真菌性角膜炎的Th反应类型调查

发布时间：2018-07-07 18:00

本文选题：BALB/C + C57BL/6　；参考：《浙江大学》2010年硕士论文

【摘要】： 目的：(1)建立BALB/C和C57BL／6两种不同品系小鼠的镰刀菌和曲霉菌性的角膜炎模型；(2)对比两种品系小鼠真菌性角膜炎的临床表现差异；(3)测定感染眼球及同侧颈部淋巴结的Th1相关细胞因子IFN-γ及Th2相关细胞因子IL-4的变化情况。方法：(1)小鼠角膜真菌感染模型的制作：用2.0mm环转在小鼠角膜中央印痕,自制基质划伤钩于印痕范围内作线形划伤至浅基质层,共5道,板层刀去除损伤区域内残余的角膜上皮,然后将5μl的真菌孢子混悬液(浓度为108CFU／ml)滴入结膜囊,自然干燥。(2)接种后用裂隙灯显微镜观察角膜浸润病灶及眼部炎症反应的变化,每天1次。(3)接种后24h,48h和72h处死小鼠,摘除模型眼及相应侧颈部淋巴结,分别制作成组织匀浆,ELISA法检测Th1相关IFN-γ及Th2相关IL-4的变化。结果：(1)BALB／c小鼠在接种镰刀菌和曲霉菌后仅出现轻、中度角膜浸润混浊和炎症反应,无组织化脓坏死,5-7d内所有角膜浸润消失而痊愈,角膜恢复透明或仅出现少量疤痕；而C57BL／6小鼠在接种真菌后1-2d内即出现浓密的角膜浸润混浊,重度基质水肿和前房积脓,3d后组织发生化脓坏死,角膜穿孔,眼内容突出,最终以严重的角膜疤痕混浊而愈合,部分小鼠出现眼球萎缩。(2)BALB／c小鼠在感染镰刀菌和曲霉菌后眼球IFN-γ均明显升高,高峰值分别为正常的2.9和2.6倍；同时,颈淋巴结IFN-γ也明显升高,但低于感染局部的升高幅度,高峰值分别为正常的2.0和1.5倍。而C57BL／6在感染镰刀菌和曲霉菌后眼球IFN-γ仅轻度升高,且短时间内下降至正常,高峰值仅为正常的1.3和1.1倍,但颈淋巴结IFN-γ则明显升高,在72h分别达到正常的1.6和1.5倍。(3)BALB/c在感染镰刀菌和曲霉菌后眼球IL-4均明显降低,低谷水平仅为正常的0.6和0.5；而颈淋巴结IL-4则逐渐升高,高峰值为正常的1.2和1.5倍。而C57BL／6在感染镰刀菌和曲霉菌后眼球IL-4也明显降低,低谷水平为正常的0.5和0.6,且持续至72h仍维持低水平；但颈淋巴结IL-4也逐渐升高,高峰值为正常的1.9和1.6倍。结论：(1)真菌性角膜炎是以Th1介导为主的免疫炎症反应过程；(2)感染局部Th1反应的强弱是真菌性角膜炎预后的重要因素；(3)BALB/c小鼠感染局部的Th1反应明显强于C57BL／6小鼠；(4)在角膜真菌感染过程中,Th1极化在感染局部和颈淋巴结同步出现,而Th2极化则在感染局部和颈淋巴结出现分离现象。
[Abstract]:Objective: (1) to establish Fusarium and aspergillus keratitis models of BALB / C and C57BL / 6 mice, (2) to compare the clinical manifestations of fungal keratitis between the two strains. (3) the changes of Th1-associated cytokine IFN- 纬 and Th2-associated cytokine IL-4 in infected eyeball and ipsilateral cervical lymph nodes were measured. Methods: (1) the model of corneal fungal infection in mice: the 2.0mm loop was applied to the central impression of the cornea of the mice, and the self-made matrix laceration hook was used for linear scratching to the superficial stromal layer in the range of the mark, for a total of 5 passages. The residual corneal epithelium in the injured area was removed by lamellar knife, and then 5 渭 l fungal spore suspension (concentration 108 CFU / ml) was dripped into conjunctival sac. (2) the corneal infiltrating focus and ocular inflammatory reaction were observed by slit lamp microscope after inoculation. (3) the mice were killed at 24 h, 48 h and 72 h after inoculation, the model eyes and the corresponding cervical lymph nodes were removed, and the changes of Th1-related IFN- 纬 and Th2-related IL-4 were detected by Elisa. Results: (1) after inoculation with Fusarium and Aspergillus, BALB / c mice only showed mild and moderate corneal infiltration, turbid and inflammatory reaction. All corneal infiltration disappeared and recovered within 5-7 days without tissue purulent necrosis, and the cornea recovered transparently or only a few scars appeared. However, C57BL / 6 mice had dense corneal infiltration and opacity within 1 to 2 days after inoculation of fungi, severe stromal edema and anterior chamber pyosis occurred 3 days later, corneal perforation, prominent eye content, and finally healed with severe corneal scar opacity. (2) after infection with Fusarium and Aspergillus, IFN- 纬 of BALB / c mice increased significantly, the peak value of IFN- 纬 was 2.9 and 2.6 times of normal value, meanwhile, IFN- 纬 of cervical lymph nodes was also significantly increased, but lower than that of local infection. The peak values were 2.0 and 1.5 times of normal values, respectively. After infection with Fusarium and Aspergillus, IFN- 纬 increased slightly and decreased to normal in a short time. The peak value of IFN- 纬 in C57BL / 6 was only 1.3 and 1.1 times of normal, but IFN- 纬 in cervical lymph nodes was significantly increased. (3) BALB / c decreased significantly after infection with Fusarium and Aspergillus, and the low level was only 0.6 and 0.5, while the level of IL-4 in cervical lymph nodes increased gradually, with a peak value of 1.2 and 1.5 times higher than that of normal. After infection with Fusarium and Aspergillus, C57BL / 6 decreased the level of IL-4 in eyeball, the low level of IL-4 was 0.5 and 0.6 of normal, and maintained a low level until 72 hours, but the IL-4 of cervical lymph node increased gradually, the peak value of IL-4 was 1.9 and 1.6 times of normal. Conclusion: (1) fungal keratitis is a Th1-mediated immune inflammatory process, (2) the intensity of local Th1 response is an important factor in prognosis of fungal keratitis, (3) the Th1 response in BALB / c mice is significantly stronger than that in C57BL / 6 mice. (4) Th1 polarization appeared simultaneously in infected area and cervical lymph node during corneal fungal infection, while Th2 polarization appeared separation phenomenon in infected area and cervical lymph node.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R772.21

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