[Banxia Xiexin Decoction inhibiting colitis-associated colorectal cancer infected with Fusobacterium nucleatum by regulating Wnt/ß-catenin pathway].

This paper investigates the intervention effect and mechanism of Banxia Xiexin Decoction(BXD) on colitis-associated colorectal cancer(CAC) infected with Fusobacterium nucleatum(Fn). C57BL/6 mice were randomly divided into a control group, Fn group, CAC group [azoxymethane(AOM)/dextran sulfate sodium salt(DSS)](AOM/DSS), model group, and BXD group. Except for the control and AOM/DSS groups, the mice in the other groups were orally administered with Fn suspension twice a week. The AOM/DSS group, model group, and BXD group were also injected with a single dose of 10 mg·kg~(-1) AOM combined with three cycles of 2.5% DSS taken intragastrically. The BXD group received oral administration of BXD starting from the second cycle until the end of the experiment. The general condition and weight changes of the mice were monitored during the experiment, and the disease activity index(DAI) was calculated. At the end of the experiment, the colon length and weight of the mice in each group were compared. Hematoxylin-eosin(HE) staining was used to observe the pathological changes in the colon tissue. Enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of interleukin(IL)-2, IL-4, and IL-6 inflammatory factors in the serum. Immunohistochemistry(IHC) was used to detect the expression of Ki67, E-cadherin, and ß-catenin in the colon tissue. Western blot was used to detect the protein content of Wnt3a, ß-catenin, E-cadherin, annexin A1, cyclin D1, and glycogen synthase kinase-3ß(GSK-3ß) in the colon tissue. The results showed that compared with the control group, the Fn group had no significant lesions. The mice in the AOM/DSS group and model group had decreased body weight, increased DAI scores, significantly increased colon weight, and significantly shortened colon length, with more significant lesions in the model group. At the same time, the colon histology of the model group showed more severe adenomas, inflammatory infiltration, and cellular dysplasia. The levels of IL-4 and IL-6 in the serum were significantly increased, while the IL-2 content was significantly decreased. The IHC results showed low expression of E-cadherin and high expression of Ki67 and ß-catenin in the model group, with a decreased protein content of E-cadherin and GSK-3ß and an increased protein content of Wnt3a, ß-catenin, annexin A1, and cyclin D1. After intervention with BXD, the body weight of the mice increased; the DAI score decreased; the colon length increased, and the tumor decreased. The histopathology showed reduced tumor proliferation and reduced inflammatory infiltration. The levels of IL-6 and IL-4 in the serum were significantly decreased, while the IL-2 content was increased. Meanwhile, the expression of E-cadherin was upregulated, and that of Ki67 and ß-catenin was downregulated. The protein content of E-cadherin and GSK-3ß increased, while that of Wnt3a, ß-catenin, annexin A1, and cyclin D1 decreased. In conclusion, BXD can inhibit CAC infected with Fn, and its potential mechanism may be related to the inhibition of Fn binding to E-cadherin, the decrease in annexin A1 protein level, and the regulation of the Wnt/ß-catenin pathway.

Huang Qin decoction increases SLC6A4 expression and blocks the NFκB-mediated NLRP3/Caspase1/GSDMD pathway to disrupt colitis-associated carcinogenesis.

Huang Qin decoction (HQD) is a traditional Chinese medicine formula for treating colitis, but the effects and molecular mechanism of action of HQD in colitis-associated carcinogenesis (CAC) are still unclear. Therefore, we aimed to determine the beneficial effects of HQD on CAC in mice and to reveal the underlying mechanism involved. AOM/DSS was used to induce CAC in mice, and the effects of HQD on tumorigenesis in mice were examined (with mesalazine serving as a positive control). Mesalazine or HQD treatment alleviated body weight loss and decreased the disease activity index in mice induced by AOM/DSS. Mesalazine or HQD treatment also suppressed the shortening of colon tissue length, the number of tumors, and the infiltration of inflammatory cells. The genes targeted by HQD were predicted and verified, followed by knockout experiments. Elevated SLC6A4 and inhibited serotonin production and inflammation were observed in HQD-treated mice. HQD inhibited the NFκB and NLRP3/caspase1/GSDMD pathways. The therapeutic effect of HQD was diminished in SLC6A4-deficient AOM/DSS mice. Additionally, the downregulation of SLC6A4 mitigated the inhibitory effect of HQD-containing serum on MODE-K cell pyroptosis. Our findings suggest that SLC6A4 is a pivotal regulator of HQD-alleviated CAC via its modulation of the NLRP3/caspase1/GSDMD pathway.

Huangqin Decoction Delays Progress of Colitis-Associated Carcinogenesis by Regulating Nrf2/HO-1 Antioxidant Signal Pathway in Mice.

