Article(id=1194344007871459430, tenantId=1146029695717560320, journalId=1190317699101192196, issueId=1194344006382486063, articleNumber=1001-2494(2025)10-1013-06, orderNo=null, doi=10.11669/cpj.2025.10.002, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1737302400000, receivedDateStr=2025-01-20, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1762683401314, onlineDateStr=2025-11-09, pubDate=1746028800000, pubDateStr=2025-05-01, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1762683401314, onlineIssueDateStr=2025-11-09, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1762683401314, creator=13701087609, updateTime=1762683401314, updator=13701087609, issue=Issue{id=1194344006382486063, tenantId=1146029695717560320, journalId=1190317699101192196, year='2025', volume='60', issue='10', pageStart='1005', pageEnd='1102', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=1, specialIssue=0, createTime=1762683400960, creator=13701087609, updateTime=1762844794786, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1195020941253128793, tenantId=1146029695717560320, journalId=1190317699101192196, issueId=1194344006382486063, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1195020941253128794, tenantId=1146029695717560320, journalId=1190317699101192196, issueId=1194344006382486063, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=1013, endPage=1018, ext={EN=ArticleExt(id=1194344008756457578, articleId=1194344007871459430, tenantId=1146029695717560320, journalId=1190317699101192196, language=EN, title=Immunogenicity and Protective Efficacy of Influenza A H1N1 mRNA Vaccines in Mice, columnId=null, journalTitle=Chinese Pharmaceutical Journal, columnName=null, runingTitle=null, highlight=null, articleAbstract=

OBJECTIVE To investigate the immunogenicity and immune-protective efficiency of influenza A mRNA vaccines based on mRNA vaccine technology platform and provide new research strategies for the development of multivalent seasonal influenza mRNA vaccines. METHODS mRNA-H1N1 encapsulated by lipid nanoparticles(LNP) was transfected into HEK 293T cells, and the expression of LNP/mRNA-H1N1 was analyzed by Western blot and immunofluorescence microscopy in vitro, respectively. Furthermore, LNP/mRNA H1N1 was administered via intramuscular injection to SPF grade BALB/c mice, and the HA specific serum IgG titers induced by LNP/mRNA H1N1 were detected by ELISA. The secretion level of cytokine IFN-γ induced by LNP/mRNA-H1N1 was detected by ELISpot, HI titers induced by LNP/mRNA-H1N1 was measured by hemagglutination inhibition (HI) assay. Finally, the immune-protective efficacy LNP/mRNA-H1N1 was detected by the homologous H1N1 virus challenge experiment. RESULTS The specific expression of antigen protein HA could be detected by Western blot and fluorescence microscope, its molecular weight was approximately 75 000. Furthermore, LNP/mRNA-NH1N1 could induce significant humoral immune response, cellular immune response, and HI titers in BALB/c mice after prime and boost immunization. More importantly, LNP/mRNA-NH1N1 could provide 100% immune protection efficiency against H1N1 virus challenge. CONCLUSION LNP/mRNA-NH1N1 is an effective influenza mRNA candidate vaccine, laying the foundation for future preclinical safety evaluation trials in large animals and providing research ideas for the development of mRNA vaccines for other subtypes of influenza or other infectious disease vaccines.

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目的 基于mRNA疫苗技术平台考察甲型流感mRNA疫苗的免疫原性和免疫保护效率,为开发多价季节性流感mRNA疫苗提供新的研究策略。方法 利用即用型脂质纳米颗粒(lipid nanoparticles,LNP)对mRNA-H1N1进行包封,将LNP/mRNA-H1N1转染至HEK 293T细胞,通过蛋白质印迹法(Western blot)和免疫荧光显微镜对LNP/mRNA-H1N1的体外表达进行分析。进一步地,LNP/mRNA-H1N1以肌内注射途径免疫SPF级BALB/c小鼠,通过酶联免疫吸附实验(enzyme linked immunosorbent assay,ELISA)检测LNP/mRNA-H1N1诱导的血凝素(hemagglutinin,HA)特异性血清免疫球蛋白G(immunoglobulin G,IgG)效价。通过血凝抑制(hemagglutination inhibition,HI)实验检测LNP/mRNA-H1N1诱导的HI效价。通过酶联免疫斑点法(enzyme-linked immunosorbent spot assay,ELISpot)检测LNP/mRNA-H1N1诱导的细胞因子γ干扰素(interferon-gamma, IFN-γ)分泌水平,最后,通过同型H1N1病毒攻击实验检测LNP/mRNA-H1N1的免疫保护效率。结果 通过Western blot和免疫荧光显微镜均能检测到抗原蛋白HA的特异性表达,其相对分子质量约为75 000。LNP/mRNA-H1N1经初次与加强免疫BALB/c小鼠后诱导产生了有意义的体液免疫应答、细胞免疫应答和HI效价。更为重要的是,LNP/mRNA-H1N1能提供100%的免疫保护效率。结论 LNP/mRNA-H1N1是一种有效的甲型流感mRNA候选疫苗,为将来开展临床前大型动物的安评试验奠定了基础,也为开发其他亚型的流感mRNA疫苗或其他传染病疫苗提供了研究思路。

