Portulaca oleracea extract can inhibit nodule formation of colon cancer stem cells by regulating gene expression of the Notch signal transduction pathway.

To investigate whether Portulaca oleracea extract affects tumor formation in colon cancer stem cells and its chemotherapy sensitivity. In addition, to analyze associated genetic changes within the Notch signal transduction pathway. Serum-free cultures of colon cancer cells (HT-29) and HT-29 cancer stem cells were treated with the chemotherapeutic drug 5-fluorouracil to assess sensitivity. Injections of the stem cells were also given to BALB/c mice to confirm tumor growth and note its characteristics. In addition, the effect of different concentrations of P. oleracea extract was tested on the growth of HT-29 colon cancer cells and HT-29 cancer stem cells, as determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. The effects of P. oleracea extract on the expression of ß-catenin, Notch1, and Notch2 in the HT-29 cells were studied using reverse transcription polymerase chain reaction and Western blotting. The tumor volume of the HT29 cells was two times larger than that of HT29 cancer stem cells. Treatment with P. oleracea extract inhibited the proliferation of both HT-29 cancer cells and HT-29 cancer stem cells at doses from 0.07 to 2.25 µg/mL. Apoptosis of HT-29 cancer cells and HT-29 cancer stem cells was assessed by flow cytometry; it was enhanced by the addition of P. oleracea extract. Finally, treatment with P. oleracea extract significantly downregulated the expression of the Notch1 and ß-catenin genes in both cell types. The results of this study show that P. oleracea extract inhibits the growth of colon cancer stem cells in a dose-dependent manner. Furthermore, it inhibits the expression of the Notch1 and ß-catenin genes. Taken together, this suggests that it may elicit its effects through regulatory and target genes that mediate the Notch signal transduction pathway.

C-Myc-activated long noncoding RNA CCAT1 promotes colon cancer cell proliferation and invasion.

Recently, more and more evidence are rapidly accumulating that long noncoding RNAs (lncRNAs) are involved in human tumorigenesis and misregulated in many cancers, including colon cancer. LncRNA could regulate essential pathways that contribute to tumor initiation and progression with their tissue specificity, which indicates that lncRNA would be valuable biomarkers and therapeutic targets. Colon cancer-associated transcript 1 (CCAT1) is a 2628 nucleotide-lncRNA and located in the vicinity of a well-known transcription factor c-Myc. CCAT1 has been found to be upregulated in many cancers, including gastric carcinoma and colonic adenoma-carcinoma. However, its roles in colon cancer are still not well documented and need to be investigated. In this study, we aim to investigate the prognostic value and biological function of CCAT1 and discover which factors may contribute to the deregulation of CCAT1 in colon cancer. Our results revealed that CCAT1 was significantly overexpressed in colon cancer tissues when compared with normal tissues, and its increased expression was correlated with patients' clinical stage, lymph nodes metastasis, and survival time after surgery. Moreover, c-Myc could promote CCAT1 transcription by directly binding to its promoter region, and upregulation of CCAT1 expression in colon cancer cells promoted cell proliferation and invasion. These data suggest that c-Myc-activated lncRNA CCAT1 expression contribute to colon cancer tumorigenesis and the metastatic process and could predict the clinical outcome of colon cancer and be a potential target for lncRNA direct therapy.

The specific methylation characteristics of cancer related genes in Chinese colorectal cancer patients.

