促凝血小板在感染性血栓形成中的调控作用

doi:10.3969/j.issn.1674-8115.2026.01.003

上海交通大学学报（医学版） ›› 2026, Vol. 46 ›› Issue (1): 25-33.doi: 10.3969/j.issn.1674-8115.2026.01.003

促凝血小板在感染性血栓形成中的调控作用

殷旭颖¹, 王炫佳², 杨咪娜³^,⁴, 姜豪杰¹, 戴菁³, 孙晓玉²(), 徐艳艳³^,⁴()

^1.上海交通大学基础医学院生物化学与分子细胞生物学系，上海 200025
^2.复旦大学上海市重大传染病和生物安全研究院，上海 200030
^3.上海交通大学医学院附属瑞金医院检验科，上海 200025
^4.上海交通大学医学院医学技术学院，上海 200025

收稿日期:2025-08-19 接受日期:2025-10-09 出版日期:2026-01-28 发布日期:2026-01-30
通讯作者: 徐艳艳，研究员，博士；电子信箱：xuyanyan901@163.com
孙晓玉，青年研究员，博士；电子信箱：sunxiaoyu@fudan.edu.cn。
作者简介:第一联系人：殷旭颖、徐艳艳、孙晓玉、戴菁、姜豪杰参与实验设计。殷旭颖、王炫佳、杨咪娜参与实验操作。殷旭颖、徐艳艳参与论文的写作和修改。所有作者均阅读并同意最终稿件的提交。
为共同第一作者（co-first authors）。
基金资助:
国家重点研发计划(2023YFC2507801);国家自然科学基金(82322004);国家自然科学基金(82170126);国家自然科学基金(82400153);国家自然科学基金(82200131);国家自然科学基金(82470128);上海市自然科学基金(25ZR1402094)

Procoagulant platelets regulate H1N1-induced thrombosis formation

Yin Xuying¹, Wang Xuanjia², Yang Mina³^,⁴, Jiang Haojie¹, Dai Jing³, Sun Xiaoyu²(), Xu Yanyan³^,⁴()

^1.Department of Biochemistry and Molecular Cell Biology, Shanghai Jiao Tong University College of Basic Medical Sciences, Shanghai 200025, China
^2.Shanghai Institute of Infectious Disease and Biosecurity, Fudan University, Shanghai 200030, China
^3.Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
^4.College of Health Sciences and Technology, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China

Received:2025-08-19 Accepted:2025-10-09 Online:2026-01-28 Published:2026-01-30
Contact: Xu Yanyan, E-mail: xuyanyan901@163.com
Sun Xiaoyu, E-mail: sunxiaoyu@fudan.edu.cn.
About author:First author contact:The study was designed by Yin Xuying, Xu Yanyan, Sun Xiaoyu, Dai Jing, and Jiang Haojie. The experiments were conducted by Yin Xuying, Wang Xuanjia, and Yang Mina. The manuscript was drafted and revised by Yin Xuying and Xu Yanyan. All authors have read the final version of the manuscript and consented to its submission.
Supported by:
National Key Research and Development Program of China(2023YFC2507801);National Natural Science Foundation of China(82322004);Natural Science Foundation of Shanghai(25ZR1402094)

