Article(id=1194643390219395233, tenantId=1146029695717560320, journalId=1189873630562394117, issueId=1194643387904136153, articleNumber=null, orderNo=null, doi=10.11855/j.issn.0577-7402.0054.2024.0913, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1705075200000, receivedDateStr=2024-01-13, revisedDate=null, revisedDateStr=null, acceptedDate=1714233600000, acceptedDateStr=2024-04-28, onlineDate=1762754779629, onlineDateStr=2025-11-10, pubDate=1737993600000, pubDateStr=2025-01-28, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1762754779629, onlineIssueDateStr=2025-11-10, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1762754779629, creator=13701087609, updateTime=1762754779629, updator=13701087609, issue=Issue{id=1194643387904136153, tenantId=1146029695717560320, journalId=1189873630562394117, year='2025', volume='50', issue='1', pageStart='1', pageEnd='120', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=0, createTime=1762754779076, creator=13701087609, updateTime=1762756450259, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1194650397408203370, tenantId=1146029695717560320, journalId=1189873630562394117, issueId=1194643387904136153, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1194650397408203371, tenantId=1146029695717560320, journalId=1189873630562394117, issueId=1194643387904136153, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=35, endPage=43, ext={EN=ArticleExt(id=1194643390420721828, articleId=1194643390219395233, tenantId=1146029695717560320, journalId=1189873630562394117, language=EN, title=Expression and diagnostic value of CYBB and CSF1R in chronic rhinosinusitis with nasal polyps, columnId=1190310109000602400, journalTitle=Medical Journal of Chinese People’s Liberation Army, columnName=Clinical Research, runingTitle=null, highlight=null, articleAbstract=

Objective To analyze the gene expression characteristics of chronic rhinosinusitis with nasal polyps (CRSwNP) using bioinformatics methods, aim to investigate the potential biomarkers and their diagnostic value of CRSwNP. Methods (1) The CRSwNP Gene expression data set was downloaded from the American Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) between CRSwNP patients and healthy controls were screened through data analysis. Gene Ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed on the identified DEGs. Protein-protein interaction (PPI) networks were constructed utilizing the STRING database, and the key genes were identified by using the cytoHubba plugin. The "Cibersort" package was used to analyze the influence of key genes on common immune cells. (2) Thirty-two patients diagnosed with CRSwNP in the First Affiliated Hospital of Hebei North University from June 2022 to June 2023 were selected as the CRSwNP group, and 21 patients with simple deviation of nasal septum without a history of sinusitis during the same period were selected as control group. The pathological characteristics of specimens in the two groups were examined using hematoxylin-eosin (HE) staining. Immunohistochemistry and Western blotting were used to detect the expression levels of key genes in CRSwNP. The levels of key proteins in plasma were detected using ELISA, and ROC curve was used to analyze its efficacy in diagnosing CRSwNP. Results (1) Analysis of three gene expression database sets (GSE36830, GSE23552, and GSE194282) showed that there were 156 DEGs in CRSwNP. GO functional enrichment and KEGG pathway analysis indicated that the functions of the above DEGs were mostly related to immune functions. Key genes such as cytochrome b-245 β chain (CYBB) and colony-stimulating factor 1 receptor (CSF1R) were identified. (2) The results of HE staining revealed that the epithelial of CRSwNP tissue was metaplastic into stratified squamous epithelium with interstitial edema. Both immunohistochemistry and Western blotting analyses indicated that the expression levels of CYBB and CSF1R in the CRSwNP group were significantly increased compared to control group (P<0.05). ELISA results demonstrated that CYBB [(21.20±3.00) μg/ml vs. (17.66±1.66) μg/ml, P<0.05] and CSF1 [(477.37±86.63) pg/ml vs. (370.71±66.24) pg/ml, P<0.05] in CRSwNP group were significantly increased compare to control group. ROC curve analysis showed that plasma concentrations of CYBB and CSF1 had AUCs of 0.888 (95%CI 0.802-0.974) and 0.821 (95%CI 0.711-0.931) for diagnosing of CRSwNP, respectively; their combined AUC was 0.927 (95%CI 0.851-1.000). Conclusions CYBB and CSF1R may be involved in the occurrence and development of CRSwNP. Plasma CYBB and CSF1 have high diagnostic value for CRSwNP.

