- Email: [email protected]
842 Eosinophils as effector cells in inflammatory and immune reactions
350
B41
Abstracts
:. ALLERGY
EOSINOPHIL CHEMOTACTIC ACTIVITY IN THE SUPERNATANT OF MONONUCLEAR CELL CULTURE STIMULATED WITH SPECIFIC ANTIGEN. Y. Fukushima,M.D., T. Fukuda,M.D., T. Numao,M.D., I. Akutsu,M.D. and S. Makino, M.D., Tochigi, Japan. To evaluate the role of lymphokine(s1 for the recruitment of eosinophils into the airways in chronic asthma, we measured eosinophil chemotactic activity in the suvernatant of peripheral blood mononuclear cell culture. -Peripheral blood mononuclear cells (PBMC) separated from patients with asthma who were sensitive to Dermatophagoides farinae(DF) were cultured in d-medium for 5 davs at 37OC in 5% CO in-the presence or absence of 10 tic/ml o $ Df antiqen. Eosinophil chemotactic activity (%A) was tested using modified Boyden chamber method. ECA was detectable after 24 hrs, peaked at 72 hrs and continued throughout the experiment. ECA was not observed when PBMC separated from subjects who were not sensitive to Df were cultured. This activity was unchanged by heating at 56OC for 30 min. but was inactivated by 100°C for 10 min. The molecular weight of this chemotactic factor was greater than 30,000 daltons by ultrafiltration-study. These-data suqqest that mononuclear cells stimulated with related antigen produce one of lymphokines which possesses ECA. This substance may play an important role in the recruitment of eosinophils in chronic
843
WT!ZNTlAL FQLE OF THE HEbAATOPOlETlC QNA-BINWG PRQMN #TM IN GGt&X GF EGBIWHIL-SPECIFIC GENEtw\FJscR3ptlm. jelallH.F.a Harvard Medical BaMlett.uBohool, Dfvlafon of Hematology. Chifdren’s Hospital, Howard Hughes Me&oaf Institute. and Infectious Disease Dhrision, Department of Mediine, Beth lsraet Hospital. 6cs.to~, MA. GATA-1 (GF-1, NF-El, Eryf-1) is a hemafopoietio-specific DNA-hiiing protein which was initially characterized for its role as a transcriptional activator of eryfhroid gene expression, It is also present in bone marrow-derfved maat cells and megakafyocjfes. MTA-1 was deteofecf by RNAse protection anafyals in RNA prepared from highly purified eosfnophfls iaofeteo from patients with hypereosinophll& syndrome. GATA-1 was similarly detected in RNA obtained from purified basophils from a patient with chronic myefogenous leukemia with basophilia and also in the rat basophhlii leukemia cell line, ABL-1. In an effort lo understand the role of GATA-1 in fhe hemafopofetio development of eosinophils and basophlfs, we have utilized a cloned subline of HL60 oommllted to eosinophil dlfferentiatlon. This cell line has the potential to produoe eoeinophil proteins such as CLC protein, eosinophlffo oatbnlc protein, and eoafnophilderlved neurofoxin on stimulation with BCGF-II as a source for IL-S. GATA-1 RNA transcripts were not detected in the unsfimulated HL-60 subline based on RNAse protection analysis. After 3 days of stimulafbn wffh BCGF-II, GATA-1 was detected at a low level whfoh in&aaed after five days. Based on the proposed role of GATA-1 as a transcriptional activator of genes eJtpressed during differentiation of hematopoletio progenitors. eosinophll-specitic genes are likely targets for GATA-I.
844
CBllpdRIsoll CAP-fm~ A.O’Connor, Asthma OPC,
asthma.
f#&? EOSINOPPICS INF~~~~~~~~
AS
EFFECTOR
CELLS
IN
R6aACTIQNS c I& M. D., Ph.. D., V. de1 PQpo. B. S.. B. de Andr&Q:. S., E. Mmrth 0. S., B. C$rdaba, B. S. , P. Trw5n,
t
T. A., P. ~~~nQ, ph. D. Madrid, Spain. effector cells in Eosinophils are hypersensitivity disorders and parasitic infections. In this report we present data of a possible rote of eoeinophiIs in the immune response as antigen presenting cells (APC) and the mechanisms implicated in the activation of these cells for mediator release (LTC,), in the inflammatory reactions. %usin~phil
in the
Immune
Retqxmaa:
We have
previously shown that this cell has mRNA for IL1 (J Immlinol 144: 3117-22, 1990). Also murine eosinoohile treated with x-GM-CSF express MHC class II molecules. In this report we dkmonstrate that antigen specific T cell clones proliferate when to the plates containing eosinophile added pretreated with GM-CSF and pulsed with the antigen. The proliferation of the T cells is dependent on the number of eeainophiis used as APC. Eosinophil presentation is restricted by Class II and inhibited by lysosomotropic agents. Looking at the role of eoain~PI&a in the irdlmmatiry reactioni we investigated the involvement of phoaphoinoaitide breakdown in the .activation of 5'-Hpoxygenase. Using neomycin, we have demon&rated a decrease in the LTC released. [Caa'I, elevation has been observe d using
Fura-B/AM,
following
stimulation
with
the
anti-Fc-gamma RI1 mAb. It is likely that the activation of murine wainophils by a Fc-&unum receptor mechanism etimuletea phoaphainoaitide breakdown.
