Gansu Engineering Laboratory for New Products of Traditional Chinese Medicine, Gansu Key Laboratory of TCM Excavation and Innovative Transformation, Gansu University of Chinese Medicine, Lanzhou, Gansu, 730000, China.
J Ethnopharmacol. 2024 Dec 5;335:118599. doi: 10.1016/j.jep.2024.118599. Epub 2024 Jul 21.
ETHNOPHARMACOLOGICAL RELEVANCE: Shenlin Baizhu Decoction (SLBZD), which comes from 'Taiping Huimin Heji Ju Fang', belongs to a classical prescription for treating spleen deficiency and dampness obstruction (SQDDS)-type ulcerative colitis (UC) in traditional Chinese medicine. However, the mechanism of SLBZD in treating UC with SQDDS remains unclear. AIM OF THE STUDY: This study aims to investigate the mechanism of SLBZD against SQDDS-type UC of based on the "gut microbiota and metabolism - bone marrow" axis to induce endogenous bone marrow mesenchymal stem cells (BMSCs) homing. MATERIALS AND METHODS: Ultra-performance liquid chromatography-mass spectrometry was used to analysis of SLBZD qualitatively. The efficacy of SLBZD in SQDDS-type UC was evaluated based on the following indicators: the body weight, colon length, disease activity index (DAI) score, Haemotoxylin and Eosin (H&E) pathological sections, and intestinal permeability proteins (occluding and ZO-1). 16S rRNA gene sequencing and non-target metabolomics were performed to identify gut microbiota changes and its metabolites in feces, respectively. BMSCs in each group was collected, cultured, and analyzed. Optimal passaged BMSCs were injected by tail vein into UC rats of SQDDS types. BMSCs homing to the colonic mucosal tissue was observed by immunofluorescent. Finally, the repairing effect of BMSCs homing to the colonic mucosal tissue after SLBZD treatment was analyzed by transmission electron microscopy, qRT-PCR, and immunohistochemistry. RESULTS: SLBZD effectively improved the colonic length and the body weight, reduced DAI and H&E scores, and increased the expression of the intestinal permeability proteins, including occluding and ZO-1, to treat SQDDS-type UC. After SLBZD treatment, the α-diversity and β-diversity of the gut microbiota were improved. The differential microbiota was screened as Aeromonadaceae, Lactobacillaceae, and Clostridiaceae at the family level, and Aeromonas, Lactobacillus, Clostridium_sensu_stricto_1 at the genus level. Meanwhile, the main metabolic pathway was the galactose metabolism pathway. SLBZD treatment timely corrected the aberrant levels of β-galactose in peripheral blood and bone marrow, senescence-associate-β-galactosidase in BMSCs, and galactose kinase-2, galactose mutase, and galactosidase beta-1 in peripheral blood to further elevate the expression levels of senescence-associated (SA) proteins (p16, p53, p21, and p27) in BMSCs. The Spearman's correlation analysis demonstrated the relationship between microbiota and metabolism, and the relationship between the galactose metabolism pathway and SA proteins. After BMSCs in each group injection via the tail vein, the pharmacodynamic effects were consistent with those of SLBZD in SQDDS-type UC rats. Furthermore, BMSCs have been homing to colonic mucosal tissue. BMSCs from the SLBZD treatment group had stronger restorative effects on intestinal permeability function due to increasing protein and mRNA expressions of occludin and ZO-1, and decreasing the proteins and mRNA expressions of SDF-1 and CXCR4 in colon. CONCLUSIONS: SLBZD alleviated the damaged structure of gut microbiota and regulated their metabolism, specifically the galactose metabolism, to treat UC of SDDOS types. SLBZD treatment promotes endogenous BMSCs homing to colonic mucosal tissue to repaire the intestinal permeability. The current exploration revealed an underlying mechanism wherein SLBZD activates endogenous BMSCs by targeting 'the gut microbiota and its metabolism-bone marrow' axis and repairs colonic mucosal damage to treat SDDOS-type UC.
民族药理学相关性：参苓白术汤（SLBZD）源自《太平惠民和剂局方》，属于中医治疗脾虚湿阻（SQDDS）型溃疡性结肠炎（UC）的经典方剂。然而，SLBZD 治疗 SQDDS 型 UC 的机制尚不清楚。
研究目的：本研究旨在基于“肠道微生物群和代谢-骨髓”轴诱导内源性骨髓间充质干细胞（BMSCs）归巢，探讨 SLBZD 治疗 SQDDS 型 UC 的机制。
材料和方法：采用超高效液相色谱-质谱联用技术对 SLBZD 进行定性分析。根据以下指标评价 SLBZD 在 SQDDS 型 UC 中的疗效：体重、结肠长度、疾病活动指数（DAI）评分、苏木精和伊红（H&E）病理切片和肠道通透性蛋白（occluding 和 ZO-1）。通过 16S rRNA 基因测序和非靶向代谢组学分别鉴定粪便中肠道微生物群的变化及其代谢物。收集各组 BMSCs，培养并分析。将最佳传代的 BMSCs 通过尾静脉注入 SQDDS 型 UC 大鼠体内。通过免疫荧光观察 BMSCs 向结肠黏膜组织的归巢情况。最后，通过透射电镜、qRT-PCR 和免疫组织化学分析 SLBZD 治疗后 BMSCs 归巢结肠黏膜组织的修复效果。
结果：SLBZD 有效改善了结肠长度和体重，降低了 DAI 和 H&E 评分，增加了肠道通透性蛋白，包括 occluding 和 ZO-1，从而治疗 SQDDS 型 UC。SLBZD 治疗后，肠道微生物群的 α-多样性和 β-多样性得到改善。筛选出的差异菌群为 Aeromonadaceae、Lactobacillaceae 和 Clostridiaceae 科水平，Aeromonas、Lactobacillus、Clostridium_sensu_stricto_1 属水平。同时，主要代谢途径是半乳糖代谢途径。SLBZD 治疗及时纠正了外周血和骨髓中β-半乳糖的异常水平、BMSCs 中的衰老相关-β-半乳糖苷酶以及外周血中的半乳糖激酶-2、半乳糖变位酶和半乳糖苷酶β-1，进一步提高了 BMSCs 中衰老相关（SA）蛋白（p16、p53、p21 和 p27）的表达水平。Spearman 相关分析表明了微生物群和代谢物之间的关系，以及半乳糖代谢途径和 SA 蛋白之间的关系。各组 BMSCs 经尾静脉注射后，药效与 SLBZD 在 SQDDS 型 UC 大鼠中的药效一致。此外，BMSCs 已归巢至结肠黏膜组织。与 SLBZD 治疗组相比，BMSCs 对肠道通透性功能的恢复作用更强，这是由于增加了 occludin 和 ZO-1 的蛋白和 mRNA 表达，降低了 SDF-1 和 CXCR4 的蛋白和 mRNA 表达。
结论：SLBZD 减轻了 SQDDS 型 UC 受损的肠道微生物群结构，并调节了其代谢，特别是半乳糖代谢，以治疗 UC。SLBZD 治疗促进内源性 BMSCs 归巢至结肠黏膜组织，修复肠道通透性。目前的研究揭示了一种潜在的机制，即 SLBZD 通过靶向“肠道微生物群及其代谢-骨髓”轴激活内源性 BMSCs，修复结肠黏膜损伤，从而治疗 SQDDS 型 UC。
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