, correspAuthors=Gang Xue, Xu Lin, authorNote=null, correspAuthorsNote=
Xue Gang, E-mail:
Lin Xu, E-mail:
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目的 采用生物信息学方法分析慢性鼻窦炎鼻息肉(CRSwNP)基因表达的特点,探寻CRSwNP潜在的生物标志物及其诊断价值。方法 (1)从美国GEO基因表达数据库下载CRSwNP基因表达数据集,通过数据分析筛选CRSwNP与健康对照的差异表达基因(DEGs);对获取的DEGs进行基因本体论(GO)功能和京都基因与基因组百科全书(KEGG)通路分析,利用STRING数据库构建蛋白-蛋白互作(PPI)网络,利用cytoHubba插件筛选关键基因。利用Cibersort包分析关键基因对常见免疫细胞的影响。(2)选取2022年6月-2023年6月在河北北方学院附属第一医院确诊的32例CRSwNP患者为CRSwNP组,另选取同期无鼻窦炎病史的单纯鼻中隔偏曲患者21例作为对照组。HE染色观察两组标本的病理学特点,免疫组化和Western blotting检测CRSwNP关键基因的表达水平,ELISA法检测血浆关键蛋白的水平,并绘制ROC曲线分析关键蛋白诊断CRSwNP的效能。结果 (1)对获取的3个基因表达数据库集(GSE36830、GSE23552和GSE194282)进行分析显示,CRSwNP有156个DEGs;GO功能和KEGG通路分析显示,上述DEGs的作用多与免疫功能有关;筛选出细胞色素b-245 β链(CYBB)和集落刺激因子1受体(CSF1R)等关键基因。(2)HE染色结果显示,CRSwNP组织上皮化生为复层鳞状上皮,间质水肿;免疫组化和Western blotting检测结果显示,与对照组比较,CRSwNP组CYBB、CSF1R表达水平均明显升高(P<0.05)。ELISA检测结果显示,与对照组比较,CRSwNP组血浆CYBB[(21.20±3.00) μg/ml vs. (17.66±1.66) μg/ml,P<0.001]和CSF1[(477.37±86.63) pg/ml vs. (370.71±66.24) pg/ml,P<0.001]水平均明显升高。ROC曲线分析显示,血浆CYBB和CSF1水平诊断CRSwNP的曲线下面积(AUC)分别为0.888(95%CI 0.802~0.974)、0.821(95%CI 0.711~0.931),二者联合诊断CRSwNP的AUC为0.927(95%CI 0.851~1.000)。结论 CYBB、CSF1R可能参与了CRSwNP的发生发展,血浆CYBB、CSF1水平可能成为CRSwNP的诊断标志物。

, correspAuthors=薛刚, 林旭, authorNote=null, correspAuthorsNote=
薛刚,E-mail:
林旭,E-mail:
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马宇龙,主要从事耳鼻咽喉疾病生物信息学方面的研究

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马宇龙,主要从事耳鼻咽喉疾病生物信息学方面的研究

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马宇龙,主要从事耳鼻咽喉疾病生物信息学方面的研究

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CRSwNP. 慢性鼻窦炎鼻息肉;DEGs. 差异表达基因;A. 主坐标分析(PCoA)图;B. DEGs的火山图;C. DEGs的热图(Top20)

, figureFileSmall=X0CWjwhiWPn0eDkn0KsvnA==, figureFileBig=rVg03YC0YRdYp+jr3eUPAg==, tableContent=null), ArticleFig(id=1194662028930421719, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=EN, label=Fig.2, caption=Enrichment analysis of related functions in CRSwNP, figureFileSmall=iMyhyTyu88+MDRNd8GFPmw==, figureFileBig=ilhA2yC22pdxqg+05yas5Q==, tableContent=null), ArticleFig(id=1194662028989141977, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=CN, label=图2, caption=CRSwNP中相关基因的功能富集分析

CRSwNP. 慢性鼻窦炎鼻息肉;NES. 归一化后的富集分数;FDR. 伪发现率;GO. 基因本体论;KEGG. 京都基因与基因组百科全书;GSEA. 基因集富集分析;A. CRSwNP中差异表达基因的GO功能富集分析和KEGG通路分析结果;B. CRSwNP中相关基因的GSEA分析结果

, figureFileSmall=iMyhyTyu88+MDRNd8GFPmw==, figureFileBig=ilhA2yC22pdxqg+05yas5Q==, tableContent=null), ArticleFig(id=1194662029052056539, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=EN, label=Fig.3, caption=PPI network diagram of DEGs in CRSwNP, figureFileSmall=cakShpJvWcCfIwbVxr7ULA==, figureFileBig=vA8fGC61RiEJNp3+1wkpDg==, tableContent=null), ArticleFig(id=1194662030113215453, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=CN, label=图3, caption=CRSwNP中DEGs的PPI网络图

CRSwNP. 慢性鼻窦炎鼻息肉;DEGs. 差异表达基因;PPI. 蛋白质-蛋白质相互作用;CYBB. 细胞色素b-245 β链;CSF1R. 集落刺激因子1受体;A. PPI网络;B. 采用cytoHubba筛选的核心基因

