从微管蛋白 α-tubulin探讨激素性白内障的发病机制

Glucocorticoid-induced cataract (GIC) is a common ocular complication secondary to long-term use of glucocorticoid and impairs visual acuity in the early stage. Histological analysis shows that the cause of lens opacity of GIC originates from abnormal lens cell proliferation and differentiation, resulting in accumulation of undifferentiated cells between the posterior cortex and capsular. Alpha-tubulin is an important cytoskeletal protein and plays an important role in lens cell differentiation. The previous study in our.group showed that Dexamethsone (DEX, a typical glucocorticoid) could enhance cell proliferation, but inhibit cell differentiation by disturbing the redistribution of α-tubulin. Therefore, for further study, the current study aims to explore the effect of DEX on α-tubulin during lens cell differentiation. With the technologies of immuno-TEM (transmitted electronic mricroscopy), image analysis, western blot, RT-PCR and etc, the morphology, redistribution and expression of α-tubulin in differentiated lens cells will be studied. In addition, we will also analyze the status of glucocorticoid receptor and pERK1/2 signal pathway, for the purpose of understanding the potential mechanism of the effect of DEX on α-tubulin and thereafter on cell differentiation. Therefore, the current study is essential in the analysis of the etiology of GIC in cellular and molecular levels.

激素性白内障是长期使用糖皮质激素引起的常见眼部并发症，发病初期即可严重损害视力。病理研究提示：晶状体上皮细胞异常增殖、分化并聚集于后囊膜下，是引起晶状体混浊的主要原因。α-tubulin是一种主要的细胞骨架蛋白，在上皮细胞分化过程中发挥重要作用。课题组前期研究发现：糖皮质激素地塞米松（DEX）可通过干扰α-tubulin的重构而抑制细胞分化。因此，本项目拟在此基础上，从晶状体细胞和器官水平进一步探讨DEX对分化细胞中α-tubulin的影响，借助于免疫电镜、图像分析、免疫印记及RT-PCR等技术，检测α-tubulin在细胞内的形态分布、聚合状态及表达量的变化，并通过检测糖皮质激素受体激活状态、pERK1/2活化状态等深入探讨DEX引起α-tubulin改变的分子机制，以明确α-tubulin与DEX引起细胞分化紊乱之间的关系，为在细胞和分子水平研究激素性白内障的发病机制提出新思路。

激素性白内障是由于长期局部或全身使用糖皮质激素引起的常见眼部并发症，确切发病机制不明。在本项目中，课题组通过建立大鼠晶状体上皮细胞正常分化和激素诱导的大鼠晶状体后囊膜下浑浊等体系，探索地塞米松引起激素性白内障的建模方法和发病机制；借助microarray等技术，由细胞和分子水平阐述激素性白内障与细胞骨架蛋白、经典MAPK信号通路之间的潜在联系和机制。本项目的研究内容主要包括以下几部分：1）明确DEX对大鼠晶状体细胞及晶状体内α-tubulin分布和表达的影响，以及这种影响与细胞异常分化和激素性白内障发生的关系；2）明确 DEX 抑制晶状体细胞正常分化的分子机制；3）明确DEX引起离体晶状体后囊下浑浊与骨架蛋白表达及关键信号通路的联系。课题组基本完成上述目标。本项目结果主要发现：DEX干扰大鼠晶状体上皮细胞向纤维细胞的正常分化，这种干扰作用主要体现在降低tubulin的表达，干扰骨架蛋白和晶体蛋白在细胞内分布。分子水平检测发现骨架蛋白actin-1/2，α/β3/γ-tubulin等，信号通路MAPK/ERK1/2、Wnt等参与了DEX引起的晶状体囊膜下浑浊。本项目的创新之处在于从疾病发生的直接原因出发，在结构相对简单的晶状体中模拟病变发生的形态和结构，并以此为基础，探索疾病发生的分子机制，为激素性白内障的药物治疗开辟新的思路。