OBJECTIVE: To investigate the effect of Huangqin Decoction (HQD) on nuclear factor erythroid 2 related-factor 2 (Nrf2)/heme oxygenase (HO-1) signaling pathway by inducing the colitis-associated carcinogenesis (CAC) model mice with azoxymethane (AOM)/dextran sodium sulfate (DSS). METHODS: The chemical components of HQD were analyzed by liquid chromatography-quadrupole-time-of-flight mass spectrometry (LC-Q-TOF-MS/MS) to determine the molecular constituents of HQD. Totally 48 C57BL/6J mice were randomly divided into 6 groups by a random number table, including control, model (AOM/DSS), mesalazine (MS), low-, medium-, and high-dose HQD (HQD-L, HQD-M, and HQD-H) groups, 8 mice in each group. Except for the control group, the mice in the other groups were intraperitoneally injected with AOM (10 mg/kg) and administrated with 2.5% DSS orally for 1 week every two weeks (totally 3 rounds of DSS) to construct a colitis-associated carcinogenesis mouse model. The mice in the HQD-L, HQD-M and HQD-H groups were given HQD by gavage at doses of 2.925, 5.85, and 11.7 g/kg, respectively; the mice in the MS group was given a suspension of MS at a dose of 0.043 g/kg (totally 11 weeks). The serum levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were measured by enzyme-linked immunosorbent assay. The mRNA and protein expression levels of Nrf2, HO-1, and inhibitory KELCH like ECH-related protein 1 (Keap1) in colon tissue were detected by quantitative real-time PCR, immunohistochemistry, and Western blot, respectively. RESULTS: LC-Q-TOF-MS/MS analysis revealed that the chemical constituents of HQD include baicalin, paeoniflorin, and glycyrrhizic acid. Compared to the control group, significantly higher MDA levels and lower SOD levels were observed in the model group (P<0.05), whereas the expressions of Nrf2 and HO-1 were significantly decreased, and the expression of Keap1 increased (P<0.01). Compared with the model group, serum MDA level was decreased and SOD level was increased in the HQD-M, HQD-H and MS groups (P<0.05). Higher expressions of Nrf2 and HO-1 were observed in the HQD groups. CONCLUSION: HQD may regulate the expression of Nrf2 and HO-1 in colon tissue, reduce the expression of MDA and increase the expression of SOD in serum, thus delaying the progress of CAC in AOM/DSS mice.

Electroacupuncture at Sensitized Acupoints Relieves Somatic Referred Pain in Colitis Rats by Inhibiting Sympathetic-Sensory Coupling to Interfere with 5-HT Signaling Pathway.

OBJECTIVE: To investigate whether electroacupuncture (EA) at sensitized acupoints could reduce sympathetic-sensory coupling (SSC) and neurogenic inflammatory response by interfering with 5-hydroxytryptamine (5-HT)ergic neural pathways to relieve colitis and somatic referred pain, and explore the underlying mechanisms. METHODS: Rats were treated with 5% dextran sodium sulfate (DSS) solution for 7 days to establish a colitis model. Twelve rats were randomly divided into the control and model groups according to a random number table (n=6). According to the "Research on Rat Acupoint Atlas", sensitized acupoints and non-sensitized acupoints were determined. Rats were randomly divided into the control, model, Zusanli-EA (ST 36), Dachangshu-EA (BL 25), and Xinshu (BL 15) groups (n=6), as well as the control, model, EA, and EA + GR113808 (a 5-HT inhibitor) groups (n=6). The rats in the control group received no treatment. Acupuncture was administered on 2 days after modeling using the stimulation pavameters: 1 mA, 2 Hz, for 30 min, with sparse and dense waves, for 14 consecutive days. GR113808 was injected into the tail vein at 5 mg/kg before EA for 10 min for 7 consecutive days. Mechanical sensitivity was assessed with von Frey filaments. Body weight and disease activity index (DAI) scores of rats were determined. Hematoxylin and eosin staining was performed to observe colon histopathology. SSC was analyzed by immunofluorescence staining. Immunohistochemical staining was performed to detect 5-HT and substance P (SP) expressions. The calcitonin gene-related peptide (CGRP) in skin tissue and tyrosine hydroxylase (TH) protein levels in DRG were detected by Western blot. The levels of hyaluronic acid (HA), bradykinin (BK), prostaglandin I2 (PGI2) in skin tissue, 5-HT, tryptophan hydroxylase 1 (TPH1), serotonin transporters (SERT), 5-HT 3 receptor (5-HT3R), and 5-HT 4 receptor (5-HT4R) in colon tissue were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: BL 25 and ST 36 acupoints were determined as sensitized acupoints, and BL 15 acupoint was used as a non-sensitized acupoint. EA at sensitized acupoints improved the DAI score, increased mechanical withdrawal thresholds, and alleviated colonic pathological damage of rats. EA at sensitized acupoints reduced SSC structures and decreased TH and CGRP expression levels (P<0.05). Furthermore, EA at sensitized acupoints reduced BK, PGI2, 5-HT, 5-HT3R and TPH1 levels, and increased HA, 5-HT4R and SERT levels in colitis rats (P<0.05). GR113808 treatment diminished the protective effect of EA at sensitized acupoints in colitis rats (P<0.05). CONCLUSION: EA at sensitized acupoints alleviated DSS-induced somatic referred pain in colitis rats by interfering with 5-HTergic neural pathway, and reducing SSC inflammatory response.

Intestinal Microbiota and Metabolomics Reveal the Role of Auricularia delicate in Regulating Colitis-Associated Colorectal Cancer.

