整合生物信息学分析痤疮的关键基因与免疫浸润水平

 目的 通过整合分析多芯片数据，探究与痤疮发病相关的关键基因及免疫细胞浸润水平。方法 从GEO数据库获取基因芯片数据集，采用“limma” R包和加权基因共表达网络分析(WGCNA)筛选与痤疮表型相关基因模块中的差异表达基因(DEGs)，并进行GO功能富集和KEGG通路分析；利用STRING在线数据库及Cytoscape软件构建蛋白质互作网络，并筛选出关键基因；基于CIBERSORT算法分析免疫细胞浸润情况。 结果 共筛选到154个DEGs。DEGs与信号受体结合、细胞因子活性、趋化因子受体结合等有关，主要集中在病毒蛋白与细胞因子和细胞因子受体的相互作用、细胞因子-细胞因子受体的相互作用、趋化因子信号传导、NF-κB信号传导、IL-17信号传导等通路。构建的PPI网络中，得到139个节点，1 597条边，筛选到两个关键基因(CCR5、CXCL8）。22种免疫细胞浸润中，相对于正常对照组，皮损组中有更多的中性粒细胞、活化的肥大细胞、活化的树突状细胞、活化的CD4记忆T细胞等，而调节性T细胞、静息的树突状细胞、静息的肥大细胞等相对更少。 结论 本研究利用生物信息学方法筛选的关键基因及免疫细胞浸润水平的变化可能在痤疮发病中起到重要作用。     

光化性角化病与皮肤鳞状细胞癌差异表达基因和通路的生物信息学分析

 目的:通过生物信息学分析的方法探究光化性角化病（AK）向皮肤鳞状细胞癌（cSCC）发展过程中可能起重要作用的关键基因和通路。方法：从GEO数据库中获取GSE32979和GSE45216数据集，共比较23个AK组织和43个cSCC组织。运用DAVID数据库对差异基因进行基因本体论（GO）分析以及基因组百科全书关键通路（KEGG）分析，建立差异基因的蛋白互作网络，获取枢纽基因和重要的模块。结果：研究发现了AK和cSCC中40个上调差异基因和35个下调差异基因。KEGG通路分析显示这些差异基因主要富集在黏着斑、 肥厚型心肌病、上皮细胞的细菌入侵等通路。此外研究建立的PPI网络包含了80个节点以及10个枢纽基因（SERPINE1、 ACTN1、 ITGA5、 MMP1、 CXCL1、 PLAUR、 GRB2、 MMP10、 TSG101、 VPS37B）。结论：本研究发现的枢纽基因和关键通路可能是AK向cSCC发展机制中可以进行干预的新靶点。     

慢性自发性荨麻疹发病机制的生物信息学研究

目的：利用生物信息学技术筛选慢性自发性荨麻疹（CSU）的关键差异表达基因,为研究CSU的生物学机制与治疗靶点提供新的思路。方法：从Gene Expression Omnibus（GEO）数据库下载GSE72541数据集,使用R语言limma包筛选CSU组与健康对照组间的差异表达基因,并通过clusterProfiler R包、STRING在线软件、Cytoscape3.8.2软件的MCODE插件分别对差异表达基因的功能、通路富集与蛋白相互作用进行分析。结果：研究发现CSU组与健康对照组共有86个差异表达基因,其中上调基因64个,下调基因22个(|logFC|>1 & P.Val<0.05),GO分析显示,差异表达基因主要富集在血小板活化、血液凝固、中性粒细胞脱颗粒等生物过程;KEGG分析显示,主要与造血细胞系、移植物抗宿主病、金黄色葡萄球菌等相关。此外,建立的PPI网络筛选出62个节点及15个关键基因（CXCL10, MMP9, SELP, MME, ITGA2B, ITGB3, CLU, GP6, C5AR1, C6orf25, GP1BB, VWF, CR1, GP9, PLAU）。结论：ITGB3,ITGA2B,MMP9及CR1等基因可能与CSU的发病密切相关。     