摘要/Abstract

摘要：

目的·探讨促凝血小板在H1N1流行性感冒（流感）病毒感染的小鼠体内血栓形成中的作用。方法·利用鼻滴注H1N1流感病毒构建小鼠病毒感染模型，持续监测病毒感染后小鼠的体质量，通过眼眶后静脉丛采集全血测定血小板数量。腹主动脉抽血后制备小鼠血浆样本，采用酶联免疫吸附试验（enzyme linked immunosorbent assay，ELISA）测定小鼠血浆凝血酶时间（thrombin time，TT），及纤维蛋白原、D-二聚体、白细胞介素-6（interleukin-6，IL-6）、肿瘤坏死因子α（tumor necrosis factor-α，TNF-α）水平。制备感染小鼠的肺和肝组织切片，进行苏木精-伊红（hematoxylin and eosin，HE）染色，观察小鼠肺和肝组织内血栓形成情况；利用酪酰胺信号放大（tyramide signal amplification，TSA）技术对肺和肝组织进行多重荧光免疫组织化学染色，分析血栓的组成成分。利用流式细胞术监测病毒感染后小鼠体内单核细胞-血小板聚集物水平和促凝血小板亚群的比例变化。通过尾静脉注射小鼠GP1b抗体（R300）构建小鼠血小板清除模型，持续监测小鼠体质量和血小板数量，测定小鼠血浆TT及纤维蛋白原、D-二聚体水平。结果·H1N1病毒感染后，小鼠体质量持续性下降，凝血相关指标异常，表现为TT延迟、纤维蛋白原水平降低、D-二聚体水平升高等，血浆IL-6和TNF-α水平也升高（均P<0.05）；HE染色结果显示，病毒感染后的小鼠肺和肝组织血管内可见明显的血栓形成（均P<0.05）。流式分析结果表明H1N1病毒感染后促凝血小板比例显著升高，促凝血小板的比例与病毒使用滴度呈正相关，且与D-二聚体、纤维蛋白原水平以及血栓面积具有相关性（均P<0.05）。TSA染色结果发现血管内血栓主要由大量纤维蛋白和促凝血小板组成。将小鼠体内血小板清除后，感染H1N1病毒引发的肺和肝组织血栓形成水平明显降低，并且凝血异常程度得到缓解（均P<0.05）。结论·H1N1病毒可能通过增加促凝血小板比例，导致凝血功能异常，从而诱发小鼠多组织脏器中的血栓形成。

关键词: 血小板, 促凝血小板, 血栓, 感染

Abstract:

Objective ·To investigate the role of procoagulant platelets in thrombosis induced by H1N1 influenza virus infection in mice. Methods ·A viral infection model was established in mice by intranasal instillation of the H1N1 influenza virus. Body weight was continuously monitored post-infection, and whole blood was collected from the retro-orbital venous plexus to measure platelet counts. Plasma samples were prepared from abdominal aortic blood. Thrombin time (TT), fibrinogen, D-dimer, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) levels were measured by using enzyme-linked immunosorbent assay (ELISA). Lung and liver tissue sections from infected mice were prepared, and hematoxylin-eosin (HE) staining was employed to observe thrombus formation in lung and liver tissues of mice. Tyramide signal amplification (TSA)-based multiplex immunohistochemistry was performed on lung and liver tissues to analyze thrombus composition. Flow cytometry was used to monitor the monocyte-platelet aggregates and the proportion of procoagulant platelets in mice after viral infection. A platelet depletion model was established by tail vein injection of anti-mouse GP1b (R300). Body weight and platelet counts were continuously monitored in infected mice. Plasma TT, fibrinogen, and D-dimer levels were measured to evaluate coagulation function in mice. Results ·Following H1N1 infection, mice exhibited progressive weight loss and abnormal coagulation parameters, including prolonged TT, decreased fibrinogen levels, and elevated D-dimer levels (all P<0.05).Plasma levels of IL-6 and TNF-α were also elevated. HE staining revealed prominent thrombus formation in the pulmonary and hepatic vasculature of infected mice (all P<0.05). Flow cytometry demonstrated a significant increase in the proportion of procoagulant platelets after infection. The proportion of procoagulant platelets showed a positive correlation with viral dose, and was associated with D-dimer levels, fibrinogen levels, and thrombus area (all P<0.05). TSA staining further confirmed that the thrombi primarily consisted of fibrin and procoagulant platelets. Depletion of platelets in mice prior to H1N1 infection markedly reduced thrombosis in lung and liver tissues and alleviated coagulation abnormalities (all P<0.05). Conclusion ·H1N1 virus infection may significantly elevate the proportion of procoagulant platelets in mice, leading to coagulation dysfunction and subsequent thrombosis in multiple tissues.