, figureFileSmall=cakShpJvWcCfIwbVxr7ULA==, figureFileBig=vA8fGC61RiEJNp3+1wkpDg==, tableContent=null), ArticleFig(id=1194662030171935711, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=EN, label=Fig.4, caption=Immune infiltration results of CYBB and CSF1R in CRSwNP in GEO database, figureFileSmall=Jkxp80uFR41UXNvsm8fffQ==, figureFileBig=E/Lt7gYU+dBPbuIQNP/0Nw==, tableContent=null), ArticleFig(id=1194662030260016097, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=CN, label=图4, caption=基于GEO数据库的CRSwNP中CYBB、CSF1R免疫浸润分析结果

CRSwNP. 慢性鼻窦炎鼻息肉;CYBB. 细胞色素b-245 β链;CSF1R. 集落刺激因子1受体;A. CYBB的免疫浸润结果;B. CSF1R的免疫浸润结果;*P<0.05,**P<0.01,***P<0.001

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CRSwNP. 慢性鼻窦炎鼻息肉;A. 对照组;B. Eos-CRSwNP组;C. nonEos-CRSwNP组

, figureFileSmall=OFMQi7gWc0VEMrgBVYmMAg==, figureFileBig=LdlKsCGsSUFcy7W74/gYUg==, tableContent=null), ArticleFig(id=1194662030448759783, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=EN, label=Fig.6, caption=The expression of CYBB and CSF1R in CRSwNP (Immunohistochemical staining), figureFileSmall=f7gXDsI0SxjCb4LJ/NJX7w==, figureFileBig=iDF9H4rR0Dd0Rox2el2vFw==, tableContent=null), ArticleFig(id=1194662030520062953, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=CN, label=图6, caption=CRSwNP组织中CYBB和CSF1R的表达情况(免疫组织化学染色)

CRSwNP. 慢性鼻窦炎鼻息肉;CYBB. 细胞色素b-245 β链;CSF1R. 集落刺激因子1受体

, figureFileSmall=f7gXDsI0SxjCb4LJ/NJX7w==, figureFileBig=iDF9H4rR0Dd0Rox2el2vFw==, tableContent=null), ArticleFig(id=1194662030595560427, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=EN, label=Fig.7, caption=The expressions of CYBB and CSF1R in CRSwNP (Western blotting), figureFileSmall=AW87X+P7bNF9Yzzu/AX8kw==, figureFileBig=g/JX+4haAQpx5kEPC66VwQ==, tableContent=null), ArticleFig(id=1194662030654280685, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=CN, label=图7, caption=CRSwNP组织中CYBB、CSF1R的表达情况 (Western blotting)

CRSwNP. 慢性鼻窦炎鼻息肉;CYBB. 细胞色素b-245 β链;CSF1R. 集落刺激因子1受体;***P<0.001

, figureFileSmall=AW87X+P7bNF9Yzzu/AX8kw==, figureFileBig=g/JX+4haAQpx5kEPC66VwQ==, tableContent=null), ArticleFig(id=1194662030725583855, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=EN, label=Fig.8, caption=ROC curves of the diagnostic value of CYBB and CSF1 levels in prediction of CRSwNP, figureFileSmall=Oln4A4IW18OLezCw21534A==, figureFileBig=qQ3ncmTKwJxoJU+Xbx6IKg==, tableContent=null), ArticleFig(id=1194662030817858545, tenantId=1146029695717560320, journalId=1189873630562394117, articleId=1194643390219395233, language=CN, label=图8, caption=血浆CYBB和CSF1水平诊断CRSwNP的ROC曲线分析

CRSwNP. 慢性鼻窦炎鼻息肉;CYBB. 细胞色素b-245 β链;CSF1R. 集落刺激因子1受体;AUC. 曲线下面积;A. 两组血浆CYBB、CSF1水平比较;B. 血浆CYBB、CSF1水平及两者联合诊断CRSwNP的ROC曲线;***P<0.001

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CYBBCSF1R在慢性鼻窦炎伴鼻息肉中的表达及诊断价值
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马宇龙 1 , 李庚 1 , 吴靖芳 1 , 薛刚 2, * , 林旭 1, *
解放军医学杂志 | 临床研究 2025,50(1): 35-43
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解放军医学杂志 | 临床研究 2025, 50(1): 35-43
CYBBCSF1R在慢性鼻窦炎伴鼻息肉中的表达及诊断价值
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马宇龙1, 李庚1, 吴靖芳1, 薛刚2, * , 林旭1, *
作者信息
  • 1河北北方学院基础医学院,河北张家口 075000
  • 2河北北方学院附属第一医院耳鼻咽喉头颈外科,河北张家口 075000
  • 马宇龙,主要从事耳鼻咽喉疾病生物信息学方面的研究

