A chronic stress-induced microbiome perturbation, highly enriched in Ruminococcaceae_UCG-014, promotes colorectal cancer growth and metastasis.

Purpose: Mounting evidence indicates that psychological stress adversely affects cancer progression including tumor growth and metastasis. The aim of this study was to investigate the role of chronic stress-induced microbiome perturbation in colorectal cancer (CRC) progression. Methods: Chronic restraint stress (CRS) was used to establish the chronic stress mouse model, behavioral tests were used for the CRS model evaluation. Subcutaneous xenograft model and lung metastasis model were established to investigate the growth and metastasis of CRC promoted by CRS exposure. 16S rRNA gene sequencing and liquid chromatograph-mass spectrometer (LC-MS) were applied to observe the effects of CRS exposure on the alteration of the gut microbiome and microbial metabolites. Bioinformatics analysis and correlation analyses were applied to analyse the changes in the frequency of body mass, tumor volume, inflammatory factors, neuroendocrine hormones and metabolites of the gut microbiota. Results: In this study, we identifed that CRS exposure model was appropriately constructed by achieving expected increases in disease activity index and enhanced depressive-like behaviors. CRS exposure can promote growth and metastasis of CRC. Besides, the data indicated that CRS exposure not only increased the neuro- and immune-inflammation, but also weakened the gut mucosal immunological function. The 16s rRNA gene sequencing data showed that CRS exposure increased the abundance of g_Ruminococcaceae_UCG_014. Furthermore, the LC-MS data indicated that with only 2 exceptions of carpaine and DG (15:0/20:4(5Z,8Z,11Z,14Z)/0:0), the majority of these 24 metabolites were less abundant in CRS-exposed mice. Bioinformatics analysis and correlation analyses indicated that only Ruminoscoccaceae-UCG-014 was significantly associated with inflammation (IL-6), neurotransmission (5-HT), and microbial metabolism (PS). Conclusion: CRS exposure altered diversity, composition and metabolites of the gut microbiome, with Ruminococcaceae_UCG-014 perturbation consistently correlated to inflammatory responses, suggesting a particular role of this bacterial genus in CRC growth and metastasis.

Tong-Xie-Yao-Fang strengthens intestinal feedback control of bile acid synthesis to ameliorate irritable bowel syndrome by enhancing bile salt hydrolase-expressing microbiota.

ETHNOPHARMACOLOGICAL RELEVANCE: A herbal formula Tong-Xie-Yao-Fang (TXYF) is traditionally used to treat irritable bowel syndrome (IBS), modern pharmacological evidence supports that the formula efficacy is associated with altered gut microbiota. Yet, the mechanistic role of gut microbiota in the therapy of TXYF remains unclear. We previously clarified that gut microbiota-dysregulated bile acid (BA) metabolism contribute to the pathogenesis of IBS, deriving a hypothesis that microbiota-BA metabolic axis might be a potential target of TXYF. AIM OF THE STUDY: We aim to investigate a new gut microbiota-mediated mechanism underlying anti-IBS efficacy of TXYF. MATERIALS AND METHODS: We established an IBS rat model with a combination of stressors, compared the herbal efficacy in models undergone gut bacterial manipulations, also examined BA metabolism-related microbiota, metabolites, genes and proteins by 16S rRNA gene sequencing, targeted metabolomics, qPCR and multiplex immunofluorescence staining. RESULTS: We observed that TXYF attenuated visceral hyperalgesia and diarrhea in IBS rats but not in those underwent gut bacteria depletion. Transferring gut microbiota from TXYF-treated donors also decreased visceral sensitivity and slightly relief diarrhea-like behaviors in IBS recipient rats. Fecal 16S rRNA gene sequencing revealed that TXYF modulated microbial ß-diversity and taxonomic structure of IBS rats, with a significant increase in relative abundance of bile salt hydrolase (BSH)-expressing Bacteroidaceae. qPCR and culturing data validated that TXYF had a promotive effect on the growth and BSH activity of Bacteroides species. TXYF-reshaped microbiota upregulated the expression of intestinal Fgf15, a feedback signal to control BA synthesis in the liver. As a result, the BA synthetic and excretory levels in IBS rats were decreased by TXYF, so as that colonic BA membrane receptor Tgr5 sensing and its mediated Calcitonin gene-related peptide (Cgrp)-positive neuronal response were attenuated. CONCLUSION: This study poses a new microbiota-driven therapeutic action for TXYF, highlighting the potential of developing new anti-IBS strategies from the herbal formula targeting BSH-expressing gut bacteria.

