Causal roles and clinical utility of cardiovascular proteins in colorectal cancer risk: a multi-modal study integrating mendelian randomization, expression profiling, and survival analysis.

PURPOSE: This comprehensive investigation delved into the intricate causal interplay existing between cardiovascular-related plasma proteins and the susceptibility to colorectal cancer, leveraging the robust framework of Mendelian randomization, and employed expression profiling and survival analysis to unravel the latent clinical worth embedded within pertinent gene expressions. METHODS: Protein quantitative trait loci (pQTLs) of 85 cardiovascular proteins were employed as instrumental variables to investigate the causal relationship between proteins and CRC risk using a Mendelian randomization approach. Causal inferences were graded as strong, intermediate or weak based on statistical checks. Drug-target MR examined VEGF receptors for their potential as therapeutic targets for colorectal cancer. Differential expression analysis, diagnostic ROC curves, and survival analyses were performed for identified proteins using RNA-seq data from The Cancer Genome Atlas (TCGA) colorectal cancer cohort. RESULTS: Using cis-pQTLs, LOX-1, VEGF-A and OPG were associated with increased CRC risk (strong evidence), while PTX3, TNF-R2 and MMP-7 were protective (strong evidence). Pan-pQTL analysis found MMP-10 increased risk (intermediate evidence) and ADM increased risk (weak evidence). Drug-target MR found VEGF R1 may be promising therapeutic targets. Differential expression analysis revealed seven genes encoding the identified proteins were dysregulated in tumors. ROC analysis showed five gene expression had high diagnostic accuracy. KM analysis showed four genes had prognostic value. CONCLUSIONS: This large-scale MR study implicates several cardiovascular proteins in CRC susceptibility and progression. Findings highlight roles for VEGF signaling and extracellular matrix regulation. Results nominate specific proteins as potential diagnostic biomarkers or therapeutic targets warranting further investigation.

Activation of free fatty acid receptors, FFAR1 and FFAR4, ameliorates ulcerative colitis by promote fatty acid metabolism and mediate macrophage polarization.

OBJECTIVE: To investigate the mechanism of action of fatty acid receptors, FFAR1 and FFAR4, on ulcerative colitis (UC) through fatty acid metabolism and macrophage polarization. METHODS: Dextran sulfate sodium (DSS)-induced mouse model of UC mice was used to evaluate the efficacy of FFAR1 (GW9508) and FFAR4 (GSK137647) agonists by analyzing body weight, colon length, disease activity index (DAI), and histological scores. Real-time PCR and immunofluorescence analysis were performed to quantify the levels of fatty acid metabolizing enzymes and macrophage makers. FFA-induced lipid accumulation in RAW264.7 cells was visualized by Oil Red O staining analysis, and cells were collected to detect macrophage polarization by flow cytometry. RESULTS: The combination of GW9508 and GSK137647 significantly improved DSS-induced UC symptoms, caused recovery in colon length, and decreased histological injury. GW9508 + GSK137647 treatment upregulated the expressions of CD206, lipid oxidation enzyme (CPT-1α) and anti-inflammatory cytokines (IL-4, IL-10, IL-13) but downregulated those of CD86, lipogenic enzymes (ACC1, FASN, SCD1), and pro-inflammatory cytokines (IL-1ß, IL-6, TNF-α). Combining the two agonists decreased FFA-induced lipid accumulation and increased CD206 expression in cell-based experiments. CONCLUSION: Activated FFAR1 and FFAR4 ameliorates DSS-induced UC by promoting fatty acid metabolism to reduce lipid accumulation and mediate M2 macrophage polarization.

