眼眶成纤维细胞局部分泌的IGF-1是甲状腺相关眼病的致病因素

发布时间：2018-08-10 21:23

【摘要】：目的：观察胰岛素样生长因子1受体(IGF-1R)在甲状腺相关眼病(TAO)患者眼眶成纤维细胞(OF)的表达及在TAO发病机制中的作用。研究TAO患者临床病情活动度与眼眶局部分泌和全身循环中IGF-1含量的相关性,及IGF-1在TAO发病机制中的作用。方法：取TAO患者及正常人球后结缔组织,体外培养OF。通过共聚焦显微镜观察,流式细胞仪检测IGF-1R的表达。此外,留取18例TAO患者及16例年龄性别匹配的正常人外周血,并将其中5例TAO患者和5例正常人的眼眶结缔组织培养成纤维细胞。ELISA法测定所有TAO患者和正常人外周血总IGF-1和游离IGF-1的含量,并测定成纤维细胞培养基中的IGF-1含量,通过MTT生长实验观察生长抑素奥曲肽对其增殖的作用。定量PCR法测定成纤维细胞IGF-1的mRNA水平。结果：经流式细胞仪测定,TAO患者的OF表达IGF-1R为72.6%±10.4,正常组的IGF-1R为27.8%±10.7,两组差异具有显著的统计学差异(P0.05)。共聚焦显微镜观察及定量分析发现TAO患者OF表达IGF-1R较正常对照增多(P0.05),且IGF-1R主要位于OF的细胞核周围及细胞质内。在观察的72小时内,TAO患者和正常人体外培养的OF均能分泌IGF-1,且TAO患者的成纤维细胞分泌IGF-1比正常人多。IGF-1的分泌作用和成纤维细胞的生长作用均能被奥曲肽抑制。定量PCR结果显示TAO患者成纤维细胞IGF-1的mRNA水平较正常人高出两倍,该差异具有显著统计学意义(P0.05),且该mRNA水平的上调能被奥曲肽抑制。TAO患者血浆中总的及游离的IGF-1与正常人无明显差异,与临床活动度无关,但TAO患者成纤维细胞分泌的IGF-1与临床活动度成正相关(r=0.97,P=0.017)。结论：通过流式细胞术和共聚焦显微镜观察发现,与正常人相比,TAO患者的OF高度表IGF-1R。眼眶局部分泌的IGF-1与TAO患者的临床病情活动度成正相关,而非系统循环中的IGF-1。后者在TAO患者和正常人之间无显著差异。因此,局部自分泌或旁分泌的IGF-1参与了TAO的病理机制。奥曲肽对TAO患者成纤维细胞生长和IGF-1分泌的抑制作用解释了生长抑素类药物治疗TAO的机制。
[Abstract]:Aim: to investigate the expression of insulin-like growth factor 1 receptor (IGF-1R) in orbital fibroblasts from patients with thyroid associated ophthalmopathy (TAO) and its role in the pathogenesis of TAO. To study the relationship between clinical activity of TAO patients and orbital regional secretion and IGF-1 content in systemic circulation and the role of IGF-1 in the pathogenesis of TAO. Methods: Retrobulbar connective tissue was taken from patients with TAO and normal subjects, and cultured in vitro. The expression of IGF-1R was detected by confocal microscope and flow cytometry. In addition, peripheral blood samples were collected from 18 TAO patients and 16 age-sex matched normal subjects. The contents of total IGF-1 and free IGF-1 in peripheral blood of all TAO patients and normal subjects were determined by Elisa, and the content of IGF-1 in fibroblast culture medium was determined by Elisa in 5 cases of TAO and 5 cases of normal controls. The effects of somatostatin octreotide on the proliferation of somatostatin were observed by MTT growth assay. The mRNA level of IGF-1 in fibroblasts was measured by quantitative PCR method. Results: the expression of IGF-1R was 72.6% 卤10.4 in the patients with Tao and 27.8% 卤10.7 in the normal group by flow cytometry. There was significant difference between the two groups (P0.05). Confocal microscopy and quantitative analysis showed that the expression of IGF-1R in TAO patients was higher than that in normal controls (P0.05), and IGF-1R was mainly located in the nucleus and cytoplasm of. IGF-1 could be secreted by the fibroblasts of TAO patients and normal subjects within 72 hours. The secretion of IGF-1 and the growth of fibroblasts in TAO patients were inhibited by octreotide. The results of quantitative PCR showed that the mRNA level of IGF-1 in fibroblasts of TAO patients was twice as high as that of normal controls. The difference was statistically significant (P0.05), and the up-regulation of the mRNA level could be inhibited by octreotide. The total and free IGF-1 in plasma of patients with octreotide had no significant difference with that of normal subjects, and had no relationship with clinical activity. However, the IGF-1 secreted by fibroblasts in TAO patients was positively correlated with clinical activity (r = 0.97, P = 0.017). Conclusion: by flow cytometry and confocal microscopy, the IGF-1R of the patients with OTAO was found to be higher than that of the normal controls. IGF-1 secreted locally in orbit was positively correlated with the clinical activity of TAO patients, but not IGF-1 in systemic circulation. There was no significant difference between TAO patients and normal subjects. Therefore, local autocrine or paracrine IGF-1 is involved in the pathological mechanism of TAO. The inhibitory effect of octreotide on fibroblast growth and IGF-1 secretion in patients with TAO explains the mechanism of somatostatin in the treatment of TAO.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R771.3

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