金黄色葡萄球菌生物膜致特应性皮炎的关键基因筛选及验证

目的 使用生物信息学方法对金黄色葡萄球菌(Staphylococcus aureus,SA)生物膜致特应性皮炎(atopic dermatitis, AD)的关键基因进行筛选、二次探索性分析,并预测潜在治疗药物。方法 从GEO数据库获得GSE32920基因芯片数据,针对SA生物膜对HaCaT细胞基因表达影响进行差异表达基因筛选、GO功能富集分析、KEGG通路富集分析、蛋白互作网络构建及hub基因筛选,筛选出的关键基因及相关通路,在GSE16161和GSE32924两个人体皮肤组织的基因芯片数据进行二次探索性分析。此外,使用Connectivity Map(CMap)平台进行治疗药物的预测。结果 本研究共筛选出SA生物膜对HaCaT细胞基因表达影响的差异表达基因共754个,其中显著上调404个、显著下调350个。GO功能富集分析、KEGG通路富集分析发现,差异表达基因功能主要富集于炎症反应、所参与的信号通路主要富集于IL-17信号通路、TNF信号通路等炎症通路。蛋白互作网络构建、hub基因筛选以及在两个人体数据集的二次探索性分析,最终确定了3个潜在关键靶点基因为：MMP1、CXCL1以及...     

采用生物信息学方法筛选和分析白癜风差异表达基因

【摘要】 目的 通过生物信息学方法探索与白癜风进展相关的信号通路和基因。方法 从GEO数据库中下载白癜风芯片检测数据集GSE75819,利用R语言软件中limma包的LMFit和eBayes函数筛选15例印度白癜风患者皮损与非皮损组织间的差异表达基因（DEG）。通过京都基因与基因组数据库（KEGG）、基因本体论（GO）分析和基因集富集分析（GSEA）评估DEG的富集途径和功能。通过蛋白-蛋白相互作用网络从DEG中筛选中心基因。2019年1 - 6月于新疆维吾尔自治区维吾尔医医院收集8例汉族寻常型白癜风患者皮损及非皮损皮肤组织标本,采用实时定量PCR法验证上述上调及下调差异最大的10个DEG的表达。结果 与15例非皮损组织相比,在15例白癜风皮损组织中共发现148个DEG,其中KRT9、CXCL10、C8ORF59、TPSAB1、RPL26为前5位上调基因,SILV、RPPH1、TYRP1、MLANA、LOC401115为前5位下调基因,且经实时定量PCR在8例汉族白癜风患者皮损及非皮损组织中验证。GO分析显示,DEG主要富集于翻译起始、细胞对脂多糖的反应、核糖体、核糖体亚基和核糖体的结构组成等。KEGG通路分析显示,DEG主要富集于酪氨酸代谢、PPAR信号通路、氧化磷酸化和Toll样受体信号通路。蛋白-蛋白相互作用分析筛选出UPF3B、SNRPG、MRPL13和RPL26L1 4个中心基因。结论 KRT9、CXCL10、C8ORF59、TPSAB1、RPL26、SILV、RPPH1、TYRP1、MLANA及LOC401115可能作为白癜风潜在的诊断标记分子和治疗靶点。     