CLIN. IMMUNOL. JANUARY 1991
OF THE Pff&WM&-WT Tg T~~F~~ Ill VfsPm mz&Y. G.Kunkel. Dpt. of FC’in. &&ol. Free University Berlin, FRG
and
Determination of specific IgE with RAST (antigens bound to paper disc as solid phase) is the @ost conmonly used in vitro diagnostic method in the practice of allergy. In this study we coarpared, a nowel test system which is based on a new type of solid phase, consisting of a flexible hydrophilic carrier polymer encased in a capsule, the lagunoCAP to the RAST. We investibated 123 sera fratn ill vospld venom allergic pi;tients (74% frtaale). All patients were diagtlosed on the basis of their history, skin test and RRST. In addition, the standard serum of the RAST-kit (hi h titer of birch pollen specific IgE) was eva 3 uated with the CAP-kit. Results: The patients’ age ranged from 8 to 80, with a mean% of 44.2a14.6 years. The total serum IoE levels ranoed from 4-1712 kU/l, with a median”of 108 kU/l.“The results of specific IgE for vespid.venoIA (i ), as determined by RAST ranged from class O-4 wi *i! h a atean*% of 1.87kl.07 (median: 2.1). The CAP systetn ranging from O-6 with a meanas of 2,78*1.38 (median: 3.0). The correlation coefficient betwaen the fWT and CAP systeai was r=g&l. The two sysstglas cowpared here, showed a higher sensitivity for the CAP system by almost one class, compared to the g&ST. It seemed that low sensitized patients were not detected by the MST (MST class 0), whereas the saare sera showed CAP classes be&man 1 aad 2. We would therefore rocgggtrsl~to cat1 the scoriz of the CAP system “CAP-clata%” imtead.of RMT classes so that it is obvious wfth which method the in vitro data were obtained.
B41
Abstracts
:. ALLERGY
EOSINOPHIL CHEMOTACTIC ACTIVITY IN THE SUPERNATANT OF MONONUCLEAR CELL CULTURE STIMULATED WITH SPECIFIC ANTIGEN. Y. Fukushima,M.D., T. Fukuda,M.D., T. Numao,M.D., I. Akutsu,M.D. and S. Makino, M.D., Tochigi, Japan. To evaluate the role of lymphokine(s1 for the recruitment of eosinophils into the airways in chronic asthma, we measured eosinophil chemotactic activity in the suvernatant of peripheral blood mononuclear cell culture. -Peripheral blood mononuclear cells (PBMC) separated from patients with asthma who were sensitive to Dermatophagoides farinae(DF) were cultured in d-medium for 5 davs at 37OC in 5% CO in-the presence or absence of 10 tic/ml o $ Df antiqen. Eosinophil chemotactic activity (%A) was tested using modified Boyden chamber method. ECA was detectable after 24 hrs, peaked at 72 hrs and continued throughout the experiment. ECA was not observed when PBMC separated from subjects who were not sensitive to Df were cultured. This activity was unchanged by heating at 56OC for 30 min. but was inactivated by 100°C for 10 min. The molecular weight of this chemotactic factor was greater than 30,000 daltons by ultrafiltration-study. These-data suqqest that mononuclear cells stimulated with related antigen produce one of lymphokines which possesses ECA. This substance may play an important role in the recruitment of eosinophils in chronic
843
WT!ZNTlAL FQLE OF THE HEbAATOPOlETlC QNA-BINWG PRQMN #TM IN GGt&X GF EGBIWHIL-SPECIFIC GENEtw\FJscR3ptlm. jelallH.F.a Harvard Medical BaMlett.uBohool, Dfvlafon of Hematology. Chifdren’s Hospital, Howard Hughes Me&oaf Institute. and Infectious Disease Dhrision, Department of Mediine, Beth lsraet Hospital. 6cs.to~, MA. GATA-1 (GF-1, NF-El, Eryf-1) is a hemafopoietio-specific DNA-hiiing protein which was initially characterized for its role as a transcriptional activator of eryfhroid gene expression, It is also present in bone marrow-derfved maat cells and megakafyocjfes. MTA-1 was deteofecf by RNAse protection anafyals in RNA prepared from highly purified eosfnophfls iaofeteo from patients with hypereosinophll& syndrome. GATA-1 was similarly detected in RNA obtained from purified basophils from a patient with chronic myefogenous leukemia with basophilia and also in the rat basophhlii leukemia cell line, ABL-1. In an effort lo understand the role of GATA-1 in fhe hemafopofetio development of eosinophils and basophlfs, we have utilized a cloned subline of HL60 oommllted to eosinophil dlfferentiatlon. This cell line has the potential to produoe eoeinophil proteins such as CLC protein, eosinophlffo oatbnlc protein, and eoafnophilderlved neurofoxin on stimulation with BCGF-II as a source for IL-S. GATA-1 RNA transcripts were not detected in the unsfimulated HL-60 subline based on RNAse protection analysis. After 3 days of stimulafbn wffh BCGF-II, GATA-1 was detected at a low level whfoh in&aaed after five days. Based on the proposed role of GATA-1 as a transcriptional activator of genes eJtpressed during differentiation of hematopoletio progenitors. eosinophll-specitic genes are likely targets for GATA-I.
