Agents and Actions, vol. 30, 1/2 (1990)
0065-4299/90/020294-03$1.50+ 0.20/0 9 1990 Birkh~iuser Verlag, Basel
Cytokine-indueed release of histamine from basophil leukocytes from AIDS patients M. Pedersen, H. Permin, K. Bendtzen 1 and S. N o r n 2 Laboratory of Immunology,University Clinic for Infectious Diseases M, Rigshospitalet, Tagensvej 20, DK-2200 Copenhagen N, 1Laboratoryof Medical Immunology7544,Rigshospitalet, Tagensvej20, DK-2200CopenhagenN, and 2 Department of Pharmacology, University of Copenhagen, Juliane Maries Vej 20, DK-2100 CopenhagenO, Denmark
Abstract
Cytokine-induced histamine release from basophil leukocytes was examined in cell suspension from A I D S patients and compared with healthy controls. Cells from approximately half o f the AIDS patients, in contrast to none from the control group, showed histamine release after stimulation with interleukin-4 (IL-4), tumor necrosis factor alpha (TNF alpha), lymphotoxin (LT) and interferon gamma ( I F N gamma). These cytokines seem to induce histamine release from cells from A I D S patients by interaction with the cell surface immunoglobulins, since removal of the immunoglobulins prior to the exposure of the cytokines completely abolished the response to the cytokines. IL-I alpha, IL-1 beta, IL-3, colony stimulating factor (CSF) and granulocyte-macrophage-CSF (GM-CSF) caused significant histamine release from cells from a similar number of AIDS patients and controls.
Introduction
The acquired immunodeficiency syndrome (AIDS) is caused by human immunodeficiency virus (HIV) and is based on severe cellular immunodeficiency [1]. A high incidence of opportunistic infections is seen and some of the microorganisms such as Candida albicans, Pneumocystis carinii, and Herpes simplex virus, as well as HIV itself, release histamine from basophil leukocytes obtained from A I D S patients, but not from healthy controls [2-4]. Since cytokines participate in a variety of cellular responses including the regulation of the immune system [5, 6], we examined whether some of these cytokines were able to release histamine in cell suspensions obtained from A I D S patients and whether the response was different from that of normal individual.
Materials and methods
Patients The study included 15 patients fulfilling the C D C A I D S criteria IV B, C-I and E [1]. Healthy, non-allergic, age and sex matched volunteers without evidence of HIV infection served as a control group.
Cytokines The human cytokines were all recombinantderived and tested at the final concentrations specified below and in at least 4 consecutive 10-fold dilutions thereof. Interleukin (IL)-I alpha (max. conc. 10 5 pg/ml) was kindly donated by Dr. S. Gillis (Seattle, WA, USA). IL-I beta (max. conc.
295
Agents a n d Actions, vol. 30, 1/2 (1990)
2 x l 0 s pg/ml) was a kind gift from Dr. C.A. Dinarello (Boston, M A , USA). IL-3 (max. conc. 5 x 10 2 pg/ml) and IL-4 (max. conc. 5 x 10 a pg/ml) were from Genzyme Corp. (Boston, M A , USA). IL-6 (max. conc. 2 x 10 4 pg/ml) was a generous gift from Dr. T. H i r a n o (Osaka, Japan). Colony-stimulating factor (CSF) and granulocyte-macrophage C S F ( G M - C S F ) (both at max. conc. 2 • 10 3 pg/ ml) were from Genzyme Corp. (Boston, M A , USA). Tumour necrosis factor ( T N F alpha), lymphotoxin (LT) and interferon g a m m a ( I F N gamma) were kindly d o n a t e d by Dr. G. R. A d o l f (Vienna, Austria) and tested at max. conc. 10 4, 10 ~ and 3 x 103 pg/ml, respectively.
Histamine release M o n o n u c l e a r cells containing approximately 2~ basophil leukocytes were isolated from whole blood by the Ficoll-Hypaque gradient method [7]. The cells were suspended in ' D i s - A M C and incubated for 40 min at 37'>C with the concentrations of the cytokines mentioned above. The release o f histamine was assayed spectrofluometrically by determination of the residual histamine in the washed cell sediment, and the released histamine was expressed as a percentage of the total histamine in the sample [7]. Histamine release above 10% was considered significant, hmnunoglobulins (Ig) were removed from the cell surface by exposure of the cells to p H 3.8 lk)r 5 min at 4"C followed by washing and rcsuspension of the cells in Tris-AMC [2].