45例麻风病患者外周血CD4+ T细胞差异表达基因筛查

【摘要】 目的 检测麻风病患者外周血CD4+ T细胞中mRNA表达谱,筛选并验证可能与麻风病发病过程密切相关的基因。方法 2018年7月至2020年5月在湖南省收集45例麻风病患者及45例健康人,以磁珠法分离外周血CD4+ T细胞,提取RNA。在上述受试者中,随机选取6例患者及6例健康人,采用Solexa测序法筛查两组间差异表达基因,将差异倍数2倍以上者（P < 0.05）定义为差异表达基因,并进行KEGG Pathway富集分析,再以实时荧光定量PCR验证基因的表达水平。结果 基因筛查发现4 831个转录本,属于新基因且具有蛋白编码潜能,在两组间筛选出8个差异表达基因,其中,CXCL8、PPBP、RPS18及IL-1β基因mRNA表达水平上调,RNH1、RPL39、RPL15及AMBRA1基因mRNA表达水平下调。实时荧光定量PCR验证结果与上述基因筛查结果一致。KEGG分析显示,麻风病患者与健康对照组差异表达基因主要富集在线粒体自噬及细胞自噬相关通路和人乳头状瘤病毒感染通路。结论 麻风病患者AMBRA1、RNH1基因mRNA表达水平下调,CXCL8、PPBP、IL-1β基因mRNA表达水平上调,可能分别通过线粒体自噬通路和趋化因子介导的信号通路参与麻风病发病。     

Th22细胞与银屑病

Th22细胞是最近发现的CD4＋T细胞功能亚群，表达CCR6、CCR4和CCRl0，产生IL-22和IL-13，但不产生IFN-7、IL-4和IL-17，是独立于1111、Th2和Thl7外的细胞亚群。Th22细胞在许多慢性炎症性疾病，如银屑病的发病中发挥重要作用。本文主要介绍Th22细胞和IL-22以及它们在银屑病发病中的作用及应用前景。     

系统性红斑狼疮患者CCR7+CD8+CD45RO+T细胞诱导CD4＋T细胞向Th2分化

目的探讨系统性红斑狼疮(SLE)患者外周血CCR7 CD8 CD45RO 记忆性T细胞对CD4 T细胞的诱导分化作用及其与SLE发病的关系。方法流式细胞仪、实时定量RT-PCR和RNA印迹检测同系CCR7 CD8 CD45RO T细胞和树突细胞协同刺激CD4 T细胞分泌细胞因子。结果活动期SLE患者CCR7 CD8 CD45RO 记忆性T细胞诱导CD4 T细胞表达Th2类细胞因子:白介素4的表达效率显著高于正常人对照组和非活动期SLE患者组(P<0.01),1型调节性T细胞(Tr1)源性细胞因子:白介素10和转化生长因子β的表达效率均低于正常对照组和非活动期SLE患者组(P<0.01);而活动期和非活动期SLE患者干扰素γ的表达效率显著低于正常人对照组(P<0.01)。结论活动期SLE患者外周血CCR7 中央型记忆性T细胞可与树突细胞相互作用,诱导同系CD4 T细胞向Th2分化,发挥CCR7-CD45RO 效应性记忆性T细胞的功能。     

RRS1对皮肤鳞状细胞癌细胞增殖、转移和侵袭的影响

目的：明确RRS1(核糖体合成调节因子1)对皮肤鳞状细胞癌(cSCC)细胞增殖、转移和侵袭的影响。方法：通过免疫组化和GEO数据库明确RRS1在cSCC与正常皮肤中表达的差异。慢病毒感染法建立RRS1敲低的cSCC细胞系SCL-1细胞。MTT实验、Celigo划痕实验、Transwell实验研究RRS1敲低后SCL-1细胞增殖、转移和侵袭性的变化。QRT-PCR法检测增殖、转移和侵袭相关基因表达。结果：RRS1在cSCC细胞中表达量明显高于正常皮肤,RRS1敲低后SCL-1细胞增殖、转移和侵袭性均降低。RRS1敲低后,增殖相关基因FGF2,AREG,cyclin E1显著降低。转移和侵袭相关基因VIM,CXCL1,CXCL3,CXCL8,MMP1表达显著降低。 结论：RRS1可能参与了cSCC细胞的增殖、转移和侵袭过程。     

