防己诺林碱通过调节T淋巴细胞亚群治疗干燥综合征的机制研究

doi:10.3969/j.issn.1674-8115.2026.06.004

上海交通大学学报（医学版） ›› 2026, Vol. 46 ›› Issue (6): 721-731.doi: 10.3969/j.issn.1674-8115.2026.06.004

• 论著 · 基础研究 • 上一篇

防己诺林碱通过调节T淋巴细胞亚群治疗干燥综合征的机制研究

钮疏桐, 石欢, 俞创奇()

上海交通大学医学院附属第九人民医院口腔外科，上海交通大学口腔医学院，国家口腔医学中心，口腔疾病国家临床医学研究中心，上海市口腔医学重点实验室，上海市口腔医学研究所，上海 200011

收稿日期:2025-11-21 接受日期:2026-03-10 出版日期:2026-06-28 发布日期:2026-06-29
通讯作者: 俞创奇，主任医师，博士；电子信箱：yuchuangqi2017@163.com。
基金资助:
国家自然科学基金(82174041);上海交通大学医学院附属第九人民医院生物样本库项目(YBKB202212)

Study on the effects of fangchinoline in regulating T lymphocyte subsets in an animal model of Sjogren's syndrome

Niu Shutong, Shi Huan, Yu Chuangqi()

Department of Oral Surgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine; College of Stomatology, Shanghai Jiao Tong University; National Center for Stomatology; National Clinical Research Center for Oral Diseases; Shanghai Key Laboratory of Stomatology; Shanghai Research Institute of Stomatology, Shanghai 200011, China

Received:2025-11-21 Accepted:2026-03-10 Online:2026-06-28 Published:2026-06-29
Contact: Yu Chuangqi, E-mail: yuchuangqi2017@163.com.
Supported by:
National Natural Science Foundation of China(82174041);Biobank Project of Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine(YBKB202212)

摘要/Abstract

摘要：

目的·阐明防己诺林碱（fangchinoline，FAN）对Nod/LtJ小鼠干燥综合征的疗效，对T细胞亚群的调控作用及潜在分子靶点。方法·将小鼠分为健康对照组、疾病组、羟氯喹治疗组、FAN治疗组。羟氯喹治疗组隔天灌胃给药，FAN治疗组隔日腹腔注射，均给药4周。给药后第5周检测体质量、唾液流率、下颌下腺指数、脾脏指数。用酶联免疫吸附试验（enzyme-linked immunosorbent assay，ELISA）检测外周血抗Ro/SSA抗体、抗La/SSB抗体及免疫球蛋白G（immunoglobulin G，IgG）抗体水平，苏木精-伊红（hematoxylin-eosin，H-E）染色观察下颌下腺淋巴细胞浸润情况，流式细胞术分析脾脏T淋巴细胞亚群分布情况。通过靶点垂钓、PAGE富集、质谱检测寻找FAN的潜在作用靶点，分子模拟对接分析结合能及预测结合位点。结果·接受FAN给药治疗后，Nod/LtJ小鼠唾液流率降低症状得到改善（P=0.046），下颌下腺内淋巴细胞灶性浸润情况显著改善（P=0.044），外周血抗Ro/SSA抗体水平显著降低（P=0.041）。流式细胞术结果提示FAN改变了小鼠脾脏中T淋巴细胞亚群分布，辅助性T细胞1（T helper cell 1，Th1细胞）（P=0.014）、Th17细胞（P=0.011）、滤泡辅助性T细胞（follicular helper T cell，Tfh细胞）（P=0.002）、Th2细胞（P=0.018）分布减少，调节性T细胞（regulatory T cell，Treg细胞）分布增加（P=0.014）。同时，靶点垂钓及质谱鉴定结果均指向丙酮酸激酶M2型（pyruvate kinase M2，PKM2）是FAN的潜在靶点，分子模拟对接提示FAN与PKM2具有稳定的相互作用。结论·FAN可有效缓解Nod/LtJ小鼠的干燥综合征症状，其作用可能与其对特定的T细胞亚群分布的调控相关，FAN可能通过靶向PKM2发挥作用。

