Absence of Toll-like receptor 4 explains endotoxin hyporesponsiveness in human intestinal epithelium

Background: The Toll protein in Drosophila regulates dorsal ventral pattern ing during embryogenesis, and participates in antibacterial and antifungal host defense. Mammalian homologues are termed Toll-like receptors and, to d ate, nine have been cloned (TLR1-9) in humans. They are characterized by ex tracellular leucine-rich repeats and a cytoplasmic domain similar to the in terleukin 1 receptor. Both TLR2 and TLR4 recognize various bacterial cell w all components including lipopolysaccharide (LPS). This results in the acti vation of the NF kappaB pathway. Peripheral blood mononuclear cells (PBMCs) express both TLR2 and TLR4, The authors hypothesized that the expression o f TLR 2 and TLR4 in human intestinal epithelial cells differs from PBMCs be cause of the abundance of LPS in the intestinal lumen. Methods: Epithelial cells were isolated from Caco-2 cells, fetal gut explan ts, and small bowel resection specimens using Hanks/ethylenediamine tetraac etic acid solution. PBMCs were used as positive controls. Ribonucleic acid (RNA) was isolated using the TRIzol method. Standard reverse transcriptionp olymerase chain reaction examined TLR2 and TLR4 messenger RNA (mRNA) expres sion. NF kappaB expression was determined using a luciferase reporter assay . Results: TLR2 mRNA was highly expressed in PBMCs and was present in all hum an intestinal epithelial cells. TLR4 mRNA was detected only in PBMCs. TLR4 is not present in epithelium from children with inflammatory bowel disease. In Caco-2 cells, significant NF kappaB activation in response to LPS occur red only in the presence of TLR4 introduced by complementary deoxyribonucle ic acid transfection. Conclusion: Absence of TLR4 is associated with endotoxin hyporesponsiveness of intestinal epithelial cells. TLR4 is not directly involved in inflammat ion of the intestinal epithelium. Although TLR2 is normally present in the epithelial cell, it plays a limited role in inflammation. It may be activat ed during conditions in which bacterial cell wall concentrations within the intestine are pathologically high.