Commentary: The effect of Ganpichengpi diarrhea and Tongxie Yaofang intervention on the intestinal mucosal microbiota and neurochemical substances

Rui LengYichuan Xv^*Jiang Lin

• Department of Gastroenterology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China

Diarrhea is a common gastrointestinal disorder characterized by increased bowel movement frequency, loose stools, and watery stools. In traditional Chinese medicine (TCM), diarrhea associated with Ganqichengpi syndrome (GQCP) is often accompanied by mood disorders and abdominal pain. Current research suggests a close link between this type of diarrhea and intestinal dysbiosis. Recent studies have shown that intestinal flora plays an important role in functional gastrointestinal disorders (Xu et al., 2022;Xv et al., 2024). It has been reported that changes in the composition, structure, and function of the microbiota play a key role in the pathogenesis of diarrhea associated with GQCP (Xu et al., 2022;Zhang et al., 2020).The intestinal mucosa is crucial for nutrient absorption, maintaining microbial balance, and providing barrier protection. The diarrhea with Ganqichengpi syndrom is closely related to the damaged mucosal integrity (Liang et al., 2022;Jia et al., 2024).Tongxie Yaofang (TXYF) is a classic Chinese medicine formula that is widely used to treat GQCP (Liang et al., 2022). Previous studies have revealed its probable pharmacological mechanism like regulating certain intestinal flora, reducing intestinal permeability, and enhancing mucosal barrier function (Xu et al., 2022;Jia et al., 2024).However, it remains unclear how this formula affects the local mucosal microenvironment and neuroimmune interactions.Therefore, investigating how TXYF treatment change the structure, diversity, and abundance of the mucosal microbiota in a mouse model of GQCP diarrhea will help elucidate its mechanism of action. It will also provide valuable insights into identifying potential biomarkers and improving diagnostic and therapeutic strategies. Microbiology exploring the therapeutic mechanisms of TXYF in a mouse model of GQCP. The research focused on the regulatory effects of TXYF on the intestinal mucosal microbiota and key neurochemical factors (Long et al., 2025). To establish a mouse model of GQCP, the researchers used a combined approach: gavage of Folium sennae extract plus physical stressors (restraint and tail-clamping). Results After model establishment, VIP and BDNF levels increased significantly, while 5-HT level decreased significantly. After TXYF intervention, BDNF level remained at a high level. Regarding the intestinal mucosal microbiota, TXYF significantly increased the abundance of Lactobacillus, reduced the abundance of lumen Clostridium and Streptococcus, and restored the balance of the Firmicutes/Bacteroidetes ratio.The study has two notable strengths. First, it innovatively focuses on intestinal mucosal microbiota-a component interacting closely with the host's intestinal homeostasis-which is critical for understanding the pathogenesis of GQCP-related diarrhea and TXYF's efficacy. Second, it adopts an integrative methodological framework, including microbial diversity analysis, taxonomic composition identification, functional prediction via Phylogenetic Investigation of Communities by Reconstruction of Unobserved States 2, and neurochemical detection. This multi-level design strongly supports the idea that TXYF may regulate the gut-brain axis and aid intestinal barrier repair by modulating the microbial ecosystem.However, the study has limitations affecting result interpretability and generalizability, and its discussion on TXYF's action mechanism and related signaling pathways remains superficial. Further exploration of the following mechanisms is needed, combined with existing studies: 1. Short-chain fatty acids (SCFAs) and mechanisms of intestinal barrier function. The TXYF-induced increase in Lactobacillus and decrease in Clostridiaceae and Streptococci may affect the production of microbial metabolites, especially SCFAs such as butyrate, propionate, and acetate. Lactobacillus is a known SCFA producer, and its increased abundance may raise colonic SCFA concentrations. SCFAs serve as intestinal epithelial cell energy sources, enhance barrier function by upregulating tight junction protein (e.g., zonula occludens-1, Occludin) expression, and regulate intestinal inflammatory responses and neuroendocrine function via activating G protein-coupled receptors (e.g., free fatty acid receptor 2/free fatty acid receptor 3) (Dalile et al., 2019;Qiao et al., 2023).Regrettably, the study did not measure SCFA content or intestinal barrier-related protein expression, so it cannot confirm if TXYF acts via the "microbe-SCFA-barrier axis." Subsequent studies should supplement metabolomic (SCFA detection) and immunohistochemical (tight junction protein expression assessment) experiments. 2. 5-HT synthesis and tryptophan metabolism pathway. The model group showed significantly decreased 5-HT levels, which TXYF did not reverse significantly-suggesting 5-HT may not be TXYF's main target, or TXYF regulates serum 5-HT via indirect pathways. Approximately 90% of intestinal 5-HT is synthesized by enterochromaffin cells and is regulated by the gut microbiota (particularly spore-forming bacteria such as Clostridium) (Yano et al., 2015).Tryptophan, the precursor of 5-HT, is metabolized through two pathways: 5-HT synthesis and epinephrine (which is associated with inflammation). Because TXYF alters the gut microbiota, it may interfere with tryptophan metabolism, affecting 5-HT levels. Future studies should examine key molecules in the tryptophan pathway (such as indoleamine 2,3-dioxygenase 1) and colonic 5-HT content. 