Expression Analysis of WDR36 and WRN Genes of Borrelia Burgdorferi in HMC3 Cell Line Infection Model
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摘要:
目的 为探索神经莱姆病的分子机制，通过转录组学与伯氏疏螺旋体感染发现相关的2个基因 WDR36 和 WRN，以证明其具有靶点治疗的潜力。 方法 通过建立非人灵长类动物模型，利用高通量测序技术获取转录组学数据。通过GO富集分析，识别出 2 个具有研究价值的WDR36和WRN差异表达基因。为进一步验证这2个基因在Bb感染神经莱姆病中的作用，采用人脑胶质细胞HMC3细胞株作为模型。实验组细胞接受了不同MOI（1和10）的Bb接种，并在感染后6 h、12 h和24 h收集细胞悬液。使用trizol法提取RNA，并通过qPCR法测定WDR36和WRN基因的mRNA相对表达水平。 结果 在Bb感染的HMC3细胞中，发现Bb MOI=1为适宜感染浓度。结果表明WDR36的表达量在24 h与12 h对比有显著的上调（P < 0.01），且6 h的PBS组与试验组基因表达含量对比上升但不明显（P > 0.05），12 h、24 h上升明显，差异有统计学意义（P < 0.01），WRN的表达量在24 h与12 h对比有显著上调，且6 h、12 h、24 h的实验组分别比其PBS组基因表达含量呈上升趋势，差异有统计学意义（P < 0.01）。 结论 WDR36和WRN的上调可能与LNB的神经病理过程相关，这些发现为进一步研究LNB的分子机制提供了新的视角，并可能为开发针对Bb感染的新型治疗方案提供潜在的分子靶点。 Abstract:Objective The aim is to explore the molecular mechanism of neurological Lyme disease, particularly by identifying two differentially expressed genes WDR36 and WRN associated with Burkholderia infection through transcriptome analysis. Secondly, to verify their association in the pathogenesis of Borrelia burgdorferi infection and demonstrate their potential therapeutic targets. Methods By establishing a non-human primate model and utilizing high-throughput sequencing technology to obtain transcriptomic data. Through GO enrichment analysis, two differentially expressed genes with research value, WDR36 and WRN, were identified. To further validate the role of these two genes in Bb infection of Lyme disease, the HMC3 cell line of human glial cells was used as a model. The experimental group cells received Bb inoculation with different MOI （1 and 10）, and cell suspensions were collected at 6 h, 12 h, and 24 h after infection. Extract RNA using the triazol method and determine the relative mRNA expression levels of WDR36 and WRN genes using qPCR. Result It was found that Bb MOI=1 was the appropriate infection concentration in HMC3 cells infected with Bb. The results showed that the expression level of WDR36 was significantly upregulated at 24 h and 12 h, and the gene expression content of the 6 h PBS group increased but not significantly compared to the experimental group. The increase was significant at 12 h and 24 h, and there was a significant statistical difference with P < 0.01. The expression level of WRN was significantly upregulated at 24 h and 12 h, and the gene expression content of the 6 h, 12 h, and 24 h experimental groups showed an upward trend compared to their PBS group, and there was a significant statistical difference with P < 0.01. Conclusion The upregulation of WDR36 and WRN may be associated with the neuropathological processes of LNB. These findings provide a new perspective for further studying the molecular mechanisms of LNB and may provide potential molecular targets for developing novel treatment strategies for Bb infection. -
Key words:
- Borrelia burgdorferi /
- WDR36 /
- WRN /
- HMC3 cell line
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图 1 23个差异表达基因（DEGs）的筛选
Figure 1. Screening of 23 differentially expressed genes （DEGs）
