Immune-enhancing effect and mechanism of natural plant-derived immunostimulatory molecule ophiopogonin

LIU Shulin; WEI Jing; ZHU Baohang; YE Yan; PAN Jiale; ZHAO Anni; SONG Zhen; PENG Liusheng; LI Haibo; SUN Hongwu; ZOU Quanming

doi:10.16016/j.2097-0927.202412021

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Immune-enhancing effect and mechanism of natural plant-derived immunostimulatory molecule ophiopogonin

Basic Medicine | 更新时间：2025-02-27

- Immune-enhancing effect and mechanism of natural plant-derived immunostimulatory molecule ophiopogonin
- Journal of Army Medical University Vol. 47, Issue 4, Pages: 350-359(2025)
- 作者机构：
  
  1.陆军军医大学（第三军医大学）药学与检验医学系微生物与生化药学教研室，国家免疫生物制品工程技术研究中心
  2.佳木斯大学临床医学，黑龙江佳木斯
  3.重庆理工大学药学院，重庆
- 作者简介：
- 基金信息：
  
  the National Key Research and Development Program of China(2021YFC2302603);the General Program of National Natural Science Foundation of China(32070924)
- DOI：10.16016/j.2097-0927.202412021
  CLC： R392.1;R392.5;R931.6
- Received：06 December 2024，
  
  Revised：16 December 2024，
  
  Published：28 February 2025
- 稿件说明：
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刘姝琳,魏静,朱宝行,等.植物源性免疫刺激分子麦冬皂苷的免疫增强效应及初步机制[J].陆军军医大学学报,2025,47(04):350-359.

LIU Shulin,WEI Jing,ZHU Baohang,et al.Immune-enhancing effect and mechanism of natural plant-derived immunostimulatory molecule ophiopogonin[J].J Army Med Univ,2025,47(04):350-359.
刘姝琳,魏静,朱宝行,等.植物源性免疫刺激分子麦冬皂苷的免疫增强效应及初步机制[J].陆军军医大学学报,2025,47(04):350-359. DOI：10.16016/j.2097-0927.202412021.

LIU Shulin,WEI Jing,ZHU Baohang,et al.Immune-enhancing effect and mechanism of natural plant-derived immunostimulatory molecule ophiopogonin[J].J Army Med Univ,2025,47(04):350-359. DOI：10.16016/j.2097-0927.202412021.

摘要

目的

探究植物源性免疫刺激分子麦冬皂苷对以冠状病毒刺突蛋白受体结合域（receptor binding domain， RBD）为抗原的亚单位疫苗的免疫应答增强效应及初步机制。

方法

用CCK-8法评价麦冬皂苷D’（ophiopogonin D’， OPD’）对骨髓来源的树突状细胞（bone marrow-derived dendritic cell， BMDCs）的细胞毒性；将Balb/c雌性小鼠按照随机数字表法分成4组：Control组、RBD组、RBD/OPD’组、RBD/Alum组，使用免疫剂量为抗原5 μg/只、佐剂100 μg/只，按照第 0、21、42天肌肉注射免疫程序进行免疫；监测小鼠体质量、生化指标等评价其体内安全性；用ELISA检测特异性IgG及其亚型抗体滴度；用ELISA试剂盒检测脾细胞上清中细胞因子水平； ELISpot检测脾细胞分泌γ干扰素（interferon-gamma ，IFN-γ）的细胞数量；用激光共聚焦显微镜观察BMDCs对抗原的摄取；流式细胞术定量分析BMDCs对抗原的吞噬能力；用转录组学技术结合生物信息学研究方法，初步探究其增强免疫效应的机制。

结果

当OPD’浓度

5 μg/mL时BMDCs存活率为100%；小鼠单次肌肉注射后除体质量略有下降外，生化指标结果均处于正常范围；疫苗肌肉注射免疫后，RBD/OPD’组血清特异性IgG、IgG1和IgG2a滴度与RBD组相比均显著升高（

0.05）；与RBD组相比， RBD/OPD’组Th1细胞免疫应答的细胞因子IL-1β、TNF-α、IFN-γ显著增高（

0.01），促进淋巴细胞分泌IFN-γ的细胞数量显著增

加（

0.001）；激光共聚焦显微镜发现，OPD’处理后的BMDCs能摄取更多抗原，流式细胞术定量分析结果显示OPD’能显著提高BMDCs的抗原摄取率（

0.01）；转录组学结果显示，与RBD组相比RBD/OPD’组PPAR信号通路显著富集。

结论

麦冬皂苷D’可有效增强RBD亚单位疫苗的免疫应答水平，且其作用机制可能与激活PPAR信号通路有关。

Abstract

Objective

To explore the effect and preliminary mechanism of the plant-derived immunostimulatory molecule

ophiopogonin

on enhancing the immune response of a subunit vaccine with the receptor-binding domain (RBD) of coronavirus spike protein as the antigen.

Methods

CCK-8 assay was used to determine the cytotoxicity of ophiopogonin D' (OPD') on bone marrow-derived dendritic cells (BMDCs). Female Balb/c mice were randomly divided into RBD

RBD/OPD'

RBD/Alum

and control groups. The immunization dose was 5 μg of antigen per mouse and 100 μg of adjuvant per mouse

and immunization was carried out according to the intramuscular injection immunization procedure on days 0

and 42. The titers of specific IgG and its subtype antibodies were detected by ELISA. The cytokine levels in the supernatant of splenocytes were detected using ELISA. The number of splenocytes secreting IFN-γ was detected by ELISpot. Laser confocal microscopy was employed to observe the uptake of antigen by BMDCs. The phagocytic ability of BMDCs for antigen was quantitatively analyzed by flow cytometry. The mechanism of its enhanced immune effect was preliminarily explored using transcriptomics technology combined with bioinformatics research.

Results

When the concentration of OPD' was less than 5 μg/mL

the survival rate of BMDCs was 100%. After a single intramuscular injection in mice

except for a slight decrease in body weight

the other biochemical indicators were within corresponding normal ranges. After intramuscular injection immunization of the vaccine

the titers of serum-specific IgG

IgG1

and

IgG2a in the RBD/OPD' group were significantly higher than those in the RBD group (

0.05). Compared with the RBD group

the RBD/OPD' group induced a high-level Th1 cell immune response of IL-1β

TNF-α

and IFN-γ (

0.01) and had more lymphocytes secreting IFN-γ (

0.001). Laser confocal microscopy displayed that BMDCs took up more antigens after OPD' treatment

which was further confirmed with flow cytometry in quantitative analysis on antigen uptake rate (

0.01). Transcriptomics results indicated that there was more significant enrichment of the PPAR signaling pathway in the RBD/OPD' group than the RBD group

suggesting that OPD' may activate the PPAR signaling pathway to exert its adjuvant effect. Conclusion‍　‍OPD' effectively enhances the immune response of the RBD subunit vaccine

and its action mechanism may be related to the activation of the PPAR signaling pathway.

关键词

Keywords

references

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