Key words: platelet, procoagulant platelet, thrombosis, infection

中图分类号:

R364

殷旭颖, 王炫佳, 杨咪娜, 姜豪杰, 戴菁, 孙晓玉, 徐艳艳. 促凝血小板在感染性血栓形成中的调控作用[J]. 上海交通大学学报（医学版）, 2026, 46(1): 25-33.

Yin Xuying, Wang Xuanjia, Yang Mina, Jiang Haojie, Dai Jing, Sun Xiaoyu, Xu Yanyan. Procoagulant platelets regulate H1N1-induced thrombosis formation[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2026, 46(1): 25-33.

图/表 4

图1 病毒感染状态下的小鼠凝血功能异常Note： Sixteen 8-week-old C57BL/6J wild-type female mice were randomly divided into four groups (n=4 per group) and intranasally instilled with equal volumes containing 0, 250, 1 000, or 2 000 pfu of H1N1, respectively. A. Body weight changes in each group of mice were measured every 12 h post-viral infection. B. Platelet counts in each group of mice were monitored via orbital blood sampling every 12 h post-viral infection. C‒E. At 96 h post-viral infection, plasma was collected from each group of mice to measure thrombin time (TT) (C), fibrinogen (D), and D-dimer levels (E). F. Detection of TNF-α and IL-6 levels in the plasma of infected mice by ELISA. G. Flow cytometry was used to detect the levels of monocyte-platelet aggregates in infected mice.

Fig 1 Coagulation dysfunction in mice under viral infection

图2 病毒感染小鼠肺组织和肝组织的血栓形成Note： At 96 h post-infection, lung and liver tissues were collected from the four groups of virus-infected mice (n=4 per group) for HE staining. A. HE staining of lung and liver tissues from the four experimental groups at 96 h post-viral infection. B. Quantitative analysis of thrombus area in lung tissues across the four groups. C. Quantitative analysis of thrombus area in liver tissues across the four groups.

Fig 2 Thrombus formation in the lung and liver tissues of virus-infected mice

图3 病毒感染状态下的促凝血小板比例上升Note： A. Proportion of procoagulant platelets in the four groups at 60 h post-infection (flow cytometry plots). B. Temporal dynamics of procoagulant platelet proportions in the four groups at 0, 12, 48, and 60 h post-infection (quantitative statistical analysis). n=4 per group. C. Multiplex fluorescence immunohistochemical staining of intravascular thrombi in mouse lung tissue. D. Co-localization of CD41 and PS. E. Statistical analysis of the percentage of overlapping area between CD41 and PS. F. Linear correlation analysis of the procoagulant platelets with D-dimer levels, fibrinogen levels, and thrombus area in lung and liver tissues.

Fig 3 Increase in the proportion of procoagulant platelets under viral infection

图4 清除血小板状态下血栓形成和凝血指标异常的改变Note： Eight 8-week-old female mice were randomly divided into two groups (n=4 per group). A. Platelet depletion and H1N1 viral infection strategy. B. Body weight changes in each group of mice were measured every day. C. Platelet counts in each group of mice were monitored via orbital blood sampling every day. D. HE staining of lung and liver tissues in both groups. E. Quantitative analysis of thrombus area in lung and liver tissues between the two groups. F. At 96 h post-viral infection, plasma was collected from both groups to measure the TT, fibrinogen, and D-dimer levels by ELISA.

