Inflammation-causing effects and mechanism of natural killer cells in experimental autoimmune uveitis rats / 中华实验眼科杂志

ABSTRACT

Background Experimental autoimmune uveitis (EAU) is a common animal model of uveitis.Natural killer (NK) cells have been confirmed to be a type of strong inflammation-causing cells,but its role in EAU is still studing.Objective This study was designed to explore the role and mechanism of NK cells in the pathogenesis of EAU.Methods Thirty-six SPF Lewis rats were randomly divided into expeimental control group and EAU 6-,9-,12-,16-,and 21-day groups (6 rats for each group).Rats in EAU group received subcutaneous injection interphotoreceptor retinoid binding protein (IRBP) combining 5 mg/ml tubercle bacillus with complete Freund's adjuvant (CFA) emulsion in foot pads,and then 400 ng pertussis toxin was intraperitoneally injected to extablish EAU models in the EAU 6-,9-,12-,16-,and 21-day group,and normal saline solution combined with CFA and 400 ng pertussis toxin was used in the same way in the experimental control group.The inflammatory response was observed by slit lamp daily after modeling and scored based on Caspi criteria.The eyeballs were extracted in 6,9,12,16 and 21 days after modeling for retinal histopathological examination,Immunofluorescent double-staining was employed to detect and locate the expression of NK cells in the retina.In addition,25 model rats were divided into EAU 0-,3-,6-,9-and 12-day groups,with 5 rats for each group,and eyeballs were extracted to prepare tissue homogenate.The expression of CXCL10 mRNA,and CXCL12 mRNA NK cell chemokines,in the tissue homogenate was assayed by real-time quantitative PCR.The use and care of the rats followed Regulations for the Administration of Affair Concerning Experimental Animal by State Science and Technology Commission.Results No inflammatory sign in ocular anterior segment of the rats was seen in the experimental control group.The expansion of rat iris vessels was found in the EAU 6-day group,and exudes and hypopyon of the anterior chamber occurred in the EAU 9-day group and the inflammation peaked in the EAU 12-day gorup.The rat retinal structure was normal in the experimental control group,and the arrangement disorder of retinal structure,the cell separation in outer nuclear layer and damage of photoreceptors were found under the optical microscope in different degree in various EAU groups,with the most serious change in the EAU 12-day group.Immunofluorescent double staining showed normally arranged nucleus in the experimental control group,and a lot of NK infiltration was seen in the EAU 6-day group and peaked in the EAU 9-day group.The expression level of CXCL10 mRNA in the EAU 9-day group was 34.298 ± 16.689,which was significantly higher than that in the EAU 3-,6-and 12-day group,respectively (1.390 ± 0.660,3.359 ± 2.581,4.711 ±1.387) (all at P＜0.01).No significant differences were found in the relative expression of CXCL12 mRNA among different EAU groups (F=2.851,P＞0.05).Conclusions Retinal NK cell infiltration occurs in the early stage of EAU,and the severity of NK cell infiltration is consistent with the inflammatory process and CXCL10 expression,suggesting NK cells play an important role in the early stage of EAU,and CXCL10 is an important chemokine of NK cells in EAU rats.