Abstract
Necrotizing enterocolitis (NEC) is an important and sometimes fatal intestinal disease that primarily affects premature neonates and although it has been extensively investigated, the complete pathogenesis of NEC remains unclear. Important immunological factors such as interleukin 8 (IL-8), toll like receptor 4 (TLR4), cluster of differentiation 14 (CD14), lipopolysaccharide (LPS), and also bacteria from the Enterobacteriaceae family have been implicated in the pathogenesis of NEC while probiotics have been used clinically as a prophylactic treatment. In order to produce more effective treatments, it is important to understand the pathophysiology of NEC and also the mechanism of action of probiotics. We demonstrate here that human colonic epithelial cells (HT-29 cells) are able to produce significantly increased amounts of IL-8 in response to purified bacterial LPS and 12 strains of Enterobacteriaceae, isolated from clinical NEC specimens, compared to unstimulated controls. This response was time and dose-dependent, and was confirmed by the use of another colonic epithelial cell line (Caco-2 cells). Two strains of clinically utilised probiotic bacteria (Bifidobacterium infantis and Lactobacillus acidophilus) did not induce IL-8 production by either intestinal epithelial cell (IEC). The addition of live B. infantis or a Gram positive cell wall component, lipoteichoic acid (LTA), was able to significantly reduce the IL-8 response seen during an LPS stimulation of HT-29 cells. No reduction in IL-8 production was demonstrated by heat inactivated probiotics or live L. acidophilus. CD14 and TLR4 were expressed by HT-29 cells and it was found that LPS induced IL-8 production was CD14-dependent. It has previously been shown that LPS and bacteria can induce an IL-8 response from intestinal epithelial cells and that probiotic bacteria are effective at reducing both this response and also the incidence of NEC. However, this is the first report of an immunological mechanism using these NEC-associated Enterobacteriaceae and probiotic strains and intestinal epithelial cells.