Statistics The rank sum test was used with a significance level of less than 0.05. Results
Cells from A I D S patients and controls were examined for histamine release following stimuhttion with biologically relevant concentrations of 10 different cytokines.
Effects qf IL-4, TNF alpha, L T and IFN gamma Table I shows that cells from 6 of the 14 A I D S patients responded to I L-4 with significant histamine release ( > 10%) while none of the cell sus-
Table 1
Histamine release response to cytokines o f cell suspensions f r o m A I D S patients a n d n o r m a l individuals. Stimulator
AIDS
Controls
IL-I alpha IL-1 beta IL-3 IL-4 IL-6 CSF GM-CSF T N F alpha UI IFN g a m m a
3 6 4 6 0 2 1 8 9 8
3 4 I 0 0 4 3 0 0 0
(7) (8) (8) (14)* (10) (7) (8) (13)* (15)* (i3)*
(12) (/2) (12) (11) (11) (12) (12) (10) (11) (10)
N u m b e r s of persons, in w h o m cells responded with significant histamine release. The figures represent the results of experiments using the optimal concentrations o f cytokines in each individual. The total n u m b e r of A I D S patients a n d controls are given in parentheses. Interleukin (ILL colony-stimulating factor (CSF), g r a n u l o c y t e - m a c r o p h a g e C S F ( G M - C S F ) , t u m o r necrosis factor ( T N F alpha), lymphotoxin (LT) a n d interferon (IFN gamma). * p < 0.02 between A I D S patients a n d controls.
Table 2
Individual response to cytokines in cell suspensions from A I D S patients. Patient
IL-4
T N F alpha
LT
IFN gamnra
1
+
+
q
+
2 3 4 5 6 7 8 9 10 11 12 13 14 15
+ } + +
+ + + NI)
+ + -+ + +
+ + + + -+
+ +
+ +
+ + t +
-
+
NI)
ND
-
~
Histamine release ( I ), no histamine release ( determined (NI)).
-
NI) ), and release not
pensions from the 10 normal individuals showed release of histamine. The median histamine release was 7(88 ranging from 0 22%, which was statistically different from the controls (median 3%, range 0 9~ (/7
0065-4299/90/020294-03$1.50+ 0.20/0 9 1990 Birkh~iuser Verlag, Basel
Cytokine-indueed release of histamine from basophil leukocytes from AIDS patients M. Pedersen, H. Permin, K. Bendtzen 1 and S. N o r n 2 Laboratory of Immunology,University Clinic for Infectious Diseases M, Rigshospitalet, Tagensvej 20, DK-2200 Copenhagen N, 1Laboratoryof Medical Immunology7544,Rigshospitalet, Tagensvej20, DK-2200CopenhagenN, and 2 Department of Pharmacology, University of Copenhagen, Juliane Maries Vej 20, DK-2100 CopenhagenO, Denmark
Abstract
Cytokine-induced histamine release from basophil leukocytes was examined in cell suspension from A I D S patients and compared with healthy controls. Cells from approximately half o f the AIDS patients, in contrast to none from the control group, showed histamine release after stimulation with interleukin-4 (IL-4), tumor necrosis factor alpha (TNF alpha), lymphotoxin (LT) and interferon gamma ( I F N gamma). These cytokines seem to induce histamine release from cells from A I D S patients by interaction with the cell surface immunoglobulins, since removal of the immunoglobulins prior to the exposure of the cytokines completely abolished the response to the cytokines. IL-I alpha, IL-1 beta, IL-3, colony stimulating factor (CSF) and granulocyte-macrophage-CSF (GM-CSF) caused significant histamine release from cells from a similar number of AIDS patients and controls.
Introduction
The acquired immunodeficiency syndrome (AIDS) is caused by human immunodeficiency virus (HIV) and is based on severe cellular immunodeficiency [1]. A high incidence of opportunistic infections is seen and some of the microorganisms such as Candida albicans, Pneumocystis carinii, and Herpes simplex virus, as well as HIV itself, release histamine from basophil leukocytes obtained from A I D S patients, but not from healthy controls [2-4]. Since cytokines participate in a variety of cellular responses including the regulation of the immune system [5, 6], we examined whether some of these cytokines were able to release histamine in cell suspensions obtained from A I D S patients and whether the response was different from that of normal individual.
