell-dependent feedback circuit that ultimately suppresses the production of other TABMs with similar (isotype-like) features. This form of regulation that has been identified in contact hypersensitivity and in delayed-type hypersensitivity (DTH) responses to tumor cells, was shown not to be antigen-specific but to be DTH-specific. Injection of mice with the TABM called picryl chloride factor (PCl-F) induced suppression of the production of DTH-initiating TABMs of other antigenic specificities. In this study, we report that intravenous injection of mice with Tric-F or PC1-F, 8 days before an oral infection with T. spiralis, induced suppressor cells that inhibited the T-cell-dependent influx into the gut of inflammatory cells, comprising mast cells and eosinophils. Similar results were obtained when the mice were skin sensitized with PC1 8 days prior to a T. spiralis infection, i.e. in a system where TABMs are known to be produced. The phenotype of these suppressor cells was Lyt-1––2+. This suppression preferentially affected the parasite-induced DTH-like response in the gut. In contrast, increased levels of IgA plasma cells in the gut, and worm expulsion were not affected by these treatments. In reciprocal experiments, intravenous injection of Tric-F, or PC1-F, or an oral infection with T. spiralis (that results in the production of TABMs) given 8 days before contact sensitizing mice with PC1, resulted in a suppression of elicitation of cutaneous DTH, as measured by ear swelling. In contrast, pretreatment with anti-dinitrophenyl IgE antibody did not interfere with intestinal inflammation to T. spiralis nor with DTH to PC1. Our results suggest that similar to cutaneous DTH, T. spiralis-specific T-cell factors are involved in the initiation and regulation of the DTH-like mast cell and eosinophil-rich intestinal inflammation that accompanies T. spiralis infections in the gut. Since both Tric-F and PCl-F induce suppression of cellular immune responses in vivo, independent of antigen specificity, it is concluded that Tric-F belongs to the same isotype of TABMs as PC1-F that therefore can be regulated by a non-antigen-specific, isotype-like, T-cell-dependent feedback mechanism.

Adjuvants differ in capacity to induce interferon (IFN). As a consequence IFN induction by adjuvants may influence their effectiveness in enhancement of delayed type hypersensitivity (DH) reactions. In this study, the lipophilic amine dimethyl dioctadecyl ammonium bromide (DDA), the synthetic double-stranded polynucleotide polyinosinic polycytidylic acid (poly I:C), liposomes and the polyols L 101 and L 121 were compared in BALB/c mice as inducers of IFN and also as adjuvants for DH to both lysozyme and inactivated Semliki Forest virus (SFV). The antigens were injected intracutaneously, alone or mixed with adjuvant. At day 6 after the immunization, DH was elicited and measured 24 h later as increase in footpad thickness. Negatively charged liposomes and polyol L 121 were unable to enhance DH to SFV and also lack the capacity to induce IFN. Polyol L 101 induced low levels of IFN and showed only slight adjuvanticity for DH to SFV. In contrast, DDA, a moderate IFN inducer, was shown to be a very effective adjuvant for induction of DH against both lysozyme and SFV. The excellent IFN inducer, poly I:C, at the tested dose range (0.03–3.0 mg/kg), displayed only a relatively weak adjuvant activity. But low doses of poly I:C (0.03 and 0.1 mg/kg) showed still adjuvanticity in contrast to equal doses of DDA. This might be related to sufficient induction of IFN by low doses of poly I:C but not by DDA. The discrepancy observed in the relation between IFN induction and a maximal DH induction suggests that IFN is not the only factor which enhances the effectiveness of adjuvants in induction of DH. Furthermore, various inbred mouse strains display considerable differences in DDA-induced enhancement of DH to both lysozyme and SFV, whereas no significant differences in DDA-induced IFN titers were detected. In conclusion, the capacity of an adjuvant to induce IFN is an important feature to achieve enhancement of DH reactions. The effectiveness of an adjuvant, however, is also determined by its other intrinsic properties and by factors determined by the host.

Different lymphocyte separation procedures, Percoll and Ficoll-Paque density sedimentation and gravity sedimentation, were tested in order to get optimal results in the lymphocyte transformation test in diagnosis of nickel allergy. Percoll at both investigated times, 3 and 6 days, gave the highest lymphocyte DNA synthesis response to nickel sulfate in nickel-allergic subjects and the most significant difference against control subjects. With all tested separation procedures there was an overlap between the patient and control group as regards the individual responses.

