Transact. Colleg. int. Allerg., 10th Syinp., Copenhagen 1974 Int. Archs Allergy appl. Immun. 49: 255-270 (1975)
Cellular Mechanisms of IgE Antibody Response1 K im ish ig e I shizaka The Johns Hopkins University, Baltimore, Md.
1 This work was supported by Research Grant AI-11202, from United States Public Health Service and GB-41443 from National Science Foundation. This is publication No. 157 from the O’Neill Laboratories at the Good Samaritan Hospital.
Downloaded by: Univ. of California Santa Barbara 128.111.121.42 - 3/28/2018 3:48:03 AM
As you know, IgE antibody is a cause of hay fever and probably in volved in other allergic diseases. In this situation, it is reasonable to ex pect that prevention or suppression of IgE antibody response would be a useful treatment for allergic disease. To achieve this goal, we have set up both a cell culture system and an adoptive transfer system to analyze fun damental mechanisms involved in the IgE antibody response and studied the immunological effect of hyposensitization treatment in hay fever pa tients. Although we have not reached our goal, some progress has been made to learn regulatory mechanisms for IgE antibody response. I would like to summarize the findings obtained by Drs. K ish im o to and O k ud aira in my laboratory and discuss several topics in this field. In order to analyze cellular basis of IgE antibody formation we have set up a cell culture system in which anamnestic reaginic antibody re sponse can be observed in vitro (fig. 1). After immunization of rabbits by two intraperitoneal injections of dinitrophenyl derivatives of Ascaris ex tract included in 10 mg alum, rabbits giving the PCA titer of 1:160 or more were sacrificed at 2 weeks after the booster injection and their mes enteric lymph node cells were obtained. The cell suspension was first in cubated with homologous antigen for 24 h at 37 °C. After washing to re move free antigen, 1 ml of the cell suspension containing 10-20 million mononuclear cells was cultured for 6 days by the method of M a r b r o o k . Anti-DNP IgG and IgM antibodies in the culture fluid were measured by radioimmunoassay and reaginic anti-DNP antibody was determined by
256
lSHIZAKA Immunization with DNP -Ase ♦ alum
j 4 weeks Booster 2 weeks Bleeding
Mesenteric lymph nodes
,
I
2 -3 x l0 7 c e lls * DNP-Asc (0. 0.1, 1.0 pg/mt) 37°C 24h Wash , * 1-2x10 cells culture for 6days Supernatant Anti-hapten antibody measurement
IgG Ab I
IgMAb I
Reagin by PCA with DNP-HSA
Radioimmunoassay with DNP-HSA immunosorbent
the homologous PCA reaction using DNP-HSA as a challenging antigen. By this procedure, primed cells stimulated with the homologous antigen formed skin-sensitizing antibody together with the IgG and IgM antibod ies specific for DNP group. The physicochemical properties of the skinsensitizing antibody in the culture indicated that the antibody belonged to IgE. It is well established in the mouse as well as in the guinea pig and rab bit that collaboration between carrier specific cells and hapten specific precursors are required for maximal anti-hapten IgG and IgM antibody responses. Participation of two cell lines, i.e. carrier-specific helper cells and hapten-specific precursors were shown in the in vitro anti-hapten IgE antibody response as well. The anti-hapten antibody response of the DNP-Asc-primed lymph node cells to homologous antigen was sup pressed by adding either r-DNP lysine or free Ascaris antigen together with DNP-Asc for stimulation. The suppressive effect of free hapten or free carrier was comparable for IgG, IgM and IgE antibody responses. In view of such findings, we have analyzed cellular basis of IgE anti body response in the in vitro system. The first question we asked was whether the B cells involved in the IgE antibody response are different
Downloaded by: Univ. of California Santa Barbara 128.111.121.42 - 3/28/2018 3:48:03 AM
Fig. 1. Experimental design for anti-hapten antibody response in vitro.
IgE Antibody Response
257
Tabic I. Effect of the depiction of immunoglobulin-bearing cells by anti-immunoglobulin column on antibody response1 Rabbit
34
52
DNP-Asc
0 0.1 0.1 0.1 0.1 0 0.1 0.1 0.1 0.1
Immunosorbent1
_ normal IgG anti-y anti-// anti-y anti-// anti-Fab
Anti-DNP formed IgE2
IgM, /zg/ml
IgG, //g/ml
1.35 4.50 4.50 1.00 1.82
0.01 11.5 11.0 1.80 5.50
Cellular Mechanisms of IgE Antibody Response1 K im ish ig e I shizaka The Johns Hopkins University, Baltimore, Md.