Aberrant DNA methylation at CpG islands has been implicated as a critical player in colorectal cancer (CRC). However, its biological role and clinical significance in carcinogenesis have not been clearly clarified in Chinese CRC patients. In order to examine the methylation status of cancer-related genes in CRC progression, 184 tumor tissues were collected from Chinese patients diagnosed with CRC during 2008-2011. Promoter methylation was assessed by combined bisulphite-restriction analysis, methylation-specific PCR, and bisulphite sequencing PCR . The relationship between the gene promoter methylation status and clinicopathological factors/CRC mortality was examined by using the chi-square test/Cox-proportional hazards models. Promoter hypermethylation of MLH1, p16, SFRP2, PHD3, KLOTHO, and IGFBP7 was observed in 1.6, 10.9, 97.3, 44.0, 59.8, and 88.6 % of CRC samples, respectively. KLOTHO promoter methylation reduced with age (P = 0.018) whereas p16 promoter methylation increased with age (P = 0.044) and was more frequent among males (P = 0.017). Tumor tissues (73.9 %) had concurrent methylation of two or more genes, with the most frequent combination as KLOTHO and IGFBP7 (53.8 %). Concurrent methylation of KLOTHO and IGFBP7 occurred more frequently among patients less than 70 years old (P = 0.035) and those with poor differentiation (P = 0.024). CRC-specific mortality was not associated with promoter methylation and clinicopathological features except for age (P = 0.038; risk ratio (RR), 1.96; 95 % confidence interval (CI), 1.04-3.70) and TNM stage (P = 0.034; RR, 3.47; 95 % CI, 1.10-10.92). Methylation frequencies of MLH1, p16, PHD3, KLOTHO, and IGFBP7 in CRC tissues were significantly higher than that in the paired normal tissues, while promoter hypermethylation of SFRP2 was widespread in normal tissues. In conclusion, we suggest that methylation of some genes (MLH1, PHD3, KLOTHO, p16, and IGFBP7) is important in CRC progression whereas SFRP2 methylation is unlikely to contribute to CRC development in Chinese patients. Besides, by identifying the characteristics of concordant methylation, we confirm the multifactorial nature of tumor progression.

Exploring the mechanism of Huanglian ointment in alleviating wound healing after anal fistula surgery through metabolomics and proteomics.

Anal fistula is a common anal and intestinal disease. The wound of anal fistula surgery is open and polluting, which is the most difficult to heal among all surgical incisions. To investigate the mechanism of Huanglian ointment (HLO) on wound healing after anal fistula incision. The S. aureus infected wound in SD rats were used to imitate poor healing wound after anal fistula surgery. SD rats with wound sites (n = 24) were randomly divided into four groups (Control group, Model group, Potassium permanganate (PP) treatment group, and HLO treatment group). The wound healing rate was evaluated, HE staining was used to evaluate the pathological changes of each group, ELISA was used to detect the secretion of inflammatory factors in each group, and the mechanism was explored through metabolomics and proteomics in plasma rat. Compared to other groups, the rate of wound healing in the HLO group was higher on days 7 and 14. Histological analysis showed that collagen and fibroblast in HLO rats were significantly increased, inflammatory cells were reduced, and vascular endothelial permeability was increased. ELISA results showed that the secretion of inflammatory factors in HLO rats was significantly lower. Significant proteins and metabolites were identified in the wound tissues of the infected rats and HLO-treated rats, which were mainly attributed to Cdc42, Ctnnb1, Actr2, Actr3, Arpc1b, Itgam, Itgb2, Cttn, Linoleic acid metabolism, d-Glutamine and d-glutamate metabolism, Phenylalanine, tyrosine and tryptophan biosynthesis, Phenylalanine metabolism, alpha-Linolenic acid metabolism, and Ascorbate and aldarate metabolism. In conclusion, this study showed that HLO can promote S. aureus infected wound healing, and the data provide a theoretical basis for the treatment of wounds after anal fistula surgery with HLO.

Shenling Baizhu Decoction (SLBZD) may play a synergistic role of tirelizumab in the treatment of colorectal cancer by influencing the imbalance of colon flora and Tumor microenvironment.