The effect of 4 immunosuppressive agents – hydrocortisone (HC), cyclophosphamide (CY), azathioprine (AZ) and methotrexate (MTX) – on cutaneous delayed-type hypersensitivity (DTH) and Arthus reactions to the intradermal injection of ovalbumin (OA) in rats sensitized to OA in Freund’s complete adjuvant (FCA) was studied. Multiple doses (daily for 4 days) were given either early, beginning on the day of sensitization, or late, beginning 9 days after sensitization. Single doses were given on the day of challenge with OA. All late multiple doses of drugs except HC depressed the DTH at 24 h in the following order of decreasing magnitude: MTX, CY, AZ. The DTH at 24 h was depressed by early multiple doses of MTX at all doses, by CY at all but the lowest dose, and by AZ at the intermediate dose; HC had no effect. When the drugs were given as single late doses, only CY at the lowest dose and MTX at the higher doses effectively (depressed the DTH at 24 h. Increased Arthus reactions occurred after early and late multiple doses of HC and after a late single dose of CY at the highest dose. After late multiple doses the Arthus reaction was unaffected by either CY or MTX but was depressed by all doses of AZ. HC administered as 3 injections around the time of challenge markedly depressed the delayed and Arthus reactions. These results show that each of the 4 immunosuppressive drugs could depress DTH and Arthus reaction to OA, but the degree of depression varied with the time of drug administration relative to sensitization and challenge, and the dose of drug used. Histologic examination of skin test sites showed that an apparently negative reaction did not necessarily imply total absence of a cellular inflammatory response.

The histology of mouse thymic tissue was examined following infection with Trichinella spiralis and/or BCG. ICR/CD-1 mice were orally infected with 200 T. spiralis 56 days preceding intravenous administration of 4x10-6 viable BCG. Following assessment of delayed hypersensitive foot pad reactions to BCG, the mice were bled and sacrificed. The thymus was removed from each animal, fixed in 10% neutral buffered formalin, embedded in paraffin and cut in 4 mum sections. Examination of serial sections indicated that T. spiralis and T. spiralis-BCG infections induced distinct anatomical changes within the thymic cortex and medulla, which may be related to previously described functional alterations of the immune system.

Following the intraperitoneal injection of cis-diamino-dichloro-platinum (II) (CDDP) into tumor-bearing mice, two immune responses were augmented. The total splenic plaque-forming cells response to sheep red blood cells and the delayed-type hypersensitivity reaction to oxazolone were higher in treated animals than in the control groups. This observation may be relevant in the establishment of clinical protocols associating CDDP and other cytotoxic drugs.

The growth of Mycobacterium bovis (BCG Montreal) and M. tuberculosis Erdman was determined in normal and T-cell depleted (THXB) mice when injected subcutaneously into a hind footpad. The bacilli multiplied only to a limited extent within the footpad itself but the infection quickly spread to the draining popliteal lymph node to eventually reach the liver, spleen and lung. The amount of systemic growth seen in the THXB mice was 10–100 times greater than in the normal controls, all of which developed a tuberculin hypersensitivity and an immune response in 14–18 days. T-cell depletion completely inhibited the expression of tuberculin sensitivity by the infected host as well as ablating the antituberculous response against both the vaccinating BCG population and a superinfecting Erdman challenge inoculum. Incorporation studies in the THXB mice indicated a striking reduction in cell division within the draining lymph node but there was an unexpected elevation in the level of incorporation by the lung cells as the BCG infection progressed. The significance of these findings is discussed in relation to the possible use of the BCG footpad model for studies of leprosy immunity.

Metronidazole administered orally in doses of 20 and 200 mg/kg daily suppressed granuloma formation around Schistosoma mansoni eggs which were injected intravenously and lodged in the pulmonary microvasculature of mice. The same doses did not suppress granuloma formation in animals which had previously been sensitized to the eggs. Nonspecific granulomatous inflammation around divinyl benzene copolymer beads was unaffected by the drug. In a daily dose of 20 mg/kg, metronidazole inhibited delayed footpad reactions to soluble schistosome egg antigen, but 200 mg/kg on alternate days failed to suppress skin allograft rejection. The drug appears to suppress selectively some aspects of cell-mediated immunity.