关键词: 干燥综合征, 防己诺林碱, T淋巴细胞, Nod/LtJ小鼠, 丙酮酸激酶M2型

Abstract:

Objective ·To elucidate the therapeutic effects of fangchinoline (FAN) on Sjogren's syndrome in Nod/LtJ mice, its regulatory effects on T cell subsets, and its potential molecular targets. Methods ·The mice were divided into four groups: control group, disease group, hydroxychloroquine (HCQ)-treated group, and FAN-treated group. The HCQ-treated group received oral gavage every other day, and the FAN-treated group received intraperitoneal injections every other day, both for 4 weeks. Body weight, salivary flow rate, submandibular gland index, and splenic index were measured in the fifth week. Levels of anti-Ro/SSA, anti-La/SSB, and immunoglobulin G (IgG) antibodies in the peripheral blood were detected by enzyme-linked immunosorbent assay (ELISA). Lymphocyte infiltration in the submandibular glands was assessed by hematoxylin-eosin (HE) staining. The distribution of splenic T lymphocyte subsets was analyzed by flow cytometry. Potential targets of FAN were identified by target fishing, PAGE enrichment, and mass spectrometry. Molecular docking was performed to analyze binding affinity and predict binding sites. Results ·FAN treatment significantly improved salivary flow reduction in Nod/LtJ mice (P=0.046) and markedly alleviated lymphocyte infiltration in the submandibular glands (P=0.044). The levels of anti-Ro/SSA antibodies in peripheral blood were significantly reduced (P=0.041). Flow cytometry results indicated that FAN altered the distribution of splenic T lymphocyte subsets. T helper 1 (Th1) (P=0.014), T helper 17 (Th17) (P=0.011), T follicular helper (Tfh) (P=0.002), and T helper 2 (Th2) (P=0.018) cells were decreased, and regulatory T cells (Treg) (P=0.014) were increased. Furthermore, target fishing coupled with mass spectrometry identified pyruvate kinase M2 (PKM2) as a potential target for FAN. Molecular docking simulations indicated a stable interaction between FAN and PKM2. Conclusion ·FAN exerts discernible therapeutic effects on Sjogren's syndrome symptoms in Nod/LtJ mice. These effects are associated with modulation of T lymphocyte subset distribution and may be mediated through PKM2. These findings highlight the promising therapeutic potential of FAN in managing Sjogren's syndrome.

Key words: Sjogren's syndrome, fangchinoline (FAN), T lymphocytes, Nod/LtJ mice, pyruvate kinase M2 (PKM2)

中图分类号:

R593.2

钮疏桐, 石欢, 俞创奇. 防己诺林碱通过调节T淋巴细胞亚群治疗干燥综合征的机制研究[J]. 上海交通大学学报（医学版）, 2026, 46(6): 721-731.

Niu Shutong, Shi Huan, Yu Chuangqi. Study on the effects of fangchinoline in regulating T lymphocyte subsets in an animal model of Sjogren's syndrome[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2026, 46(6): 721-731.

图/表 7

图1 各组小鼠脾脏T淋巴细胞亚群荧光标记及圈门策略

Fig 1 Fluorescence labeling and gating strategy for splenic T lymphocyte subsets in different groups

表1 RT-qPCR引物序列

Tab 1 Primer sequences for RT-qPCR

Primer	Sequence
β-actin-Forward	CACCATTGGCAATGAGCGGTTC
β-actin-Reverse	AGGTCTTTGCGGATGTCCACGT
LDHA-Forward	GGATCTCCAACATGGCAGCCTT
LDHA-Reverse	AGACGGCTTTCTCCCTCTTGCT
DLD-Forward	CTCATGGCCTACAGGGACTTT
DLD-Reverse	GCATGTTCCACCAAGTGTTTCAT
ACAT1-Forward	CCAGCCACTAAGCTTGGTTCCA
ACAT1-Reverse	GTAGGAGCTTGTCCTTCACCTC

