葡萄膜炎糖皮质激素治疗分析及胡黄连苷Ⅱ在内毒素诱导大鼠葡萄膜炎模型中的抗炎作用

发布时间：2018-06-22 22:48

本文选题：胡黄连苷Ⅱ + 脂多糖（LPS）　；参考：《大连医科大学》2013年硕士论文

【摘要】：目的:葡萄膜炎是常见眼病，也是主要致盲眼病之一，其中以前葡萄膜炎为最多，其次是全葡萄膜炎。在发达国家，5-20%的患者由于葡萄膜炎致盲，而在发展中国家这个比例可能要超过25%，因为发展中国家影响眼部的传染病机会较多，如弓形虫病和结核。但是目前葡萄膜炎的治疗方法主要是使用激素，激素治疗副作用大，眼科方面可能导致激素源性角膜炎、青光眼、白内障；全身性副作用包括诱发高血压、糖尿病、骨质疏松、消化道出血、癫痫发作等，，这些都导致激素在应用过程中具有相当的局限性。所以我们寻找一种可以有效治疗葡萄膜炎并且毒副作用小的药物，中国传统医学在此方面为我们提供了新的思路和方法。胡黄连作为一种传统保肝中药，具有抗氧化、抗炎症反应、免疫调节等多种药理学作用，胡黄连苷Ⅱ是从胡黄连的干根茎中提取的有效成分，分子量小，提纯纯度高，已经发现其有抗炎症作用。内毒素诱导的葡萄膜炎(endotoxin induced uveitis，EIU)是用内毒素中的脂多糖通过与脂多糖结合蛋白结合，提高血管通透性，通过激活补体系统和刺激血小板聚合因子释放，从而引起组织炎症。实验室常用的内毒素为伤寒沙门氏菌或大肠杆菌内毒素。实验动物一般为大鼠或小鼠，大鼠一般采用Lewis或SD鼠系。EIU模型是应用最广泛的葡萄膜炎动物模型，本模型是通过在实验动物脚掌皮下、腹腔或玻璃体腔注射细菌脂多糖，建立天然免疫机制诱导，释放炎症因子，也可有细胞介导的免疫反应参与，EIU引起的葡萄膜炎持续时间短，表现为急性起病前葡萄膜炎。4小时出现炎症，24小时达到高峰，之后炎症反应减轻。可能机制为毒素的脂多糖进入动物血液后形成脂多糖结合蛋白，然后再与Toll样受体结合随后传递到核转录因子NF—κB进而启动下游基冈的表达，最终引起组织炎症反应。我们即采用EIU葡萄膜炎模型来观察胡黄连苷Ⅱ的抗炎作用。方法:首先我们回顾分析了2012年03月至2013年09月于沈阳市第四人民医院眼科就诊并住院治疗的葡萄膜炎患者195例235眼，患者均予以局部或全身糖皮质激素治疗，所有患者均随访6个月以上，观察患者治疗前后视力变化、炎症反应变化及局部全身出现的副作用。随后进行基础实验，研究胡黄连苷Ⅱ的抗炎作用。36只体重200~250g成年雄性SPF级SD大鼠随机分成四组，每组9只，分别用20mg/kg和10mg/kg的胡黄连苷Ⅱ鼠尾静脉注射两次，其中3组第二次给药后半小时予单侧足底注射100μl浓度为100μg/100μl的LPS（Lipopolysaccharides），给药后24小时前节裂隙灯照相，观察炎症变化并且评分，30gauge针收集（15-20μl）房水，细胞计数器计数，其余房水BCA试剂盒测房水蛋白浓度，取出眼球去核，OCT包埋冰冻切片，HE染色。结果:病例回顾性分析中我们发现，195例葡萄膜炎患者的235眼中，男性70例80眼，女性125例155只眼，年龄9~78（平均37±23）岁，其中前葡萄膜炎109例（46.38%）、中间葡萄膜炎12例（5.12%）、后葡萄膜炎46例（19.57%）、全葡萄膜炎68例（28.93%）。治疗后视力：眼前指数1眼（0.43%）、0.02~0.114眼（5.96%）、0.1~0.5101眼（42.98%）、0.5~0.8122眼（51.91%）、0.8~1.0及1.0以上7眼（2.98%）。血糖超过正常值41例（17.45%），停药后血糖逐渐恢复正常；高血压62例（26.38%），通过治疗可以控制在正常血压范围内；便潜血3例（1.28%），给与胃黏膜保护剂、糖皮质激素减量停药；癫痫发作1例（0.43%），经治疗后症状缓解；白细胞下降1例（0.43%），经治疗后痊愈；激素性青光眼3眼（1.28%），经糖皮质激素减量或停用局部激素眼药水，并联合降眼压治疗把眼压控制在范围内；激素性白内障9眼（3.83%），待葡萄膜炎稳定后行白内障超声乳化术，术后视力恢复良好。基础实验中，细胞计数结果显示20mg/kg组房水细胞少于LPS单纯造模组（P0.05)）；10mg/kg组房水中细胞与LPS单纯造模组比较无统计学差异（P0.05）。蛋白浓度测定结果20mg/kg组房水细胞少于LPS单纯造模组（P0.05)）；10mg/kg组房水中蛋白浓度与LPS单纯造模组比较无统计学差异（P0.05）。结论:应用激素治疗各种非感染性葡萄膜炎具有一定效果，大多数患者病情好转，但是糖皮质激素治疗期间均出现不同类型不同程度的副作用。胡黄连苷Ⅱ对于内毒素诱导的大鼠葡萄膜模型中具有抗炎作用。
[Abstract]:Objective: uveitis is one of the common ophthalmopathy and also one of the main blinding ophthalmopathy. Former uveitis is the most, followed by full uveitis. In developed countries, 5-20% patients are blinded by uveitis, and the proportion in developing countries may exceed 25%, because there are more opportunities for infectious diseases in the developing countries, such as arched. But at present, the main treatment for uveitis is the use of hormones, hormone treatment has a large side effect, and the ophthalmology may lead to hormone derived keratitis, glaucoma, and cataract. Systemic side effects include induced hypertension, diabetes, osteoporosis, gastrointestinal bleeding, seizures and so on, which all lead to the use of hormones. So we look for a drug that can effectively treat uveitis with small side effects. Chinese traditional medicine provides us with new ideas and methods in this field. Hu Huanglian, as a traditional Chinese medicine, has many pharmacological effects, such as antioxidation, anti-inflammatory reaction and immunoregulation. Lipopolysin II is an effective component extracted from rhizome of Rhizoma Coptidis, which has small molecular weight and high purity. It has been found that it has anti inflammatory effect. Endotoxin induced uveitis (EIU), which is induced by endotoxin, is a combination of lipopolysaccharide in endotoxin by binding with lipopolysaccharide binding protein to improve the permeability of blood vessels by activating a supplementary system. The release of platelet aggregation factors and stimulation of platelet aggregation causes inflammation of the tissues. The commonly used endotoxin in the laboratory is Salmonella typhi or Escherichia Coli Endotoxin. Experimental animals are generally rats or mice. Rats generally use the Lewis or SD mouse.EIU model to be the most widely used animal model of Vitis. This model is used in experimental animals. Subcutaneous, intraperitoneal or intravitreal injection of