BACKGROUND: The edible fungus Auricularia delicate (ADe) is commonly employed in traditional medicine for intestinal disorders; however, its inhibitory effect on colitis-associated colorectal cancer (CAC) and the underlying mechanisms remain unexplored. (2) Methods: The inhibitory effect of ADe on CAC was investigated using a mouse model induced by azoxymethane/dextran sulfate sodium. RESULTS: ADe effectively suppressed the growth and number of intestinal tumors in mice. Intestinal microbiota analyses revealed that ADe treatment increased Akkermansia and Parabacteroides while it decreased Clostridium, Turicibacter, Oscillospira, and Desulfovibrio. ADe regulated the levels of 2'-deoxyridine, creatinine, 1-palmitoyl lysophosphatidylcholine, and choline in serum. Furthermore, the levels of these metabolites were associated with the abundance of Oscillospira and Paraacteroides. ADe up-regulated the free fatty acid receptor 2 and ß-Arrestin 2, inhibited the nuclear factor kappa B (NF-κB) pathway, and significantly attenuated the levels of inflammatory cytokines, thereby mitigating the inflammatory in CAC mice. CONCLUSIONS: The protective effect of ADe in CAC mice is associated with the regulation of intestinal microbiota, which leads to the inhibition of NF-kB pathway and regulation of inflammation.

Chemopreventive Effects of Polysaccharides and Flavonoids from Okra Flowers in Azomethane/Dextran Sulfate Sodium-Induced Murine Colitis-Associated Cancer.

Okra flowers are a good source of polysaccharides and flavonoids, with biological activities of anti-inflammatory action and modulation of the gut microbiota. Previously, we reported that flavonoid-rich extracts from okra flowers (AFE) presented effective anti-colorectal cancer (CRC) activity in CRC cells as well as xenograft models, but their role in colitis-associated cancer (CAC) is unidentified. In this study, we aimed to evaluate the effects of AFE and APE (polysaccharides extracted from okra flowers) on the CAC symptoms of azoxymethane (AOM)/dextran sodium sulfate (DSS)-intervened mice. The results showed that APE and AFE exert potent efficacy in inhibiting colitis and colorectal tumorigenesis stimulated by AOM/DSS, characterized by decreased colonic shortening, DAI score, and tumor numbers. Compared with the control group, APE/AFE alleviated the microbiota dysbiosis driven by AOM/DSS. In addition, AFE elicited its anticancer activity through regulation of NFκB/IL-6/Stat3, JAK2/Stat3, MAPKs, PI3K/AKT, and Wnt/ß-catenin signal transductions in AOM/DSS mice, which was consistent with a vitro model of CT26 cells, while APE treatment exhibited anticancer activity through regulation of Nrf2/IL-6, MAPKs, PI3K/AKT, and Wnt/ß-catenin signal transductions in the AOM/DSS mouse model. Collectively, our studies revealed, for the first time, that flavonoids and polysaccharides from okra flowers possess the ability to attenuate colitis and colorectal tumorigenesis, with them having great potential to become promising candidates against CRC.

Anti-Hyperalgesic Efficacy of Acetyl L-Carnitine (ALCAR) Against Visceral Pain Induced by Colitis: Involvement of Glia in the Enteric and Central Nervous System.

The management of abdominal pain in patients affected by inflammatory bowel diseases (IBDs) still represents a problem because of the lack of effective treatments. Acetyl L-carnitine (ALCAR) has proved useful in the treatment of different types of chronic pain with excellent tolerability. The present work aimed at evaluating the anti-hyperalgesic efficacy of ALCAR in a model of persistent visceral pain associated with colitis induced by 2,4-dinitrobenzene sulfonic acid (DNBS) injection. Two different protocols were applied. In the preventive protocol, ALCAR was administered daily starting 14 days to 24 h before the delivery of DNBS. In the interventive protocol, ALCAR was daily administered starting the same day of DNBS injection, and the treatment was continued for 14 days. In both cases, ALCAR significantly reduced the establishment of visceral hyperalgesia in DNBS-treated animals, though the interventive protocol showed a greater efficacy than the preventive one. The interventive protocol partially reduced colon damage in rats, counteracting enteric glia and spinal astrocyte activation resulting from colitis, as analyzed by immunofluorescence. On the other hand, the preventive protocol effectively protected enteric neurons from the inflammatory insult. These findings suggest the putative usefulness of ALCAR as a food supplement for patients suffering from IBDs.

Refined fiber inulin promotes inflammation-associated colon tumorigenesis by modulating microbial succinate production.