图2 各组Nod/LtJ小鼠的一般指标Note： A.Weight of ICR mice and Nod/LtJ mice from 8th week to 12th week. B. The salivary flow rate in the 12th week. C/D. The gland index (C) and splenic index (D) of mice in the indicated groups. E—G. The levels of anti-Ro/SSA (E), anti-La/SSB (F), and IgG (G) antibodies in the plasma of mice(n=4).

Fig 2 General indicators of Nod/LtJ mice in the indicated groups

图3 各组Nod/LtJ小鼠的下颌下腺局部炎性浸润情况Note： A. Histological assessment of submandibular glands by HE staining. B/C. Quantification of lymphocyte infiltration foci (B) and histological scores (C) in submandibular glands among the indicated groups(n=4).

Fig 3 Inflammatory infiltration in the submandibular glands of Nod/LtJ mice in the indicated groups

图4 各组Nod/LtJ小鼠的脾脏T淋巴细胞分布情况Note： A. Representative flow cytometry images of T cell subsets in the indicated groups. B. The percentages of CD4+T cells, Treg cells, Tfh cells, Th1 cells, Th2 cells, and Th17 cells in the indicated groups.

Fig 4 Distribution of splenic T lymphocytes in Nod/LtJ mice in the indicated groups.

图5 FAN对过度活化T淋巴细胞的作用通路Note： A. IC50 value of FAN. B. Principal component analysis. C. Counts of differentially expressed genes (DEGs). D. KEGG enrichment analysis. E. GO enrichment analysis. F‒H. mRNA expression levels of LDHA (F), DLD (G), and ACAT1 (H) in Jurkat T cells.

Fig 5 Signaling pathways of FAN in overactivated T lymphocytes

图6 FAN作用于过度活化T淋巴细胞的潜在靶点Note： A. Chemical synthesis of the biotinylated probe Biotin-FAN. B. CCK-8 assay of Jurkat T cells in the three groups. C. Schematic diagram of molecular docking between FAN and PKM2.