Fig 4 Changes in thrombus formation and coagulation abnormalities under platelet depletion

参考文献 39

[1]	Ksiazek T G, Erdman D, Goldsmith C S, et al. A novel coronavirus associated with severe acute respiratory syndrome[J]. N Engl J Med, 2003, 348(20): 1953-1966.
[2]	Sullivan S J, Jacobson R M, Dowdle W R, et al. 2009 H1N1 influenza[J]. Mayo Clin Proc, 2010, 85(1): 64-76.
[3]	Van Wissen M, Keller T T, Ronkes B, et al. Influenza infection and risk of acute pulmonary embolism[J]. Thromb J, 2007, 5: 16.
[4]	Al-Samkari H, Karp Leaf R S, Dzik W H, et al. COVID-19 and coagulation: bleeding and thrombotic manifestations of SARS-CoV-2 infection[J]. Blood, 2020, 136(4): 489-500.
[5]	Evans P C, Rainger G E, Mason J C, et al. Endothelial dysfunction in COVID-19: a position paper of the ESC working group for atherosclerosis and vascular biology, and the ESC council of basic cardiovascular science[J]. Cardiovasc Res, 2020, 116(14): 2177-2184.
[6]	Mackman N, Antoniak S, Wolberg A S, et al. Coagulation abnormalities and thrombosis in patients infected with SARS-CoV-2 and other pandemic viruses[J]. Arterioscler Thromb Vasc Biol, 2020, 40(9): 2033-2044.
[7]	GóMez-Mesa J E, Galindo-Coral S, Montes M C, et al. Thrombosis and coagulopathy in COVID-19[J]. Curr Probl Cardiol, 2021, 46(3): 100742.
[8]	Zerangian N, Erabi G, Poudineh M, et al. Venous thromboembolism in viral diseases: a comprehensive literature review[J]. Health Sci Rep, 2023, 6(2): e1085.
[9]	Avnon L S, Munteanu D, Smoliakov A, et al. Thromboembolic events in patients with severe pandemic influenza A/H1N1[J]. Eur J Intern Med, 2015, 26(8): 596-598.
[10]	Repsold L, Joubert A M. Platelet function, role in thrombosis, inflammation, and consequences in chronic myeloproliferative disorders[J]. Cells, 2021, 10(11): 3034.
[11]	Gremmel T, Frelinger A L, Michelson A D. Platelet physiology[J]. Semin Thromb Hemost, 2024, 50(8): 1173-1186.
[12]	Koupenova M, Kehrel B E, Corkrey H A, et al. Thrombosis and platelets: an update[J]. Eur Heart J, 2017, 38(11): 785-791.
[13]	Holinstat M. Normal platelet function[J]. Cancer Metastasis Rev, 2017, 36(2): 195-198.
[14]	Ali M A M, Spinler S A. COVID-19 and thrombosis: from bench to bedside[J]. Trends Cardiovasc Med, 2021, 31(3): 143-160.
[15]	Ding J, Hostallero D E, El Khili M R, et al. A network-informed analysis of SARS-CoV-2 and hemophagocytic lymphohistiocytosis genes′ interactions points to neutrophil extracellular traps as mediators of thrombosis in COVID-19[J]. PLoS Comput Biol, 2021, 17(3): e1008810.
[16]	Ackermann M, Verleden S E, Kuehnel M, et al. Pulmonary vascular endothelialitis, thrombosis, and angiogenesis in covid-19[J]. N Engl J Med, 2020, 383(2): 120-128.
[17]	Heemskerk J M, Mattheij N A, Cosemans J M. Platelet-based coagulation: different populations, different functions[J]. J Thromb Haemost, 2013, 11(1): 2-16.
[18]	Reddy E C, Rand M L. Procoagulant phosphatidylserine-exposing platelets in vitro and in vivo[J]. Front Cardiovasc Med, 2020, 7: 15.
[19]	Bevers E M, Williamson P L. Getting to the outer leaflet: physiology of phosphatidylserine exposure at the plasma membrane[J]. Physiol Rev, 2016, 96(2): 605-645.
