11β-HSD1及皮质类固醇受体在兔糖皮质激素性高眼压模型睫状体组织中的表达

Expressions of 11β-hydroxysteroid dehydrogenase type 1 and steroids receptors in ciliary body with glucocorticoid-induced ocular hypertension rabbit model

背景糖皮质激素的长期应用可能诱发敏感个体的眼压升高，研究证实11β羟类固醇脱氢酶1（11β—HSD1）及糖皮质激素受体（GR）、盐皮质激素受体（MR）可能影响房水的产生，这种机制是否与糖皮质激素性高眼压的发生有关目前国内外研究甚少。目的检测11β-HSDl及GR、MR在兔糖皮质激素性高眼压模型睫状体组织中的表达变化，探讨其在糖皮质激素性高眼压发病中的作用。方法采用随机数字表法将12～16周龄新西兰白兔13只随机分为对照组5只和实验组8只。实验组兔隔日在固定时间及固定地点于左眼球结膜下注射质量分数0．5％地塞米松磷酸钠注射液5mg（1ml），注射次日用质量分数0．5％地塞米松滴眼液点眼3次。每次注射后用氧氟沙星滴眼液点眼3次以预防感染，持续2个月；对照组兔以同法注射等容积无菌生理盐水，实验组兔眼压升高至18mmHg（1mmHg=0．133kPa）以上并能持续1周者为造模成功。实验组注射5周后以过量麻醉法处死各组实验兔，分离模型眼睫状体组织，采用免疫组织化学法检测和定位11β-HSD1蛋白在兔模型眼睫状体中的表达；采用逆转录-PCR（RT—PCR）法定量检测11β-HSDlmRNA、GRmRNA及MRmRNA在实验组兔模型眼睫状体组织中的表达，并与对照组兔眼进行比较。结果实验组采用地塞米松局部注射联合点眼后前2周眼压无明显升高，第1周与第2周问眼压差异无统计学意义（q=0．469，P〉0．05）；第3、4、5周眼压逐渐升高，与对照组比较差异均有统计学意义（q=10．535、20．353、28．681，P〈0．01），第5周时达（18．87±0．77）mmHg。免疫组织化学法检测显示，11β—HSDl蛋白在睫状体中呈阳性表达，位于非色素上皮细胞，但模型眼11β—HSDl的阳性染色强度明显弱于对照组。RT—PCR法检测显示，实验：组兔模型眼睫状体中MRmRNA、GRmRNA及11β-HSDlmRNA的相对表达量（以GADPH为内参）分别为2．22±0．78、0．64±0．11、0．47±0．16，而对照组分别为0．94±0．27、1．88±0．74、2．68±1．28，差异均有统计学意义（t=6．070，P=0．004；t=5．170，P=0．007；t=5．540，P=0．005）。结论外源性糖皮质激素性高眼压的发病可能与睫状体中MR表达的上调及11β—HSDl和GR表达的下调有关。

Background Long-term administration of glucocorticoid drugs induces ocular hypertension in susceptible individuals probably. It has been verified that 11β-hydroxysteroid dchydrogenase type 1 （11β-HSD1）, glucocortieoid receptor （GR） and mineralocorticoid receptor （MR） can affect the generating of aqueous humor,but how they play the role in glucocorticoid-induced ocular hypertension is unclear. Objective This study was to investigate the relationship of expressions of 11β-HSD1 and steroids receptors in ciliary body and steroid-induced ocular hypertension. Methods Thirteen 12-16 week-old New Zealand albino rabbits were randomized to control group （5 rabbits ） and experimental group （8 rabbits ）. Steroid-induced glaucoma models were induced by administration of subconjunctival injection of 5 mg dexamethasone solution（ 1 ml） and 0.5% dexamethasone eye drops on alternate days in the left eyes for consecutive two months in the experimental group, and the equal volume of sterile normal saline solution was used in the same way in the control group. The successful criteria of model eyes was defined as rising of intraocular pressure （IOP） to ≥18 mmHg for over one week. Then, the animals were sacrificed by excessive anesthesia and the ciliary tissues were isolated for the assay of expressions of 11β-HSD1 protein by immunochemistry,and the expressions of 11β-HSD1 mRNA, GR mRNA and MR mRNA in ciliary body were semi-quantitatively detected by reverse transcription-PCR （RT-PCR）. The experimental results were compared between the two groups. Results The lOP was normal in the first two weeks after administration of drugs, and no significant difference was found in IOP between the first week and the second week in the experimental group （q = 0. 469 ,P 〉0.05）. From 3 through 5 weeks after injection, the lOP was gradually elevated, with the highest value of （ 18.87±0. 77） mmHg in the fifth week. Significant differences were seen between the two groups at mentioned-above time points （ q = 10. 535,20. 353,28. 681, all at P 〈 0. 01 ）. 11β-HSD1 protein was positively expressed in non- pigmented epithelial cells of ciliary tissue of rabbits in both groups, however, the expression intensity was weaker in the experimental group compared with the control group. The relative expressional values of MR mRNA, GR mRNA and 11β-HSD1 mRNA in the ciliary tissue were 2.22±0. 78,0.64±0. 11 and 0. 47±0. 16 in the experimental group, and those in the control group were 0. 94±0. 27 ,1. 85±0. 74 and 2.68±1.28 ,with significant differences between the two groups （ t = 6. 070, P = 0. 004 ; t = 5. 170, P = 0. 007 ; t = 5. 540, P = 0. 005 ）. Conclusions Corticosteroidinduced glaucoma probably is associated with the up-regulation of MR level and down-regulations of GR and 11β- HSD1 in ciliary body.