通讯作者:

薛刚,E-mail:
林旭,E-mail:
Expression and diagnostic value of CYBB and CSF1R in chronic rhinosinusitis with nasal polyps
Yu-Long Ma1, Geng Li1, Jing-Fang Wu1, Gang Xue2, * , Xu Lin1, *
Affiliations
  • 1School of Basic Medical Sciences, Hebei North University, Zhangjiakou, Hebei 075000, China
  • 2Department of Otolaryngology, Head and Neck Surgery, the First Affiliated Hospital of Hebei North University, Zhangjiakou, Hebei 075000, China
出版时间: 2025-01-28 doi: 10.11855/j.issn.0577-7402.0054.2024.0913
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目的 采用生物信息学方法分析慢性鼻窦炎鼻息肉(CRSwNP)基因表达的特点,探寻CRSwNP潜在的生物标志物及其诊断价值。方法 (1)从美国GEO基因表达数据库下载CRSwNP基因表达数据集,通过数据分析筛选CRSwNP与健康对照的差异表达基因(DEGs);对获取的DEGs进行基因本体论(GO)功能和京都基因与基因组百科全书(KEGG)通路分析,利用STRING数据库构建蛋白-蛋白互作(PPI)网络,利用cytoHubba插件筛选关键基因。利用Cibersort包分析关键基因对常见免疫细胞的影响。(2)选取2022年6月-2023年6月在河北北方学院附属第一医院确诊的32例CRSwNP患者为CRSwNP组,另选取同期无鼻窦炎病史的单纯鼻中隔偏曲患者21例作为对照组。HE染色观察两组标本的病理学特点,免疫组化和Western blotting检测CRSwNP关键基因的表达水平,ELISA法检测血浆关键蛋白的水平,并绘制ROC曲线分析关键蛋白诊断CRSwNP的效能。结果 (1)对获取的3个基因表达数据库集(GSE36830、GSE23552和GSE194282)进行分析显示,CRSwNP有156个DEGs;GO功能和KEGG通路分析显示,上述DEGs的作用多与免疫功能有关;筛选出细胞色素b-245 β链(CYBB)和集落刺激因子1受体(CSF1R)等关键基因。(2)HE染色结果显示,CRSwNP组织上皮化生为复层鳞状上皮,间质水肿;免疫组化和Western blotting检测结果显示,与对照组比较,CRSwNP组CYBB、CSF1R表达水平均明显升高(P<0.05)。ELISA检测结果显示,与对照组比较,CRSwNP组血浆CYBB[(21.20±3.00) μg/ml vs. (17.66±1.66) μg/ml,P<0.001]和CSF1[(477.37±86.63) pg/ml vs. (370.71±66.24) pg/ml,P<0.001]水平均明显升高。ROC曲线分析显示,血浆CYBB和CSF1水平诊断CRSwNP的曲线下面积(AUC)分别为0.888(95%CI 0.802~0.974)、0.821(95%CI 0.711~0.931),二者联合诊断CRSwNP的AUC为0.927(95%CI 0.851~1.000)。结论 CYBB、CSF1R可能参与了CRSwNP的发生发展,血浆CYBB、CSF1水平可能成为CRSwNP的诊断标志物。

