ScRNA‐Seq Reveals Jiangfu Zhixie Formula Alleviates IBS‐D by Inhibiting Macrophage M1 Polarization through HIF‐1α‐Glycolysis Axis
Xiaosong Xu, Yao Du, Lipeng Du, Wanning Zhang, Bowen Guo, Yanru Cai, Tianyi ZhangABSTRACT
Background
Irritable bowel syndrome with diarrhea (IBS‐D) remains a challenging condition to manage due to its chronic nature and the limited long‐term efficacy of current treatments. The traditional Chinese herbal formula Jiangfu Zhixie (JFZX) has shown clinical benefits, but its underlying mechanisms remain unclear.
Aim
This study aimed to investigate the therapeutic effects of JFZX on IBS‐D and elucidate its molecular mechanisms, focusing on macrophage polarization and the HIF‐1α‐glycolysis axis.
Methods
An IBS‐D mouse model was established by senna leaf infusion combined with restraint stress. Mice were treated with varying doses of JFZX or pinaverium bromide (positive control) for 14 days. Diarrhea symptoms, visceral hypersensitivity, and intestinal barrier integrity were assessed. Single‐cell RNA sequencing (scRNA‐seq) was performed on colon tissues to analyze cellular composition and intercellular interactions. In vitro, LPS‐stimulated RAW264.7 macrophages were treated with JFZX‐medicated serum to evaluate M1 polarization, cytokine secretion (IL‐1β, TNF‐α), and expression of HIF‐1α and glycolysis‐related proteins (GLUT1, LDHA).
Results
JFZX treatment significantly ameliorated diarrhea, reduced visceral hypersensitivity, and restored intestinal barrier integrity in IBS‐D mice, accompanied by upregulation of tight junction proteins ZO‐1 and Occludin. scRNA‐seq revealed that JFZX reshaped the colonic cellular landscape, attenuating myeloid‐epithelial interactions and reducing the proportion of pro‐inflammatory M1‐like macrophages. Mechanistically, HIF‐1α was identified as a key driver of M1 polarization in IBS‐D. JFZX suppressed HIF‐1α expression and its downstream glycolytic pathway (GLUT1, LDHA) both in vivo and in vitro, thereby inhibiting M1 macrophage polarization and decreasing IL‐1β and TNF‐α secretion in LPS‐stimulated macrophages.
Conclusion
JFZX alleviates IBS‐D by inhibiting M1 macrophage polarization through suppression of the HIF‐1α‐glycolysis axis, highlighting its potential as a therapeutic agent for IBS‐D and identifying HIF‐1α as a promising target for intervention.