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Curr Pharm Des, 2011, 17(17):1720-1739., articleTitle=Computational studies of influenza A virus at three important targets: hemagglutinin, neuraminidase and M2 protein, refAbstract=null)], funds=[Fund(id=1194372477083153019, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, awardId=32070920, language=CN, fundingSource=国家自然科学基金面上项目资助(32070920), fundOrder=null, country=null), Fund(id=1194372477141873276, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, awardId=2682023ZTPY063, language=CN, fundingSource=中央高校基本科研业务费专项资金资助(2682023ZTPY063), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1194372474189083209, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, xref=null, ext=[AuthorCompanyExt(id=1194372474197471818, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, companyId=1194372474189083209, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=College of Medicine, Institute of Biomedical Engineering, Southwest Jiaotong University, Chengdu 610031, China), AuthorCompanyExt(id=1194372474201666123, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, companyId=1194372474189083209, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=西南交通大学医学院, 生物医学工程研究院, 成都 610031)])], figs=[ArticleFig(id=1194372476189766255, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=EN, label=Fig.1, caption=HA protein expression after LNP/mRNA-H1N1 transfected into HEK 293T cells was detected by Western blot analysis

Lane 1-20 μL of control (HEK 293T cells); Lane 2-10 μL of the cell lysates after LNP/mRNA-H1N1 was transfected into HEK 293T cells at 24 h; Lane 3-20 μL of the cell lysates after LNP/mRNA-H1N1 was transfected into HEK 293T cells at 24 h; Lane 4-protein marker; β-actin-internal control.

, figureFileSmall=KRyT6/1hwTe+4iQ8tvLqkw==, figureFileBig=ULLef521M2oZQxy9lM0WVA==, tableContent=null), ArticleFig(id=1194372476277846640, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=CN, label=图1, caption=Western blot检测即用型脂质纳米颗粒包封mRNA-H1N1(LNP/mRNA-H1N1)转染HEK 293T细胞后表达血凝素(HA)蛋白的结果

Lane 1-20 μL空白组(HEK 293T细胞)的裂解液;Lane 2-10 μL LNP/mRNA-H1N1转染HEK 293T细胞24 h后的裂解液;Lane 3-20 μL LNP/mRNA-H1N1转染HEK 293T细胞24 h后的裂解液; Lane 4-蛋白质分子量标准;β-actin-内参。

, figureFileSmall=KRyT6/1hwTe+4iQ8tvLqkw==, figureFileBig=ULLef521M2oZQxy9lM0WVA==, tableContent=null), ArticleFig(id=1194372476361732721, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=EN, label=Fig.2, caption=The results of LNP/mRNA-H1N1 transfected with HEK 293T cells were detected by immunofluorescence microscopy assay(200×)

A-negative control(HEK 293T cells); B-immunofluorescence image of LNP/mRNA-H1N1 transfected into HEK 293T cells at 24 h.