[20]	Millington-Burgess S L, Harper M T. Cytosolic and mitochondrial Ca²⁺ signaling in procoagulant platelets[J]. Platelets, 2021, 32(7): 855-862.
[21]	Lentz B R. Exposure of platelet membrane phosphatidylserine regulates blood coagulation[J]. Prog Lipid Res, 2003, 42(5): 423-438.
[22]	Colicchia M, Schrottmaier W C, Perrella G, et al. S100A8/A9 drives the formation of procoagulant platelets through GPIbα[J]. Blood, 2022, 140(24): 2626-2643.
[23]	Meyer N J, Gattinoni L, Calfee C S. Acute respiratory distress syndrome[J]. Lancet, 2021, 398(10300): 622-637.
[24]	Kwong J C, Schwartz K L, Campitelli M A, et al. Acute myocardial infarction after laboratory-confirmed influenza infection[J]. N Engl J Med, 2018, 378(4): 345-353.
[25]	Goldberg R, Ye W, Johns K, et al. Comparison of thrombotic and clinical outcomes in SARS-CoV-2-pneumonia versus other viral pneumonia in an urban academic medical center[J]. Heart Lung, 2023, 61: 153-157.
[26]	Snell J. SARS-CoV-2 infection and its association with thrombosis and ischemic stroke: a review[J]. Am J Emerg Med, 2021, 40: 188-192.
[27]	Mandel J, Casari M, Stepanyan M, et al. Beyond hemostasis: platelet innate immune interactions and thromboinflammation[J]. Int J Mol Sci, 2022, 23(7): 3868.
[28]	Thomas M R, Storey R F. The role of platelets in inflammation[J]. Thromb Haemost, 2015, 114(3): 449-458.
[29]	Franco A T, Corken A, Ware J. Platelets at the interface of thrombosis, inflammation, and cancer[J]. Blood, 2015, 126(5): 582-588.
[30]	Jenne C N, Kubes P. Platelets in inflammation and infection[J]. Platelets, 2015, 26(4): 286-292.
[31]	Li S P, Lu Z F, Wu S Y, et al. The dynamic role of platelets in cancer progression and their therapeutic implications[J]. Nat Rev Cancer, 2024, 24(1): 72-87.
[32]	Koupenova M, Corkrey H A, Vitseva O, et al. The role of platelets in mediating a response to human influenza infection[J]. Nat Commun, 2019, 10(1): 1780.
[33]	Rolling C C, Barrett T J, Berger J S. Platelet-monocyte aggregates: molecular mediators of thromboinflammation[J]. Front Cardiovasc Med, 2023, 10: 960398.
[34]	Liang H, Duan Z J, Li D, et al. Higher levels of circulating monocyte-platelet aggregates are correlated with viremia and increased sCD163 levels in HIV-1 infection[J]. Cell Mol Immunol, 2015, 12(4): 435-443.
[35]	Shantsila E, Lip G Y H. The role of monocytes in thrombotic disorders. Insights from tissue factor, monocyte-platelet aggregates and novel mechanisms[J]. Thromb Haemost, 2009, 102(5): 916-924.
[36]	Kappelmayer J, Kunapuli S P, Wyshock E G, et al. Characterization of monocyte-associated factor V[J]. Thromb Haemost, 1993, 70(2): 273-280.
[37]	Le Guyader A, Davis-Gorman G, Copeland J G, et al. A flow cytometric method for determining the binding of coagulation factor X to monocytes in whole human blood[J]. J Immunol Methods, 2004, 292(1/2): 207-215.
[38]	Toti F, Satta N, Fressinaud E, et al. Scott syndrome, characterized by impaired transmembrane migration of procoagulant phosphatidylserine and hemorrhagic complications, is an inherited disorder[J]. Blood, 1996, 87(4): 1409-1415.
[39]	Yuan Y P, Alwis I, Wu M C L, et al. Neutrophil macroaggregates promote widespread pulmonary thrombosis after gut ischemia[J]. Sci Transl Med, 2017, 9(409): eaam5861.