Fig 6 Potential targets of FAN in overactivated T lymphocytes

参考文献 37

[1]	Fox R I. Sjögren's syndrome[J]. Lancet, 2005, 366(9482): 321-331.
[2]	Mavragani C P, Moutsopoulos H M. Sjögren's syndrome: old and new therapeutic targets[J]. J Autoimmun, 2020, 110: 102364.
[3]	Zhan Q P, Zhang J N, Lin Y B, et al. Pathogenesis and treatment of Sjogren's syndrome: review and update[J]. Front Immunol, 2023, 14: 1127417.
[4]	Ramos-Casals M, Brito-Zerón P, Sisó-Almirall A, et al. Primary sjögren syndrome[J]. Praxis, 2012, 101(24): 1565-1571.
[5]	Baldini C, Fulvio G, La Rocca G, et al. Update on the pathophysiology and treatment of primary Sjögren syndrome[J]. Nat Rev Rheumatol, 2024, 20(8): 473-491.
[6]	Beydon M, McCoy S, Nguyen Y, et al. Epidemiology of sjögren syndrome[J]. Nat Rev Rheumatol, 2024, 20(3): 158-169.
[7]	Yao Y, Ma J F, Chang C, et al. Immunobiology of T cells in sjögren's syndrome[J]. Clin Rev Allergy Immunol, 2021, 60(1): 111-131.
[8]	Singh N, Cohen P L. The T cell in Sjogren's syndrome: force majeure, not spectateur[J]. J Autoimmun, 2012, 39(3): 229-233.
[9]	Wang Y H, Li W Y, McDermott M, et al. IFN-γ-producing T_H1 cells and dysfunctional regulatory T cells contribute to the pathogenesis of Sjögren's disease[J]. Sci Transl Med, 2024, 16(778): eado4856.
[10]	Xu D, Zhao S, Li Q, et al. Characteristics of Chinese patients with primary Sjögren's syndrome: preliminary report of a multi-centre registration study[J]. Lupus, 2020, 29(1): 45-51.
[11]	Wang X, Zhang T, Guo Z Z, et al. The efficiency of hydroxychloroquine for the treatment of primary Sjögren's syndrome: a systematic review and meta-analysis[J]. Front Pharmacol, 2021, 12: 693796.
[12]	Saraux A, Pers J O, Devauchelle-Pensec V. Treatment of primary sjögren syndrome[J]. Nat Rev Rheumatol, 2016, 12(8): 456-471.
[13]	Sun P, Zhu L L, Yu Y, et al. Combination of Astragalus-Salvia and Ophiopogon-Dendrobium herb pairs alleviates Sjögren's Syndrome via inhibiting the JAK1/STAT3 and PI3K/AKT pathways in NOD/Ltj mice[J]. Chin J Nat Med, 2025, 23(6): 733-741.
[14]	王蓉, 马腾茂, 刘飞, 等. 防己的药理作用及临床应用研究进展[J]. 中国中药杂志, 2017, 42(4): 634-639.
	Wang R, Ma T M, Liu F, et al. Research progress on pharmacological action and clinical application of Stephania Tetrandrae Radix[J]. China J Chin Mater Med, 2017, 42(4): 634-639.
[15]	Jiang Y P, Liu M, Liu H T, et al. A critical review: traditional uses, phytochemistry, pharmacology and toxicology of Stephania tetrandra S. Moore (Fen Fang Ji)[J]. Phytochem Rev, 2020, 19(2): 449-489.
[16]	Shan L, Tong L, Hang L, et al. Fangchinoline supplementation attenuates inflammatory markers in experimental rheumatoid arthritis-induced rats[J]. Biomedecine Pharmacother, 2019, 111: 142-150.
[17]	Shao Y X, Fu J Y, Zhan T L, et al. Inhibition of CD4⁺ T cells by fanchinoline via miR506-3p/NFATc1 in Sjögren's syndrome[J]. Inflammopharmacology, 2023, 31(5): 2431-2443.
[18]	刘维, 岳青云, 陈常青, 等. 中医药在干燥综合征治疗中的应用[J]. 中草药, 2024, 55(10): 3516-3528.
	Liu W, Yue Q Y, Chen C Q, et al. Application of traditional Chinese medicine in treatment of Sjögren's syndrome[J]. Chin Tradit Herb Drugs, 2024, 55(10): 3516-3528.
[19]	曹策, 李玲美, 訾明杰, 等. 医学研究中动物实验样本量的确定方法[J]. 中国比较医学杂志, 2023, 33(2): 99-105.
	Cao C, Li L M, Zi M J, et al. Methods for determining animal sample sizes in medical experimental studies[J]. Chin J Comp Med, 2023, 33(2): 99-105.
