远视离焦对虚证豚鼠巩膜重塑的影响及其机制研究

Axial myopia accounted for the 95% myopia in young patients. Sclera remodeling regulated by various factors is usually considered as the main mechanism of myopic pathogenesis. According to the clinical observation on TCM, the incidence of myopia is closely related to people's physical. People with syndrome of deficiency are more likely suffering from myopia. In view of this, we speculated that the sclera remodeling vary between people with deficiency and with normal physical. At present, the studies are mostly limited to clinical observation of traditional Chinese medicine on myopia. The research analyzing the nature of myopia which is based on the theory of syndrome of TCM has not yet been reported. It is very important to establish a new method for studying myopia in TCM.In this study ,the model will be obtained by hyperopic defocus combining with intraperitoneal injectiong of cyclophosphamide or propylthiouracil. The expression of TGF-β, bFGF, MMP-2 and TIMP-2 in retina and sclera will be examined with methods of histology, immunohistochemistry and real-time PCR to analyze the changing discipline of different model's refraction and scleral remodeling at the level of tissue, cells, protein and molecule. The difference of refraction, scleral remodeling and relative factors' expression of guinea pigs with different physique according to the performance of treadmill test will be analyzed to elucidate the influence and mechanism of deficiency and physical factors on myopia. The results of the research will provide theoretical basis for developing new methods and theory of TCM in preventing and treating myopia.

近视95%以上为轴性近视，巩膜重塑是轴性近视发生发展的重要病理基础。祖国医学认为，过用目力、劳瞻竭视为近视发病主要诱因，肝肾气血不足之虚证为主要病机，虚证与正常体质者是否在近视形成过程中存在差异？从中医证候学角度分析和认识近视发生发展的基础研究尚属空白，建立病证结合的证候学研究近视的新方法至关重要。针对巩膜重塑在近视发病中的关键作用，本研究采用远视离焦法结合环磷酰胺与丙基硫氧嘧啶腹腔注射制作脾气虚和肾阳虚豚鼠近视模型，利用组织学、免疫组化、实时定量PCR等分子生物学技术检测视网膜和巩膜基质中TGF-β、bFGF、MMP-2、TIMP-2等相关因子表达的变化，从组织、细胞及分子水平分析正常与虚证近视豚鼠眼球屈光度和巩膜重塑的变化规律，还将根据豚鼠跑台实验结果评价不同体质豚鼠屈光变化、巩膜重塑及相关因子表达的差异，探讨虚证和体质因素对近视发生发展的作用及机制，为中医防治青少年近视提供理论依据。

近视是危害我国青少年视觉健康的主要疾病，已成为广受关注的社会与医学问题。近视95%以上为轴性近视，巩膜重塑是轴性近视发生发展的重要病理基础。祖国医学认为，“阳不足, 阴有余”是近视发病主要病机，但从中医证候学角度分析虚证在近视发生发展中作用的基础研究尚属空白。本研究通过动物实验分析负透镜诱导对肾阳虚豚鼠眼球屈光度、眼轴长度及相关生物学参数的影响，研究肾阳虚近视豚鼠后极部巩膜基质金属蛋白酶-2（MMP-2）及其抑制剂（TIMP-2）和后极部视网膜AREG的表达变化，探讨肾阳虚与近视发生发展的关系，为中医防治青少年近视提供理论依据。.1. 通过腹腔注射低、中、高剂量的氢化可的松注射液，探讨制作肾阳虚豚鼠动物模型的最佳药物浓度。结果发现，中剂量（10 mg/kg）的氢化可的松注射液腹腔注射豚鼠，其体重明显下降，甲状腺及性腺功能明显受到抑制，且死亡率较低，呈典型肾阳虚表现，成功制作肾阳虚豚鼠近视模型，为下一步研究肾阳虚证与近视发病机制的相关实验研究奠定了基础。.2. 分析肾阳虚近视豚鼠眼球屈光度、眼轴长度及相关生物学参数的变化后发现，肾阳虚豚鼠较正常豚鼠更容易发生近视，表现为近视屈光度增加，眼轴延长，后极部巩膜干重及其厚度降低。但近视豚鼠运动能力与屈光参数无关。研究证实了肾阳虚对巩膜重塑和近视发展的显著影响，为进一步的机制研究提供了证据。.3. 分子生物学研究发现，肾阳虚近视豚鼠后极部巩膜MMP-2mRNA和蛋白水平的表达明显增加，TIMP-2 mRNA和蛋白水平的表达明显降低，MMP-2和TIMP-2之间的平衡严重失调，这与全身激素水平失调有关，可能是导致肾阳虚豚鼠较正常豚鼠近视程度严重的重要机制。.4. 采用免疫荧光法检测并分析肾阳虚近视豚鼠后极部视网膜表皮生长因子受体的配体双调蛋白（AREG）的表达，发现肾阳虚近视豚鼠后极部视网膜双调蛋白的表达明显增加，证实AREG可能是参与肾阳虚近视发病的重要作用因子。