Huangqin Decoction ameliorates ulcerative colitis by regulating fatty acid metabolism to mediate macrophage polarization via activating FFAR4-AMPK-PPARα pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Huangqin Decoction (HQD), a traditional Chinese medicine (TCM) formula chronicled in Shang Han Lun, is safe and effective for treatment of ulcerative colitis (UC). AIM OF THE STUDY: To investigate the effect of HQD against dextran sulfate sodium (DSS)-induced UC mice by regulating gut microbiota and metabolites, and further explore the mechanism of fatty acid metabolism on macrophage polarization. MATERIALS AND METHODS: Based on 3% dextran sulfate sodium (DSS)-induced UC mice model, clinical symptoms observation (body weight, DAI, and colon length) and histological inspection were used to evaluate the efficacy of HQD and fecal microbiota transplantation (FMT) from HQD-treated mice. The gut microbiota and metabolites were detected by 16S rRNA sequencing and metabolomics analysis. The parameters of fatty acid metabolism, macrophage polarization, and FFAR1/FFAR4-AMPK-PPARα pathway were analyzed by immunofluorescence analysis, western blotting, and real-time PCR. Then, the effects of FFAR1 and FFAR4 on macrophage polarization were examined by agonists based on LPS-induced RAW264.7 cell model. RESULTS: The results showed that FMT, like HQD, ameliorated UC by improving weight loss, restoring colon length, and reducing DAI scores and histopathological scores. Besides, HQD and FMT both enhanced the richness of gut microbiota, and modulated intestinal bacteria and metabolites to achieve a new balance. Untargeted metabolomics analysis revealed that fatty acids, especially long-chain fatty acids (LCFAs), dominated in HQD against DSS-induced UC by regulating the gut microenvironment. Further, FMT and HQD recovered the expression of fatty acid metabolism-related enzymes, and simultaneously activated FFAR1/FFAR4-AMPK-PPARα pathway but suppressed NF-κB pathway. Combined with cell experiment, HQD and FMT promoted macrophage polarization from M1 toward M2, which were well associated with anti-inflammatory cytokines and combined with the activated FFAR4. CONCLUSIONS: The mechanism of HQD against UC was related to regulating fatty acid metabolism to mediate M2 macrophage polarization by activating the FFAR4-AMPK-PPARα pathway.

Huangqin decoction ameliorates DSS-induced ulcerative colitis: Role of gut microbiota and amino acid metabolism, mTOR pathway and intestinal epithelial barrier.

BACKGROUND: The clinical treatment of ulcerative colitis (UC) is limited. A traditional Chinese medicinal formula, Huangqin decoction (HQD), is chronicled in Shang Han Lun and is widely used to ameliorate gastrointestinal disorders, such as UC; however, its mechanism is yet to be clarified. PURPOSE: The present study aimed to investigate the effect of HQD on 7-day colitis induced by 3% dextran sulfate sodium (DSS) in mice and further explore the inhibitory effect of metabolites on DSS-damaged FHC cells. METHODS: The therapeutic efficacy of HQD was evaluated in a well-established DSS-induced colitis mice model. The clinical symptoms were analyzed, and biological samples were collected for microscopic examination, metabolomics, metagenomics, and the evaluation of the epithelial barrier function. The mechanism of metabolites regulated by HQD was evaluated in the DSS-induced FHC cell damage model. The samples were collected to detect the physiological functions of the cells. RESULTS: HQD suppressed the inflammation of DSS-induced colitis in vivo, attenuated DSS-induced clinical manifestations, reversed colon length reduction, and reduced histological injury. After HQD treatment, the DSS-induced gut dysbiosis was modulated, and the gut microbiota achieved a new equilibrium state. In addition, HQD activated the mTOR signaling pathway by upregulating amino acid metabolism. Significant phosphorylation of S6 and 4E-BP1 ameliorated intestinal epithelial barrier dysfunction. Moreover, HQD-regulated metabolites protected the epithelial barrier integrity by inhibiting DSS-induced apoptosis of FHC cells and regulating the proteins affecting apoptosis and cell-cell junction. CONCLUSIONS: These findings indicated that the mechanism of HQD was related to regulating the gut microbiota and amino acid metabolism, activating the mTOR signaling pathway, and protecting the intestinal mucosal barrier integrity.

Shaoyao Decoction Inhibits Inflammation and Improves Intestinal Barrier Function in Mice With Dextran Sulfate Sodium-Induced Colitis.

Shaoyao decoction (SYD), a classical traditional Chinese medicine formula, is effective for the treatment of inflammatory bowel disease (IBD). This study was designed to investigate the therapeutic effects of SYD on IBD and possible mechanisms. Dextran sulfate sodium (DSS, 3.5%) was used to induce colitis in C57BL/6 mice. Disease phenotypes were investigated based on disease activity index (DAI), colon length, and microscopic and macroscopic scores. Additionally, the presence of proinflammatory cytokines, immune cell infiltrates, intestinal cell proliferation, apoptosis, epithelial permeability, signal transducer and activator of transcription 3 (STAT3), and nuclear factor-κB (NF-κB) signaling, as well as the intestinal mucosal barrier function, were investigated. The administration of SYD significantly ameliorated the clinical signs, suppressed the levels of proinflammatory cytokines, and reduced immune cell infiltrates into colonic tissues of DSS-induced colitis model mice. SYD also significantly reduced the DSS-induced activation of STAT3 and NF-κB signaling. Furthermore, SYD promoted epithelial integrity by regulating epithelial cell apoptosis and epithelial permeability. Finally, we demonstrated that SYD protected the intestinal barrier function by significantly regulating the mucus layer genes Muc1, Muc2, Muc4, and Tff3, as well as the epithelial barrier genes Z O -1 and Occludin. Our results indicate that SYD has a protective effect on DSS-induced colitis, which is attributable to its anti-inflammatory activity and intestinal barrier function-enhancing effects. These results provide valuable insights into the pharmacological actions of SYD for the treatment of IBD.