BACKGROUND AND AIM: There is an increased risk of colon cancer associated with inflammatory bowel disease (IBD). Dietary fibers (DFs) naturally present in vegetables and whole grains offer numerous beneficial effects on intestinal health. However, the effects of refined DFs on intestinal health remain unclear. Therefore, we elucidated the impact of the refined DF inulin on colonic inflammation and tumorigenesis. METHODS: Four-week-old wild-type (WT) mice were fed diets containing insoluble DF cellulose (control) or refined DF inulin for 4 weeks. A subgroup of mice was then switched to drinking water containing dextran sulfate sodium (DSS, 1.4% wt/vol) for colitis induction. In another subgroup of mice, colitis-associated colorectal cancer (CRC) was initiated with three 7-day alternate cycles of DSS following an initial dose of mutagenic substance azoxymethane (AOM; 7.5 mg/kg body weight; i.p.). Post 7 weeks of AOM treatment, mice were euthanized and examined for CRC development. RESULTS: Mice consuming inulin-containing diet exhibited severe colitis upon DSS administration, as evidenced by more body weight loss, rectal bleeding, and increased colonic inflammation than the DSS-treated control group. Correspondingly, histological analysis revealed extensive disruption of colon architecture and massive infiltration of immune cells in the inulin-fed group. We next examined the effect of inulin on CRC development. Surprisingly, significant mortality (~50%) was observed in the inulin-fed but not in the control group during the DSS cycle. Consequently, the remaining inulin-fed mice, which completed the study exhibited extensive colon tumorigenesis. Immunohistochemical characterization showed comparatively high expression of the cell proliferation marker Ki67 and activation of the Wnt signaling in tumor sections obtained from the inulin-fed group. Gut microbiota and metabolite analysis revealed expansion of succinate producers and elevated cecal succinate in inulin-fed mice. Human colorectal carcinoma cells (HCT116) proliferated more rapidly when supplemented with succinate in an inflamed environment, suggesting that elevated luminal succinate may contribute to tumorigenesis. CONCLUSIONS: Our study uncovers that supplementation of diet with refined inulin induces abnormal succinate accumulation in the intestinal lumen, which in part contributes to promoting colon inflammation and tumorigenesis.

[Anemoside B4 regulates fatty acid metabolism reprogramming in mice with colitis-associated cancer].

The study aimed to investigate the effect of anemoside B4(B4) on fatty acid metabolism in mice with colitis-associated cancer(CAC). The CAC model was established by azoxymethane(AOM)/dextran sodium sulfate(DSS) in mice. Mice were randomly divided into a normal group, a model group, and low-, medium-, and high-dose anemoside B4 groups. After the experiment, the length of the mouse colon and the size of the tumor were measured, and the pathological alterations in the mouse colon were observed using hematoxylin-eosin(HE) staining. The slices of the colon tumor were obtained for spatial metabolome analysis to analyze the distribution of fatty acid metabolism-related substances in the tumor. The mRNA levels of SREBP-1, FAS, ACCα, SCD-1, PPARα, ACOX, UCP-2, and CPT-1 were determined by real-time quantitative PCR(RT-qPCR). The results revealed that the model group showed decreased body weight(P<0.05) and colon length(P<0.001), increased number of tumors, and increased pathological score(P<0.01). Spatial metabolome analysis revealed that the content of fatty acids and their derivatives, carnitine, and phospholipid in the colon tumor was increased. RT-qPCR results indicated that fatty acid de novo synthesis and ß-oxidation-related genes, such as SREBP-1, FASN, ACCα, SCD-1, ACOX, UCP-2, and CPT-1 mRNA expression levels increased considerably(P<0.05, P<0.001). After anemoside B4 administration, the colon length increased(P<0.01), and the number of tumors decreased in the high-dose anemoside B4 group(P<0.05). Additionally, spatial metabolome analysis showed that anemoside B4 could decrease the content of fatty acids and their derivatives, carnitine, and phospholipids in colon tumors. Meanwhile, anemoside B4 could also down-regulate the expression of FASN, ACCα, SCD-1, PPARα, ACOX, UCP-2, and CPT-1 in the colon(P<0.05, P<0.01, P<0.001). The findings of this study show that anemoside B4 may inhibit CAC via regulating fatty acid metabolism reprogramming.

Xue-Jie-San prevents the early development of colitis-associated intestinal fibrosis by blocking Notch1 and FGL1 signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Xue-Jie-San (XJS), as a traditional Chinese herb prescription, has satisfactory effects on improving clinical symptoms and facilitating the healing of intestinal ulcers in patients with Crohn's disease (CD). This motivates the application of XJS on CD-associated complications. AIM OF THE STUDY: Intestinal fibrosis is a debilitating complication of CD. Currently, there is no effective medication available for preventing or reversing CD-related intestinal fibrosis. This study aimed to assess the efficacy and underlying mechanisms of XJS in the treatment of colitis-associated intestinal fibrosis. MATERIALS AND METHODS: A rat model of CD-related intestinal fibrosis was induced by 2,4,6-trinitrobenzene sulfonic acid administration and treated with XJS. The pathological changes of intestinal fibrosis were evaluated using Masson staining. Collagen deposition and epithelial-to-mesenchymal transition (EMT) were verified by immunohistochemical staining and Western blot analysis. Endothelial-to-mesenchymal transition (EndoMT) was assessed with immunofluorescence and immunohistochemical staining as well as Western blot analysis. Transmission electron microscopy was utilized to observe autophagosomes. The levels of autophagy-related proteins were detected via immunofluorescence staining and Western blot. Finally, the mTOR/ULK1 signaling pathway regulated by Notch1 or FGL1 was analyzed by Western blot. RESULTS: The results found that XJS ameliorated intestinal fibrosis through reducing the deposition of collagens such as Collagen 1 and Collagen 3. XJS inhibited the EMT process by increasing E-cadherin levels and decreasing the expressions of N-cadherin, Vimentin and Snail, which played a crucial role in collagen secretion and intestinal fibrosis. In addition, XJS also repressed the EndoMT process as reflected by the upregulation of CD31 and VE-cadherin levels and the downregulation of FSP1 and α-SMA expressions. Autophagy was activated following XJS treatment via suppression of the mTOR/ULK1 signaling pathway. Furthermore, XJS acted as an inhibitor of Notch1 and FGL1 signals, both of which regulated the mTOR signaling. CONCLUSIONS: Our findings validated that XJS prevented the early development of CD-related intestinal fibrosis by blocking the Notch1 and FGL1 signaling pathways to activate autophagy and thereby inhibit EMT and EndoMT.

