Integrated microbiome-metabolomics analysis reveals the potential therapeutic mechanism of Zuo-Jin-Wan in ulcerative colitis.

BACKGROUND: Dysregulation in gut microbiota and host cometabolome contributes to the complicated pathology of ulcerative colitis (UC), while Zuo-Jin-Wan (ZJW), a traditional Chinese medicine has shown therapeutic effects against UC with its underlying mechanism remains elusive. PURPOSE: This study utilized an integrated analysis combining gut microbiome and host cometabolism to disclose the potential therapeutic mechanism of ZJW on dextran sulfate sodium (DSS)-induced UC in rats. METHODS: We first evaluated the therapeutic effects of ZJW treatment in DSS-induced rat model. 16S rRNA sequencing, 1H NMR spectroscopy-based metabolomics and Spearman correlation analysis were conducted to explore the potential therapeutic mechanism during the treatment. RESULTS: Our results showed that UC symptoms in ZJW rats were significantly attenuated. Marked decline in microbial diversity in ZJW group was accompanied by its correspondent function adjustment. Specific enrichment of genus Bacteroides, Sutterella, Akkermansia and Roseburia along with the major varying amino acid metabolism and lipid metabolism were observed meantime. Metabolic data further corroborated that ZJW-related metabolic changes were basically gathered in amino acid metabolism, carbohydrate/energy metabolism and lipid metabolism. Of note, some biochemical parameters were deeply implicated with the discriminative microbial genera and metabolites involved in tricarboxylic acid (TCA) cycle and amino acid metabolism, indicating the microbiome-metabolome association in gut microbiota-metabolite-phenotype axis during UC treatment of ZJW. CONCLUSION: For the first time, integrated microbiome-metabolome analysis depicted that ZJW could alleviate DSS-induced UC in rats via a crosstalk between gut microbiota and host cometabolites.

In-vitro anti-Helicobacter pylori activity and preliminary mechanism of action of Canarium album Raeusch. fruit extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Canarium album Raeusch. belongs to the Burseraceae family. Its ripe fruits, known as Qing Guo (QG) in Traditional Chinese Medicine (TCM), are used to treat sore throat, cough, and fish or crab poisoning. QG was reported to have antibacterial activity, and it exerted excellent anti-Helicobacter pylori (H. pylori) activity in our screening of abundant TCM. However, few studies have reported its anti-H. pylori activity and mechanism. AIM OF STUDY: The commonly used eradication therapies for H. pylori infection are antibiotic-based therapies. With the increasing antibiotic resistance of H. pylori, interest in finding alternative therapies has been aroused. This study investigated the phytochemistry profile, in vitro anti-H. pylori activity and possible anti-bacterial mechanism of QG extracts. MATERIALS AND METHODS: QG extracts were obtained by heat reflux extraction, ultrasonic extraction or liquid-liquid extraction with different solvents. The quantitative and qualitative phytochemical analyses were performed by colorimetric determination, high-performance liquid chromatography (HPLC), and UPLC-electrospray ionization mass spectrometry (ESI-MS). In vitro anti- H. pylori activity was assessed by broth micro-dilution method. Mechanism of action studies included morphological observation using electron microscopy, urease inhibition assay and determination of expression of virulence genes by RT-qPCR. RESULTS: All QG extracts especially ethyl acetate extract (QGEAE) were rich in phenolic components, with the minimum inhibitory concentrations (MICs) on H.pylori of 39-625 µg/ml and minimum bactericidal concentrations (MBCs) of 78-1250 µg/ml. Both aqueous extract (QGAE) and QGEAE could induce the morphological and structural changes of H. pylori, inhibit urease activity with IC50 of 1093 µg/ml and 332.90 µg/ml, respectively, and down-regulate the virulence genes, such as vacA and cagA. CONCLUSIONS: QG may exhibit in vitro anti-H. pylori activity by inhibiting growth, destroying the bacterial structure and down-regulating the expression of virulence factors. Moreover, QG is the homology of food and TCM, which can be considered as a safe and convenient agent against H. pylori infection.

Sanguisorba officinalis L. suppresses 5-fluorouracil-sensitive and-resistant colorectal cancer growth and metastasis via inhibition of the Wnt/ß-catenin pathway.