3. BDNF regulation and microbiota-gut-brain axis signaling. TXYF inhibited the decrease of BDNF levels in the model group, but the regulatory mechanism is unclear. Recent studies have shown that probiotics (e.g., Lactobacillus) can promote BDNF expression via SCFA production or inflammatory response downregulation, affecting central nervous system plasticity and exerting anti-anxiety effects (Suda & Matsuda, 2022). Although studies have detected changes in the gut microbiota and BDNF, whether Lactobacillus mediated changes in BDNF remains unknow. Introducing fecal microbiota transplantation (FMT) or antibiotic-induced microbiota depletion models will help verify the causal relationship between the gut microbiota and BDNF levels. shown a positive correlation between the relative abundance of Lactobacillus and plasma gamma-aminobutyric acid (GABA) levels (Zou et al., 2024). Another study confirmed that administering GABA replicated the effects of vagotomy, which modulates the expression of inflammatory factors such as IL-6 and IL-1β in the gastrointestinal tract and brain, and reduced intestinal inflammation and anxiety-like symptoms in infected mice (Zhao et al., 2022). This suggests that Lactobacillus may affect vagal nerve function by increasing plasma GABA, thereby achieving the goal of treating diarrhea and anxiety. Future investigations should incorporate cytokine profiling and spatial transcriptomics to elucidate localized immune responses.In addition to the insufficient mechanistic investigation, the study also has other problems related to study design. First, the mouse model assessment relied heavily on behavioral symptoms (e.g., irritability, diarrhea) without examining objective molecular markers (e.g., corticosterone, proinflammatory cytokines), which impaired the reproducibility of the model and the reliability of the results. Second, there was a lack of causal validation experiments (e.g., FMT, antibiotic-induced microbiota depletion) to confirm whether changes in the gut microbiota were necessary for the effectiveness of TXYF.To address these limitations and improve our understanding of the mechanisms of TXYF, future studies should: (1) use larger sample cohorts to improve statistical power and generalizability; (2) validate the mouse model of GQCP diarrhea with objective biochemical markers (e.g., cortisol, proinflammatory cytokines) to enhance reliability and reproducibility; (3) use causal designs (e.g., microbiota transplantation, germ-free models) to confirm the causal relationship between the gut microbiota and the efficacy of TXYF; (4) apply multi-omics approaches to identify key functional pathways (e.g., SCFA-receptor [free fatty acid receptor 2/free fatty acid receptor 3] signaling) and clarify how TXYF regulates gut-brain communication through microbial metabolites. Multi-omics strategies can systematically link microbial composition to metabolic output and host responses. For instance, metagenomic sequencing can identify functional genes involved in SCFA production or tryptophan metabolism; metabolomics can quantify associated metabolites (e.g., acetate, butyrate, 5-HT, kynurenine); and transcriptomic/proteomic profiling can reveal downstream host pathways related to barrier function, inflammation, and neuroprotection. However, several methodological limitations should be noted. The evaluation of the animal model relied heavily on behavioral indicators, lacking corroboration through objective molecular or physiological biomarkers. The small sample size may further undermine the statistical power of the conclusions. Although the study reported correlations between microbial composition and BDNF levels, the causal mechanisms remain unaddressed. Importantly, it is still unclear whether TXYF exerts pharmacological effects via specific microbial regulation, such as through Lactobacillus. Moreover, the investigation into TXYF's mechanistic pathways was incomplete. Key aspects such as the role of SCFAs in intestinal barrier function and the link between microbial changes and tryptophan metabolism-a crucial aspect for serotonin synthesis-were not explored. The lack of data on inflammatory markers and a comprehensive analysis of neuro-endocrine-microbial interactions also limited the depth of mechanistic exploration.Future studies should employ multi-omics strategies-such as metagenomics and metabolomics-to identify functionally relevant bacteria and metabolites, including SCFAs. Complementary techniques such as mmunofluorescence can help elucidate intestinal barrier integrity and neural signaling pathways. Additionally, dose-response studies and validation in clinical settings will fertilize standardizing and personalizing TXYF therapy. Finally, mechanistic studies employing FMT and antibiotic-induced depletion models are needed to verify causal relationships, such as the "microbiota-SCFA-barrier" axis, which will strengthen the evidence base for TCM modernization in treating functional gastrointestinal disorders.