男男性行为者CCR5、CCR2、SDF1、CXCR4基因多态性与HIV-1感染关联性分析

目的：研究男男性行为者(MSM)艾滋病病毒感染相关辅助受体CCR5、CCR2、CXCR4和SDF1基因多态性与HIV-1感染的相关性。方法：收集HIV-1抗体确证阳性者纳入病例组,HIV抗体筛查阴性者作为对照组,应用聚合酶链反应（PCR）和DNA测序法,检测两组MSM样本CCR5基因△32(rs333)和59029A/G(rs1799987)、CCR2、SDF1、CXCR4基因外显子区域的等位基因多态性,分析病例组和对照组基因型分布差异与HIV-1感染的相关性。结果：共收集病例组102例,对照组91例,两组样本CCR5基因rs333位点未检出杂合子和纯合子突变,rs1799987A/G位点突变频率为59.59%,病例组和对照组基因型差异分析显示,rs1799987A/G位点变异可能为影响HIV感染的风险因素［比值比(OR)=2.998,95%可信区间(CI）：1.034～8.696］;rs1799987A/G、rs1799864、rs1799865、CXCR4、SDF1基因突变频率分别为59.59%、19.95%、26.61%、11.92%和0.26%;CCR2第三外显子测序产物多态性分析发现,病例组和对照组差异有统计学意义（P<0.05）,经多因素回归分析未发现与HIV-1感染明显关联。结论：本地区MSM人群未发现CCR5基因△32突变以及对HIV-1感染的保护作用,59029A/G突变频率较高（为59.59%）,CCR5、CCR2、SDF1和CXCR4基因多态性与HIV-1感染未发现明显关联。     

Gene Expression Patterns of Cutaneous Squamous Cell Carcinoma and Actinic Keratosis: Biomarkers Screening for Skin Disease Diagnosis

BackgroundActinic keratosis (AK) was an intraepidermal tumor which caused by ultraviolet irradiation-induced skin damage.ObjectiveThe aim was to screen biomarkers for development of skin disease by comparing the gene expression profiles between cutaneous squamous cell carcinoma (CSCC) and AK.Methods{"type":"entrez-geo","attrs":{"text":"GSE45216","term_id":"45216"}}GSE45216 with 30 cutaneous squamous cell carcinoma patients and 10 actinic keratosis patients were downloaded and significance analysis of microarrays was processed to screen differently expressed genes (DEGs). Fisher''s exact test was processed for DEGs enrichment. Pathway relationship network systematically reflected the signal conduction and synergism between enriched pathways based on Kyoto Encyclopedia of Genes and Genomes database. Gene co-expression network was constructed according to gene expression data. Quantitative real-time-PCR was used to verify screened biomarkers.ResultsTotal 410 DEGs were screened and enriched into various functions, such as signal transduction and negative regulation of apoptotic process. They also participated into cytokine-cytokine receptor interaction and focal adhesion. The pathway relationship network was constructed with 27 nodes. Hub nodes with higher degree of this network were mitogen-activated protein kinase signaling pathway and apoptosis. The gene co-expression network was constructed with 39 nodes. Thereinto, hub node was ELOVL fatty acid elongase. The expression levels of ELOVL4 and HPGD were significantly higher in CSCC samples than that in AK samples, while the expression levels of INHBA and LAMC2 in CSCC samples were significantly lower than that in AK samples.ConclusionThese screened genes, including ELOVL4, HPGD, INHBA and LAMC2, played important roles in transformation from AK to CSCC.     

不同抗银屑病药物对实验性动物Th1/Th2基因表达的影响

目的分析不同抗银屑病药物作用前后实验性自身免疫性甲状腺炎(EAT)大鼠外周血单一核细胞Th1/Th2基因表达谱的差异性,寻找不同药物调节Th1/Th2平衡的相关基因并分析其可能机制。方法建立EAT模型,随机分成三组,给予灌胃治疗[模型组给予生理盐水,环孢素组予环孢素A 25 mg/(kg.d),雷公藤组予雪公藤2.75 mg/(kg.d)],4周后处死大鼠。分别提取各组外周血单一核细胞,提取总RNA,反转录成cRNA,用含115个目的基因的Th1/Th2功能分类基因芯片进行杂交,GEArray软件分析,筛选出治疗组与未治疗组(模型组)表达有差异的基因。结果筛选出两治疗组共同表达基因,其中有1条基因(IL-10)共同显著上调,有9条基因(CD28,CD69,Fadd,IL18,Socs3,Socs6,Stat1,Tnfsf5,Tnfsf6)共同下调。结论各抗银屑病药物均通过上调Th2细胞因子IL-10表达,下调Th1型相关基因表达以调节Th1/Th2平衡,其中Socs3,Socs6,Stat1,Tnfsf6等基因可能参与调节Th1/Th2平衡。     