Objective: To explore whether SLBZD can play a synergistic role in promoting the efficacy of PD-1 inhibitors in the treatment of colorectal cancer by influencing the intestinal microenvironment and Tumor microenvironment. Method: Shenling Baizhu Decoction (SLBZD) and tirelizumab (TLzmab) treated the colorectal mouse model. The tumor growth rate, tumor weight, and tumor growth inhibition rate were evaluated. Fecal microbiota was detected by 16S rDNA sequencing and immune cell was detected by the flow cytometry analysis. Result: Compared to tumor weight, there exist significant differences between each group among the three groups. Compared to tumor volume, there was no statistically significant difference in tumor size between the control group and the TLzmab group at 7 days. However, there was a statistical difference in tumor size among the three groups at 18 days. By analyzing the diversity of the Gut microbiota, the diversity decreased after TLzmab treatment with a statistically significant difference. Compared with the control group, the diversity of the TLzmab+SLBZD group increased. The proportion of lymphocytes in the blood was analyzed by flow cytometry. Compared with the control group, Myeloid-derived suppressor cells (MDSCs) decreased and T regulatory cells (Treg) increased significantly in the TLzmab group. Compared with the control group and TLzmab group, the TLzmab+SLBZD group showed a significant increase in M1 type macrophages, while the M2 type macrophages, MDSCs, and Treg showed a significant decrease. Conclusion: An imbalance of Gut microbiota and imbalance of tumor immune microenvironment will occur during TLzmab treatment, which will lead to poor therapeutic effect of TLzmab or drug resistance. SLBZD will increase the abundance of Gut microbiota, which will lead to the increase of M1 macrophages in the tumor immune microenvironment and the decrease of M2 macrophages and Treg cells, thus exerting the synergistic effect of TLzmab. This study provides a new way to explore the improvement of ICIs through traditional Chinese medicine.

The Potential Mechanism of the Anti-Liver Fibrotic Effect of Curcumin in the Gut-Liver Axis.

This study aimed to explore the curative effect of curcumin on liver fibrosis and its correlation with the gut-liver axis in animal models. Histological staining was utilized to conduct histological analysis of the liver and intestine. An automatic biochemical analyzer or enzyme-linked immunosorbent assay system was utilized for analyzing the biochemical indexes in mice. Western blotting was employed to examine the level of relevant proteins. Furthermore, 16S rRNA high-throughput sequencing was performed to explore the impact of curcumin on intestinal microorganisms in rats with liver fibrosis. Ultrahigh-performance liquid chromatography with quadrupole-orbitrap mass spectrometry was utilized to analyze the effect of curcumin on rat feces metabolites. Our results showed that curcumin reduced the formation of collagen fibers caused by carbon tetrachloride in a dose-dependent manner. In addition, curcumin was able to restore intestinal permeability in rats with liver fibrosis. By adopting α diversity analysis (Chao 1 index, Shannon index, and Simpson index), we observed that both the diversity and the abundance of intestinal flora in rats with liver fibrosis were increased. The principal component analysis diagram demonstrated that curcumin could enhance the abundance and diversity of intestinal flora, and also restore the composition of model rat flora, which was similar to that in normal rats, thereby correcting the imbalance of flora in rats with liver fibrosis. In addition, curcumin regulated feces metabolites and their signaling pathways, including glycerophospholipid metabolism, pantothenate and CoA biosynthesis. Our findings suggest that curcumin exhibits antiliver fibrosis effects, and its antiliver fibrosis effects might correlate with gut-liver axis.

Treatment of adult congenital anal atresia with rectovestibular fistula: A rare case report.

BACKGROUND: Female anorectal malformation is a correctable congenital defect. Delayed manifestations in patients with anal deformities are uncommon, especially after adolescence. CASE SUMMARY: The clinical data of a 19-year-old adult female patient with congenital anal atresia accompanied by rectovestibular fistula as the main manifestation was retrospectively analyzed. Diagnosis was made based on the patient's clinical symptoms, signs, imaging showing the fistula, X-ray and magnetic resonance imaging results. The preoperative examination was improved. Anorectoplasty was performed. The patient exhibited an improvement in quality of life and presented no evidence of fecal incontinence during the 6-mo follow-up. CONCLUSION: Transfistula anorectoplasty is a reasonable and reliable surgical method for the treatment of adult congenital anal atresia and rectovestibular fistula.

Integrated microbiome and metabolome analysis reveals the interaction between intestinal flora and serum metabolites as potential biomarkers in hepatocellular carcinoma patients.

