壳聚糖-苯二酚修饰灭活PEDV的口服免疫效果评价
作者:
作者单位:

四川农业大学动物医学院,成都 611130

作者简介:

杨云涵,E-mail: yhyang123000@163.com;

通讯作者:

万红平,E-mail: hpwan@sicau.edu.cn

中图分类号:

S852.659.6

基金项目:

国家自然科学基金项目(2122999007);中央引导地方科技发展专项(2212129410)


Evaluating oral immunization of inactivated PEDV vaccine modified with chitosan-catechol
Author:
Affiliation:

College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130, China

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    摘要:

    为评估壳聚糖-苯二酚(chitosan-catechol,Chic)修饰的灭活猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)疫苗的口服免疫效果,通过控制Chic的量制备出由不同质量浓度Chic(0.2、0.4 mg/mL)包裹的灭活PEDV(0.5 mg/mL),命名为C1+P和C2+P,建立小鼠模型进行免疫试验。采用间接酶联免疫吸附试验(ELISA)分别检测小鼠黏膜sIgA和血清IgG、IgG1、IgG2a抗体水平,并通过流式细胞术和CCK-8试验检测脾细胞的分型和增殖活化情况。结果显示,经2次口服免疫后,C2+P组诱导的免疫效果与昂贵的霍乱毒素B亚单位(CTB)佐剂组效果相当;与未经Chic修饰的PEDV组相比,C1+P、C2+P组诱导的鼻黏膜、肺泡、小肠黏膜特异性sIgA的抗体效价分别提高了6.09、6.37倍,5.35、5.65倍和5.64、5.89倍,血清特异性IgG、IgG1、IgG2a分别提高了8.83、9.65倍,13.54、15.07倍和4.27、5.07倍;CD4+T淋巴细胞、CD8+T淋巴细胞比例分别提升了1.15、1.17倍和1.11、1.11倍;脾细胞的增殖活化情况也显著增加(P<0.05)。结果表明,口服免疫C1+P和C2+P成功诱导了小鼠体内黏膜免疫和系统性免疫应答效应,显示出Chic作为黏膜免疫佐剂的潜力,表明Chic修饰的灭活PEDV疫苗在有效诱导免疫应答方面具有可行性。

    Abstract:

    Inactivated PEDV (0.5 mg/mL) encapsulated with different concentrations of chitosan-catechol(Chic) including 0.2 mg/mL and 0.4 mg/mL named as C1+P and C2+P was prepared by controlling the amount of Chic, and a mouse model was established for immunization testing to evaluate the oral immunization efficacy of inactivated porcine epidemic diarrhea virus (PEDV) vaccine modified with Chic. Indirect enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of sIgA in mouse mucosa and IgG, IgG1, IgG2a antibodies in serum. Flow cytometry and CCK-8 assay were used to detect the phenotyping and proliferation activation of spleen cells. The results showed that the immune response induced by the C2+P group was comparable to that of the expensive cholera toxin B subunit (CTB) adjuvant group after two oral immunizations. Compared with the PEDV group without Chic modification, the antibody titer of nasal,lung,and small intestine mucosa-specific sIgA in the C1+P group and C2+P group was increased by 6.09, 6.37 times, and 5.35, 5.65 times, and 5.64, 5.89 times, respectively. The serum specific IgG, IgG1, and IgG2a in the C1+P group and C2+P group increased by 8.83, 9.65 times, 13.54, 15.07 times, and 4.27, 5.07 times, respectively. The proportion of CD4+ T lymphocytes and CD8+ T lymphocytes in the C1+P group and C2+P group increased by 1.15, 1.17 times and 1.11,1.11 times, respectively. The proliferation and activation of spleen cells significantly increased as well (P<0.05). It is indicated that oral immunization with C1+P and C2+P successfully induced mucosal and systemic immune responses in mice, demonstrating the potential of Chic as a mucosal immune adjuvant and the feasibility of inactivated PEDV vaccines modified with Chic in effectively inducing immune responses.

    图1 PEDV的分离纯化及PEDV、C1+P、C2+P透射电镜图Fig.1 Purification of PEDV and TEM images of PEDV,C1+P and C2+P
    图2 黏膜sIgA抗体效价Fig.2 Mucosa sIgA titers
    图3 血清IgG抗体效价Fig.3 Serum IgG titers
    图4 流式细胞术检测脾脏T淋巴细胞分型Fig.4 Spleen T lymphocyte typing test by FCM
    图5 脾细胞增殖活化情况Fig.5 Splenocyte proliferation
    表 1 PEDV、C1+P、C2+P 粒径及电位Table 1 Diameters and zeta potentials of PEDV,C1+P and C2+P
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杨云涵,邓锴,涂志文,赵兴洪,万红平.壳聚糖-苯二酚修饰灭活PEDV的口服免疫效果评价[J].华中农业大学学报,2025,44(1):217-224

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  • 收稿日期:2024-07-28
  • 在线发布日期: 2025-03-03
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