Effects of Huangqin Decoction on ulcerative colitis by targeting estrogen receptor alpha and ameliorating endothelial dysfunction based on system pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Huangqin Decoction (HQD), a traditional Chinese medicinal (TCM) formula chronicled in Shang Han Lun, has been used to treat gastrointestinal diseases for nearly 1800 years. OBJECTIVE: To investigate the effects and underlying mechanisms of HQD on ulcerative colitis (UC). METHODS: The bioactive compounds in HQD were obtained from the traditional Chinese medicine systems pharmacology database. Then, the HQD and UC-related targets were analyzed by establishing HQD-Compounds-Targets (H-C-T) and protein-protein interaction (PPI) networks. Enrichment analysis was used for further study. The candidate targets for the effects of HQD on UC were validated using a dextran sulfate sodium-induced UC mouse experiment. RESULTS: The results showed that 51 key targets were gained by matching 284 HQD-related targets and 837 UC-related targets. Combined with H-C-T and PPI network analyses, the key targets were divided into endothelial growth, inflammation and signal transcription-related targets. Further experimental validation showed that HQD targeted estrogen receptor alpha (ESR1) and endothelial growth factor receptors to relieve endothelial dysfunction, thereby improving intestinal barrier function. The expression of inflammatory cytokines and signal transducers was suppressed by HQD treatment and inflammation was inhibited. CONCLUSIONS: HQD may acts on UC via the regulation of targets and pathways related to improving the intestinal mucosal barrier and ameliorating endothelial dysfunction. Additionally, ERS1 may be a new target to explore the mechanisms of UC.

Herbal pair Huangqin-Baishao: mechanisms underlying inflammatory bowel disease by combined system pharmacology and cell experiment approach.

BACKGROUND: Inflammatory bowel disease (IBD) is a severe digestive system condition, characterized by chronic and relapsing inflammation of the gastrointestinal tract. Scutellaria baicalensis Georgi (Huangqin, HQ) and Paeonia lactiflora Pall (Baishao, BS) from a typical herbal synergic pair in traditional Chinese medicine (TCM) for IBD treatments. However, the mechanisms of action for the synergy are still unclear. Therefore, this paper aimed to predict the anti-IBD targets and the main active ingredients of the HQ-BS herbal pair. METHODS: A systems pharmacology approach was used to identify the bioactive compounds and to delineate the molecular targets and potential pathways of HQ-BS herbal pair. Then, the characteristics of the candidates were analyzed according to their oral bioavailability and drug-likeness indices. Finally, gene enrichment analysis with DAVID Bioinformatics Resources was performed to identify the potential pathways associated with the candidate targets. RESULTS: The results showed that, a total of 38 active compounds were obtained from HQ-BS herbal pair, and 54 targets associated with IBD were identified. Gene Ontology and pathway enrichment analysis yielded the top 20 significant results with 54 targets. Furthermore, the integrated IBD pathway revealed that the HQ-BS herbal pair probably acted in patients with IBD through multiple mechanisms of regulation of the nitric oxide biosynthetic process and anti-inflammatory effects. In addition, cell experiments were carried out to verify that the HQ-BS herbal pair and their Q-markers could attenuate the levels of nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in lipopolysaccharide (LPS)-stimulated THP-1-derived macrophage inflammation. In particular, the crude materials exerted a much better anti-inflammatory effect than their Q-markers, which might be due to their synergistic effect. CONCLUSION: This study provides novel insight into the molecular pathways involved in the mechanisms of the HQ-BS herbal pair acting on IBD.

Deoxyshikonin isolated from Arnebia euchroma inhibits colorectal cancer by down-regulating the PI3K/Akt/mTOR pathway.

