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内质网应激对肝泡型包虫病部分炎症介质的影响
DOI: 10.3969/j.issn.1001-5256.2021.01.026
利益冲突声明：本研究不存在研究者、伦理委员会成员、受试者监护人以及与公开研究成果有关的利益冲突，特此声明。
作者贡献声明：李姚、王靖超、袁加琪、温浩、周瀛、樊海宁、王志鑫负责课题设计、资料分析; 李姚、王靖超负责收集数据与撰写论文; 李姚、王靖超、王志鑫参与修改论文; 温浩、周瀛、樊海宁、王志鑫责拟定写作思路，指导撰写论文并最后定稿。
Influence of endoplasmic reticulum stress on the secretion of some inflammatory mediators in hepatic alveolar echinococcosis
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摘要:
目的 探究内质网应激(ERS)在肝泡型包虫病(HAE)病情发展过程中对部分炎症介质的影响，初步探讨其临床意义。 方法 选取2018年6月—2019年9月因HAE在青海大学附属医院接受肝脏部分切除术的患者15例，切除HAE病灶边缘带作为AE组，同期肝脏正常组织15例作为对照组。通过蛋白质印迹法、实时荧光定量PCR检测AE组和对照组中的蛋白激酶R样激酶(PERK)、CCAAT/增强子结合蛋白同源蛋白(CHOP)、含半胱氨酸的天冬氨酸蛋白水解酶12(caspase-12)和葡萄糖调节蛋白78(GRP-78)表达水平以及IL-1β、IL-6和TNF的mRNA表达水平。两组间不满足正态性的计量资料比较采用Mann-Whitney U检验; 满足正态性的计量资料两组间分析采用t检验; 双变量相关性分析采用Pearson相关分析法。 结果 AE组中PERK、CHOP、caspase-12、GRP-78的蛋白表达量(U值分别为4.165、3.461、2.577、3.344，P值均＜0.001)和mRNA表达水平(t值分别为34.003、4.461、53.573、55.224，P值均＜0.001)均明显高于对照组，差异均有统计学意义。AE组中IL-1β、IL-6和TNF的mRNA表达水平明显高于对照组(t值分别为6.090、12.578、53.573，P值均＜0.001)。PERK、CHOP、caspase-12、GRP-78的蛋白表达水平与IL-1β、IL-6和TNF的mRNA表达水平均呈正相关(r值均＞0.700，P值均＜0.05)。 结论 HAE中ERS的激活与炎症介质的活化存在正相关性，ERS的过度激活能够改变部分炎症介质的分泌来加剧肝脏的损伤，具体机制有待进一步研究。 Abstract:Objective To investigate the influence of endoplasmic reticulum stress (ERS) on some inflammatory mediators during the progression of hepatic alveolar echinococcosis (HAE) and its clinical significance. Methods A total of 15 patients with HAE who underwent partial liver resection in Qinghai University Affiliated Hospital from June 2018 to September 2019 were enrolled, and the marginal zone of HAE lesion was resected as AE group; 15 normal liver tissue samples collected during the same period of time were selected as control group. Western blot and qRT-PCR were used to measure the protein and mRNA expression of protein kinase R-like ER kinase (PERK), CCAAT/enhancer-binding protein homologous protein (CHOP), caspase-12, and glucose-regulated protein-78 (GRP-78), and q-PCR was used to measure the mRNA expression of interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor (TNF). The Mann-Whitney U test was used for comparison of non-normally distributed continuous data between two groups, and the t-test was used for comparison of normally distributed continuous data between two groups; a Pearson correlation analysis was performed to investigate the correlation between two variables. Results Compared with the control group, the AE group had significantly higher protein expression levels of PERK, CHOP, caspase-12, and GRP78 (U=4.165, 3.461, 2.577, and 3.344, all P < 0.001) and their mRNA expression levels (t= 34.003, 4.461, 53.573, and 55.224, all P < 0.001). The AE group had significantly higher mRNA expression levels of IL-1β, IL-6, and TNF than the control group (t=6.090, 12.578, and 53.573, all P < 0.001). The protein expression levels of PERK, CHOP, caspase-12, and GRP-78 were positively correlated with the mRNA expression levels of IL-1β, IL-6, and TNF (all r > 0.700, all P≤0.05). Conclusion Positive correlation is observed between the activation of ERS and inflammatory mediators in HAE, and excessive activation of ERS can change the secretion of several inflammatory mediators to exacerbate liver injury, while further studies are needed to clarify the specific mechanism. -
Key words:
- Echinococcosis, Hepatic /
- Endoplasmic Reticulum Stress /
- Inflammation /
- Cytokines
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表 1 PCR引物序列
基因 引物序列(5′-3′) 引物长度(bp) IL-1β R：CGATCACTGAACTGCACGCTC 168 F：ACAAAGGACATGGAGAACACCACTT IL-6 R：CAATGAGGAGACTTGCCTGGTG 203 F：TGGCATTTGTGGTTGGGTCA 
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表 2 两组间ERS相关蛋白表达水平的比较
组别 例数 PERK CHOP caspase-12 GRP-78 AE组 15 27.63(25.32~29.32) 37.36(35.34~39.52) 31.41(29.34~34.63) 30.89(28.34~33.12) 对照组 15 19.89(18.34~21.28) 28.63(24.52~32.14) 20.36(17.62~24.83) 20.45(16.34~23.66) U值 4.165 3.461 2.577 3.344 P值 ＜0.001 ＜0.001 ＜0.001 ＜0.001
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表 3 两组间ERS相关指标mRNA表达水平的比较
组别 例数 PERK CHOP caspase-12 GRP-78 AE组 15 27.445±0.553 37.007±1.364 31.252±0.568 30.927±0.536 对照组 15 19.892±0.658 27.752±2.539 20.227±0.559 20.110±0.138 t值 34.003 4.461 53.573 55.224 P值 ＜0.001 ＜0.001 ＜0.001 ＜0.001
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表 4 两组间相关炎症介质mRNA表达水平的比较
组别 例数 IL-1β IL-6 TNF AE组 15 3.265±1.215 33.461±5.991 26.368±2.472 对照组 15 1.294±0.311 12.548±2.361 10.563±0.618 t值 6.090 12.578 53.573 P值 ＜0.001 ＜0.001 ＜0.001
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表 5 ERS相关蛋白与炎症介质的相关性
指标 PERK CHOP caspase-12 GRP-78 r值 P值 r值 P值 r值 P值 r值 P值 IL-1β 0.811 0.016 0.782 0.008 0.778 ＜0.001 0.786 0.005 IL-6 0.922 0.020 0.879 0.032 0.934 0.012 0.917 ＜0.001 TNF 0.784 ＜0.001 0.943 0.011 0.935 0.006 0.885 0.031
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