Globally, liver cancer poses a serious threat to human health and quality of life. Despite numerous studies on the microbial composition of the gut in hepatocellular carcinoma (HCC), little is known about the interactions of the gut microbiota and metabolites and their role in HCC. This study examined the composition of the gut microbiota and serum metabolic profiles in 68 patients with HCC, 33 patients with liver cirrhosis (LC), and 34 healthy individuals (NC) using a combination of metagenome sequencing and liquid chromatography-mass spectrometry (LC-MS). The composition of the serum metabolites and the structure of the intestinal microbiota were found to be significantly altered in HCC patients compared to non-HCC patients. LEfSe and metabolic pathway enrichment analysis were used to identify two key species (Odoribacter splanchnicus and Ruminococcus bicirculans) and five key metabolites (ouabain, taurochenodeoxycholic acid, glycochenodeoxycholate, theophylline, and xanthine) associated with HCC, which then were combined to create panels for HCC diagnosis. The study discovered that the diagnostic performance of the metabolome was superior to that of the microbiome, and a panel comprised of key species and key metabolites outperformed alpha-fetoprotein (AFP) in terms of diagnostic value. Spearman's rank correlation test was used to determine the relationship between the intestinal flora and serum metabolites and their impact on hepatocarcinogenesis and progression. A random forest model was used to assess the diagnostic performance of the different histologies alone and in combination. In summary, this study describes the characteristics of HCC patients' intestinal flora and serum metabolism, demonstrates that HCC is caused by the interaction of intestinal flora and serum metabolites, and suggests that two key species and five key metabolites may be potential markers for the diagnosis of HCC.

Expression and clinical significance of Notch signaling genes in colorectal cancer.

The objective of this study was to explore the expression and clinical significance of Notch signaling genes in colorectal cancer. Colorectal cancer samples were prospectively collected from patients post-surgery at the 3rd Affiliated Hospital, Nanjing University of Traditional Chinese Medicine. Immunohistochemistry of tissue arrays was used to analyze the samples and genes involved in the Notch signaling pathway. Microsatellite instability (MSI) was detected by fluorescence multiplex polymerase chain reaction. A total of 146 colorectal cancer samples were collected, including 84 men (57.7%) and 62 women (42.5%). The average age of the study population was 60.8 ± 10.5 years. Notch1 and Notch2 gene expression correlated with tumor pathology type and degree of differentiation. In addition, Jagged 1 (JAG1) and hairy enhancer of split 1 gene expression correlated with the degree of tumor differentiation. Delta-like 1 gene expression varied significantly with tumor location. There was a significant difference between gene expression and MSI. Of the 138 patients, 134 (91.8%) participated in on-site visits, and the average follow-up time was 42.3 ± 13.3 months. During this period, 86 patients (71.6%) were tumor-free. At 1 year post-surgery, 93% of patients were alive, 74% of patients lived for 3 years, and 67% of patients lived for 5 years. The log-rank test was used to perform univariate analysis, and the COX proportional hazards model was used for the multivariate analysis. Based on these analyses, tumor prognosis correlated with the TNM stage, pathological type, microsatellite status as well as Notch2 and JAG1 gene expression. Patients expressing high levels of Notch2 and JAG1 presented with a significantly better prognosis compared to patients expressing negative or weak levels of Notch2 and JAG1. The expression levels of genes associated with the Notch signaling pathway correlated with tumor pathology and the degree of differentiation. In addition, Notch2 and JAG1 expression levels correlated with survival; however, the underlying mechanism for these correlations remains unclear.

Dietary flavonoid for preventing colorectal neoplasms.