Context: Shikonins, a series of natural occurring naphthoquinones extracted from Arnebia euchroma (Royle) Jonst. (Boraginaceae), have antitumor activities and low toxicity. Objective: To illuminate potential activity and mechanism of shikonins against colorectal cancer (CRC). Materials and methods: Five shikonins were isolated from A. euchroma, and elucidated by extensive spectroscopic analysis. Anti-proliferative activities of shikonins (0-100 µg/mL) on human colorectal cells were evaluated by MTT and CCK-8 for 24 or 48 h. Cell apoptosis and cycle distribution were examined by FCM analysis. The expression of PI3K/Akt/mTOR pathway mRNAs and proteins was analysed by RT-PCR and Western blot, respectively. Cell viability, cell apoptosis, cell cycle and protein expression were measured, when co-treated with PI3K/Akt/mTOR pathway inhibitors. The in vivo activity of deoxyshikonin was evaluated using xenograft tumour model. Results: Deoxyshikonin and another four shikonins were isolated and identified. Deoxyshikonin exhibited anti-proliferative activity with IC50 of 10.97 µM against HT29 cells. Moreover, the percentage of early apoptotic cells and G0/G1 cells increased from 1 to 29% and 44 to 67% with 0-50 µg/mL deoxyshikonin, respectively. Deoxyshikonin also down-regulated the expression of PI3K, p-PI3K, Akt, p-Akt308 and mTOR proteins in HT29 and DLD-1 cells. Moreover, LY294002, NVP-BEZ235 and MK-2206 can make deoxyshikonin more cell proliferation inhibited, cell cycle arrested at G0/G1 and apoptosis promoted. In vivo study, the weight of tumour tissues at deoxyshikonin groups was significantly reduced compared with the control group, and PI3K, p-PI3K, Akt, p-Akt308 and mTOR expression was decreased. Discussion and conclusions: We can conclude that deoxyshikonin isolated from Arnebia euchroma inhibited CRC through the PI3K/Akt/mTOR pathway.

[Effect of Shaoyao Tang on ulcerative colitis in rats via regulation of TLR4/NF-κB signal pathway].

The aim of this paper was to investigate the effect of Shaoyao Tang on ulcerative colitis(UC) in rats via regulation of TLR4/NF-κB signal pathway. A total of 56 Wistar rats were randomly divided into 6 groups: normal control group(double distilled water), model group(double distilled water), mesalazine group(10 mL·kg~(-1)), high dose, middle dose and low dose Shaoyao Tang groups(2.4, 1.2, and 0.6 g·mL~(-1)). After UC rat models were established by 2, 4-dinitrochlorobenzene(DNCB)/ethanol enema, the rats received double distilled water or corresponding drugs twice a day for 7 days. After the treatment cycle, the general performance and disease activity index(DAI) of rats were observed on the next day. Then the rats were sacrificed. The length of colon was measured. Macroscopic and histological score of colon were evaluated. Histopathological changes of colon were observed by HE staining. Ultraviolet spectrophotometry detection was used to detect the content of myeloperoxidase(MPO) in blood and colon tissues. The levels of P-selectin, macrophage migration inhibitory factor(MIF) and thromboxane B_2(TXB_2) in blood and colon tissues were determined by ELISA. Immunohistochemistry and Western blot analysis were performed to detect the protein expressions of TLR4 and NF-κB in colon tissues. The results showed that as compared with the model group, Shaoyao Tang of different doses improved the general performance of UC rats. Moreover, high-dose Shaoyao Tang group showed the most obvious effect in scoring of disease activity index(P<0.001); both medium and high doses of Shaoyao Tang significantly inhibited the colon shortening and pathological injury, with significantly decreased expression levels of MPO, P-selectin, MIF and TXB_(2 )in serum and colon tissues of UC rats(P<0.001). Immunohistochemistry and Western blot assay showed that the levels of TLR4 and NF-κB protein expression in the colon tissues of Shaoyao Tang high-dose group were remarkably lower than that in the model group(P<0.001). This study shows that Shaoyao Tang has protective and repairing effects on UC, and its possible mechanism is achieved probably by regulating the TLR4/NF-κB pathway and inhibiting the expressions of MPO, P-selectin, MIF and TXB_2.

Anti-Inflammatory Effects of Huangqin Decoction on Dextran Sulfate Sodium-Induced Ulcerative Colitis in Mice Through Regulation of the Gut Microbiota and Suppression of the Ras-PI3K-Akt-HIF-1α and NF-κB Pathways.

