Blockade of transforming growth factor beta upregulates T-box transcription factor T-bet, and increases T helper cell type 1 cytokine and matrix metalloproteinase-3 production in the human gut mucosa.
OBJECTIVE
The role of transforming growth factor beta (TGFbeta) in inhibiting T cell function in the normal gut has been studied in animal models. However, the impact of TGFbeta inhibition on T cells in the normal human gut remains poorly understood. The effect of TGFbeta blockade in normal intestinal biopsies grown ex vivo and lamina propria mononuclear cells (LPMCs) on T-bet, a T-box transcription factor required for T helper cell type (Th)1 differentiation, interferon gamma (IFN gamma) production, T cell apoptosis and matrix metalloproteinase (MMP)-3 production has therefore been tested.


METHODS
TGFbeta transcripts were determined by quantitative reverse transcription-PCR in laser-captured gut epithelium and lamina propria. Biopsies and LPMCs were cultured with anti-TGFbeta neutralising antibody. After 24 h culture, T-bet was determined by immunoblotting, and T cell apoptosis was assessed by flow cytometry. IFN gamma, tumour necrosis factor alpha (TNFalpha), interleukin (IL) 2, IL6, IL8, IL10, IL12p70 and IL17 were measured by ELISA. MMP-3 and tissue inhibitor of matrix metalloproteinase (TIMP)-1 were assessed by immunoblotting.


RESULTS
A higher number of TGFbeta transcripts was found in the lamina propria than in the epithelium in normal gut. T-bet expression was significantly higher in biopsies and LPMCs cultured with anti-TGFbeta antibody than in those cultured with control antibody. TGFbeta blockade downregulated T cell apoptosis, and induced a significant increase in IFN gamma, TNFalpha, IL2, IL6, IL8 and IL17 production. A higher expression of MMP-3, but not TIMP-1, was observed in the tissue and supernatant of biopsies treated with anti-TGFbeta antibody.


CONCLUSIONS
The findings support a crucial role for TGFbeta in dampening T cell-mediated tissue-damaging responses in the human gut.
Di Sabatino
A
A
Centre for Infectious Disease, Institute of Cell and Molecular Science, Barts, London School of Medicine and Dentistry, Whitechapel, London E1 2AT, UK.
Pickard
K M
KM
Rampton
D
D
Kruidenier
L
L
Rovedatti
L
L
Leakey
N A B
NA
Corazza
G R
GR
Monteleone
G
G
MacDonald
T T
TT
D000328:Adult; D016207:Cytokines / Q000096:biosynthesis*; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D015212:Inflammatory Bowel Diseases / Q000378:metabolism*; D007371:Interferon-gamma / Q000378:metabolism; D007378:Interleukins / Q000378:metabolism; D008297:Male; D019278:Matrix Metalloproteinase 3 / Q000096:biosynthesis*; D008875:Middle Aged; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020825:T-Box Domain Proteins / Q000378:metabolism*; D018417:Th1 Cells / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors*; D015854:Up-Regulation; D000098815:T-bet Transcription Factor
D016207:Cytokines; D007378:Interleukins; D020825:T-Box Domain Proteins; D000098815:T-bet Transcription Factor; D016212:Transforming Growth Factor beta; D007371:Interferon-gamma; D019278:Matrix Metalloproteinase 3