The Landscape of Gut Microbiota and Its Metabolites: A Key to Understanding the Pathophysiology of Pattern in Chinese Medicine.

Liver Stagnation and Spleen Deficiency (LSSD) is a Chinese Medicine (CM) pattern commonly observed in gastrointestinal (GI) diseases, yet its biological nature remains unknown. This limits the global use of CM medications for treating GI diseases. Recent studies emphasize the role of gut microbiota and their metabolites in the pathogenesis and treatment of LSSD-associated GI diseases. There is increasing evidence supporting that an altered gut microbiome in LSSD patients or animals contributes to GI and extra-intestinal symptoms and affects the effectiveness of CM therapies. The gut microbiota is considered to be an essential component of the biological basis of LSSD. This study aims to provide an overview of existing research findings and gaps for the pathophysiological study of LSSD from the gut microbiota perspective in order to understand the relationship between the CM pattern and disease progression and to optimize CM-based diagnosis, prevention, and therapy.

Altered gut microbiota-host bile acid metabolism in IBS-D patients with liver depression and spleen deficiency pattern.

BACKGROUND: Dysregulation of gut microbiota-host bile acid (BA) co-metabolism is a critical pathogenic factor of diarrhea-predominant irritable bowel syndrome (IBS-D). Traditional Chinese Medicine (TCM), instructed by pattern differentiation, is effective in treating IBS-D, in which liver depression and spleen deficiency (LDSD) is the most prevalent pattern. Still, it is unclear the linkage between the LDSD pattern and the BA metabolic phenotype. PURPOSE: This study aimed to uncover the biological basis of the LDSD pattern from the BA metabolic perspective. METHODS: Patients with IBS-D completed questionnaires regarding the irritable bowel severity scoring system (IBS-SSS), stool frequency, Stool Bristol scale, and Self-Rating Scales of mental health. Fasting blood and morning feces were collected to analyze the gut metagenome and BA-related indices/metabolites. RESULTS: IBS-D patients with LDSD had a higher incidence of BA overexcretion (41% vs. 23% non-LDSD) with significant elevations in fecal total BAs and serum BA precursor 7α-hydroxy-4-cholesten-3-one levels. Compared to controls or non-LDSD patients, LDSD patients had a featured fecal BA profile, with higher proportions of deoxycholic acid (DCA), 7-ketodeoxycholic acid, and lithocholic acid. It is consistent with the BA-metabolizing genomic changes in the LDSD gut microbiota characterized by overabundances of 7-dehydroxylating bacteria and BA-inducible genes (baiCD/E/H). The score of bowel symptoms (stool frequency and abdominal pain) showing greater severity in the LDSD pattern were positively correlated with bai-expressing bacterial abundances and fecal DCA levels separately. CONCLUSION: We clarified a differed BA metabolic phenotype in IBS patients with LDSD, which closely correlates with the severity of bowel symptoms. It demonstrates that gut microbiota and host co-metabolism of BAs would provide crucial insight into the biology of the LDSD pattern and its internal relationship with IBS progression.

A High Hepatic Uptake of Conjugated Bile Acids Promotes Colorectal Cancer-Associated Liver Metastasis.