1 This work was supported by Research Grant AI-11202, from United States Public Health Service and GB-41443 from National Science Foundation. This is publication No. 157 from the O’Neill Laboratories at the Good Samaritan Hospital.
Downloaded by: Univ. of California Santa Barbara 128.111.121.42 - 3/28/2018 3:48:03 AM
As you know, IgE antibody is a cause of hay fever and probably in volved in other allergic diseases. In this situation, it is reasonable to ex pect that prevention or suppression of IgE antibody response would be a useful treatment for allergic disease. To achieve this goal, we have set up both a cell culture system and an adoptive transfer system to analyze fun damental mechanisms involved in the IgE antibody response and studied the immunological effect of hyposensitization treatment in hay fever pa tients. Although we have not reached our goal, some progress has been made to learn regulatory mechanisms for IgE antibody response. I would like to summarize the findings obtained by Drs. K ish im o to and O k ud aira in my laboratory and discuss several topics in this field. In order to analyze cellular basis of IgE antibody formation we have set up a cell culture system in which anamnestic reaginic antibody re sponse can be observed in vitro (fig. 1). After immunization of rabbits by two intraperitoneal injections of dinitrophenyl derivatives of Ascaris ex tract included in 10 mg alum, rabbits giving the PCA titer of 1:160 or more were sacrificed at 2 weeks after the booster injection and their mes enteric lymph node cells were obtained. The cell suspension was first in cubated with homologous antigen for 24 h at 37 °C. After washing to re move free antigen, 1 ml of the cell suspension containing 10-20 million mononuclear cells was cultured for 6 days by the method of M a r b r o o k . Anti-DNP IgG and IgM antibodies in the culture fluid were measured by radioimmunoassay and reaginic anti-DNP antibody was determined by
256
lSHIZAKA Immunization with DNP -Ase ♦ alum
j 4 weeks Booster 2 weeks Bleeding
Mesenteric lymph nodes
,
I
2 -3 x l0 7 c e lls * DNP-Asc (0. 0.1, 1.0 pg/mt) 37°C 24h Wash , * 1-2x10 cells culture for 6days Supernatant Anti-hapten antibody measurement
IgG Ab I
IgMAb I
Reagin by PCA with DNP-HSA
Radioimmunoassay with DNP-HSA immunosorbent
the homologous PCA reaction using DNP-HSA as a challenging antigen. By this procedure, primed cells stimulated with the homologous antigen formed skin-sensitizing antibody together with the IgG and IgM antibod ies specific for DNP group. The physicochemical properties of the skinsensitizing antibody in the culture indicated that the antibody belonged to IgE. It is well established in the mouse as well as in the guinea pig and rab bit that collaboration between carrier specific cells and hapten specific precursors are required for maximal anti-hapten IgG and IgM antibody responses. Participation of two cell lines, i.e. carrier-specific helper cells and hapten-specific precursors were shown in the in vitro anti-hapten IgE antibody response as well. The anti-hapten antibody response of the DNP-Asc-primed lymph node cells to homologous antigen was sup pressed by adding either r-DNP lysine or free Ascaris antigen together with DNP-Asc for stimulation. The suppressive effect of free hapten or free carrier was comparable for IgG, IgM and IgE antibody responses. In view of such findings, we have analyzed cellular basis of IgE anti body response in the in vitro system. The first question we asked was whether the B cells involved in the IgE antibody response are different
Downloaded by: Univ. of California Santa Barbara 128.111.121.42 - 3/28/2018 3:48:03 AM
Fig. 1. Experimental design for anti-hapten antibody response in vitro.
IgE Antibody Response
257
Tabic I. Effect of the depiction of immunoglobulin-bearing cells by anti-immunoglobulin column on antibody response1 Rabbit
34
52
DNP-Asc
0 0.1 0.1 0.1 0.1 0 0.1 0.1 0.1 0.1
Immunosorbent1
_ normal IgG anti-y anti-// anti-y anti-// anti-Fab
Anti-DNP formed IgE2
IgM, /zg/ml
IgG, //g/ml
1.35 4.50 4.50 1.00 1.82
0.01 11.5 11.0 1.80 5.50