Stercoral Colitis in the Emergency Department: A Retrospective Review of Presentation, Management, and Outcomes.

STUDY OBJECTIVE: Stercoral colitis is inflammation of the bowel wall caused by fecal impaction. Despite reported high morbidity and risk of perforation, little research assessing outcomes is available. This study characterizes the presentation, management, and outcomes of emergency department patients with stercoral colitis. METHODS: We performed a retrospective chart review of ED patients with stercoral colitis identified on computed tomography (CT) scan. Of 814, 522 visits to multiple EDs across the US, 269 met the inclusion criteria. Variables regarding patient presentation, management, and outcomes were extracted from electronic medical records. Results were analyzed with percentages and 95% confidence intervals (CIs). RESULTS: Of 269 patients, the median age was 76 years. The most common chief concern was abdominal pain/distension (33.8%). However, abdominal pain was documented as absent in 62.1% of cases. The most common CT findings included fecal impaction (96.7%), bowel wall inflammation (72.9%), and fat stranding (48.3%). Eighty-four (31.2%) patients were discharged home from the ED, and over half of these (45/84, 53.6%) received no enema, laxatives, or disimpaction. Overall, 9 patients (3.3%, 95% CI 1.6% to 6.5%) required surgical management of a related complication within 3 months, 27 (10.0%, 95% CI 6.8% to 14.4%) returned to the ED within 72 hours, and 9 (3.3%, 95% CI 1.6% to 6.5%) died from a cause related to stercoral colitis within 3 months. CONCLUSION: Patients with stercoral colitis often present in a nonspecific manner, and short-term mortality is substantial. In this study, most discharged patients did not receive recommended treatment. This represents the largest ED study of stercoral colitis and provides further evidence linking this diagnosis with adverse outcomes.

Dioscin modulates macrophages polarization and MDSCs differentiation to inhibit tumorigenesis of colitis-associated colorectal cancer.

It has been reported that colitis is one of risk factors in colorectal cancer (CRC). Intervention of intestinal inflammation and in the early stage of tumorigenesis is of great significance to control the incidence and mortality of CRC. In recent years, natural active products of traditional Chinese medicine have been confirmed that they had made great progress in disease prevention. Here, we showed that Dioscin, a natural active product of Dioscorea nipponica Makino, inhibited initiation and tumorigenesis of AOM/DSS-induced colitis-associated colon cancer (CAC), including alleviating colonic inflammation, improving intestinal barrier function and decreasing tumor burden. In addition, we also explored the immunoregulatory effect of Dioscin on mice. The results showed that Dioscin modulated M1/M2 macrophages phenotype in spleen and decreased monocytic myeloid-derived suppressor cells (M-MDSCs) population in blood and spleen of mice. The in vitro assay demonstrated that Dioscin promoted M1 as well as inhibited M2 macrophages phenotype in LPS- or IL-4-induced bone marrow-derived macrophages (BMDMs) model. Based on the plasticity of MDSCs and its ability to differentiate into M1/M2 macrophages, we here found that Dioscin increased M1- and decreased M2-like phenotype during the process of MDSCs differentiation in vitro, suggesting Dioscin promoted MDSCs differentiate into M1 as well as inhibited its differentiation into M2 macrophages. Taken together, our study indicated that Dioscin had the inhibitory effect on the initial of tumorigenesis at early stage of CAC via the ant-inflammatory effect, which provided a natural active candidate for effective prevention of CAC.

Role of Combination Treatment of Aspirin and Zinc in DMH-DSS-induced Colon Inflammation, Oxidative Stress and Tumour Progression in Male BALB/c Mice.

