Introduction :
Systemic lupus erythematosus (SLE) is a complex autoimmune disease manifested by the damage of multiple organs and systems[11[] Relle, Manfred et al. “Epigenetic Aspects of Systemic Lupus Erythematosus.” Rheumatology and therapy vol. 2,1 (2015): 33-46. doi:10.1007/s40744-015-0014-y]. The pathogenesis of SLE is characterized by the abnormal activation of T and B lymphocytes, and the formation of autoantibodies[22[] Lou, Hantao et al. “Autoantibodies in systemic lupus erythematosus: From immunopathology to therapeutic target.” Journal of autoimmunity vol. 132 (2022): 102861. doi:10.1016/j.jaut.2022.102861]. Among which, the imbalance between regulatory T (Treg) cells and follicular helper T (Tfh) cells has attracted much attention[33[] He, Jing et al. “Low-dose interleukin-2 treatment selectively modulates CD4(+) T cell subsets in patients with systemic lupus erythematosus.” Nature medicine vol. 22,9 (2016): 991-3. doi:10.1038/nm.4148]. Tfh cells represent a distinct subset of CD4+ T cells specialized in providing help to B cells and are required for the formation of germinal centers. In lupus, abnormal proliferation and function of Tfh cells will lead to pathological processes like autoantibody production and tissue damage, and thus become a promising therapeutic target [44[] Feng, Xuebing et al. “Inhibition of aberrant circulating Tfh cell proportions by corticosteroids in patients with systemic lupus erythematosus.” PloS one vol. 7,12 (2012): e51982. doi:10.1371/journal.pone.0051982][55[] Wei, Xindi, and Xiaoyin Niu. “T follicular helper cells in autoimmune diseases.” Journal of autoimmunity vol. 134 (2023): 102976. doi:10.1016/j.jaut.2022.102976].
Our previous studies have shown that MSCs could suppress Tfh cell differentiation and proliferation, which was mediated by inducible nitric oxide synthase (iNOS) [6]. Recently, it has been proven that bone marrow-derived mesenchymal stem cells (MSCs) from SLE patients were inefficient to maintain Treg and Tfh balance, while transplantation of healthy MSCs effectively inhibited the expansion of Tfh cells and alleviate lupus symptoms[66[] Zhang, Zhuoya et al. “Human Umbilical Cord Mesenchymal Stem Cells Inhibit T Follicular Helper Cell Expansion Through the Activation of iNOS in Lupus-Prone B6.MRL-Fas Mice.” Cell transplantation vol. 26,6 (2017): 1031-1042. doi:10.3727/096368917X694660][77[] Jang, Eunkyeong et al. “Infusion of Human Bone Marrow-Derived Mesenchymal Stem Cells Alleviates Autoimmune Nephritis in a Lupus Model by Suppressing Follicular Helper T-Cell Development.” Cell transplantation vol. 25,1 (2016): 1-15. doi:10.3727/096368915X688173]. However, this treatment is only effective for a subset of patients[88[] Wang, Dandan et al. “A Long-Term Follow-Up Study of Allogeneic Mesenchymal Stem/Stromal Cell Transplantation in Patients with Drug-Resistant Systemic Lupus Erythematosus.” Stem cell reports vol. 10,3 (2018): 933-941. doi:10.1016/j.stemcr.2018.01.029], for reasons that may be related to the insufficient inhibitory effect or short persistence time[99[] Nie M, Chen G, Zhao C, Gan J, Alip M, Zhao Y, Sun L. Bio-inspired adhesive porous particles with human MSCs encapsulation for systemic lupus erythematosus treatment. Bioact Mater. 2020 Aug 10;6(1):84-90. ]. Therefore, it is necessary to find effective strategies to maximize the therapeutic effect of MSCs.
Silybin (SBI), a flavonoid extracted from the seeds and fruits of silymarin in the compositae[1010[] Křen V, Valentová K. Silybin and its congeners: from traditional medicine to molecular effects. Nat Prod Rep. 2022 Jun 22;39(6):1264-1281. doi: 10.1039/d2np00013j.], is mainly used as a liver-protective drug in clinical practice[1111[] ederico, Alessandro et al. “Evaluation of the Effect Derived from Silybin with Vitamin D and Vitamin E Administration on Clinical, Metabolic, Endothelial Dysfunction, Oxidative Stress Parameters, and Serological Worsening Markers in Nonalcoholic Fatty Liver Disease Patients.” Oxidative medicine and cellular longevity vol. 2019 8742075. 15 Oct. 2019, doi:10.1155/2019/8742075], while studies have shown it has anti-tumor and anti-diabetic effects[1212[] Wei, Zibo et al. “Silybin suppresses ovarian cancer cell proliferation by inhibiting isocitrate dehydrogenase 1 activity.” Cancer science vol. 113,9 (2022): 3032-3043. doi:10.1111/cas.15470][1313[] Liu, Panwen et al. “Silibinin ameliorates STZ-induced impairment of memory and learning by up- regulating insulin signaling pathway and attenuating apoptosis.” Physiology & behavior vol. 213 (2020): 112689. doi:10.1016/j.physbeh.2019.112689]. Recently, evidence has suggested that SBI has a therapeutic effect on collagen-induced arthritis models and patients with rheumatoid arthritis[1414[] Tong, W W et al. “Silibinin alleviates inflammation and induces apoptosis in human rheumatoid arthritis fibroblast-like synoviocytes and has a therapeutic effect on arthritis in rats.” Scientific reports vol. 8,1 3241. 19 Feb. 2018, doi:10.1038/s41598-018-21674-6][1515[] Xie, Ying et al. “Suppression of up-regulated LXRα by silybin ameliorates experimental rheumatoid arthritis and abnormal lipid metabolism.” Phytomedicine : international journal of phytotherapy and phytopharmacology vol. 80 (2021): 153339. doi:10.1016/j.phymed.2020.153339], which opens new perspectives for the treatment of systemic autoimmune diseases. Moreover, SBI could up-regulate the gene expression of MSC-related markers in a pulmonary arterial hypertension rat model[1616[] Zhang, Tingting et al. “Silibinin Upregulates CXCR4 Expression in Cultured Bone Marrow Cells (BMCs) Especially in Pulmonary Arterial Hypertension Rat Model.” Cells vol. 9,5 1276. 21 May. 2020, doi:10.3390/cells9051276], suggesting that it may also have an impact on the function of MSCs. In this study, we systematically observed the efficacy of SBI alone and in combination with MSCs in the treatment of lupus mice for the first time and elaborated its regulatory mechanisms on Tfh cells.