眼部应用国产曲安奈德混悬液安全性的实验研究

Characteristic of ocular safety profile of triamcinolone made in China

目的 评价国产曲安奈德及醋酸曲安奈德混悬液眼部应用的安全性.方法 实验研究.通过体外将曲安奈德和醋酸曲安奈德混悬液的上清液稀释成不同比例后,用MTT比色法评价其上清液中的防腐剂和赋形剂对3种人眼细胞株的细胞毒性;选取28只健康豚鼠,根据计算机（SAS软件）生成的随机数字均分为4组,分别在豚鼠右眼的玻璃体腔注射曲安奈德或醋酸曲安奈德的上清液18μl和混悬液6μl.给药前后通过裂隙灯、间接检眼镜、眼压计、眼电生理检查及组织学石蜡切片来评价其玻璃体腔应用的安全性.体外细胞毒性研究采用Dunnett-t检验,眼电生理数据采用配对t检验,眼压数据比较采用重复测量统计方法广义估计方程.结果 未经稀释的曲安奈德和醋酸曲安奈德上清液对人眼后巩膜成纤维细胞的MTT值分别为0.046±0.036和0.044±0.05,与其各自的标准品上清液对照组（0.367±0.106和0.413±0.128）和BSS空白对照组（0.368±0.106和0.441±0.137）相比,差异均有统计学意义（P值均＜0.01）.醋酸曲安奈德的上清液经1∶15或更高倍数的稀释后对人RPE细胞无细胞毒性（P＞0.05）,但1∶15稀释的上清液对人Müller细胞有细胞毒性（MTT值为0.366±0.062,BSS对照组和上清液对照组的MTT值分别为0.417±0.042和0.399±0.045）（P=0.03）;曲安奈德的上清液经1∶15或更高倍数的稀释对RPE及Müller细胞均未显示细胞毒性.豚鼠玻璃体腔注射这两种制剂的混悬液6μl（含药物0.25 mg,相当于人眼玻璃体腔注射0.1 ml含4 mg曲安奈德的混悬液）或上清液18μl（相当于兔眼玻璃体腔注射0.1 ml含4 mg曲安奈德混悬液中含有的防腐剂浓度）均未见眼内毒性.结论 豚鼠玻璃体腔注射6μl曲安奈德和醋酸曲安奈德混悬液均是安全的,该剂量相当于人眼玻璃体腔注射0.1 ml（含4 mg）曲安奈德混悬液.

Objective To characterize the safety profile of triamcinolone acetonide made in China （Transton） and triamcinolone acetonide acetate （Tongyong） for their ocular application.Methods Experimental study.In vitro cell viability assay was performed on 3 types of human ocular cells to evaluate the cytotoxicity of the simulated vitreal concentrations （from a 1 ∶ 15 dilution as if injected into 1.5 ml of rabbit vitreous to 1∶50 dilution as if injected into 5 ml of human vitreous） of Transton and Tongyong using MTT method.In vivo 28 guinea pigs,randomly divided into four groups,were used for evaluating either intravitreal 6 μl of the two types of triamcinolone suspension or 18 μ1 of their supernatant.Following the injections,the eyes were monitored by biomicroscopy,ophthalmoscopy,tonometry,electroretinography,and histology.The Dunnett＇s test was used to analyze in vitro cell viability.Paired sample t-test and Generalized Estimating Equations （GEE） were respectively used to compare electroretinography data and intraocular pressure between experimental eyes and the control eyes.Results The undiluted supernatant of Transton and Tongyong was toxic to human scleral fibroblasts when compared with their control groups（MTT values =0.046 ±0.036 and 0.044 ±0.05 versus 0.367 ± 0.106 and 0.413 ± 0.128） （P 〈 0.01） or with the BSS group （0.368 ±0.106 and 0.441 ±0.137） （P 〈0.01）.1∶15 or greater dilution of supernatant of Transton and Tongyong did not show cytotoxicity on cultured human retinal pigment epithelium cells or Müller cell （P 〉 0.05）,but 1∶15 dilution of Tongyong supematant showed cytotoxicity on the Müller cells （MTT value =0.366 ± 0.062 versus 0.417 ± 0.042 for BSS） （P =0.03）.In vivo,neither intravitreal 6 μl （0.25 mg,equivalent to 4 mg in 0.1 ml for human eyes） of the suspension nor 18 μl（equivalent resultant preservative concentration to intravitreal 4 mg in 0.1 ml for rabbit eyes） of the supernatant of Transton and Tongyong showed ocular toxicity.Conclusion The equivalent dose （6 μl） to 4 mg in 0.1 ml intravitreal suspension for human eye or equivalent resultant preservative concentration to 0.1 ml intravitreal suspension for rabbit eye is found safe in guinea pig eyes.