Nickel-responding T cells are CD4+ CLA+ CD45RO+ and express chemokine receptors CXCR3, CCR4 and CCR10

BACKGROUND: Whereas T lymphocytes are widely accepted as effector cells determining the pathogenesis of allergic contact dermatitis, contradictory results have been found regarding the roles of different T-cell subsets. The use of various experimental models, involving long-term cultured T-cell lines or clones, may explain these contradictory results. OBJECTIVE: To investigate the involvement of distinct T-cell subsets in patients with nickel contact allergy. METHODS: Different T-cell subsets were directly isolated from peripheral blood mononuclear cells (PBMCs) of nickel-allergic patients, and their proliferative capacity, type-1 or type-2 cytokine secretion [measured by interferon (IFN)-gamma or interleukin (IL)-5 release] and phenotypical marker expression were analysed after stimulation with nickel. RESULTS: Only CD4+ CLA+ CD45RO+ and not CD8+ T cells proliferate and produce both type-1 (IFN-gamma) and type-2 (IL-5) cytokines in response to nickel. Moreover, cells expressing the marker CLA in combination with CD4, CD45RO or CD69 are increased after nickel-specific stimulation. Interestingly, in addition, CD45RA+ CLA+ cells showed an increased frequency after allergen-specific stimulation. Analysis of nickel-reactive T cells for expression of distinct chemokine receptors showed that both proliferative capacity and cytokine production are restricted to subsets expressing CXCR3, CCR4 but not CCR6. Fluorescence-activated cell sorting analysis of chemokine receptors expressed on nickel-stimulated T cells confirmed these results; a subset of T cells expressing CLA and CXCR3, CCR4 and, most importantly, CCR10 increased in response to allergen, while these CLA+ nickel-reactive T cells were all negative for CCR6. CONCLUSIONS: These findings demonstrate that freshly isolated nickel-reactive T cells can be characterized as CD4+ CLA+ memory T cells which express the chemokine receptors CXCR3, CCR4 and CCR10, but not CCR6.     

Expression pattern of chemokine receptors and chemokine release in inflammatory erythroderma and Sézary syndrome

BACKGROUND: Erythroderma can be caused by inflammatory dermatoses or cutaneous T-cell lymphoma. Even if chemokines and their receptors are involved in the skin-selective lymphocyte recruitment, their role in inflammatory erythroderma is yet unclear. OBJECTIVE: To evaluate the chemokine release (TARC, MDC, IP-10) and to define the expression pattern of Th1- (CCR5, CXCR3) and Th2-related (CCR4) chemokine receptors in inflammatory erythroderma and Sézary syndrome (SS). MATERIALS AND METHODS: Flow cytometry has been carried out on both circulating and skin-infiltrating T lymphocytes; serum chemokine levels have been evaluated using ELISA techniques. RESULTS: CCR4, CCR5 and CXCR3 were expressed on about 40% of peripheral blood lymphocytes and on the majority of skin-infiltrating lymphocytes in the inflammatory erythroderma patients, whereas the leukemic CD4+CD26- subpopulation in SS was characterized by a high CCR4 expression without a concurrent increase in CCR5 or CXCR3. TARC, MDC and IP-10 serum levels were significantly increased in both erythrodermic and SS patients. CONCLUSIONS: Our results confirm that SS is a Th2 disorder with a selective expression of CCR4, whereas inflammatory erythroderma shares an overexpression of both Th1- and Th2-related chemokine receptors, suggesting an activation of different pathways driving reactive lymphocytes to the skin.