[20]	Gu F, Xu S X, Zhang P Y, et al. CP-25 alleviates experimental Sjögren's syndrome features in NOD/LtJ mice and modulates T lymphocyte subsets[J]. Basic Clin Pharmacol Toxicol, 2018, 123(4): 423-434.
[21]	Song Y, Li J, Wu Y Z. Evolving understanding of autoimmune mechanisms and new therapeutic strategies of autoimmune disorders[J]. Signal Transduct Target Ther, 2024, 9(1): 263.
[22]	Ríos-Ríos W J, Sosa-Luis S A, Torres-Aguilar H. T cells subsets in the immunopathology and treatment of sjogren's syndrome[J]. Biomolecules, 2020, 10(11): 1539.
[23]	He J, Chen J L, Miao M, et al. Efficacy and safety of low-dose interleukin 2 for primary Sjögren syndrome: a randomized clinical trial[J]. JAMA Netw Open, 2022, 5(11): e2241451.
[24]	Vendel A C, Calemine-Fenaux J, Izrael-Tomasevic A, et al. B and T lymphocyte attenuator regulates B cell receptor signaling by targeting Syk and BLNK[J]. J Immunol, 2009, 182(3): 1509-1517.
[25]	Choi H S, Kim H S, Min K R, et al. Anti-inflammatory effects of fangchinoline and tetrandrine[J]. J Ethnopharmacol, 2000, 69(2): 173-179.
[26]	Shen Y C, Chou C J, Chiou W F, et al. Anti-inflammatory effects of the partially purified extract of Radix Stephaniae tetrandrae: comparative studies of its active principles tetrandrine and fangchinoline on human polymorphonuclear leukocyte functions[J]. Mol Pharmacol, 2001, 60(5): 1083-1090.
[27]	Jiang F Q, Ren S, Chen Y D, et al. Fangchinoline exerts antitumour activity by suppressing the EGFR-PI3K/AKT signalling pathway in colon adenocarcinoma[J]. Oncol Rep, 2021, 45(1): 139-150.
[28]	Gülçin I, Elias R, Gepdiremen A, et al. Antioxidant activity of bisbenzylisoquinoline alkaloids from Stephania rotunda: cepharanthine and fangchinoline[J]. J Enzyme Inhib Med Chem, 2010, 25(1): 44-53.
[29]	Kim D E, Min J S, Jang M S, et al. Natural bis-benzylisoquinoline alkaloids-tetrandrine, fangchinoline, and cepharanthine, inhibit human coronavirus OC43 infection of MRC-5 human lung cells[J]. Biomolecules, 2019, 9(11): 696.
[30]	Villa T, Kim M, Oh S. Fangchinoline has an anti-arthritic effect in two animal models and in IL-1β-stimulated human FLS cells[J]. Biomol Ther, 2020, 28(5): 414-422.
[31]	Psianou K, Panagoulias I, Papanastasiou A D, et al. Clinical and immunological parameters of Sjögren's syndrome[J]. Autoimmun Rev, 2018, 17(10): 1053-1064.
[32]	Maehara T, Moriyama M, Hayashida J N, et al. Selective localization of T helper subsets in labial salivary glands from primary Sjögren's syndrome patients[J]. Clin Exp Immunol, 2012, 169(2): 89-99.
[33]	Katsifis G E, Moutsopoulos N M, Wahl S M. T lymphocytes in Sjögren's syndrome: contributors to and regulators of pathophysiology[J]. Clin Rev Allergy Immunol, 2007, 32(3): 252-264.
[34]	Bustamante M F, Oliveira P G, Garcia-Carbonell R, et al. Hexokinase 2 as a novel selective metabolic target for rheumatoid arthritis[J]. Ann Rheum Dis, 2018, 77(11): 1636-1643.
[35]	Sharabi A, Tsokos G C. T cell metabolism: new insights in systemic lupus erythematosus pathogenesis and therapy[J]. Nat Rev Rheumatol, 2020, 16(2): 100-112.
[36]	Angiari S, Runtsch M C, Sutton C E, et al. Pharmacological activation of pyruvate kinase M2 inhibits CD4⁺ T cell pathogenicity and suppresses autoimmunity[J]. Cell Metab, 2020, 31(2): 391-405.e8.
[37]	Zhao Z H, Wang Y, Gao Y H, et al. The PRAK-NRF2 axis promotes the differentiation of Th17 cells by mediating the redox homeostasis and glycolysis[J]. Proc Natl Acad Sci U S A, 2023, 120(19): e2212613120.