The liver is the most common site for colorectal cancer (CRC)-associated metastasis. There remain unsatisfactory medications in liver metastasis given the incomplete understanding of pathogenic mechanisms. Herein, with an orthotopic implantation model fed either regular or high-fat diets (HFD), more liver metastases were associated with an expansion of conjugated bile acids (BAs), particularly taurocholic acid (TCA) in the liver, and an increased gene expression of Na+-taurocholate cotransporting polypeptide (NTCP). Such hepatic BA change was more apparently shown in the HFD group. In the same model, TCA was proven to promote liver metastases and induce a tumor-favorable microenvironment in the liver, characterizing a high level of fibroblast activation and increased proportions of myeloid-derived immune cells. Hepatic stellate cells, a liver-residing source of fibroblasts, were dose-dependently activated by TCA, and their conditioned medium significantly enhanced the migration capability of CRC cells. Blocking hepatic BA uptake with NTCP neutralized antibody can effectively repress TCA-triggered liver metastases, with an evident suppression of tumor microenvironment niche formation. This study points to a new BA-driven mechanism of CRC-associated liver metastases, suggesting that a reduction of TCA overexposure by limiting liver uptake is a potential therapeutic option for CRC-associated liver metastasis.

A method to establish a chronic restraint stress mouse model with colorectal cancer xenografts.

Colorectal cancer (CRC) remains one of the most common clinical cancers of digestive tract. Recently, a large number of researches have shown that chronic stress can actively participate in the development of CRC. The proposed method successfully established the model of chronic stress mouse model with colorectal cancer.•Chronic restraint stress (CRS) was used to establish chronic stress model.•CRS was combined with a colorectal cancer xenografts model.•Behavioural tests and tumour growth were used to evaluate model construction.

Long noncoding RNA NEAT1 promotes tumorigenesis in H. pylori gastric cancer by sponging miR-30a to regulate COX-2/BCL9 pathway.

BACKGROUND: Helicobacter pylori (H. pylori) is a carcinogenic factor for gastric cancer. Our previous study demonstrated that H. pylori decreased the expression of micro-RNA (miRNA)-30a to promote the tumorigenesis of gastric cancer. However, the upstream regulatory molecules of miR-30a are not well elucidated. In this study, we found the long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) may sponge miR-30a to regulate COX-2/BCL9 pathway. METHODS: The expression of NEAT1 was detected in gastric cancer tissues and tumor-adjacent tissues by fluorescence in situ hybridization (FISH) analysis and RT-qPCR. LncRNA-miRNA interaction networks were constructed using the RNAhybrid and starBase v.2.0. and then validated using a dual-luciferase reporter assay. The effects of NEAT1 dysregulation on the proliferative, migratory, and invasive abilities of H. pylori filtrate-infected gastric cancer cells were observed by cell counting kit-8 (CCK-8), colony formation, wound healing test, and transwell assays. Western blot and RT-qPCR were performed to detect protein and RNA expression. Immunohistochemistry (IHC) was carried out to analyze the localization and expression of COX-2 and BCL9. RESULTS: FISH and RT-qPCR demonstrated that the expression of NEAT1 was up-regulated in gastric cancer tissues, especially in H. pylori-infected gastric cancer tissues, and the expression of NEAT1 was negatively correlated with miR-30a (miR-30a-3p and miR-30a-5p). The upregulation of NEAT1 enhanced proliferation, migration, and invasion of H. pylori filtrate-infected gastric cancer cells, while the downregulation of NEAT1 decreased these abilities, and miR-30a could reverse the effect of NEAT1 on these abilities. The dual-luciferase reporter assay identified that NEAT1 directly targeted miR-30a (miR-30a-3p and miR-30a-5p). Because miR-30a (miR-30a-3p and miR-30a-5p) negatively regulates the expression of downstream COX-2 and BCL9, NEAT1 was identified to upregulate indirectly the expression of COX-2 and BCL9. IHC showed that the expression of COX-2 and BCL9 was increased in H. pylori gastric cancer tissues. CONCLUSION: The study demonstrated that lncRNA NEAT1 may act as a promoter of tumorigenesis in H. pylori gastric cancer, by sponging miR-30a (miR-30a-3p and miR-30a-5p) to regulate the COX-2/BCL9 pathway.