Colitis-associated colorectal cancer serves as a prototype of inflammation-associated cancers which is linked with repeated cycles of inflammation and DNA repair deficits. Several preclinical and clinical data reported that aspirin has a chemo-preventive effect in colorectal cancer and is associated with dose-dependent side effects. Furthermore, it has been reported that zinc supplementation improves the quality of life in patients undergoing chemotherapy by alteration of colonic cancer cell gene expression. However, explication of the detailed molecular mechanisms involved in the combined administration of aspirin and zinc-mediated protection against colitis-associated colorectal cancer deserves further investigation. For the induction of colitis-associated colorectal cancer, male BALB/c mice were administered 1,2-dimethylhydrazine dihydrochloride (DMH) 20 mg/kg/bw thrice before the initiation of every DSS cycle (3%w/v in drinking water). One week after the initiation of DSS treatment, aspirin (40 mg/kg; p.o.) and zinc in the form of zinc sulphate (3 mg/kg; p.o.) were administered for 8 weeks. Combination of aspirin and zinc as intervention significantly ameliorated DAI score, myeloperoxidase activity, histological score, apoptotic cells and protein expression of various inflammatory markers including nuclear factor kappa light chain enhancer of activated B cells (NFκBp65), cycloxygenase-2 (COX-2) and interleukin-6 (IL-6); proliferation markers such as proliferating cell nuclear antigen (PCNA), signal transducer and activator of transcription 3 (STAT3) expression significantly decreased, and antioxidant enzymes nuclear factor erythroid 2-related factor 2 (Nrf-2), metallothionein, catalase and superoxide dismutase (SOD) significantly increased as evaluated by immunohistochemistry and western blot analysis.

Dietary Supplementation with Black Raspberries Altered the Gut Microbiome Composition in a Mouse Model of Colitis-Associated Colorectal Cancer, although with Differing Effects for a Healthy versus a Western Basal Diet.

Black raspberries (BRB) are rich in anthocyanins with purported anti-inflammatory properties. However, it is not known whether dietary supplementation would ameliorate Western-diet enhanced gut inflammation and colon tumorigenesis. We employed a mouse model of colitis-associated colorectal cancer (CAC) to determine the effects of dietary supplementation with 5 to 10% (w/w) whole, freeze-dried BRB in male C57BL/6J mice fed either a standard healthy diet (AIN93G) or the total Western diet (TWD). In a pilot study, BRB suppressed colitis and colon tumorigenesis while also shifting the composition of the fecal microbiome in favor of taxa with purported health benefits, including Bifidobacterium pseudolongum. In a follow-up experiment using a 2 × 2 factorial design with AIN and TWD basal diets with and without 10% (w/w) BRB, supplementation with BRB reduced tumor multiplicity and increased colon length, irrespective of the basal diet, but it did not apparently affect colitis symptoms, colon inflammation or mucosal injury based on histopathological findings. However, BRB intake increased alpha diversity, altered beta diversity and changed the relative abundance of Erysipelotrichaceae, Bifidobacteriaceae, Streptococcaceae, Rikenellaceae, Ruminococcaceae and Akkermansiaceae, among others, of the fecal microbiome. Notably, changes in microbiome profiles were inconsistent with respect to the basal diet consumed. Overall, these studies provide equivocal evidence for in vivo anti-inflammatory effects of BRB on colitis and colon tumorigenesis; yet, BRB supplementation led to dynamic changes in the fecal microbiome composition over the course of disease development.

Quxie Capsule Alleviates Colitis-associated Colorectal Cancer Through Modulating the Gut Microbiota and Suppressing A. fumigatus-induced Aerobic Glycolysis.

AIM: Quxie capsule (QX), a compound of 21 kinds of Traditional Chinese Medicine (TCM) herbs, has been used to treat patients with metastatic colorectal cancer (mCRC) and could suppress the growth of colon cancer. However, the mechanisms of QX inhibiting colorectal cancer remain unclear. In current study, we attempted to determine the anti-colorectal cancer (CRC) effects of QX and the mechanisms of QX in alleviating colorectal cancer. METHODS: A colitis-associated colon cancer (CAC) model was established by intraperitoneally injecting mice with AOM followed by 3 cycles of 2% DSS in water. During establishment of CAC model, we orally gavaged mice with QX. Hematoxylin and eosin (H&E) and immunohistochemistry were performed to assess lesion of the colonic tumors. The expression of pro-inflammatory cytokines in colonic tumors was measured by qPCR. The proportion of immune cells in colonic tumors was analyzed by flow cytometry. Internal transcribed spacer (ITS) sequencing and 16S rRNA gene sequencing were performed to detect intestinal microbiota. The expression of glycolytic related enzymes, lactate production, and extracellular acidification rate (ECAR) were used to assess the level of aerobic glycolysis. RESULTS: QX markedly inhibited intestinal tumorigenesis by decreasing the expression of pro-inflammatory cytokines and the proportion of myeloid-derived suppressor cells (MDSCs), and increasing the proportion of CD8+ T cells in colon tumors. Fecal microbiota sequencing revealed that QX increased the relative abundances of intestinal symbiotic probiotics, such as, Lactobacillus, Bifidobacterium and Faecalibacterium genera. What's more, opportunistic pathogens, Bacteroides genera and Aspergillus-Aspergillus fumigatus, exhibited remarkably reduced abundances in mice treated with QX compared with untreated CAC mice. Further experiments showed that QX significantly reduced glycolysis of colon tumor and suppressed A. fumigatus-induced glycolytic metabolism of colon cancer cells. CONCLUSIONS: QX alleviates the development of CRC at least in part through modulating intestinal microbiota and reducing A. fumigatus-induced aerobic glycolysis of colon cancer cells.