OBJECTIVE: Huangqin decoction (HQD), a classical traditional Chinese medicinal formula, has been commonly used to treat gastrointestinal diseases for thousands of years. We investigated the anti-inflammatory effects and underlying mechanisms of HQD on dextran sulfate sodium (DSS)-induced ulcerative colitis (UC). METHODS: Experimental mice were given 3% DSS, and HQD (2.275, 4.55, and 9.1 g/kg), or mesalazine (ME, 200 mg/kg) orally for 7 days. Body weight loss, disease activity index (DAI), colon length, histology, and levels of inflammatory cytokines were measured to evaluate the effects of HQD on colitis. The effects of HQD on the Ras-phosphoinositide-3-kinase (PI3K)-Akt-hypoxia inducible factor 1 alpha (HIF-1α) and nuclear factor-kappa B (NF-κB) pathways were evaluated by Western blot analysis. In addition, the gut microbiota was characterized using high-throughput Illumina MiSeq sequencing. RESULTS: The results showed that HQD significantly reduced the body weight loss, ameliorated DAI, restored colon length, and improved the intestinal epithelial cell barrier in mice with DSS-induced colitis. The messenger RNA (mRNA) expression levels of inflammatory mediators were decreased following HQD treatment. Furthermore, the Ras-PI3K-Akt-HIF-1α and NF-κB pathways were significantly inhibited by HQD. Finally, treatment with HQD resulted in recovery of gut microbiota diversity. CONCLUSIONS: HQD ameliorates DSS-induced colitis through regulation of the gut microbiota, and suppression of Ras-PI3K-Akt-HIF-1α and NF-κB pathways. Our results suggested that HQD may be a potential candidate for treatment of UC.

[Study of isobutyrylshikonin inhibiting proliferation of colon carcinoma cells through PI3K/Akt/m-TOR pathway].

To investigate the inhibitory effect of isobutyrylshikonin on the growth of human colon carcinoma cells in vitro and its effect on the PI3K/Akt/m-TOR pathway. MTT assay was used to detect the inhibitory effect of different concentrations (0, 6.25, 12.5, 25, 50, 100 mg·L⁻¹) of isobutyrylshikonin on the proliferation of human colon carcinoma cell HT29 at 24, 48 h. CCK-8 method was used to detect the inhibitory effect of isobutyrylshikonin on HT29, HCT116, DLD-1 and Caco-2 at 48 h. AnnexinV/propidium iodide staining was applied in detecting the apoptoticrate of HT29 cells treated with different concentrations of isobutyrylshikonin at 24 h and 48 h. Cycletest plus DNA was employed to analyze HT29 apoptosis and cell cycle after 48 h treatment with isobutyrylshikonin at different concentrations. Western blot and RT-PCR assay were used to examine the protein and mRNA expressions of PI3K, p-PI3K, Akt, p-Akt and m-TOR. The results showed that isobutyrylshikonin inhibited the proliferation of different human colon carcinoma cells, and the inhibition rate was in a dose-dependent manner. Isobutyrylshikonin induced apoptosis mainly in the early stage and blocked cells in the G0/G1 or G2/M phase. Isobutyrylshikonin reduced the protein expressions of PI3K, p-PI3K, Akt, p-Akt, m-TOR and the mRNA expressions of PI3K, Akt, m-TOR in a dose-dependent manner. Isobutyrylshikonin can significantly inhibit the proliferation, induce the early apoptosis and change the cycle distribution in colon carcinoma cells.This biological effect may be correlated with the inhibition of PI3K/AKT/m-TOR pathway.

Aloperine Protects Mice against DSS-Induced Colitis by PP2A-Mediated PI3K/Akt/mTOR Signaling Suppression.

Colitis is a major form of inflammatory bowel disease which involved mucosal immune dysfunction. Aloperine is an alkaloid isolated from the shrub Sophora alopecuroides L. and has been recognized as an effective treatment for inflammatory and allergic diseases. The present study aimed to examine the molecular mechanisms underlying aloperine-mediated colitis protection. We found that aloperine treatment improved colitis induced by dextran sodium sulfate (DSS) based on body weight, disease activity index, colonic length, and spleen index. Aloperine also effectively attenuated DSS-induced intestinal inflammation based on the pathological score and myeloperoxidase expression and activity in colon tissues. In addition, aloperine regulated T-cell proportions and promoted Foxp3 expression in the spleens and mesenteric lymph nodes of DSS-induced colitis mice and in the spleens of the Foxp3GFP mice. Aloperine inhibited Jurkat and mouse naïve T-cell apoptosis. Furthermore, aloperine inhibited PI3K/Akt/mTOR signaling and upregulated PP2A expression in the DSS-induced colitis mice and in Jurkat cells, but LB-100 (PP2A inhibitor) resulted in an elevated Akt activity in Jurkat cells, activated T-cells, and human splenic mononuclear cells. Aloperine inhibited T-cell and lymphocyte proliferation, but LB-100 reverse these effects. In conclusion, aloperine regulates inflammatory responses in colitis by inhibiting the PI3K/Akt/mTOR signaling in a PP2A-dependent manner.