促凝血小板在感染性血栓形成中的调控作用

Procoagulant platelets regulate H1N1-induced thrombosis formation

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

图/表 4

参考文献 39

相关文章 15

编辑推荐

Metrics

[1]	严治, 吴星玥, 姚卫芹, 颜灵芝, 金松, 商京晶, 施晓兰, 吴德沛, 傅琤琤. 免疫不全麻痹在新诊断多发性骨髓瘤患者中的动态变化及预后意义[J]. 上海交通大学学报（医学版）, 2025, 45(7): 807-814.
[2]	朱晗懿, 石欢, 俞创奇, 郑凌艳. 白介素-1β在极重症口腔颌面部感染中的预警价值及机制初探[J]. 上海交通大学学报（医学版）, 2025, 45(6): 661-672.
[3]	梁效宁, 石亭旺, 陈云丰. 小菌落变异株的致病机制及治疗研究进展[J]. 上海交通大学学报（医学版）, 2025, 45(6): 784-791.
[4]	连明珠, 张常晓, 盛凯, 郭梦, 方姝予. 老年营养风险指数对住院老年2型糖尿病患者发生肺部感染的预测价值[J]. 上海交通大学学报（医学版）, 2025, 45(4): 452-458.
[5]	黄周轩, 邵静波. 慢性原发性免疫性血小板减少症的治疗研究进展[J]. 上海交通大学学报（医学版）, 2025, 45(4): 508-516.
[6]	徐斐翔, 俞凤, 王瑞兰, 宋振举, 童朝阳, 朱长清. 病原宏基因组二代测序在肺部感染所致脓毒症患者中的应用[J]. 上海交通大学学报（医学版）, 2025, 45(2): 169-178.
[7]	高稳, 潘杰松, 赵奕凯, 罗心平, 李剑. 黄嘌呤氧化酶在高尿酸血症患者血小板高反应性中的作用及机制[J]. 上海交通大学学报（医学版）, 2025, 45(12): 1568-1577.
[8]	乔靓, 张婷娟, 冯媛, 杨磊, 钱军, 周静东. 发热伴血小板减少综合征合并噬血细胞综合征2例诊疗分析[J]. 上海交通大学学报（医学版）, 2025, 45(10): 1400-1406.
[9]	PANDIT Roshan, 卢君瑶, 何立珩, 包玉洁, 季萍, 陈颖盈, 许洁, 王颖. 肿瘤坏死因子-α在新型冠状病毒感染合并肾损伤中的作用[J]. 上海交通大学学报（医学版）, 2025, 45(1): 1-10.
[10]	李一白, 崔瑞及, 高珊, 胡嘉晋, 郭晓英. 婴儿晚发型B族链球菌感染及其预防的研究进展[J]. 上海交通大学学报（医学版）, 2024, 44(8): 1044-1049.
[11]	王珂璇, 刘芳, 涂佳佳, 茅一萍. 基于断面时点流行病学调查方法探究医院抗菌药物的使用[J]. 上海交通大学学报（医学版）, 2024, 44(3): 365-372.
[12]	刘嘉榆, 黄方, 郝思国. 慢性粒-单核细胞白血病合并免疫性血小板减少症1例[J]. 上海交通大学学报（医学版）, 2024, 44(2): 287-290.
[13]	朱泽宇, 吕成奇, 刘旭凌, 陈昱璐, 邹德荣, 陆家瑜. A-PRF促进兔膝关节骨软骨损伤愈合的观察[J]. 上海交通大学学报（医学版）, 2024, 44(1): 13-22.
[14]	吴淇琦, 汪豪, 林砺, 晏博, 张舒林. miR-185-5p通过抑制巨噬细胞自噬促进胞内分枝杆菌生长[J]. 上海交通大学学报（医学版）, 2023, 43(6): 699-708.
[15]	闫文月, 李强. 发热伴血小板减少综合征与肾综合征出血热的诊断与鉴别诊断的研究进展[J]. 上海交通大学学报（医学版）, 2023, 43(11): 1457-1462.