Huang Qin Decoction inhibits the initiation of experimental colitis associated carcinogenesis by controlling the PAD4 dependent NETs.

BACKGROUND: Colorectal cancer is associated with ulcerative colitis (UC). The infiltration of neutrophils is the main cause of DNA damage produced by inflammation in the intestinal epithelium. Under the action of peptidyl arginine deaminase 4 (PAD4), neutrophils dissociate chromatin and form neutrophil extracellular traps (NETs), which can aggravate tissue inflammation and encourage tumor development. Although Huang Qin Decoction (HQD) was found to be useful in treating UC and was used to gradually prevent and treat digestive tract cancers, the underlying reasons were unclear. METHODS: To demonstrate HQD could inhibits the initiation of colitis associated carcinogenesis by controlling NETs related inflammation, we first performed an AOM/DSS-generated colitis-associated carcinogenesis model to assess the efficacy of HQD in reducing neutrophil infiltration and anti-tumor activity. Then, using network pharmacology research, we investigated the potential mechanisms underlying those medicinal effects, as demonstrated by the detection of NETs aggregation and PAD4 expression changes in the colon. RESULTS: HQD substantially reduced the number of colon cancers and the expression of Ki67, restored the level of intestinal tight junction protein occludin and ZO-1, and relieved the intestinal inflammation caused by TNF-α, IL-1ß. At the same time, it inhibited neutrophil infiltration in the colon and improved the immunosurveillance of CD8+T cells. The potential mechanisms of HQD intervention against UC and UC with neoplasia (UCN) were studied using network pharmacology, and 156 conjunct genes as well as numerous inflammation-related pathways were identified. Protein-protein interaction (PPI) analysis indicated that HQD inhibition of intestinal tumors might be related to the deactivation of PAD4, which was verified by the down-regulation of NETs, MPO-DNA complex levels, and PAD4 expression after HQD treatment. CONCLUSION: Huang Qin Decoction inhibits the initiation of colitis associated carcinogenesis by controlling PAD4-dependent neutrophil extracellular traps.

Wumei Pill Ameliorates AOM/DSS-Induced Colitis-Associated Colon Cancer through Inhibition of Inflammation and Oxidative Stress by Regulating S-Adenosylhomocysteine Hydrolase- (AHCY-) Mediated Hedgehog Signaling in Mice.

Wumei Pill (WMP) is a traditional Chinese herbal formulation and widely used to treat digestive system diseases in clinical. S-Adenosylhomocysteine hydrolase (AHCY) can catalyze the hydrolysis of S-adenosylhomocysteine to adenosine and homocysteine in living organisms, and its abnormal expression is linked to the pathogenesis of many diseases including colorectal cancer (CRC). A previous study reported that WMP could prevent CRC in mice; however, the underlying mechanisms especially the roles of AHCY in WMP-induced anti-CRC remain largely unknown. Here, we investigated the regulatory roles and potential mechanisms of AHCY in WMP-induced anti-CRC. WMP notably alleviated the azoxymethane/dextran sulfate sodium- (AOM/DSS-) induced colitis-associated colon cancer (CAC) in mice. Besides, WMP inhibited the inflammation and oxidative stress in AOM/DSS-induced CAC mice. AHCY was high expression in clinical samples of colon cancer compared to the adjacent tissues. WMP inhibited the AHCY expression in AOM/DSS-induced CAC mice. An in vitro study found that AHCY overexpression induced cell proliferation, colony formation, invasion, and tumor angiogenesis, whereas its knockdown impaired its oncogenic function. AHCY overexpression enhanced, while its knockdown weakened the inflammation and oxidative stress in colon cancer cells. Interestingly, WMP potently suppressed the hedgehog (Hh) signaling in AOM/DSS-induced CAC mice. A further study showed that AHCY overexpression activated the Hh signaling while AHCY knockdown inactivated the Hh signaling. Moreover, activation of the Hh signaling reversed the effect of AHCY silencing on inflammation and oxidative stress in vitro. In conclusion, WMP alleviated the AOM/DSS-induced CAC through inhibition of inflammation and oxidative stress by regulating AHCY-mediated hedgehog signaling in mice. These findings uncovered a potential molecular mechanism underlying the anti-CAC effect of WMP and suggested WMP as a promising therapeutic candidate for CRC.

Cortex periplocae modulates the gut microbiota to restrict colitis and colitis-associated colorectal cancer via suppression of pathogenic Th17 cells.

Aberrant microbe-immune cell interaction is a predisposing factor in inflammatory bowel disease (IBD) and colitis-associated cancer (CAC). Cortex Periplocae is a famous traditional Chinese medicine with putative anti-rheumatoid arthritis and anti-dyspepsia effects. Here, we show that the Periploca sepium periplosides (PePs), a cardiac glycosides-free pregnane glycosides extract from root bark of Cortex Periplocae, alleviates colon inflammation, improves intestinal epithelial barrier function, and prevents colitis-associated tumorigenesis in mice with colitis and CAC. Mechanistically, PePs treatment modulates abnormal gut microbiota composition in model mice, especially enriches an anti-inflammatory commensal bacterium A. muciniphila BAA-835. We further demonstrate that the altered gut microbiota following PePs treatment plays an important role in modulation of intestinal Type 17 immunity in both colitis and CAC mouse model. Our results indicate that PePs may be used as a potential gut microbiota modulator to treat IBD and CAC.