慢性鼻窦炎伴鼻息肉  /  关键基因  /  生物信息学  /  筛查诊断

Objective To analyze the gene expression characteristics of chronic rhinosinusitis with nasal polyps (CRSwNP) using bioinformatics methods, aim to investigate the potential biomarkers and their diagnostic value of CRSwNP. Methods (1) The CRSwNP Gene expression data set was downloaded from the American Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) between CRSwNP patients and healthy controls were screened through data analysis. Gene Ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed on the identified DEGs. Protein-protein interaction (PPI) networks were constructed utilizing the STRING database, and the key genes were identified by using the cytoHubba plugin. The "Cibersort" package was used to analyze the influence of key genes on common immune cells. (2) Thirty-two patients diagnosed with CRSwNP in the First Affiliated Hospital of Hebei North University from June 2022 to June 2023 were selected as the CRSwNP group, and 21 patients with simple deviation of nasal septum without a history of sinusitis during the same period were selected as control group. The pathological characteristics of specimens in the two groups were examined using hematoxylin-eosin (HE) staining. Immunohistochemistry and Western blotting were used to detect the expression levels of key genes in CRSwNP. The levels of key proteins in plasma were detected using ELISA, and ROC curve was used to analyze its efficacy in diagnosing CRSwNP. Results (1) Analysis of three gene expression database sets (GSE36830, GSE23552, and GSE194282) showed that there were 156 DEGs in CRSwNP. GO functional enrichment and KEGG pathway analysis indicated that the functions of the above DEGs were mostly related to immune functions. Key genes such as cytochrome b-245 β chain (CYBB) and colony-stimulating factor 1 receptor (CSF1R) were identified. (2) The results of HE staining revealed that the epithelial of CRSwNP tissue was metaplastic into stratified squamous epithelium with interstitial edema. Both immunohistochemistry and Western blotting analyses indicated that the expression levels of CYBB and CSF1R in the CRSwNP group were significantly increased compared to control group (P<0.05). ELISA results demonstrated that CYBB [(21.20±3.00) μg/ml vs. (17.66±1.66) μg/ml, P<0.05] and CSF1 [(477.37±86.63) pg/ml vs. (370.71±66.24) pg/ml, P<0.05] in CRSwNP group were significantly increased compare to control group. ROC curve analysis showed that plasma concentrations of CYBB and CSF1 had AUCs of 0.888 (95%CI 0.802-0.974) and 0.821 (95%CI 0.711-0.931) for diagnosing of CRSwNP, respectively; their combined AUC was 0.927 (95%CI 0.851-1.000). Conclusions CYBB and CSF1R may be involved in the occurrence and development of CRSwNP. Plasma CYBB and CSF1 have high diagnostic value for CRSwNP.