, figureFileSmall=0DITIZqgasL+yg7Hw/2mTw==, figureFileBig=LYP0AJZfJaXj7/PPRLIYcg==, tableContent=null), ArticleFig(id=1194372476416258674, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=CN, label=图2, caption=免疫荧光分析LNP/mRNA-H1N1转染HEK 293T细胞的结果(200×)

A-阴性对照组(HEK 293T细胞);B-LNP/mRNA-H1N1转染HEK 293T细胞24 h后免疫荧光结果。

, figureFileSmall=0DITIZqgasL+yg7Hw/2mTw==, figureFileBig=LYP0AJZfJaXj7/PPRLIYcg==, tableContent=null), ArticleFig(id=1194372476470784627, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=EN, label=Fig.3, caption=HA specific serum IgG titers induced by LNP/mRNA-H1N1 were detected by ELISA. n=10,$\stackrel{-}{x}$±s

PBS-phosphate buffered solution; LNP-lipid nanoparticle; ¹⁾P<0.05, compared with the PBS and LNP controls.

, figureFileSmall=PxFyu8BH+EwOjVJa84j/Cg==, figureFileBig=k3+5sEzZ4nm/95mHsM7a4w==, tableContent=null), ArticleFig(id=1194372476533699188, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=CN, label=图3, caption=ELISA检测LNP/mRNA-H1N1诱导的HA特异性血清免疫球蛋白G(IgG)效价。 n=10,$\stackrel{-}{x}$±s

PBS-磷酸缓冲盐溶液;LNP-即用型脂质的纳米颗粒;与对照组(PBS组和LNP组)相比,¹⁾P<0.05。

, figureFileSmall=PxFyu8BH+EwOjVJa84j/Cg==, figureFileBig=k3+5sEzZ4nm/95mHsM7a4w==, tableContent=null), ArticleFig(id=1194372476605002357, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=EN, label=Fig.4, caption=The secretion levels of cytokine IFN-γ detected by ELISpot.n=5,$\stackrel{-}{x}$±s

¹⁾P<0.05,compared with the PBS and LNP controls.

, figureFileSmall=sVugHk16noSneUYUu0mJ2g==, figureFileBig=G0nVAlPVAV6eoF/D95aGJg==, tableContent=null), ArticleFig(id=1194372476672111222, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=CN, label=图4, caption=ELISpot检测细胞因子干扰素(IFN-γ)分泌水平。n=5,$\stackrel{-}{x}$±s

与对照组(PBS组和LNP组)相比较,¹⁾P<0.05。

, figureFileSmall=sVugHk16noSneUYUu0mJ2g==, figureFileBig=G0nVAlPVAV6eoF/D95aGJg==, tableContent=null), ArticleFig(id=1194372476739220087, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=EN, label=Fig.5, caption=The results of immune protection with LNP/mRNA-H1N1 were detected by H1N1 virus challenge experiment. n=10,$\stackrel{-}{x}$±s

A-body mass changes of mice of post virus challenge; B-survival statistics of mice at post virus challenge; ¹⁾P<0.05, compared with the PBS and LNP controls.

, figureFileSmall=Wv7+TQw6n6lfWJfwuYjAgw==, figureFileBig=CMx09PWoZT0BKQXZdrcdLA==, tableContent=null), ArticleFig(id=1194372476802134648, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=CN, label=图5, caption=H1N1病毒攻击实验检测LNP/mRNA-H1N1的免疫保护结果。 n=10,$\stackrel{-}{x}$±s

A-攻毒后小鼠的体质量变化图;B-攻毒后小鼠的存活统计图;与对照组(PBS组和LNP组)相比,¹⁾P<0.05。

, figureFileSmall=Wv7+TQw6n6lfWJfwuYjAgw==, figureFileBig=CMx09PWoZT0BKQXZdrcdLA==, tableContent=null), ArticleFig(id=1194372476869243513, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=EN, label=Tab.1, caption=

Serum HI titers after immunization of LNP/mRNA-H1N1 (GMT). n=10

, figureFileSmall=null, figureFileBig=null, tableContent=

Groups	Day 14	Day 28
PBS	<10	<10
LNP	20	20
LNP/mRNA-H1N1	3671)	6401)

), ArticleFig(id=1194372476932158074, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1194344007871459430, language=CN, label=表1, caption=

LNP/mRNA-H1N1免疫后的血清血凝抑制(HI)效价(GMT).n=10

, figureFileSmall=null, figureFileBig=null, tableContent=

Groups	Day 14	Day 28
PBS	<10	<10
LNP	20	20
LNP/mRNA-H1N1	3671)	6401)

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