Huangqin-tang ameliorates dextran sodium sulphate-induced colitis by regulating intestinal epithelial cell homeostasis, inflammation and immune response.

Huangqin-tang (HQT) is a traditional Chinese medicine (TCM) formula widely used for the treatment of inflammatory bowel disease in China. However, the molecular mechanisms by which HQT protects the colon are unclear. We studied the protective effects of HQT and the underlying mechanisms in an experimental mouse model and in vitro. In vivo, dextran sodium sulphate (DSS)-induced acute and chronic colitis were significantly ameliorated by HQT as gauged by phenotypic, histopathologic and inflammatory manifestations of the disease. Mechanistically, DSS-induced nuclear factor-κB (NF-κB) signalling was inhibited by HQT. Moreover, HQT-treated mice demonstrated significant changes in cell apoptosis, expression of apoptosis-associated genes such as caspase-3, bax, bcl-2, and intestinal permeability. HQT also increased occluding and zonula occludens-1 (ZO-1), inhibited cell proliferation (Ki67), and increased regulatory T cells numbers, protein expression of Foxp3 and IL-10 in the colonic tissue. In vitro, HQT down-regulated production of pro-inflammatory cytokines and supressed the NF-κB signalling pathway in lipopolysaccharides-induced RAW 264.7 macrophages. Our study suggests that HQT plays a critical role in regulating intestinal epithelial cell homeostasis, inflammation and immune response in colitis and offers novel therapeutic options in the management of inflammatory bowel disease.

Comparison of 5-hydroxytryptophan signaling pathway characteristics in diarrhea-predominant irritable bowel syndrome and ulcerative colitis.

AIM: To study differences in the visceral sensitivity of the colonic mucosa between patients with diarrhea-predominant irritable bowel syndrome (IBS-D) and those with ulcerative colitis (UC) in remission and to relate these differences with changes in the 5-hydroxytryptophan (5-HT) signaling pathway. METHODS: Gastrointestinal symptoms were used to determine the clinical symptom scores and rectal visceral sensitivity of patients with IBS-D and patients with UC in remission. Blood levels of 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) were measured using an HPLC-electrochemical detection system. The levels of 5-HT 3 receptor (3R), 4R, and 7R mRNAs in colonic biopsy samples were detected using reverse transcription-polymerase chain reaction. The protein expression of TPH1 was analyzed by Western blot and immunohistochemistry. RESULTS: Abdominal pain or discomfort, stool frequency, and the scores of these symptoms in combination with gastrointestinal symptoms were higher in the IBS-D and UC groups than in the control groups. However, no significant differences were observed between the IBS-D and UC remission groups. With respect to rectal visceral sensitivity, the UC remission and IBS-D groups showed a decrease in the initial perception threshold, defecating threshold and pain threshold. However, these groups exhibited significantly increased anorectal relaxation pressure. Tests examining the main indicators of the 5-HT signaling pathway showed that the plasma 5-HT levels, 5-HIAA concentrations, TPH1 expression in the colonic mucosa, and 5-HT3R and 5-HT5R expression were increased in both the IBS-D and the UC remission groups; no increases were observed with respect to 5-HT7R expression. CONCLUSION: The IBS-D and UC groups showed similar clinical symptom scores, visceral sensitivity, and levels of serotonin signaling pathway indicators in the plasma and colonic mucosa. However, the pain threshold and 5-HT7R expression in the colonic mucosa were significantly different between these groups. The results reveal that (1) IBS-D and UC are related to visceral sensitivity pathogenesis and the clinical manifestations of these conditions and (2) the observed differences in visceral hypersensitivity are possibly due to differences in levels of the 5-HT7 receptor, a component of the 5-HT signaling pathway.

Growth inhibition effects of ent-11α-hydroxy-15-oxo-kaur-16-en-19-oic-acid on colorectal carcinoma cells and colon carcinoma-bearing mice.

