Nuclear factor translocation and acute anterior uveitis

[摘要] Purpose: To investigate the roles ofactivation of macrophages isolated from C3H/HeN and C3H/HeJ mice andstimulated by lipopolysaccharide (LPS), and toll-like receptor4-mediated signal transduction in the development of acute anterioruveitis. Methods: Establish animal models withacute anterior uveitis by intraperitoneal injection of vibrio choleraendotoxin into C3H/HeN mice (wild type) and C3H/HeJ mice (toll-likereceptor 4 (TLR4) gene defection type). Peritoneal macrophageswere obtained from C3H/HeN and C3H/HeJ mice. Immunofluorescencestaining was used to identify the F4/80+ positive cells (iris,peritoneal macrophages) and to observe the expression of TLR4, myeloiddifferentiation factor 88 (MyD88), and nuclear factor kappa B (NF-κB),with or without LPS (1 μg/ml). To investigate the importance of TLR4 inthe signal pathway, a group, blocked by anti-TLR4 antibody before LPSstimulation, was added to theC3H/HeN mice sample. Results: In vitro, in C3H/HeN mice, Irisposterior synechia was found 24 h later. However, an inflammationreaction was not found in the anterior chamber of the C3H/HeJ mice. Incell culture, TLR4 expression was observed in peritoneal macrophages ofthe C3H/HeN mice, both with and without LPS stimulation. TLR4 wasprimarily expressed in the membrane and no significant difference ininflorescence intensity (p=0.081) was found among the groups. MyD88 wasexpressed in the cytoplasm and the nucleus. There is statisticalsignificance in the fluorescence intensity among groups of C3H/HeN mice(p<0.0001). NF-κB was primarily expressed in the cytoplasm beforeLPS stimulation. However, this occurred 1 h after LPS stimulation andcould be observed in the nucleus. Three hours after LPS stimulation,the expression of NF-κB could not be detected in the cytoplasm or thenucleus. The fluorescence intensity of TLR4 and MyD88 expression showedno significant difference (p=0.113) between the anti-TLR4 antibodypretreatment group and the other groups of C3H/HeN mice. However, inthe anti-TLR4 antibody pretreatment group, 1 h to 24 h after LPSstimulation, NF-κB only expressed in the cell membrane. Peritonealmacrophages from all groups of C3H/HeJ mice showed no obvious changesin morphology and size after LPS stimulation (p=0.257). TLR4 wasprimarily expressed in the cell membrane, and fluorescence intensityshowed no statistical significance (p=0.228); MyD88 was expressed inthe cytoplasm and the nucleus and there was no significant differencein fluorescence intensity among the groups (p=0.108); NF- κB wasexpressed in the cytoplasm, without LPS stimulation; however, 1 h afterLPS stimulation, it appeared in the cell membrane and persisted until24 h. Conclusions: Acute anterior uveitis canbe induced in wild-type mice, but it cannot be induced in TLR4gene-deficient mice. The variation of expression of TLR4, and itsdownstream signal transduction molecules, MyD88 and NF-κB, after LPSstimulation in vitro, suppose the potential role of aTLR4-MyD88-dependent pathway in the pathogenesis of acute anterioruveitis. The blockage of this pathway by anti-TLR4 may signal a newdirection in the treatment of acute anterior uveitis.

[学科分类] 生物化学/生物物理