According to TCM, damp-heat syndrome is considered a significant syndrome type associated with PC. Therefore, this study utilized a mouse model of damp-heat syndrome to investigate its effects [8]. Two different modeling methods were employed [10, 11], inspired by a study related to climatic alterations in the gut flora of monkeys [23]. The CDHS group was induced with dietary factors alone, while the CC group was modeled with a combination of a high-fat, high-sugar diet and exposure to specific environmental conditions. Both groups exhibited symptoms consistent with damp-heat syndrome, including vigor deficiency, constipation, odor, abdominal fat accumulation, and slow weight gain. Interestingly, the survival rate of the CC group was higher, and colon tissue analysis revealed better outcomes in terms of colitis compared to the CDHS group, suggesting that the combined internal and external factor modeling approach may be more effective than single endogenous modeling.
Furthermore, a PC model was established using Panc02 cells [24]. Subsequent observations revealed a significant increase in tumor volume and weight among mice in the CDHS and CC groups, indicating the promotion of PC growth by damp-heat syndrome. To elucidate the underlying mechanisms, we conducted HE staining, MASSON staining, and immunohistochemistry analyses. The results demonstrated an enlargement in the proportion of cellular substances and an increase in collagenous fibers within the tumor tissue of mice affected by damp-heat syndrome. This observation aligns with the characteristic presence of a rich fibrotic stroma in PC, which is known to contain abundant collagen [25]. Notably, damp-heat syndrome was found to promote the proliferation of cancer-associated fibroblasts, leading to collagen deposition [26]. Cancer-associated fibroblasts play a pivotal role in tumor stroma composition and function, contributing to tumor homeostasis and progression through various mechanisms. These include supporting tumor cell growth by secreting inflammatory ligands, growth factors, chemokines, extracellular vesicles, and participating in extracellular matrix remodeling, which collectively regulate tumor proliferation, invasion, metastasis, therapy resistance [27], immune evasion, and relapse [28].
Subsequently, we turned our attention to the effects of damp-heat syndrome on the intestinal barrier in PC-afflicted mice. The intestinal barrier encompasses both physical and microbial components [29], with tight junction proteins serving as the primary constituents of the physical barrier. These proteins, including Occludin and ZO-1 [30], are transmembrane proteins crucial for maintaining intestinal integrity. In our investigation, histological examination of the colon tissues from the CDHS and CC groups revealed severe disruption of the intestinal mucosal structure. Moreover, a notable decrease in the fluorescence intensity of Occludin and ZO-1 was observed. Collectively, these findings suggest that damp-heat syndrome contributes to the disruption of intestinal mechanical barrier integrity, thereby promoting intestinal inflammation and increased permeability. Additionally, the intestinal biological barrier comprises a layer of the bacterial membrane formed by the attachment of intestinal microorganisms [14]. Our findings also suggest that damp-heat syndrome promotes the overgrowth and altered diversity of intestinal bacteria. Consequently, this leads to an imbalance in intestinal flora homeostasis, culminating in intestinal inflammation and impaired intestinal barrier function.
In our study, OTUs belong to Lactobacillus of control group were higher. Lactobacillus, a probiotic, was effective in normalizing body weight ratios, biochemical markers, cytokines, and intestinal tight junctions [31], and accounted for the highest percentage of intestinal flora in normal mice [32]. In model group, there was a significant increase in the relative abundance for Treponema. Treponema is positively correlated with arachidonic acid metabolism and lipopolysaccharide biosynthesis pathways [33]. Lipopolysaccharide as an important inflammatory mediator [34], can directly induce differential gene expression in PDAC cells, such as PI3K/Akt/mTOR, a known oncogenic driver [35]. In CDHS group, Allobaculum was one of potential flora markers of damp-heat syndrome. Allobaculum is associated with glycerophospholipid metabolism, taurine and hypotaurine metabolism, glycerol ester metabolism, arachidonic acid metabolism, primary bile acid synthesis, and tryptophan metabolism [9]. Coprobacillus excelled in CC group. Coprobacillus is an important butyric acid-producing bacterium and is positively correlated with the expression of immunocompetent genes [36] and lipid metabolism [37]. Therefore, the abundance of Coprobacillus increased in the gut of obese animals [38].
According to RFA and intragroup cumulative abundance, this study found that the intestinal microbial abundance of mice in the CDHS and CC groups was higher. At the order level, Enterobacterales was significantly increased in both groups. Enterobacterales is a type of Gammaproteobacteria. Recent reports have demonstrated that Gammaproteobacteria can produce the bacterial enzyme cytidine deaminase, which can metabolize active Gemcitabine into an inactive form. Gemcitabine is widely used to treat PC. Hence, Enterobacterales might contribute to the chemoresistance of PC [39, 21]. Based on this study, it is hypothesized that damp-heat syndrome might accelerate the growth of Enterobacterales, thereby promoting PC chemoresistance.
Furthermore, this study had the following inadequacies: (1) It only established a correlation, not a causal relationship, between flora and PC in damp-heat syndrome; (2) Mechanism studies related to intestinal flora were not explored in depth; (3) Due to the small sample size included in this study, the results might be biased. Therefore, more samples should be included in subsequent studies.