BACKGROUND: Colorectal cancer (CRC) is a widespread cancer with high morbidity and mortality. Chemoresistance and metastasis are the current challenges for CRC treatment. Sanguisorba officinalis Linn. (called DiYu in Chinese, DY) is a traditional Chinese medicine (TCM) whose root is long used as medicinal part. In our previous study, the aqueous extract of DY could inhibit the Wnt/ß-catenin pathway and showed great antitumor effect against CRC. The Wnt/ß-catenin pathway is involved in CRC chemoresistance and metastasis. However, there is little study on the antitumor and antimetastatic effects of DY on resistant CRC cells. The aim of this study is to explore the effect of aqueous extract of DY on the growth and metastasis of 5-fluorouracil (5-FU) sensitive and resistant CRC, and to elucidate the underlying molecular mechanism. METHODOLOGY: In this study, cell viability, cell colony formation and apoptosis analyses were performed to verify the in vitro antitumor effect of DY on 5-FU-sensitive and -resistant CRC cells. Next, transwell assays were used to test the inhibition activity of DY on CRC migration and invasion. Western Blotting assays were carried out to identify the molecular mechanism underlying the efficacy of DY extract. Xenograft CRC nude mice model and tumor metastasis model were used to confirm the in vivo antitumor and antimetastatic effects of DY. RESULTS: DY inhibited cell proliferation and apoptosis via the upregulation of Bax, cleaved-caspase3 and cleaved-PARP proteins and downregulation of Bcl-2 protein. DY also inhibited cell migration and invasion via the downregulation of N-cadherin, vimentin and snail proteins and upregulation of E-cadherin protein, demonstrating that DY suppressed cell metastasis by reversing epithelial-to-mesenchymal transition (EMT) procession. Moreover, the protein expression levels of ß-catenin in whole cell, cytoplasm and nucleus were decreased after DY treatment. Taken together, DY suppressed CRC cell growth and metastasis via inhibition of the Wnt pathway. Additionally, DY also demonstrated effective antitumor and anti-metastasis activities in vivo. CONCLUSIONS: In conclusion, DY suppressed the growth and metastasis of 5-FU-sensitive and -resistant CRC via inhibition of the Wnt pathway, which indicated that DY could be a potential drug to treat CRC patients and improve clinic outcome.

Dietary synbiotic ameliorates constipation through the modulation of gut microbiota and its metabolic function.

The purpose of this study is to investigate the mitigatory effect of a novel synbiotic (SBT) on constipation from the perspective of gut microbiome and metabolome. Here, intake of SBT effectively attenuated diphenoxylate-induced constipation, recuperated colonic epithelial integrity and increased serum levels of gastrointestinal excitatory neurotransmitters (P substance, vasoactive intestinal peptide, motilin, gastrin and serotonin). 16S rRNA sequencing showed that SBT intake rehabilitated the composition and functionality of gut microbiota. Relative abundances of short-chain fatty acids (SCFAs)-producing bacteria including Lactobacillus, Faecalibaculum and Bifidobacterium were elevated by administration of SBT. The gas chromatography-mass spectrometry analysis confirmed that fecal concentrations of propionate and butyrate were significantly increased in the rats intervened with SBT. In addition, SBT ingestion reduced the relative levels of opportunistic pathogens, such as Oscillibacter, Parasutterella and Parabacteroides. Microbial functional prediction showed that the relative abundances of lipopolysaccharide (LPS) biosynthesis and arachidonic acid metabolism were downregulated with SBT administration, which were in accordance with the serum metabolomics results. Furthermore, serum levels of LPS, tumour necrosis factor alpha and interleukin 6 were significantly decreased, indicating that SBT supplementation suppressed inflammatory responses. Therefore, this study demonstrated that consumption of SBT ameliorated constipation possibly by regulating gut microbiota, promoting the SCFAs production and inhibiting inflammatory responses in rats. Our study also indicated that SBT may provide a novel alternative strategy for the treatment of constipation clinically in future.

Mass spectrometry investigation of nucleoside adducts of fatty acid hydroperoxides from oxidation of linolenic and linoleic acids.

The widespread presence of lipid hydroperoxides in foodstuffs and biological samples has aroused great attentions in recent years, while it remains challenging for analysis of the fragility of O - O bond linkage of peroxides. In this present study, we explored the utility of electrospray ionization mass spectrometry (ESI-MS) for characterization of two fatty acid hydroperoxides from oxidation of linoleic acid and α-linolenic acid, which are the essential fatty acids abundant in many seeds and vegetable oils. The results indicated that in-source fragmentation occurred in the detection of the two fatty acid hydroperoxides in both positive and negative ion modes, which yielded characteristic fragments for ESI-MS analysis. In addition, the genotoxicity of fatty acid hydroperoxides for generation of nucleoside adducts was investigated. It was found that a variety of nucleoside adducts were formed from the reactions of fatty acid hydroperoxides and nucleosides. Furthermore, the decomposition products of the fatty acid hydroperoxides were determined, which provided evidence to elucidate the reaction mechanism for formation of nucleoside adducts.