The aim of the present study was to investigate the mechanism underlying the antitumor effects of ent-11α-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F) in colorectal cancer (CRC). 5F was isolated and used to treat C26 murine colon carcinoma cells, a xenograft tumor mouse model (induced by C26 cells) and a CRC mouse model [induced by 1,2-dimethylhydrazine (DMH)/dextran sodium sulfate (DSS)]. C26 cell growth was inhibited by 5F in a dose- and time-dependent manner in vitro. In addition, 5F induced cell apoptosis and cell cycle arrest in the G2 phase, increased the activity of caspase-3 and caspase-9, but did not affect the activity of cascase­8, suggesting that 5F induced apoptosis via activation of the mitochondrial signaling pathway rather than the death­receptor signaling pathway. Furthermore, treatment of C26 cells with 5F resulted in upregulation of cyclin­dependent kinase inhibitor 1A (p21, Cip1), Bcl­2­associated X protein, nuclear factor of κ light polypeptide gene enhancer in B­cells inhibitor, α and downregulation of B­cell lymphoma 2, nuclear factor κ­light­chain enhancer of activated B cells and survivin. In vivo animal models demonstrated that 5F treatment protected mice from carcinogenesis induced by DMH/DSS and markedly decreased the xenograft tumor weight with minimal side effects. Therefore, 5F may have potential as an anti-CRC therapeutic agent for use in the clinical setting.

Huangqin-Tang Ameliorates TNBS-Induced Colitis by Regulating Effector and Regulatory CD4(+) T Cells.

Huangqin-Tang decoction (HQT) is a classic traditional Chinese herbal formulation that is widely used to ameliorate the symptoms of gastrointestinal disorders, including inflammatory bowel disease (IBD). This study was designed to investigate the therapeutic potential and immunological regulatory activity of HQT in experimental colitis in rats. Using an animal model of colitis by intrarectally administering 2,4,6-trinitrobenzenesulfonic acid (TNBS), we found that administration of HQT significantly inhibited the severity of TNBS-induced colitis in a dose-dependent manner. In addition, treatment with HQT produced better results than that with mesalazine, as shown by improvedweight loss bleeding and diarrhoea scores, colon length, and intestinal inflammation. As for potential immunological regulation of HQT action, the percentages of Th1 and Th17 cells were reduced, but those Th2 and Treg cells were enhanced in LPMCs after HQT treatment. Additionally, HQT lowered the levels of Th1/Th17-associated cytokines but increased production of Th2/Treg-associated cytokines in the colon and MLNs. Furthermore, we observed a remarkable suppression of the Th1/Th17-associated transcription factors T-bet and ROR-γt. However, expression levels of the Th2/Treg-associated transcription factors GATA-3 and Foxp3 were enhanced during treatment with HQT. Our results suggest that HQT has the therapeutic potential to ameliorate TNBS-induced colitis symptoms. This protective effect is possibly mediated by its effects on CD4(+) T cells subsets.

[Effect of baicalin on signal transduction and activating transcription factor expression in ulcerative colitis patients].

OBJECTIVE: To explore the intervention of baicalin on signal transduction and activating transcription factor expression of ulcerative colitis (UC) patients. METHODS: Recruited were UC patients at Outpatient Department of Digestive Disease, Inpatient Department of Digestive Disease, Center for Digestive Endoscopy of College City Branch, Guangdong Provincial Hospital of Traditional Chinese Medicine, and Southern Hospital affiliated to Southern Medical University from June 2010 to January 2011. They were assigned to the UC group (33 cases) and the diarrhea-predominant irritable bowel syndrome (IBS-D) group (30 cases). Another 30 healthy subjects were recruited as a healthy control group. Peripheral blood mononuclear cells (PBMCs) in vitro intervened by different concentrations baicalin were taken from UC patients. IL23R gene expressions in vitro intervened by different concentrations baicalin were detected using Q-PCR. Expressions of signal transducer and activator of transcription 4 (STAT4) , STAT6, phosphorylated-STAT4 (p-STAT4), and p-STAT6 were detected using Western blot. Serum levels of IFN-γ, IL-4, IL-6, and IL-10 were measured by ELISA. Effects of different concentrations baicalin on expressions of PBMCs, and levels of IFN-γ, IL-4, IL-10 of UC patients were also detected. RESULTS: Compared with the negative control group, 40 µmol baicalin obviously decreased IL23R gene expression of UC patients (P <0. 01). Compared with the healthy control group and the IBS-D group, p-STAT4/STAT4 ratios increased, p-STAT6/STAT6 ratios decreased, levels of IFN-γ, IL-4, IL-10 all increased in the US group (all P <0. 05). Compared with the negative control, 5 and 10 µmol baicalin groups, 20 and 40 moL baicalin obviously decreased p-STAT4/STAT4 ratios (all P <0. 05); 20 and 40 µmoL baicalin obviously increased p-STAT6/STAT6 ratios (all P <0. 05); 20 and 40 µmoL baicalin obviously lowered levels of IFN-γ and IL-4, and elevated IL-10 levels (all P <0. 05). CONCLUSION: 40 µmoL baicalin could in vitro inhibit p-STAT4/STAT4 ratios, adjust p-STAT6/STAT6 ratios and related cytokines, thereby balancing the immunity and relieving inflammatory reactions of UC.