CRSwNP  /  hub genes  /  bioinformatics  /  screening and diagnosis
马宇龙, 李庚, 吴靖芳, 薛刚, 林旭. CYBBCSF1R在慢性鼻窦炎伴鼻息肉中的表达及诊断价值. 解放军医学杂志, 2025 , 50 (1) : 35 -43 . DOI: 10.11855/j.issn.0577-7402.0054.2024.0913
Yu-Long Ma, Geng Li, Jing-Fang Wu, Gang Xue, Xu Lin. Expression and diagnostic value of CYBB and CSF1R in chronic rhinosinusitis with nasal polyps[J]. Medical Journal of Chinese People’s Liberation Army, 2025 , 50 (1) : 35 -43 . DOI: 10.11855/j.issn.0577-7402.0054.2024.0913
慢性鼻窦炎(chronic rhinosinusitis,CRS)是一种与黏膜炎症相关的临床综合征,其患病率在全球范围持续居高位,美国和欧洲患病率超过10%,每年医疗费用超过80亿美元[1],我国CRS的患病率约为8%。CRS的主要症状包括鼻塞、流鼻涕、面部疼痛和嗅觉减退,一般通过鼻内窥镜检查或鼻窦CT进行诊断。CRS可分为两种类型:CRS伴鼻息肉(CRSwNP)和CRS无鼻息肉(CRSsNP)。CRSwNP症状较重、易复发[2],且致病因素复杂、无明确诊断标准,缺乏个性化的诊疗方案。根据炎性细胞浸润水平的差异,CRSwNP可分为嗜酸性粒细胞性慢性鼻窦炎伴鼻息肉(eosinophilic CRS with nasal polyps,Eos-CRSwNP)与非嗜酸粒细胞性慢性鼻窦炎伴鼻息肉(non-eosinophilic CRS with nasal polyps,nonEos-CRSwNP)。研究显示,鼻息肉的发病与上皮屏障功能障碍、过敏原致敏、机体免疫功能紊乱有关[3-4],但CRSwNP的促炎分子机制及免疫调控机制尚不清楚。因此,深入挖掘CRSwNP潜在的致病分子机制,探究核心基因的作用,对于CRSwNP的诊治至关重要。本研究通过整合现有公共数据集,利用综合生物信息学方法深入挖掘CRSwNP中潜在的关键基因及其作用,并采用HE染色、免疫组化染色、Western blotting验证获得的关键基因,探索其可能的分子机制,最后利用ELISA实验探究关键基因是否可作为CRSwNP的血液筛查标志物,旨在为CRSwNP的诊断与治疗提供新的生物标志物和药物靶点。
抗细胞色素b-245 β链(cytochrome b-245 beta chain,CYBB;#DF6520)、集落刺激因子1受体(colony-stimulating factor 1 receptor,CSF1R;#AF0080)、β-actin(#AF7018)抗体,HRP标记的山羊抗兔IgG二抗(#S0001)购自江苏亲科生物研究中心有限公司;DAB显色液(PV-8000)购自北京中杉金桥公司;蛋白抽提试剂盒(R0018S)购自上海碧云天生物技术有限公司;ELISA法CYBB(CB14284-Hu)、CSF1(CB17500-Hu)检测试剂盒购自上海科艾博生物技术有限公司。
从Gene Expression Omnibus(GEO)数据库(https://www.ncbi.nlm.nih.gov/geo/)中下载基因表达数据集GSE36830、GSE23552、GSE194282,剔除与CRSwNP无关及临床信息不全的样本后,3个数据集中分别包括CRSwNP样本6例、11例、6例,健康对照样本6例、17例、6例。
收集2022年6月-2023年6月在河北北方学院附属第一医院就诊、参照2018年慢性鼻窦炎诊疗指南[5]确诊的CRSwNP患者32例作为CRSwNP组;另选取同期因单纯鼻中隔偏曲就诊的患者21例作为对照组。对照组既往无鼻窦炎病史。术中组织标本经处理分为二份,一份置于液氮保存,一份经固定液固定。收集两组患者的外周血样本。CRSwNP组纳入标准:首次以鼻息肉为主要症状就诊的CRSwNP患者。排除标准:术前1个月内全身使用过类固醇激素;术前发现合并后鼻孔息肉、真菌性鼻窦炎等疾病;术后病理检查结果显示并非鼻息肉。本研究获河北北方学院附属第一医院医学伦理委员会审核批准(K2023041)。
采用Limma软件包对GEO数据集进行校正及标准化处理,去除批次效应后合并为单一表达矩阵[6]。主坐标分析后,使用Array_DEG软件包对合并后的数据集进行分析,设置阈值为|log2 fold change|>1和adjust P-value <0.05筛选差异表达基因(differentially expressed genes,DEGs),ggplot2包绘制DEGs的热图和火山图。采用clusterProfiler包进行基因集富集分析(Gene Set Enrichment Analysis,GSEA),选择带注释的基因集(c2.cp.kegg.v7.2.symbols.gmt)作为参考基因集,每个分析随机运行排列基因组1000次,ggplot2包绘制山峦图;对DEGs进行基因本体论(Gene Ontology,GO)功能分析和KEGG通路分析,设置P<0.05和adjust P-value <0.05为筛选条件,通过ggplot2软件包、igraph软件包和ggraph软件包对相似性结果进行EMAP图可视化。应用STRING数据库对DEGs进行蛋白-蛋白互作网络(PPI)构建,选择“Homo sapiens”,置信度设置为默认值(0.4),使用Cytoscape软件重新可视化PPI网络,运用cytoHubba插件通过Degree算法筛选出关键基因(hub genes)[7]。使用Cibersort包计算CRSwNP关键基因与常见免疫细胞丰度的关系。使用pROC包绘制ELISA结果的相关接受者操作特征(receiver operating characteristic,ROC)曲线。
