钝挫伤性白内障大鼠模型中晶状体可溶性蛋白和不可溶性蛋白分析及意义探讨

Study on soluble and insoluble lens proteins in contusion cataract of rat model

目的研究大鼠慢性眼拍打伤后晶状体可溶性蛋白与不溶性蛋白的改变，并研究大鼠热休克或喂饲HSP70阻滞剂Quercetin后对钝挫伤性晶状体蛋白的影响。方法实验研究。Sprague—Dawley（SD）大鼠24只（24只眼），完全随机设计分成以下4组：A组（对照组）：6只眼；B组（拍打组）：6只眼，每次以20g钢球20cm高度拍打大鼠右眼100回，每周1次，连续5周；C组（热休克组）：6只眼，温水浴（45℃）使大鼠体温提高至40．5～41．5℃8min，常温下恢复2～3h后拍打眼球同上。每周重复1次，连续5周；D组（Quercetin组）：6只眼，喂饲大鼠Quercetin100mg／kg体重，2～3h后拍打眼球同上。每周重复1次，连续5周。蛋白定量用Bradford法。对晶状体蛋白测定结果，采用完全随机设计的方差分析，并用q检验方法进行组间两两比较。对透明晶状体和混浊晶状体蛋白测定结果采用成组设计定量资料f检验进行分析。结果拍打眼球5周后，热休克组的可溶性晶状体蛋白含量为22．71±1．99，较其他3组明显升高，差异有统计学意义（F=37．82，P〈0．01）；拍打组的不可溶性蛋白含量为2．60±0．48，较其他3组高，差异有统计学意义（F=3．86，P〈0．05）。可溶性晶状体蛋白的十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS—PAGE）分析发现混浊晶状体眼在66000处出现明显的高分子量蛋白带，随白内障程度增加更显著。结论慢性拍打眼球可造成晶状体损伤，不可溶性蛋白含量增加。热休克可增加晶状体可溶性蛋白含量，减少不可溶性蛋白。可溶性蛋白SDS—PAGE分析结果可以看出慢性拍打眼球引起晶状体蛋白质的变化向高分子蛋白移动。

Objective To study the water soluble and insoluble lens proteins in contusion cataract of rat model, and to study the influence of heat shock （ thermotolerance ） or quereetin （ heat shock protein inhibitor） on the lens proteins. Methods Sprague-Dawley （SD） rats were randomized into four groups （6 in each group） ： （1）A group,control group. （2）B group, contusion group. （3）C group, heat shock group. （4） D group, quereetin group. In contusion group, contusion of eyes was induced by dropping steel ball against the eyeball （20 g ball dropped from 20 cm height for 100 times each week for 5 weeks）. In heat shock group, heat shock was induced by raising the body temperature to 40. 5-41.5℃ for 8 rain, 2-3 h before the eye contusion （as previously described ）. Heat shock and contusion was given each week for 5 weeks. In quercetin group, rats were fed with quereetin （ 100 mg/kg weight） 2-3 h before the contusion. Quercetin and contusion was given each week for 5 weeks. Amount of proteins was measured by using Bradford methods. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis （SDS-PAGE） was used for the separation and analysis of lens proteins. Results Five weeks after contusion, the mean water soluble protein of heat shock group was 22. 71 ± 1.99, increased significantly than that of the other three groups. The mean insoluble protein of contusion group was 2. 60 ± 0. 48, increased than that of the other three groups. From the analysis of SDS-PAGE, we found that there was a high molecular band near 66 KD in opaque lens and the band became more prominent with the growing opacification of the lens. Conclusions In contusion cataract, water insoluble lens protein is increased and water soluble lens protein is decreased. SDS-PAGE showed that lens injury resulted in lens proteins moving toward high molecular band.