Xiao-Chai-Hu-Tang ameliorates tumor growth in cancer comorbid depressive symptoms via modulating gut microbiota-mediated TLR4/MyD88/NF-κB signaling pathway.

BACKGROUND: Depressive symptoms are thought to promote cancer development and depressive remission has been reported to be effective for defeating cancer. The herbal formula Xiao-Chai-Hu-Tang (XCHT), that has an anti-depressive efficacy, has been widely utilized in China. However, its anti-cancer effect and underlying mechanisms remain unclear. PURPOSE: The present study aims to investigate the effects of XCHT on the depression-associated tumor and its potential mechanisms. METHODS: A placebo-controlled trial was conducted in cancer patients comorbid with depressive symptoms to evaluate the effects of XCHT on depressive scales, tumor-related immune indicators, and gut microbial composition. A xenografted colorectal cancer (CRC) mouse model exposure to chronic restraint stress (CRS) was established to examine XCHT effects on tumorigenesis in vivo. Further, by manipulating gut bacteria with fecal microbial transplantation (FMT) or antibiotics-induced bacterial elimination in CRS-associated xenografted model, gut microbiota-mediated anti-tumor mechanism was explored. RESULTS: In cancer patients comorbid with depressive symptoms, XCHT showed substantial effects on improvement of depressive scales, system inflammatory levels and gut dysbiosis. In vivo, XCHT inhibited tumor growth and prolonged survival time in addition to showing anti-depressive effect. Similarly, in our clinical trial, XCHT partially reversed gut dysbiosis, particularly through reducing abundances of Parabacteroides, Blautia and Ruminococcaceae bacterium. Manipulation of gut bacteria in CRS-associated xenografted model further proved that the inhibition of XCHT on tumor progression was mediated by gut microbiota and that the underlying mechanism involves in downregulation of TLR4/MyD88/NF-κB signaling. CONCLUSIONS: We demonstrated that gut microbiota mediates the anti-tumor action of the formula XCHT in cancer patients and models that were comorbid with depressive symptoms. This study implies a novel clinical significance of anti-depressive herbal medicine in the cancer treatment and clarifies the important role of gut microbiota in treating cancer accompanied by depressive symptoms.

Xiaoyaosan slows cancer progression and ameliorates gut dysbiosis in mice with chronic restraint stress and colorectal cancer xenografts.

Depression is a risk factor for colorectal cancer (CRC) progression. Xiaoyaosan (XYS) is a traditional Chinese medicine prescription for treating depression. Our present study aimed to investigate the effect of XYS on chronic restraint stress (CRS) in mice with CRC xenografts and explore its underlying mechanisms. XYS treatment for 21 consecutive days successfully reduced the tumour volume and tumour weight in mice and prolonged the overall survival time. In addition, the intestinal permeability in the XYS group was significantly improved after administration. The 16S rRNA high-throughput sequencing method was used to sequence stool samples to check the structure and changes of gut bacteria. XYS mainly regulated the abundance of Bacteroides, Lactobacillus, Desulfovibrio and Rikenellaceae. Taken together, these results provide direct strong evidence that XYS effectively improves the progression of CRC in CRS-handled mice, and its efficacy is associated with the modulation of gut dysbiosis. The application of XYS can be a novel therapeutic strategy for CRC patients with depression.

Tanshinone IIA reduces secretion of pro­angiogenic factors and inhibits angiogenesis in human colorectal cancer.