A critical role of mir-199a in the cell biological behaviors of colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is one of the most common cancer and the leading causes of cancer mortality worldwide. The critical role of hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) are important in the cancer development. METHODS: The purpose of this study was to investigate the association of miR-199a expression in CRC and non-tumor tissues as well as assessed the effect of miR-199a on biological behaviors including cell proliferation, apoptosis, migration and invasion of CRC cells. The expression of miR-199a was distinctly decreased in colorectal cancer tissues compared with non-neoplastic colorectal tissues. RESULTS: In this study, we found that miR-199a down-regulation was associated with the CRC and metastasis incidence. Advanced study showed that miR-199a up-regulation would lead to decreased CRC proliferation, migration and invasion. However, no significant association of miR-199a treatment and apoptosis rate and cell-cycle were detected in this study. The detection for the mechanisms of miR-199a on the development of CRC showed that the anticarcinogenic effect of miR-199a might be produced through HIF-1α/VEGF pathway. CONCLUSION: It was found that miR-199a would reduce the proliferation, migration and invasion. However, overexpression of miR-199a on the apoptosis rate and cell cycles showed no significant results. The potential functionary mechanism of miR-199a might through HIF-1α/VEGF pathway. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9806714131513041 .

CD4+CD29+T cells are blamed for the persistent inflammatory response in ulcerative colitis.

Ulcerative colitis (UC) is a chronic gastrointestinal disorder eliciting occurrence of colorectal cancer, the third most common human malignancy. The diagnosis of UC is based on clinical symptoms combined with typical findings on endoscopy, radiology, and ultimately pathology. We investigated the variation trend of CD4+CD29+T cells together with MPO, VCAM-1 in different periods of rat UC model and UC patients. We also evaluated the relationship between CD4+CD29+T cells and disease severity. UC model was induced by administering DNCB liquid and acetate solution. We found upregulated expression of CD4+CD29+T cells in both peripheral blood and colon from rats, and a similar trend for MPO and VCAM-1 in colon (P<0.05); the expression was especially enhanced in UC rats at two weeks after the model was established (P<0.01). Such upregulation was also indicated in active and remission UC patients as compared to the healthy and enteritis groups (P<0.05), with the highest expression level detected in the active UC patients (P<0.01). Pearson correlation analysis showed a positive correlation of CD4+CD29+T cells in rat and human peripheral blood with DAI score (rrat=0.712, rhuman=0.677, P<0.01), and MPO in colon (rrat=0.514, rhuman=0.682, P<0.05). These results suggest that CD4+CD29+T cells may act as major effector cell subsets in persistent inflammatory responses for UC and that infiltration into colon inflammation may be induced by the combination of VCAM-1 and CD29.

Inhibitory effect of 5F on development of lung cancer in A/J mice.

The purpose of the study is to investigate the effect of ent-11α-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F) on the model of induced A/J mice lung cancer in A/J mice. The expressions of tumor-related molecules including P65 and Bcl-2 at protein level were examined using the immunohistochemical method (IHC). Side effects of 5F were also monitored. The results indicated that 5F significantly suppressed the development of B[a]P and NNK-induced lung cancer in vivo by facilitating cell apoptosis with minimal side effects. Compared to the expressions of P65 and Bcl-2 in model group, the levels were strongly attenuated both in blank and 5F injection groups. Moreover, P65 and Bcl-2 levels varied among different groups receiving 5F treatment. The expressions of P65 and Bcl-2 were much lower in groups receiving high-concentration 5F treatment than those with low-concentration 5F injection. Findings revealed that 5F inhibited the pathogenesis of lung cancer through accelerating apoptosis in a dose-dependent manner.