IgE-dependent expression of mRNA for IL-4 and IL-5 in human lung mast cells.
By using the reverse transcriptase (RT)-PCR and in situ hybridization we have studied the expression of mRNA for IL-5 and IL-4 in human lung mast cells induced by cross-linkage of high affinity Fc epsilon Rs. Lung mast cells were purified using affinity magnetic selection with mAb YB5.B8 against c-kit to achieve a final mast cell purity > 93%. Purified mast cells were precultured with stem cell factor (SCF) (10 ng/ml) and myeloma IgE (3 micrograms/ml) for 16 h before challenge with anti-IgE (1 or 10 micrograms/ml). IgE-dependent activation of lung mast cells caused expression of IL-5 mRNA, which was evident by 2 h and persisted for up to 48-72 h in all of 12 experiments, whereas IL-4 mRNA expression was of a shorter duration and was demonstrable in 6 of 13 experiments. We confirmed that mast cells, and not T cells, were the source of these cytokine messages by using reverse transcriptase-PCR in cell preparations containing known numbers of mast cells and T cells, in situ hybridization in enriched mast cell preparations, and double in situ hybridization-immunocytochemical staining. IL-5 mRNA expression did not require the pretreatment of cells with SCF, whereas expression of IL-4 mRNA seemed to require both anti-IgE and SCF. The strength of IL-5 mRNA signal was related to anti-IgE concentration. Immunoreactive IL-5 was detectable 8 h after anti-IgE challenge, and 10(6) mast cells generated a mean of 731 +/- 400 pg of IL-5 into the supernatant during 48-h culture, but no IL-4 product was detectable. These findings demonstrate the capacity of human lung mast cells to transcribe IL-4 and IL-5 after IgE-dependent activation and to synthesize and release immunoreactive IL-5.
Okayama
Y
Y
Immunopharmacology Group, Southampton General Hospital, United Kingdom.
Petit-Frére
C
C
Kassel
O
O
Semper
A
A
Quint
D
D
Tunon-de-Lara
M J
MJ
Bradding
P
P
Holgate
S T
ST
Church
M K
MK
D001483:Base Sequence; D002478:Cells, Cultured; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D006801:Humans; D007073:Immunoglobulin E / Q000502:physiology*; D015847:Interleukin-4 / Q000235:genetics*; D015848:Interleukin-5 / Q000235:genetics*; D008168:Lung / Q000378:metabolism; D008407:Mast Cells / Q000378:metabolism*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis*; D017455:Receptors, IgE / Q000502:physiology; D019089:Stem Cell Factor
D016298:Hematopoietic Cell Growth Factors; D015848:Interleukin-5; D012333:RNA, Messenger; D017455:Receptors, IgE; D019089:Stem Cell Factor; D015847:Interleukin-4; D007073:Immunoglobulin E