图 2 免疫反应相关DEGs的相互作用关系网络
Figure 2. The interaction network of immune response related DEGs
图 4 组内HMC3在不同时间点WDR36表达量的比较
*P < 0.05，**P < 0.01。
Figure 4. Comparison of WDR36 secretion of HMC3 at different time points in the group
图 5 组间HMC3在不同时间点WDR36表达量的比较
**P < 0.01。
Figure 5. Comparison of WDR36 secretion of HMC3 at different time points between groups
图 6 组内HMC3在不同时间点WRN表达量的比较
**P < 0.01。
Figure 6. Comparison of WRN secretion of HMC3 at different time points in the group
图 7 组间HMC3在不同时间点WRN表达量的比较
**P < 0.01。
Figure 7. Comparison of WRN secretion of HMC3 at different time points between groups
表 1 主要差异表达基因BP（TMEM87A、WRN、FOLR2）和MF（BTAF1、MFN1、WRN）
Table 1. BP（TMEM87A、WRN、FOLR2）and MF（BTAF1、MFN1、WRN） selected genes
基因 GO富集BP GO富集MF CFH
OGN
BTAF1
WDR36
TMEM87A
ALCAM
MFN1
WRN
CENPC
SF3B1
CTNNAL1
ZBED5
FOLR2未选择
未选择
未选择
未选择
选中
未选择
未选择
选中
未选择
未选择
未选择
未选择
选中未选择
未选择
选中
未选择
未选择
未选择
选中
选中
未选择
未选择
未选择
未选择
未选择
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[1] Koedel U,Pfister H W. Lyme neuroborreliosis[J]. Curr Opin Infect Dis,2017,30(1):101-107. doi: 10.1097/QCO.0000000000000332 [2] Ogrinc K,Bogovic P,Maraspin V,et al. Why is the duration of erythema migrans at diagnosis longer in patients with Lyme neuroborreliosis than in those without neurologic involvement?[J]. Pathogens,2024,13(2):137. doi: 10.3390/pathogens13020137 [3] 曹文静,岳鹏,徐鑫,等. 神经莱姆病诊治进展[J]. 中国感染与化疗杂志,2021,21(3):369-373. [4] Dersch R,Toews I,Sommer H,et al. Methodological quality of guidelines for management of Lyme neuroborreliosis[J]. BMC Neurol,2015,15(11):242. [5] Halperin J. Diagnosis and management of Lyme neuroborreliosis[J]. Expert Rev Anti Infect Ther,2018,16(1):5-11. doi: 10.1080/14787210.2018.1417836 [6] Skarie J M,Link B A. The Primary open-angle glaucoma gene WDR36 functions in ribosomal RNA processing and interacts with the p53 stress–response pathway[J]. Human molecular genetics,2008,17(16):2474-2485. doi: 10.1093/hmg/ddn147 [7] Hui C W,St-Pierre M K,Detuncq J,et al. Nonfunctional mutant WRN protein leads to neurological deficits,neuronal stress,microglial alteration,and immune im-balance in a mouse model of Werner syndrome[J]. Brain Behav Immun,2018,73(10):450-469. [8] Chen L,Huang S,Lee L,et al. WRN,the protein deficient in Werner syndrome,plays a critical structural role in optimizing DNA repair[J]. Aging Cell,2003,2(4):191-199. doi: 10.1046/j.1474-9728.2003.00052.x [9] Monemi S,Spaeth G,DaSilva A,et al. Identification of a novel adult-onset primary open-angle glaucoma (POAG) gene on 5q22.1[J]. Hum Mol Genet,2005,14(6):725-733. doi: 10.1093/hmg/ddi068 [10] 王雅琴,朱益华. WDR36基因与原发性开角型青光眼[J]. 福建医药杂志,2011,33(3):136-138. doi: 10.3969/j.issn.1002-2600.2011.03.071 [11] 石珂. 江西籍原发性开角型青光眼家系的遗传学调查及与WDR36基因突变的相关性研究[D]. 南昌: 南昌大学,2011. [12] An S,Yao D,Zhang W,et al. WDR36 safeguards self-renewal and pluripotency of human extended pluripotent stem cells[J]. Front Genet,2022,22(13):905395. [13] 石珂,汪昌运. 江西籍开角型青光眼家系WDR36基因突变研究[J]. 