Berberine regulates short-chain fatty acid metabolism and alleviates the colitis-associated colorectal tumorigenesis through remodeling intestinal flora.

BACKGROUND: Colitis-associated cancer (CAC) is known to be a complex combination of tumor cells, non-tumor cells and a large intestinal flora. The increasing role of intestinal flora in CAC may represent a new approach to improving CAC treatment. Berberine can reduce colorectal adenoma recurrence and inhibit colorectal carcinogenesis. PURPOSE: Berberine has demonstrated efficacy for the control and suppression of CAC. Given the low oral absorption into the blood and large intestinal excretion of berberine, intestinal flora may be one of the important targets of berberine inhibiting the occurrence of colorectal cancer (CRC). The purpose of this study was to investigate the effects of berberine on intestinal flora in CAC mice and its ability to remodel intestinal flora to improve short-chain fatty acid metabolism. STUDY DESIGN AND METHODS: The CAC model in mice was induced by Azoxymethane/Dextran sodium sulfate (AOM/DSS). Berberine was administered daily at doses of 50 and 100 mg/kg, and aspirin was used as the positive control. The effect of berberine on colitis-associated colorectal tumorigenesis was assessed by general imaging, tumor counting, and Ki67 staining. Intestinal flora changes were detected by 16S rDNA sequencing technology. Targeted short-chain fatty acid detection was performed by GC-MS/MS, and Lipopolysaccharide (LPS) levels in feces were quantified with an ELISA kit. The signaling pathway of TLR4/NF-κB P65/IL-6/p-STAT3 was evaluated by Western blotting and immunofluorescence. The expression levels of intestinal barrier functional biomarkers Occludin and ZO-1 were detected by immunohistochemistry. Fecal flora transplantation (FMT) was used to evaluate the effect of intestinal flora in inhibiting inflammatory cancer transformation by berberine. RESULTS: Berberine reduced the number and load of tumors in CAC mice. Berberine remodeled the composition of pathogenic and beneficial bacteria in mice with colitis-associated colorectal tumorigenesis. Berberine treatment resulted in increases in fecal butyric acid, acetic acid and propionic acid levels, but did not alter isobutyric acid, isovaleric acid, valeric acid and caproic acid. In addition, berberine reduced LPS content in feces in mice with colitis-associated colorectal tumorigenesis. Occludin and ZO-1 were upregulated, and the TLR4/p-NF-κB p65/IL-6/p-STAT3 inflammatory-cancer transformation pathway was inhibited with berberine. The FMT results further verified that the berberine-treated intestinal flora was sufficient to alleviate the occurrence of colonic tumors associated with colitis in mice. CONCLUSION: Our study showed that berberine alleviated the colitis-associated colorectal tumorigenesis from three equilibrium levels: (1) Pathogenic and beneficial bacteria; (2) Short-chain fatty acids and LPS produced by intestinal flora; and (3) Inflammatory cancer transformation signaling and intestinal barrier function. This study provided a new approach and experimental basis for the application of berberine in the treatment of CAC in clinical practice.

Ojeok-san ameliorates visceral and somatic nociception in a mouse model of colitis induced colorectal cancer.

Cancer patients can develop visceral, somatic, and neuropathic pain, largely due to the malignancy itself and its treatments. Often cancer patients and survivors turn to the use of complementary and alternative medicine (CAM) to alleviate pain and fatigue. Thus, it is necessary to investigate how CAM therapies work as novel analgesics to treat cancer pain. Ojeok-san (OJS) is an herbal formula consisting of seventeen herbs. This herbal formula has been shown to possess anti-inflammatory, immunoregulatory, and analgesic properties. In this study, we examined the potential beneficial effects and mechanism of action of OJS in a preclinical model of colitis-associated colorectal cancer. Male and female C57BL/6J mice were exposed to the carcinogen, azoxymethane (AOM, 10 mg/kg) and a chemical inflammatory driver, dextran sulfate sodium (DSS1-2%), to promote tumorigenesis in the colorectum. OJS was given orally (500, 1000, and 2000 mg/kg) to determine its influence on disease activity, tumor burden, nociception, sedation, Erk signaling, and behavioral and metabolic outcomes. In addition, in vitro studies were performed to assess CT-26 cell viability, dorsal root ganglia (DRG) activation, and bone-marrow-derived macrophage (BMDM) inflammatory response to lipopolysaccharide stimulation after OJS treatment. We found that administration of 2000 mg/kg of OJS was able to mitigate mechanical somatic and visceral nociception via Erk signaling without affecting symptom score and polyp number. Moreover, we discovered that OJS has sedative properties and elicits prolonged total sleeping time in AOM/DSS mice. Our in vitro experiments showed that OJS has the capacity to reduce TNFα gene expression in LPS-stimulated BMDM, but no changes were observed in DRG spike number and CT-26 cell proliferation. Taken together, these data suggest that OJS ameliorates nociception in mice and warrants further examination as a potential CAM therapy to promote analgesia.