bacterial lipopolysaccharide to establish a natural immune mechanism, release inflammatory factors, and be involved in cell mediated immune responses. EIU induced uveitis has a short duration of uveitis, showing an acute onset of uveitis for.4 hours of inflammation, reaching a peak at 24 hours, and then alleviating the inflammatory response. The mechanism is that the lipopolysaccharide into the animal blood forms the lipopolysaccharide binding protein and then combines with the Toll like receptor and then transferred to the nuclear factor NF - kappa B and then initiates the expression of the downstream Keegan and eventually causes the inflammatory response of the tissue. We use the EIU uveitis model to observe the anti-inflammatory effect of the glucoside II.
Methods: first, we reviewed and analyzed 195 cases of uveitis in Fourth People's Hospital, Fourth People's Hospital of Shenyang from 03 months to 09 months in 2013. The patients were treated with local or systemic glucocorticoid. All patients were followed up for more than 6 months. The changes of visual acuity and the changes of inflammation were observed before and after treatment. Side effects of the local body.
Then the basic experiment was carried out to study the anti-inflammatory effect of hanthoside II,.36 only weight 200~250g adult male SPF SD rats were randomly divided into four groups, 9 rats in each group were injected two times with the tail vein of hanthoside 20mg/kg and 10mg/kg respectively, of which 3 groups were given second times after the injection of the sole plantar 100 L concentration of 100 u g/100 Mu L LPS ( Lipopolysaccharides), a slit lamp was taken 24 hours after the administration to observe the changes of inflammation and score. 30gauge needle was used to collect (15-20 L) water, cell counter count, and the rest aqueous BCA kit was used to measure the concentration of aqueous protein, remove the nucleus of the eyeball, OCT embedded frozen section, and HE staining.
Results: in the retrospective analysis, we found that 195 cases of uveitis were found in 235 eyes, male 70 cases, 80 eyes, 125 women in 155 eyes, age 9~78 (average 37 + 23) years, including 109 cases of anterior uveitis (46.38%), 12 cases of middle uveitis (5.12%), posterior Vitis 46 cases (19.57%), full uveitis. The index was 1 eyes (0.43%), 0.02~0.114 eye (5.96%), 0.1~0.5101 eye (42.98%), 0.5~0.8122 eye (51.91%), 0.8~1.0 and 7 eyes (2.98%). Blood sugar exceeded normal value in 41 cases (17.45%), blood sugar gradually returned to normal after withdrawal; hypertension 62 (26.38%) could be controlled within the normal blood pressure range; then the occult blood 3 (1.28%) was given to the gastric mucosa. 1 cases (0.43%) of epileptic seizures (0.43%) were relieved after treatment; the symptoms were relieved after treatment; 1 cases (0.43%) of leukocyte decreased (0.43%), 3 eyes (1.28%), glucocorticoid reduction or discontinuation of local hormone ophthalmic water, and intraocular pressure controlled by combined ophthalmic pressure reduction; 9 eyes of hormonal cataract (3) .83%) phacoemulsification is performed after uveitis is stable.
In the basic experiment, the cell count results showed that the water cells in the 20mg/kg group were less than the LPS model group (P0.05), and there was no statistical difference between the 10mg/kg group and the LPS simple model group (P0.05). The determination of protein concentration in the 20mg/kg group was less than that of the LPS simple model group (P0.05), and the protein concentration in the 10mg/kg group and LPS were simple. There was no statistical difference in the model (P0.05).
Conclusion: the use of hormones in the treatment of various non infectious uveitis has a certain effect, most patients have a better condition, but there are different types of side effects during the treatment of glucocorticoid.
【学位授予单位】：大连医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R773.9

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