Global Metabolomic and Lipidomic Analysis Reveal the Synergistic Effect of Bufalin in Combination with Cinobufagin against HepG2 Cells.

Chansu, which is prepared from the skin secretions of toad (Bufo bufo gargarizans Cantor), is widely used in traditional Chinese medicine (TCM). Being the principal bioactive constituents of Chansu, bufalin (BFL) and cinobufagin (CBF) have been shown to possess anticancer properties. TCM confers bioactivities through the synergistic effect between potential active ingredients, so as to interfere with the development of the disease, and ultimately achieve the therapeutic effect. We found that the anticancer effect was significantly potentiated by cotreatment with BFL and CBF compared to monotreatment, suggesting their synergistic interaction. To reveal their synergistic mechanisms, metabolomic and lipidomic profiling based on liquid chromatography-mass spectrometry (LC-MS) was utilized to delineate the responses in HepG2 cells after treatment with BFL and CBF individually or in combination. Metabolic pathways, including methionine metabolism, energy metabolism, lipid metabolism, and amino acid metabolism, were modulated and subsequently led to apoptosis and cell cycle arrest of HepG2 cells. In particular, the discrepant regulation of methionine metabolism between the monotreatment and cotreatment with BFL and CBF may account for their synergistic effect. Our study provided novel insights into the mechanistic links between cellular metabolism and the synergistic effect, which may ultimately lead to better treatments for hepatoma.

Spatial Lipidomics Reveals Anticancer Mechanisms of Bufalin in Combination with Cinobufagin in Tumor-Bearing Mice.

Bufalin (BFL) and cinobufagin (CBF) are the principal bioactive constituents of Chansu, a widely used traditional Chinese medicine (TCM). The synergistic effects of potential active components are responsible for the bioactivities of TCM. Our results showed that the cotreatment with BFL and CBF confers superior anticancer efficacy compared to monotreatment. To reveal the underlying mechanisms of their cotreatment, an integrated method composed of mass spectrometry-based lipidomics and matrix-assisted laser desorption/ionization mass spectrometry imaging was used to delineate the responses of tumor-bearing mice treated with BFL and CBF individually or in combination. The cotreatment with BFL and CBF modulated the sphingolipid metabolism and glycerophospholipid metabolism, and subsequently led to mitochondria-driven apoptosis and systemic disruption of biomembranes in tumor cells. Furthermore, we found that the disturbed lipid markers were mainly located in the non-necrotic tumor areas, the essential parts for the formation of solid tumor framework. Together, our findings revealed what occurred in tumor in response to the treatment of BFL and CBF, from lipids to enzymes, and thus provide insights into the critical role of lipid reprogramming in the satisfactory anticancer effect of BFL in combination with CBF.

Investigation of the reverse effect of Danhong injection on doxorubicin-induced cardiotoxicity in H9c2 cells: Insight by LC-MS based non-targeted metabolomic analysis.

Although Danhong injection (DHI) has been clearly shown to attenuate ischemic myocardial injury and improve heart function, there is no research regarding its role in doxorubicin (DOX)-induced cardiomyopathy. In this study, we aimed to investigate the reverse effect of DHI on DOX-induced cardiotoxicity in H9c2 cells. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay demonstrated that DHI had no cytotoxicity towards the relevant cell line unless the concentration was as high as 50 µL/mL. The satisfactory cardioprotective effect of DHI exerted at the concentration of 10 µL/mL, which agreed well with the result of real-time cell viability assay. Then non-targeted metabolomics based on LC-MS was employed to characterize metabolic alterations in DOX-induced cells with DHI treatment. Multivariate analysis, including PCA and PLS-DA, revealed 31 altered metabolites after DOX treatment that were primarily related to the disturbance of amino acids and nucleotides metabolism. While DHI could intervene in some disturbed metabolic pathways, such as the metabolism of arginine, glutathione (GSH), pantothenic acid, cytidine, inosine and 5'-methylthioadenosine. These results suggested that DHI exerted the therapeutic effect by improving energy metabolism and attenuating oxidative stress. The present study can lay a foundation for further research on the promising therapeutic effect of DHI in managing DOX-induced cardiotoxicity.