844
CBllpdRIsoll CAP-fm~ A.O’Connor, Asthma OPC,
asthma.
f#&? EOSINOPPICS INF~~~~~~~~
AS
EFFECTOR
CELLS
IN
R6aACTIQNS c I& M. D., Ph.. D., V. de1 PQpo. B. S.. B. de Andr&Q:. S., E. Mmrth 0. S., B. C$rdaba, B. S. , P. Trw5n,
t
T. A., P. ~~~nQ, ph. D. Madrid, Spain. effector cells in Eosinophils are hypersensitivity disorders and parasitic infections. In this report we present data of a possible rote of eoeinophiIs in the immune response as antigen presenting cells (APC) and the mechanisms implicated in the activation of these cells for mediator release (LTC,), in the inflammatory reactions. %usin~phil
in the
Immune
Retqxmaa:
We have
previously shown that this cell has mRNA for IL1 (J Immlinol 144: 3117-22, 1990). Also murine eosinoohile treated with x-GM-CSF express MHC class II molecules. In this report we dkmonstrate that antigen specific T cell clones proliferate when to the plates containing eosinophile added pretreated with GM-CSF and pulsed with the antigen. The proliferation of the T cells is dependent on the number of eeainophiis used as APC. Eosinophil presentation is restricted by Class II and inhibited by lysosomotropic agents. Looking at the role of eoain~PI&a in the irdlmmatiry reactioni we investigated the involvement of phoaphoinoaitide breakdown in the .activation of 5'-Hpoxygenase. Using neomycin, we have demon&rated a decrease in the LTC released. [Caa'I, elevation has been observe d using
Fura-B/AM,
following
stimulation
with
the
anti-Fc-gamma RI1 mAb. It is likely that the activation of murine wainophils by a Fc-&unum receptor mechanism etimuletea phoaphainoaitide breakdown.
CLIN. IMMUNOL. JANUARY 1991
OF THE Pff&WM&-WT Tg T~~F~~ Ill VfsPm mz&Y. G.Kunkel. Dpt. of FC’in. &&ol. Free University Berlin, FRG
and
Determination of specific IgE with RAST (antigens bound to paper disc as solid phase) is the @ost conmonly used in vitro diagnostic method in the practice of allergy. In this study we coarpared, a nowel test system which is based on a new type of solid phase, consisting of a flexible hydrophilic carrier polymer encased in a capsule, the lagunoCAP to the RAST. We investibated 123 sera fratn ill vospld venom allergic pi;tients (74% frtaale). All patients were diagtlosed on the basis of their history, skin test and RRST. In addition, the standard serum of the RAST-kit (hi h titer of birch pollen specific IgE) was eva 3 uated with the CAP-kit. Results: The patients’ age ranged from 8 to 80, with a mean% of 44.2a14.6 years. The total serum IoE levels ranoed from 4-1712 kU/l, with a median”of 108 kU/l.“The results of specific IgE for vespid.venoIA (i ), as determined by RAST ranged from class O-4 wi *i! h a atean*% of 1.87kl.07 (median: 2.1). The CAP systetn ranging from O-6 with a meanas of 2,78*1.38 (median: 3.0). The correlation coefficient betwaen the fWT and CAP systeai was r=g&l. The two sysstglas cowpared here, showed a higher sensitivity for the CAP system by almost one class, compared to the g&ST. It seemed that low sensitized patients were not detected by the MST (MST class 0), whereas the saare sera showed CAP classes be&man 1 aad 2. We would therefore rocgggtrsl~to cat1 the scoriz of the CAP system “CAP-clata%” imtead.of RMT classes so that it is obvious wfth which method the in vitro data were obtained.