| Backgrounds and Objective:The pathogenesis of uveitis is very complicated. More and more researchesdemonstrated that the pathogenesis is associated with disorders of many kinds ofcytokines. Interleukin-10 (IL-10), which is a potential anti-inflammatory agent, caninhibit the cytokine production from Th1 cells and suppress the activity of thosecytokines. Moreover, cytokines such as chemotactic factor, cell adhesion factor, activeoxygen and nitrogen medium, can also be suppressed by IL-10. Early experimentalresearch revealed that IL-10 could function as a powerful cytokine regulatory factor toprotect many kinds of inflammations. Whether IL-10 could be used in uveitis, fewliteratures have been reported abroad and no related literature was found domestically.Immuning Wistar rats with the lipopolysaccharide (LPS), and establishing a uveitismodel and this afford a worthy tool for experimental study of immunogenic andtherapy of uveitis. Endotoxin-induced uveitis (EIU) is a model for acute panuveitisseen in human. We report a study of the effect of systemic administration of IL-10 onthe the clinical presentation, histopathologic change and the level of tumor necrosisfactor (TNF-α), IL-10 and IL-6 in the serum and intraocular fluid of EIU, andinvestigate the therapeutic role of IL-10 on the EIU in the Wistar rats.Methods1. The establishment of endotoxin-induced uveitis disease modelMale Wistar rats were selected between 6～8 weeks age (150～180g). Fifteen Wistarrats were randomly divided into the healthy control group (3 rats) and the experimental group (12 rats). Lipopolysaccharides were dissolved in the sterile,pyrogen-free 0.9% saline at a concentration of 2 mg/ml, and 75μl was injected intoeach hind footpad of Wistar rats in the experimental group (300μg total dose per rat).The rats in the healthy group were injected 0.9% saline only. The clinical presentationsuch as iris hyperemia, floating cells, flaring and exudation in anterior chamber wereobserved at different times. Five rats were killed at 4 hours, 24 hours and 7 daysrespectively after injection with LPS, and histological examinations were performed.Clinical evaluation and histopathologic study were carried through consulting thestandards of Hoekzema's and Tilton's research.2. The role of interleukin-10 in the treatment of experimental EIU(1) Fifty-four Wistar rats were randomly divided into three groups, including thehealthy control group (Group A, 6 rats), EIU group (Group B, 24 rats) and IL-10group (Group C, 24 rats), 18 rats were killed at the end of 4 hours, 24 hours and 3days respectively.(2) The rats in the healthy group were injected 0.9% saline only. 300μg ofLipopolysaccharide were injected into hind footpads of Wistar rats in EIU group.Recombinant rat IL-10 at 1μg /kg were administrated twicely by intravenousinjections in the tail vein given at the time of 4.0h and 0.5h before lipopolysaccharideinjection.(3) Clinical representation and histopathologic examination: All the Wistar ratswere observed using slit lamp biomicroscope and ophthalmoscope every 2 hours afterimmunization. The onset times, peak times, durations of the inflammation, clinicalmanifestations were noted. The clinical representation was evaluated according thestandards of Hoekzema's research. We extracted the blood from angular vein at theend of 4h, 24h and 3d after immunization, then the rats were killed and their eyeballswere excised. One eye was used to extract intraocular fluid, and the other was used to make pathlolgical section conventionally. All samples were examined with lightmicroscope, and the histopathologic study was evaluated according the standards ofTilton's research.(4) The levels of TNF-α, IL-10 and IL-6 in the serum and intraocular fluid weremeasured by ELISA method.(5) Statistic methods: The data was denoted by (?)±s, and analyzed the data bySPSS 11.5 software.ResultsThe EIU disease model was established successfully in Wistar rats in this study.The disease incidence was 100%. The clinical and histological features were stable. Inthe EIU groups, the inflammation began 2～4 hours after LPS injection. The peak timeof the inflammation was 22～24 hours after injection. Obvious uveitic inflammatorysigns such as flare, cells and protein exudates in anterior chamber, occlusion of pupiland vitreous opacity were observed. Whereafter the inflammation subsidizedgradually. Histological examination revealed the vasodilatation of iris vessels,massive infiltration of polymorphonuclear cells or a few of monocytes andlymphocytes in the anterior chamber, iris, ciliary body, retina and choroids at 22～24hours after the endotoxin injection.In the IL-10 treatment group, the degree of eye damagement was significantlyattenuated compared to the EIU group, the disease scores and histopathologic levelswere also lower than those in the EIU group, and the duration was shorter. The levelof TNF-αin the serum and intraocular fluid in healthy control group were 51.03±9.59 pg/ml and 26.8 pg/ml at the beginning of the inflammation. High level of TNF-α(212.76±64.26 pg/ml and 411.45±177.34 pg/ml) was detected in the serum andintraocular fluid early after LPS injection in EIU group, and it was higher than that inthe healthy control group. But in IL-10 group, the level of TNF-αin the serum and intraocular fluid decreased to the level of the heathy control group. The level ofTNF-αin the serum (F=13.77, p＜0.01) and intraocular fluid (F=26.77, p＜0.01) wassignificantly reduced in the groups treated with IL-10. High levels of IL-6 weredetected in the serum and eye fluid early after LPS injection in EIU group, and theywere higher than those in healthy control group. The serum level of IL-6 was alsoreduced in the group treated with IL-10 compared to the EIU group (F=12.65, p＜0.01).The serum level of IL-10 was increased in the treatment group (F=8.96, p＜0.01), butthe intraocular fluid level of IL-10 (F=0.01, p＞0.05) and IL-6 (F=2.05, p＞0.05) wasnot significantly affected by IL-10 treatment. There were positive correlationsbetween the level of IL-6 in serum and intraocular fluid and the disease severity (r=0.687, p＜0.01, r=0.672, p＜0.01), between the level of TNF-αin serum and thedisease severity (r=0.707, p＜0.01).ConclusionsUveitis with marked clinical signs is successfully induced in the Wistar rats withinjection of LPS, which can act as a model for panuveitis seen in human. This mothedis simple and stable, there is a high incidence and perfect repetition, and it will be anexperimental foundation for the further study of uveitis. Furthermore, we comparedLPS and S-antigen in inducing the uveitis, discussed the immunogenic machinism ofLPS to induce uveitis.IL-10, as a potent anti-inflammation factor, could significantly inhibit thepro-inflammatory cytokines such as TNF-αand IL-6 in the uveitis, decrease theconcentration of TNF-αand IL-6 in the serum and intraocular fluid, and attenuatepathological eye damagement. Thus IL-10 contributes to keep the balance betweeninflammatory and anti-inflammatory mediators. IL-10 may be effective in thetreatment of EIU, and this provides direct evidence for immunotherapy of uveitis in clinics. It will become the main direction in the future that the cytokine regulatoryfactor adjusts the cytokine in the patients with uveitis. |
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