中国寄生虫学与寄生虫病杂志

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调节性T细胞转录因子Foxp3和IL-8在棘球蚴病患者肝组织病灶中的表达

单骄宇1,2,热比亚·努力1,李瑞1,林仁勇2,温浩2,3,李海涛2,3*   

  1. 新疆医科大学 1 基础医学院人体寄生虫学教研室,2 第一附属医院新疆包虫病基础医学重点实验室,3 第一附属医院消化血管外科中心肝胆包虫外科,乌鲁木齐 830011
  • 出版日期:2018-06-30 发布日期:2018-07-02

Expression of the regulatory T cell transcription factor Foxp3 and interleukin-8 in liver tissues of patients with echinococcosis

SHAN Jiao-yu1,2, REBIYA·Nuli1, LI Rui1, LIN Ren-yong2, WEN Hao2,3, LI Hai-tao2,3*   

  1. 1 Department of Human Parasitology, Basic Medicine College, Xinjiang Medical University; 2 Xinjiang Laboratory of Hydatid Functional Medicine, the First Affiliated Hospital; 3 The Hepatobiliary and Hydatid Surgery Department of Digestive Vascular Surgery Center, the First Affiliated Hospital, Urumqi 830011, China
  • Online:2018-06-30 Published:2018-07-02

摘要:

目的 探究调节性T细胞转录因子Foxp3和IL-8在棘球蚴病患者肝组织病灶中的表达情况。 方法 收集2013年10月-2014年10月在新疆医科大学第一附属医院肝胆包虫外科住院的多房棘球蚴病(AE)患者、细粒棘球蚴病(CE)患者及其他非感染性肝病手术患者的肝脏病变组织样品,制备切片,HE染色和免疫组织化学法观察肝脏组织中Foxp3的表达情况。蛋白质印迹(Western blotting)检测Foxp3的蛋白相对表达量。Trizol法提取肝脏组织总RNA,采用实时荧光定量PCR检测Foxp3、IL-8的mRNA相对表达水平。采用Spearman相关分析对AE组患者肝脏组织中的Foxp3与IL-8的相关性进行分析。组间两两比较采用t检验。 结果 共收集16例AE患者、23例CE患者和8例肝血管瘤手术患者的肝脏病变组织样品。HE染色后,镜下可见,AE患者肝脏组织角皮层和生发层均不完整,CE患者肝脏组织有较厚的板层状角皮层结构,肝血管瘤患者肝脏组织完整。免疫组织化学检测结果显示,AE患者肝脏组织中有大量表达Foxp3的阳性细胞,CE患者肝脏组织中有少量阳性细胞,肝血管瘤患者肝脏组织中无阳性细胞。Western blotting结果显示,AE患者、CE患者和肝血管瘤患者肝脏组织的Foxp3蛋白灰度值分别为0.977 ± 0.082、0.728 ± 0.094和0.290 ± 0.084;AE患者、CE患者的蛋白灰度值分别与肝血管瘤患者两两比较,差异均具有统计学意义(P < 0.05);AE患者与CE患者间差异无统计学意义(P > 0.05)。实时荧光定量PCR检测结果显示,AE患者、CE患者和肝血管瘤患者肝脏组织中的Foxp3 mRNA相对表达水平分别为0.013 ± 0.003、0.007 ± 0.002和0.004 ± 0.001;AE患者的Foxp3 mRNA相对表达水平与肝血管瘤患者比较,差异有统计学意义(P < 0.05);CE患者的Foxp3 mRNA相对表达水平分别与AE患者、肝血管瘤患者两两比较,差异无统计学意义(P > 0.05)。AE患者、CE患者和肝血管瘤患者肝脏组织中的IL-8 mRNA相对表达水平分别为0.402 ± 0.068、0.005 ± 0.019和0.002 ± 0.001;三者两两比较,差异均有统计学意义(P < 0.01)。Spearman相关分析结果显示,AE患者肝脏组织中的Foxp3与IL-8在mRNA相对表达水平上呈正相关(r = 0.802,P < 0.01)。  结论 AE患者肝组织中的Foxp3 mRNA和IL-8 mRNA均高于CE患者,提示AE在一定程度上刺激机体分泌大量IL-8,并与Foxp3共同作用病患部位,形成严重的炎症反应。
 

关键词: 细粒棘球蚴病, 多房棘球蚴病, 调节性T细胞, 转录因子Foxp3, 白细胞介素-8

Abstract:

Objective To explore the expression of the regulatory T cell transcription factor Foxp3 and interleukin-8(IL-8) in liver lesions in patients with echinococcosis. Methods Liver tissues were collected from 16 patients with alveolar echinococcosis(AE), 23 patients with cystic echinococcosis(CE) and 8 patients with hepatic hemangioma. HE staining and immunohistochemical(IHC) analysis were performed to examine Foxp3 in liver tissue. Western blotting was performed to detect protein levels of Foxp3. Total RNA was extracted from liver tissue by Trizol, and the mRNA levels of Foxp3 and IL-8 were tested by Real-time PCR. The correlation between Foxp3 and IL-8 was analzyed by Spearman method. Between-group comparisons were made by t test.  Results  H&E staining results showed incomplete horny layer and growth layer of the liver in AE cases, a thick horny layer in CE cases, and complete liver structures in hepatic hemangioma cases. Liver IHC results showed abundant cells positive for Foxp3 in the AE group, in contrast with few in the CE group and absence in  the hepatic hemangioma cases. Western blotting analysis revealed that the gray values of Foxp3 were (0.977 ± 0.082), (0.728 ± 0.094), and (0.290 ± 0.084), respectively in the AE, CE and hepatic hemangioma groups. There were significant differences in the gray value between AE or CE group and the hepatic hemangioma group(both P < 0.05), but there was no difference between AE and CE groups(P > 0.05). Results of real-time PCR showed that the relative levels of Foxp3 mRNA were(0.013 ± 0.003), (0.007 ± 0.002), and (0.004 ± 0.001), respectively in the AE, CE and hepatic hemangioma groups. There was a significant difference in the Foxp3 mRNA level between AE and hepatic hemangioma groups (P < 0.05), but no difference was found for CE versus AE or CE versus hepatic hemangioma(P > 0.05). The IL-8 mRNA levels were(0.402 ± 0.068), (0.005 ± 0.019), and(0.002 ± 0.001), respectively in the AE, CE and hepatic hemangioma groups, with significant differences for all the between-group comparisons(P < 0.01). The Spearman correlation analysis demonstrated that the Foxp3 and IL-8 mRNA levels were positively correlated in the AE group (r = 0.802, P < 0.01).  Conclusion Foxp3 and IL-8 mRNA in the liver tissue are both higher in AE than in CE, suggesting that AE may stimulate secretion of a large amount of IL-8, which, together with Foxp3, causes severe inflammation at the foci.

Key words: Cystic echinococcosis, Alveolar echinococcosis, Regulatory T cell, Transcription factor Foxp3, Interleukin-8