Tumor angiogenesis is an important factor which precipitates recurrence and metastasis of colorectal cancer (CRC). Angiogenesis is also a significant feature which accompanies invasion and metastasis of CRC. Tumor hypoxia activates hypoxia inducible factor (HIF), which promotes angiogenesis in CRC. HIF significantly promotes cell proliferation and angiogenesis in CRC, facilitating invasion and metastasis. Tanshinone IIA (Tan IIA) has been revealed to effectively inhibit angiogenesis in CRC, although the underlying mechanism remains to be determined. The aim of the present study was to determine the effects of HIF­1α on hypoxia induced angiogenesis in CRC cells, the effects of Tan IIA on the expression of pro­angiogenic factors in CRC cells, and on human umbilical vein endothelial cell (HUVEC) tube formation in normal and hypoxic conditions. The results of the present study revealed that Tan IIA not only decreased HIF­1α expression and inhibited the secretion level of vascular endothelial growth factor and basic fibroblast growth factor, but also efficiently decreased proliferation, tube formation and metastasis of HUVECs. The results highlight the potential of Tan IIA­mediated targeting of HIF­1α as a potential therapeutic option for treatment of patients with CRC.

Mechanisms underlying the effects of stress on tumorigenesis and metastasis (Review).

Stress is one of the fundamental survival mechanisms in nature. Although chronic or long-lasting stress can be detrimental to health, acute or short-term stress can have health benefits. The aim of the present review was to address the complexity and significance of stress in tumorigenesis. The review covers an evaluation of previously used and reported experimental animal models of stress, as well as the effects of stress on the neuroendocrine system, immune function, gut microbiota, and inflammation and multidrug resistance, all of which are closely associated with cancer occurrence, progression and treatment. The review concludes that understanding the efficacy of stress management (prevention and rehabilitation) is crucial to the development of comprehensive and individualized strategies for cancer prevention and treatment.

Metabolic engineering of Escherichia coli for the synthesis of polyhydroxyalkanoates using acetate as a main carbon source.

BACKGROUND: High production cost of bioplastics polyhydroxyalkanoates (PHA) is a major obstacle to replace traditional petro-based plastics. To address the challenges, strategies towards upstream metabolic engineering and downstream fermentation optimizations have been continuously pursued. Given that the feedstocks especially carbon sources account up to a large portion of the production cost, it is of great importance to explore low cost substrates to manufacture PHA economically. RESULTS: Escherichia coli was metabolically engineered to synthesize poly-3-hydroxybutyrate (P3HB), poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P3HB4HB), and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) using acetate as a main carbon source. Overexpression of phosphotransacetylase/acetate kinase pathway was shown to be an effective strategy for improving acetate assimilation and biopolymer production. The recombinant strain overexpressing phosphotransacetylase/acetate kinase and P3HB synthesis operon produced 1.27 g/L P3HB when grown on minimal medium supplemented with 10 g/L yeast extract and 5 g/L acetate in shake flask cultures. Further introduction succinate semialdehyde dehydrogenase, 4-hydroxybutyrate dehydrogenase, and CoA transferase lead to the accumulation of P3HB4HB, reaching a titer of 1.71 g/L with a 4-hydroxybutyrate monomer content of 5.79 mol%. When 1 g/L of α-ketoglutarate or citrate was added to the medium, P3HB4HB titer increased to 1.99 and 2.15 g/L, respectively. To achieve PHBV synthesis, acetate and propionate were simultaneously supplied and propionyl-CoA transferase was overexpressed to provide 3-hydroxyvalerate precursor. The resulting strain produced 0.33 g/L PHBV with a 3-hydroxyvalerate monomer content of 6.58 mol%. Further overexpression of propionate permease improved PHBV titer and 3-hydroxyvalerate monomer content to 1.09 g/L and 10.37 mol%, respectively. CONCLUSIONS: The application of acetate as carbon source for microbial fermentation could reduce the consumption of food and agro-based renewable bioresources for biorefineries. Our proposed metabolic engineering strategies illustrate the feasibility for producing polyhydroxyalkanoates using acetate as a main carbon source. Overall, as an abundant and renewable resource, acetate would be developed into a cost-effective feedstock to achieve low cost production of chemicals, materials, and biofuels.