An analysis of interferon gamma, IL-4, IL-5 and IL-10 production by ELISPOT and quantitative reverse transcriptase-PCR in human Peyer's patches.
The cytokine profiles of mononuclear cells freshly isolated from Peyer's patch (PPMC), adjacent ileal lamina propria lymphocytes (LPMC) and peripheral blood (PBMC) in children without histological evidence of gastrointestinal disease has been investigated by single-cell enzyme-linked immunoabsorbent spot forming assay (ELISPOT) and reverse transcriptase (RT)-PCR. In the blood, interferon gamma and IL-4 ELISPOTs were regularly detected albeit at low frequency (< 50/10(5) cells). IL-5 and IL-10 ELISPOTs were not seen in most patients. In Peyer's patches and lamina propria there was a dramatic increase in cytokine secreting cells of all types compared to blood, reaching a very high frequency for interferon gamma in the lamina propria (1000-3000/10(5) cells). IL-4 and IL-5 ELISPOTs were 20-100-fold less common in both PP and LPL. At all sites, cytokine secretion depended on protein synthesis and enrichment for CD4+ cells in PP increased the frequency of all cytokine-secreting cells. Quantification of messenger RNA for cytokines using RT-PCR demonstrated that IL-4 and IL-10 transcripts were significantly greater than interferon gamma transcripts in PP and in lamina propria, IL-4, IL-10 and interferon gamma transcripts were equivalent. IL-5 transcripts were not detected in most samples of PP and lamina propria. These results clearly show that cells secreting interferon gamma predominate in human PP and LPL. However the high mRNA concentrations for IL-4 and IL-10 shows that although these cells are quantitatively few, they are highly transcriptionally active.
Department of Paediatric Gastroenterology, St Bartholomews Hospital, London.
A C
AC
Hauer
M
M
Bajaj-Elliott
C B
CB
Williams
J A
JA
Walker-Smith
T T
TT
MacDonald
D000293:Adolescent; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D003513:Cycloheximide / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D007223:Infant; D007371:Interferon-gamma / Q000096:biosynthesis* / Q000235:genetics; D016753:Interleukin-10 / Q000096:biosynthesis* / Q000235:genetics; D015847:Interleukin-4 / Q000096:biosynthesis* / Q000235:genetics; D015848:Interleukin-5 / Q000096:biosynthesis* / Q000235:genetics; D007963:Leukocytes, Mononuclear / Q000187:drug effects; D008297:Male; D010581:Peyer's Patches / Q000378:metabolism* / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012194:RNA-Directed DNA Polymerase
D015848:Interleukin-5; D011500:Protein Synthesis Inhibitors; D016753:Interleukin-10; D015847:Interleukin-4; D007371:Interferon-gamma; D003513:Cycloheximide; D012194:RNA-Directed DNA Polymerase