鼻息肉组织及单纯鼻中隔偏曲患者部分下鼻甲组织经固定、脱水、包埋、切片后,切片依次进行脱蜡、水化、苏木精染色、盐酸乙醇分化、伊红染色、脱水、透明封片后光镜下观察并拍照。
将切片依次进行脱蜡至水、抗原修复、3%过氧化氢孵育、山羊血清封闭后,加入CYBB(1:250)与CSF1R(1:200)一抗孵育,二抗(1:5000)孵育,DAB显色、苏木精染核,脱水、透明、封片后拍照。每张切片随机选取5个高倍镜下阳性视野,用Image-Pro Plus 6.0软件进行图像分析。比较CRSwNP组与对照组CYBB、CSF1R表达水平的差异。
采用RIPA裂解液提取总蛋白,BCA定量分析蛋白浓度。变性后的蛋白经电泳、转膜、封闭,CYBB(1:650)与CSF1R(1:500)一抗孵育,二抗(1:5000)孵育后,ECL显色,采用凝胶成像系统扫描,ImageJ软件分析条带的灰度值。
收集两组患者的外周血,分离血浆,按照ELISA试剂盒操作说明分别检测CYBB、CSF1的水平,采用酶标仪在450 nm波长下测定吸光度值,通过标准曲线计算血浆中CYBB、CSF1的水平。
采用SPSS 22软件进行统计分析。符合正态分布的计量资料以$\bar{x}±s$表示,两组间比较采用独立样本t检验,多组间比较采用单因素方差分析,进一步两两比较采用Dunnett-t检验;非正态分布的计量资料以M(Q1Q3)表示,组间比较采用Mann-Whitney U检验。计数资料以例(%)表示,组间比较采用χ2检验。采用ROC曲线分析血浆CYBB、CSF1水平对CRSwNP的诊断价值。P<0.05为差异有统计学意义。
主坐标分析结果显示,合并后的基因表达数据集中CRSwNP组与对照组之间共筛选出156个DEGs,其中表达上调101个,表达下调55个(图1)。
GO功能富集和KEGG通路分析结果显示,156个DEGs的主要功能富集在白细胞趋化、与趋化因子受体CCR结合、免疫效应过程的调节、调节炎症反应、趋化因子活性、细胞因子-细胞因子受体相互作用等。KEGG通路分析结果显示这些DEGs主要通过趋化因子信号通路等发挥调节细胞因子或其他炎性因子而发挥作用(图2A)。对差异表达不显著但有重要生物学意义的基因进行GSEA分析,结果显示,这部分基因功能主要富集于细胞因子-细胞因子受体相互作用、免疫系统细胞因子信号转导、中性粒细胞脱颗粒、白细胞介素信号转导、淋巴细胞与非淋巴细胞的免疫调节相互作用等(图2B)。
将DEGs导入STRING数据库构建PPI,并通过Cytoscape软件重新可视化处理。结果显示,PPI由340个节点及1380条边构成(图3A)。cytoHubba插件最后成功筛选出PPI网络中的关键基因CYBBCSF1RITGAMCD86TYROBP(图3B)。以下对相关性居前两位的基因CYBB、CSF1R进行进一步观察。
采用GEO数据库对CYBB、CSF1R是否在CRSwNP中参与调节免疫微环境进行分析,结果显示,CYBB与10种免疫细胞相关,其中与M0型巨噬细胞在内的免疫细胞呈负相关,与M2型巨噬细胞在内的免疫细胞呈正相关(图4A);CSF1R在CRSwNP中的免疫细胞浸润结果与CYBB相近(图4B)。
HE染色结果显示,对照组上皮为假复层纤毛柱状上皮,纤毛完整,固有层中未见大量免疫细胞及水肿情况(图5A)。根据嗜酸性粒细胞浸润水平不同,将本研究纳入的CRSwNP组织分为Eos-CRSwNP型15例[嗜酸性粒细胞:(183.6±23.8)个/高倍视野]和nonEos-CRSwNP型17例[嗜酸性粒细胞:(8.1±1.7)个/高倍视野](图5B、C)。与对照组比较,Eos-CRSwNP型的上皮化生为复层鳞状上皮,间质部分轻度水肿;nonEos-CRSwNP型的上皮变厚,间质水肿,血管扩张充血,腺体高度增生,炎性细胞浸润明显。
免疫组化染色结果示,CYBB主要表达于上皮细胞质及胞膜,在胞核中弱表达,呈棕黄色;CSF1R主要定位于上皮细胞的细胞质,呈棕褐色。与对照组比较,Eos-CRSwNP组和nonEos-CRSwNP组CYBB、CSF1R表达水平均明显升高(P<0.05),而Eos-CRSwNP组与nonEos-CRSwNP组CYBB、CSF1R表达水平差异无统计学意义(P>0.05,图6)。Western blotting检测结果与免疫组化检测结果一致(图7)。
ELISA检测结果显示,与对照组比较,CRSwNP组血浆CYBB[(21.20±3.00) μg/ml vs. (17.66±1.66) μg/ml,P<0.001]和CSF1[(477.37±86.63) pg/ml vs. (370.71±66.24) pg/ml,P<0.001]水平明显升高(图8A)。ROC曲线分析结果显示,CYBB和CSF1水平诊断CRSwNP的曲线下面积分别为0.888(95%CI 0.802~0.974)、0.821(95%CI 0.711~0.931),二者联合诊断CRSwNP的曲线下面积为0.927(95%CI 0.851~1.000,图8B)。