Those scientists interested in allergic inflammatory processes have recently been focusing on the late-phase response, since it appears most similar to the chronic disease states observed in allergic patients. In this review we will focus on the pattern of mediator release and cellular traffic observed in two in vivo human models of the late-phase reaction, one involving the upper airways and the other the skin. We have observed in these models, as had been observed earlier in blood, that the late-phase reaction is associated with a second increase in the level of mediators. We also describe our studies of the secretagogues responsible for this late-phase mediator release and, in so doing, introduce the subjects of histamine-releasing factors and IgE heterogeneity.

Levamisole and D-penicillamine are not skin sensitizers in rats. Furthermore, they do not stimulate T-lymphocyte proliferation in the draining lymph node when injected in oil into the hindpaw or base of the tail. Their copper coordination compounds also do not sensitize rats though they are irritant when injected locally.

The relationship of mitogenic factor (MF) to delayed hypersensitivity was examined in the guinea pig. MF was readily produced in vitro by blood leucocytes from animals sensitized with Keyhole limpet haemocyanin in Freund's complete adjuvant. Such animals exhibited delayed skin reactions and macrophage migration inhibition. Treatment with cyclophosphamide markedly reduced serum antibody but caused no diminution of MF or delayed skin reactions. A migration inhibition response was observed 10, but not 20, days after sensitization by means of incomplete adjuvant.

The immune response to a hapten-carrier conjugate appears to be a complex phenomenon where reactions of the T-cell population are not restricted to the carrier and where the reactions of the B-cell population are not limited to the hapten determinant of the antigen molecule. To get a better understanding of the different cell interactions during the immune response to a hapten-carrier complex, the effects of immunogenic or tolerogenic injections of various hapten-containing compounds on the responses induced by immunization with the same hapten coupled to protein carriers were studied. The results indicate that T cells involved in delayed hypersensitivity and T cells involved in contact dermatitis could belong to distinct subclasses and confirm that hapten and carrier moieties of the antigen molecule could compete, probably at the macrophage level, for both delayed hypersensitivity to the carrier and antibody synthesis to the hapten.

A strong delayed-type footpad reaction was established in mice using azobenzenearsonate-acetyl bovine serum albumin (ABA-AcBSA) as an antigen. Male ddY/S mice were sensitized by subcutaneous injection of 100 μl of Freund’s complete adjuvant – saline (1:1) emulsion containing 100 μg of the antigen and challenged by subcutaneous injection of 2.5 μl of Freund’s incomplete adjuvant – saline (1:1) emulsion with 2.5 μg antigen in the footpad on the 10th day after the sensitization. The analysis of the system which fulfilled criteria for delayed-type hypersensitivity with regard to kinetics, passive transfer and histology of the footpad reaction, and the effect of dexamethasone, indomethacin and quinacrine on the footpad reaction were described.

An inbred mouse model was used to evaluate in vivo host immune response to Cryptococcus neoformans. Within 1 week of immunization, mice developed delayed type hypersensitivity reactions (DTH) to cryptococcal extracts derived from either culture filtrates or disrupted cells. There was no significant cross reactivity with extracts of other fungi. Previous immunization provided considerable protection against subsequent challenge with multiple strains of cryptococci. DTH also developed after nonimmunized mice were challenged with C. neoformans; however, in this situation DTH was not associated with prolonged survival. These studies indicate that mice can be immunized and protected against cryptococcosis and that this protection is associated with acquisition of DTH to cryptococcal antigens.

Guinea pigs infected intradermally with Treponema pallidum Nichols strain were examined for the presence of lesions and for cellular and humoral response. Of the 26 guinea pigs (77%) demonstrated darkfield positive lesions between 6 and 20 days after infection. Animals divided into six groups of 4-5 in each were sacrificed between 2 and 24 weeks. The peritoneal exudate cells, obtained 4 days after injection of oil, were examined by the direct and the cells from lymphoid organs by the indirect capillary leukocyte migration method in the presence of 10 and 30 mug/ml of Reiter antigen. The peritoneal exudate cells responded with enhancement of leukocyte migration, reaching significant values between 6 and 24 weeks after injection. The cells from the lymphoid organs showed enhancement of leukocyte migration until the 4th week of infection. Treponemal antibodies, found in all animals, started to appear at the 4th week reaching a maximum titer at the 12th week after infection. The appearance of the antibodies correlated well (p less than 0.05) with the appearance of the enhancement of leukocyte migration. Wassermann antibodies were not detected throughout the 24 weeks of infection. The unique immune responses are discussed in view of the available data concerning natural acquired and experimental syphilis.