Materials and methods
Patients The study included 15 patients fulfilling the C D C A I D S criteria IV B, C-I and E [1]. Healthy, non-allergic, age and sex matched volunteers without evidence of HIV infection served as a control group.
Cytokines The human cytokines were all recombinantderived and tested at the final concentrations specified below and in at least 4 consecutive 10-fold dilutions thereof. Interleukin (IL)-I alpha (max. conc. 10 5 pg/ml) was kindly donated by Dr. S. Gillis (Seattle, WA, USA). IL-I beta (max. conc.
295
Agents a n d Actions, vol. 30, 1/2 (1990)
2 x l 0 s pg/ml) was a kind gift from Dr. C.A. Dinarello (Boston, M A , USA). IL-3 (max. conc. 5 x 10 2 pg/ml) and IL-4 (max. conc. 5 x 10 a pg/ml) were from Genzyme Corp. (Boston, M A , USA). IL-6 (max. conc. 2 x 10 4 pg/ml) was a generous gift from Dr. T. H i r a n o (Osaka, Japan). Colony-stimulating factor (CSF) and granulocyte-macrophage C S F ( G M - C S F ) (both at max. conc. 2 • 10 3 pg/ ml) were from Genzyme Corp. (Boston, M A , USA). Tumour necrosis factor ( T N F alpha), lymphotoxin (LT) and interferon g a m m a ( I F N gamma) were kindly d o n a t e d by Dr. G. R. A d o l f (Vienna, Austria) and tested at max. conc. 10 4, 10 ~ and 3 x 103 pg/ml, respectively.
Histamine release M o n o n u c l e a r cells containing approximately 2~ basophil leukocytes were isolated from whole blood by the Ficoll-Hypaque gradient method [7]. The cells were suspended in ' D i s - A M C and incubated for 40 min at 37'>C with the concentrations of the cytokines mentioned above. The release o f histamine was assayed spectrofluometrically by determination of the residual histamine in the washed cell sediment, and the released histamine was expressed as a percentage of the total histamine in the sample [7]. Histamine release above 10% was considered significant, hmnunoglobulins (Ig) were removed from the cell surface by exposure of the cells to p H 3.8 lk)r 5 min at 4"C followed by washing and rcsuspension of the cells in Tris-AMC [2].
Statistics The rank sum test was used with a significance level of less than 0.05. Results
Cells from A I D S patients and controls were examined for histamine release following stimuhttion with biologically relevant concentrations of 10 different cytokines.
Effects qf IL-4, TNF alpha, L T and IFN gamma Table I shows that cells from 6 of the 14 A I D S patients responded to I L-4 with significant histamine release ( > 10%) while none of the cell sus-
Table 1
Histamine release response to cytokines o f cell suspensions f r o m A I D S patients a n d n o r m a l individuals. Stimulator
AIDS
Controls
IL-I alpha IL-1 beta IL-3 IL-4 IL-6 CSF GM-CSF T N F alpha UI IFN g a m m a
3 6 4 6 0 2 1 8 9 8
3 4 I 0 0 4 3 0 0 0
(7) (8) (8) (14)* (10) (7) (8) (13)* (15)* (i3)*
(12) (/2) (12) (11) (11) (12) (12) (10) (11) (10)
N u m b e r s of persons, in w h o m cells responded with significant histamine release. The figures represent the results of experiments using the optimal concentrations o f cytokines in each individual. The total n u m b e r of A I D S patients a n d controls are given in parentheses. Interleukin (ILL colony-stimulating factor (CSF), g r a n u l o c y t e - m a c r o p h a g e C S F ( G M - C S F ) , t u m o r necrosis factor ( T N F alpha), lymphotoxin (LT) a n d interferon (IFN gamma). * p < 0.02 between A I D S patients a n d controls.
Table 2
Individual response to cytokines in cell suspensions from A I D S patients. Patient
IL-4
T N F alpha
LT
IFN gamnra
1
+
+
q
+
2 3 4 5 6 7 8 9 10 11 12 13 14 15
+ } + +
+ + + NI)
+ + -+ + +
+ + + + -+
+ +
+ +
+ + t +
-
+
NI)
ND
-
~
Histamine release ( I ), no histamine release ( determined (NI)).
-
NI) ), and release not
pensions from the 10 normal individuals showed release of histamine. The median histamine release was 7(88 ranging from 0 22%, which was statistically different from the controls (median 3%, range 0 9~ (/7
