Background Nuclear factor-kappa B （NF-kB） is elevated in regulating transcription of many cytokines and inflammatory mediators. The purpose of this study was to investigate the activation and the significance NF-KB in lipopolysaccharide （LPS） induced keratitis.Methods LPS induced keratitis model was based on Wistar rats. At 0. 5, 1, 3, 6, 12, 24 or 72 hours after LPS treatment, the rat corneas were observed with a slit lamp microscope, then the rats were sacrificed and their corneas were excised for routine histological analysis. The expression of NF-kB was detected with immunohistochemical staining. The change identified by reverse transcriptase polymerase of tumour necrosis factors-α （TNF-α） mRNA expression was chain reaction （RT-PCR）. Results Histological findings demonstrated that LPS treated corneas showed significant changes in corneal structure. Corneal edema, pronounced inflammatory cells infiltration and inordinate collagen fibres were observed. Immunohistochemical results showed that the expression of NF-kB and its activation obviously increased after LPS treatment compared with the normal group and control group. Positive cells could be observed at 0. 5 hour and peak expression of NF-kB was observed between 3 hours and 12 hours after infection, but returned to or approached normal level by 72 hours. RT-PCR showed that the level of TNF-α mRNA began to increase 0. 5 hour after LPS treatment, peaked at 6 hours and then subsided by 72 hours. NF-kB had a positive correlation with the expression of TNF-α mRNA （r=0. 964, P 〈 0. 01 ） , both NF-kB and TNF-α had a strong positive correlation with the degree of inflammatory response in LPS treated corneas （ r=0. 929, P 〈 0. 01 ; r=0. 587, P 〈 0.05, respectively）. Conclusions The activation of NF-kB was increased keratitis by promoting overexpression of TNF-α mRNA. LPS-associated keratitis in rats. in LPS treated corneas and was elevated in LPS induced NF-kB may play an important role in the pathogenesis ofLPS-associated keratit
WU Xin-yi HAN Shao-ping REN Mei-yu CHANG Yuan YU Fu-xin
Background Toll-like receptors play an important role in the human immune system. This study was conducted to investigate the expression profiles and function of Toll-like receptor （TLR）1-9 in human corneal epithelium. Methods The expression of TLR1-9 mRNA in 20 human donor corneal epithelia samples abraded during photorefractive keratotomy （PRK） and cultivated telomerase-immortalized human corneal epithelial cells （THCEs） was examined by semi-quantitative reverse-transcriptase polymerase chain reaction （RT-PCR） analysis. Human peripheral blood mononuclear cells （PBMCs） were used as positive controls. The expression of the TLR2 and TLR4 proteins was detected by Western analysis. ELISA was used to detect IL-8 secretion from THCEs challenged with ligands for TLR3 and TLR4 with and without antibody blockade. Results The expression of TLR1-9 at the mRNA level was detected in the epithelia of 20 patients and in THCE. Significant differences among individuals were observed. One patient was found to lack of the expression of TLR3, 4, 6 and 8, whereas another did not express TLRS. The expression of TLR2 and TLR4 protein was detected in human corneal epithelial cells. As THCE cells express TLR1-9, cells were challenged with lipopolysaccharides （LPS） and poly I：C to determine whether TLR4 and TLR3 were functional. The results showed that secretion of IL-8 by cells stimulated with LPS and Poly I：C was 7 to 10 fold greater than secretion by unchallenged cells. Blocking TLR4 with an anti-TLR4 antibody significantly inhibited the LPS-induced IL-8 production by THCE （P〈0.05）. Conclusion Human corneal epithelial cells express multiple TLRs and are able to recognize LPS and poly I：C. Different expression profiles among individuals suggest that differences in the susceptibilities and sensitivities to bacterial and viral infection in human populations relate to different patterns of TLR expression.