防己诺林碱通过调节T淋巴细胞亚群治疗干燥综合征的机制研究

Study on the effects of fangchinoline in regulating T lymphocyte subsets in an animal model of Sjogren's syndrome

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

图/表 7

参考文献 37

相关文章 15

编辑推荐

Metrics

[1]	刘楚萱, 左佳鑫, 熊屏. 基于超声评分参数及临床指标的列线图鉴别原发性干燥综合征与IgG4相关唾液腺炎[J]. 上海交通大学学报（医学版）, 2025, 45(3): 373-380.
[2]	李燕香, 石欢, 俞创奇. 青蒿琥酯对自发性干燥综合征NOD/Ltj小鼠的治疗作用[J]. 上海交通大学学报（医学版）, 2025, 45(10): 1279-1287.
[3]	姚莉, 田沃土, 曹立. 以神经系统损伤为首发症状的2例原发性干燥综合征报道[J]. 上海交通大学学报（医学版）, 2024, 44(6): 795-800.
[4]	李博, 胡秋侠, 吴系美, 佘若男, 谭锦辉, 罗俊佳, 杨海涛, 张皓茹. 类风湿关节炎患者CD4⁺T淋巴细胞中miR-146a的表达及其与炎性细胞因子的相关性[J]. 上海交通大学学报（医学版）, 2024, 44(10): 1249-1254.
[5]	李自云, 庄新娟, 季业, 田海荣, 殷峻. 黄芪对甲状腺功能正常的桥本甲状腺炎患者外周血T淋巴细胞亚群表达的影响[J]. 上海交通大学学报（医学版）, 2024, 44(1): 108-115.
[6]	杨辰玲, 周慧芳. PD-1/PD-L1通路在自身免疫性眼病中作用的研究进展[J]. 上海交通大学学报(医学版), 2022, 42(1): 107-112.
[7]	赫玮, 左勇. 冬凌草甲素上调PLK1对Jurkat细胞细胞周期的影响[J]. 上海交通大学学报(医学版), 2021, 41(5): 603-611.
[8]	郝健亨，陈浩，高艳，徐慧超，高玉亭，苗宇船，郝慧琴，刘杨. 柴胡皂苷d对自身免疫性肝炎小鼠差异表达基因CTLA-4、IL-10和IL-17的影响[J]. 上海交通大学学报（医学版）, 2020, 40(3): 303-.
[9]	王领1, 2，张娜1，王艳玲1 ，赵福涛1. 原发性干燥综合征患者唇腺组织水通道蛋白5的表达及意义[J]. 上海交通大学学报(医学版), 2020, 40(10): 1425-1429.
[10]	赵乙汜1，余应喜1，林时辉2，徐昉1, 2. T细胞免疫在脓毒症继发侵袭性真菌感染中作用的研究进展[J]. 上海交通大学学报（医学版）, 2019, 39(11): 1325-.
[11]	吴菁 1，范凯健 1, 2，王婷玉 1. 二氢杨梅素对类风湿性关节炎大鼠脾脏免疫功能的调节作用[J]. 上海交通大学学报（医学版）, 2018, 38(12): 1401-.
[12]	汤梦婕，袁晓军，汤静燕，等. 多中心治疗儿童急性T淋巴细胞性白血病的长期疗效评价及随访报告[J]. 上海交通大学学报（医学版）, 2014, 34(11): 1605-.
[13]	卿恺, 陈芳源, 沈莉菁, 等. 转Notch1基因T淋巴细胞白血病双荧光示踪斑马鱼模型的建立及鉴定[J]. , 2013, 33(3): 257-.
[14]	余奇，许迅. 实验性自身免疫性葡萄膜炎模型的研究进展[J]. 上海交通大学学报（医学版）, 2013, 33(10): 1411-.
[15]	肖世金, 董倩, 赵爱民. 自然流产母胎界面嗜酸粒细胞趋化因子、T淋巴细胞激活上调性表达分泌因子S及其受体的表达[J]. , 2011, 31(8): 1170-.