TLR4 and MD-2 expression is regulated by immune-mediated signals in human intestinal epithelial cells

J Biol Chem. 2002 Jun 7;277(23):20431-7. doi: 10.1074/jbc.M110333200. Epub 2002 Mar 28.


The normal intestinal epithelium is not inflamed despite contact with a high density of commensal bacteria. Intestinal epithelial cells (IEC) express low levels of TLR4 and MD-2 and are lipopolysaccharide (LPS)-unresponsive. We hypothesized that immune-mediated signals regulate the expression of TLR4 and MD-2 in IEC. Expression of TLR4 and MD-2 was examined in normal colonic epithelial cells or intestinal epithelial cell lines. The effect of the cytokines interferon (IFN)-gamma, IFN-alpha, and tumor necrosis factor-alpha (TNF-alpha) on TLR4 and MD-2 expression was examined by reverse transcription-PCR and Western blot. NF-kappaB transcriptional activation and interleukin-8 secretion were used as measures of LPS responsiveness. Native colonic epithelial cells and IEC lines express a low level of TLR4 and MD-2 mRNA. IFN-gamma regulates MD-2 expression in both IEC lines, whereas IFN-gamma and TNF-alpha regulate TLR4 mRNA expression in IEC lines. Pre-incubation with IFN-gamma and/or TNF-alpha sensitizes IEC to LPS-dependent interleukin-8 secretion. To examine MD-2 transcriptional regulation, we cloned a 1-kb sequence proximal to the MD-2 gene translational start site. This promoter directed expression of a reporter gene in endothelial cells and IEC. IFN-gamma positively regulated MD-2 promoter activity in IEC. Co-expression of a STAT inhibitor, SOCS3, blocked IFN-gamma-mediated MD-2 promoter activation. T cell-derived cytokines lead to increased expression of TLR4 and MD-2 and LPS-dependent pro-inflammatory cytokine secretion in IEC. IFN-gamma regulates expression of the critical TLR4 co-receptor MD-2 through the Janus tyrosine kinase-STAT pathway. Th1 cytokines may initiate or perpetuate intestinal inflammation by altering toll-like receptor expression and bacterial reactivity.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Antigens, Surface / genetics
  • Antigens, Surface / metabolism*
  • Base Sequence
  • Cell Line
  • Cytokines / immunology
  • DNA Primers
  • Drosophila Proteins*
  • Gene Expression Regulation / immunology*
  • Humans
  • Intestinal Mucosa / cytology
  • Intestinal Mucosa / immunology
  • Intestinal Mucosa / metabolism*
  • Lymphocyte Antigen 96
  • Membrane Glycoproteins / genetics
  • Membrane Glycoproteins / metabolism*
  • Promoter Regions, Genetic
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Receptors, Cell Surface / genetics
  • Receptors, Cell Surface / metabolism*
  • Signal Transduction / immunology*
  • Toll-Like Receptor 4
  • Toll-Like Receptors


  • Antigens, Surface
  • Cytokines
  • DNA Primers
  • Drosophila Proteins
  • LY96 protein, human
  • Lymphocyte Antigen 96
  • Membrane Glycoproteins
  • RNA, Messenger
  • Receptors, Cell Surface
  • TLR4 protein, human
  • Toll-Like Receptor 4
  • Toll-Like Receptors