实用医学杂志,2014,30(8):1185-1188. [14] Lotze M T, Frana L W, Sharrow S O, et al. In vivo administration of purified human interleukin 2. I. Half-life and immunologic effects of the Jurkat cell line-derived interleukin 2[J]. J Immunol,1985,134(1):157-166. [15] 朱曦,张国伟,管怀进. WRN基因研究进展[J]. 江苏医药,2015,41(12):1430-1433. [16] 吉磊. RecQ解旋酶WRN和RecQ1酶学特性和结构研究[D]. 咸阳: 西北农林科技大学,2020. [17] Hui C W,St-PierreM K,Detuncq J,et al. Nonfunctional mutant WRN protein leads to neurological deficits,neuronal stress,microglial alteration,and immune imbalance in a mouse model of Werner syndrome[J]. Brain Behav Immun,2018,73(12):450-469. [18] Picco G, Cattaneo C M, Crisafulli G, et al. Werner Helicase Is a Synthetic-Lethal Vulnerability in Mismatch Repair-Deficient Colorectal Cancer Refractory to Targeted Therapies, Chemotherapy, and Immunotherapy[J]. Cancer Discov,2021,11(8):1923-1937. [19] Derenty C,Ellis N A. Bloom’s syndrome: Why not premature aging?A comparison of the BLM and WRNhelicases[J]. Ageing Res Rev,2017,33(6):36-51. [20] Cui X,Hou J,Wang S,et al. Werner helicase mediates the senescence and cell cycle of leukemia cells by regulating DNA repair pathways[J]. Int J Biol Macromol,2024,255(6):128305. [21] 郑宏毅,郑永唐. 非人灵长类动物: 免疫学基础研究到临床应用的桥梁[J]. 中国免疫学杂志,2018,34(8):1121-1130. [22] Sebastienp,Jacques D,Catherinep,et al. Diagnosis and treatment of "chronic Lyme": Primum non nocere[J]. BMC Infect Dis,2023,23(1):642. doi: 10.1186/s12879-023-08618-w [23] Bransfield R C. Neuropsychiatric Lyme borreliosis: An overview with a focus on a specialty psychiatrist's clinical practice[J]. Healthcare (Basel),2018,6(3):104. doi: 10.3390/healthcare6030104 [24] Bransfield R C. The psychoimmunology of lyme/tick-borne diseases and its association with neuropsychiatric symptoms[J]. Open Neurol J,2012,6(10):88-93. doi: 10.2174/1874205X01206010088 [25] 中国心血管健康疾病报告撰写委员会. 中国心血管健康与疾病报告2022要点解读[J]. 中国心血管杂志,2023,28(4): 297-312. [26] Xiao Z W,Liu Y,Li Z Y,et al. Molecular mechanism of Tongmai Yangxin Pill intervention in elderly patients with coronary heart disease[J]. Eur Rev Med Pharmacol Sci,2022,26(22):8265-8275. [27] 毕云凤. 伯氏疏螺旋体致神经细胞凋亡和炎症反应的机制研究[D]. 昆明: 昆明医科大学,2023. [28] Ramesh G,Borda J T,Dufour J. Interaction of the Lyme disease spirochete Borrelia burgdorferi with brain parenchyma elicits inflammatory mediators from glial cells as well as glial and neuronal apoptosis[J]. Am J Pathol,2008,173(5):1415-1427. doi: 10.2353/ajpath.2008.080483 [29] Ramesh,G,Borda,J T,Gill A. Possible role of glial cells in the onset and progression of Lyme neuroborreliosis[J]. J Neuroinflammation,2009,6(8):23. doi: 10.1186/1742-2094-6-23 [30] Ramesh G,Didier P J,England J D. Inflammation in the pathogenesis of Lyme neuroborreliosis[J]. Am J Pathol,2015,185(4):1344-1360. doi: 10.1016/j.ajpath.2015.01.024 [31] Franchi L,Eigenbrod T,Munoz-planillo R,et al. The inflammasome: A caspase-1-activation platform that regulates immune responses and disease pathogenesis[J]. Nat Immunol,2009,10(3):241-247. doi: 10.1038/ni.1703 [32] Sung. Death and survival from executioner caspase activation[J]. Semin Cell Dev Biol,2024,156(3):66-73. -
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