CRS是耳鼻咽喉头颈外科的常见疾病,在我国患病率约为8.0%,涉及1.07亿人,造成了沉重的经济和社会负担[8]。CRSwNP是CRS的常见类型,临床症状较重,相较于不伴鼻息肉的CRS,其术后复发率高,严重影响患者的生活质量[9]。组织病理学检查是CRSwNP分型的金标准,通过观察息肉组织中嗜酸性粒细胞的浸润程度可明确Eos-CRSwNP和nonEos-CRSwNP两种亚型。但术前行有创活检存在患者接受程度差、费用高等缺点[10]。虽然糖皮质激素治疗对大多数病例有效,但由于疾病的异质性,不同CRSwNP患者对治疗的反应差异较大,可能出现抵抗[11]。因此,探索简单易行、价格低廉的诊断指标对CRSwNP的早期诊断非常重要。对于不同分型的CRSwNP进行个体化诊疗符合精准医学的要求,必将成为该病治疗的趋势。精准医学的高速发展离不开基因测序技术的快速更新以及生物信息学技术的广泛应用。近年来,有多项研究尝试探索Eos-CRSwNP的生物标志物和诊断模型。周方伟等[12]指出,血清25-(OH)D3水平可作为区分Eos-CRSwNP与nonEos-CRSwNP的有效指标,但作为单一的预测指标,易受寄生虫感染、自身免疫性疾病等其他共病因素的影响,特异度较差;Yu等[13]建立了一个基于患者病史、症状、体征和鼻息肉评分的联合预测模型,虽然受自身因素影响较小,但一致性指数(C-index)仅为0.808,在预测CRSwNP方面的价值相对有限。深入探索CRSwNP潜在的分子机制,有可能为其提供更为准确的诊断学标志物。本研究旨在利用生物信息学技术对3个CRSwNP基因表达数据集进行深入挖掘,为其无创筛查诊断提供潜在的标志物。
本研究中,CRSwNP鼻息肉组织及正常黏膜组织转录组学数据筛选得到156个DEGs,其中表达上调101个,表达下调55个。GO功能分析结果显示,上述DEGs的功能主要与白细胞趋化、免疫反应有关;KEGG通路分析结果显示,上述DEGS可能经趋化因子信号通路调节组织中细胞因子或其他炎性因子的释放而发挥作用,与文献报道一致[14];对差异表达不显著但有重要生物学意义的基因进行GSEA分析,结果也显示,CRSwNP的相关基因与免疫调节、炎症反应存在密切联系,与GO功能和KEGG通路分析结果基本一致。构建的PPI网络提供了CRSwNP的基因调控网络,并鉴定出CYBBCSF1RITGAMCD86TYROBP共5个关键基因,提示其可能在CRSwNP发病过程发挥重要作用。ITGAM是补体C3降解产物iC3b的受体,也是细胞黏附分子-1(ICAM-1)和纤维蛋白的受体,参与上皮细胞的黏附、趋化及屏障修复作用[15]。CD86属于免疫球蛋白超家族成员,在鼻息肉形成过程中起着调节作用[16]。TYROBP是TYRO蛋白酪氨酸激酶结合蛋白,通过参与免疫应答、激活效应淋巴细胞而发挥促炎作用[17-18]
CYBB又称NOX2,是宿主抗菌防御和调节炎症的关键酶。有研究指出CYBB突变后可调控巨噬细胞引起慢性肉芽肿病[19]。CYBB可被与感染相关的因素迅速激活,进而通过调节中性粒细胞的积累及清除来抑制炎症和损伤[20]。集落刺激因子(CSF1)主要通过CSF1R激活单核细胞吞噬系统调节机体的免疫应答,特别是调节巨噬细胞表型的极化[21]。本研究结果显示,CYBB与CSF1R在CRSwNP中呈过表达,且M2型巨噬细胞在CYBB和CSF1R过表达组中较低表达组明显增多,提示二者可能通过调节免疫微环境中的肿瘤相关巨噬细胞发生M2型极化来影响CRSwNP的发生发展。CYBB、CSF1R在Eos-CRSwNP组与nonEos-CRSwNP组的表达水平均高于对照组,提示二者在CRSwNP的形成过程中发挥了促进作用。CSF1刺激产生的巨噬细胞可持续对M2型巨噬细胞诱导产生的白细胞介素-4(IL-4)作出免疫应答,维持炎症的发生[22]。CYBB作为NADPH氧化酶蛋白家族中的一员,亦可促进巨噬细胞的M2型极化[23]。考虑到CRSwNP起病隐匿,通过血液检测其潜在标志物有助于疾病的早期诊断。本研究发现,CRSwNP患者血浆CYBB与CSF1水平均较健康人群升高,二者均可作为CRSwNP诊断的标志物,特别是二者联合应用的ROC曲线下面积达0.927,诊断效能较高。
综上所述,本研究结果提示CYBB和CSF1对于CRSwNP具有较好的临床诊断价值,可为CRSwNP的早期诊断及精准化治疗方案的制订提供帮助。但本研究未对CYBB和CSF1R在CRSwNP中的具体分子机制进行深入探究,未来可进一步探讨二者是否通过调节巨噬细胞M2型极化来影响CRSwNP的发生发展。
  • 国家级大学生创新创业训练计划项目(202310092007)
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doi: 10.11855/j.issn.0577-7402.0054.2024.0913
  • 接收时间:2024-01-13
  • 首发时间:2025-11-10
  • 出版时间:2025-01-28
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  • 收稿日期:2024-01-13
  • 录用日期:2024-04-28
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National College Students' Innovative Entrepreneurial Training Plan(202310092007)
国家级大学生创新创业训练计划项目(202310092007)
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    1河北北方学院基础医学院,河北张家口 075000
    2河北北方学院附属第一医院耳鼻咽喉头颈外科,河北张家口 075000

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2种不同金属材料的力学参数

Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)

Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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