BACKGROUND: Flavonoids are polyphenolic compounds that are distributed widely in the plant kingdom; they are especially abundant in fruits and vegetables. More than 5,000 individual flavonoids have been identified and classified into more than 10 subgroups according to their chemical structure. Flavonoids have many possible biological effects that may play a role in cancer prevention. Prior studies have suggested that a high intake of flavonoids may help prevent cancer. OBJECTIVES: To assess the effect of dietary flavonoids on the incidence of colorectal adenoma and CRC. SEARCH METHODS: Eligible studies were searched up until July 2011 in the Cochrane Library, PubMed, EMBASE and other CINAHL databases and reference lists of previous reviews. SELECTION CRITERIA: All prospective, controlled interventional studies and observational studies that either assessed the association between flavonoids and risk of CRC incidence or colorectal adenoma recurrence were included. DATA COLLECTION AND ANALYSIS: At least two investigators independently reviewed the material and extracted the data according to the inclusion criteria; in addition, the methodological quality of the studies was assessed. MAIN RESULTS: Eight studies with 390,769 participants were included. Five studies used a prospective cohort design, two were case-control studies and one a randomised controlled trial (RCT). The methodological quality was measured using the Newcastle-Ottawa scale (NOS). The three prospective cohort studies were of high methodological quality, and two were of medium quality. The two case-control studies were of medium methodological quality.The results form the studies assessing associations between flavonoids, colorectal cancer and adenomas were contradictory. There was no evidence that total flavonoid intake reduced the risk of colorectal neoplasms. The evidence for Isoflavones, Flavonols, Flavones and Flavanones was conflicting. For Flavan-3-ols, the results from two studies suggested that increased intake of Flavan-3-ols reduced the risk of both CRC and colorectal adenomas. A statistically significant reduced risk of CRC was found with high intake of epicatechin. There was medium quality evidence to support that increased intake of procyanidin and phytoestrogen could reduced the incidence of CRC. There was no evidence that suggested that high anthocyanin intake had an inverse association with colorectal adenomas. AUTHORS' CONCLUSIONS: There is insufficient and conflicting evidence regarding flavonoid intake and the prevention of colorectal neoplasms. It is difficult to determine flavonoid intake. Therefore, more evidence is needed to clarify the association between flavonoids and colorectal neoplasms.

A superficial colon tumor model involving subcutaneous colon translocation and orthotopic transplantation of green fluorescent protein-expressing human colon tumor.

The orthotopic transplantation model of human tumor has been demonstrated to be more patient-like animal tumor model. However, observations of tumor progression and metastasis are limited by the deep location of the colon or limited deep penetration ability of fluorescence through tissue. The purpose of this study is to establish a superficial orthotopic model to allow easier real-time visualization and more sensitive monitoring of fluorescent orthotopic colon tumor. Human colon cancer HT-29 cells were transduced with a pLPCX expression retroviral vector containing green fluorescent protein and neomycin resistance genes. For superficial orthotopic transplantation model, the cecum was identified and pulled out of the peritoneal cavity, the space between the cecum and peritoneum was sutured, the cecum was pulled to subcutaneous tissue, and incision was made on the cecal serosa followed by the implantation of a 1-mm tumor tissue to the cecum. For comparison, a conventional orthotopic transplantation model was established in a separate group of mice simultaneously. When tumor sizes reached 5 mm in diameter, half the mice in each model received 5-FU treatment. Primary tumor and metastases were monitored by fluorescent imaging or caliber measurement. Tumor fluorescence was observed as early as 3 days (median time of 4.7 ± 1.3 days) post-transplantation in the superficial orthotopic transplantation model, which was much earlier than 21 days (median time of 26.2 ± 9.9 days) in conventional orthotopic transplantation model. Although tumor growth of 5-FU-treated mice in conventional orthotopic model was lower than those of the untreated mice, the difference was not significant. However, in superficial orthotopic model, tumor growth was significantly inhibited in 5-FU-treated mice relative to the untreated mice. Fluorescence imaging showed similar metastasis incidence between the superficial and conventional orthotopic transplantation models. The fluorescent superficial orthotopic transplantation colon model allows easier real-time visualization and more sensitive monitoring of tumor growth as well as convenient repeated sampling. It is a valuable orthotopic implantation model for study of colon cancer and evaluation of new anti-cancer therapy.

Separation and purification of Escherichia coli-expressed human thymosin-α1 using affinity chromatography and high-performance liquid chromatography.

In this study, a human thymosin-α1 (hTα1) fusion protein was overexpressed in Escherichia coli (E. coli). The hexahistidine-tagged hTα1 fusion protein was obtained in soluble form in cells of the engineered E. coli strain BL21 (DE3)/pET-28a-hTα1 that had been induced with isopropyl -D-1-thiogalactopyranoside (IPTG). The recombinant protein accounted for approximately 50-60% of the total protein. We then developed and validated a separation method for hTα1 from E. coli cells based on thermal denaturation, nickel-resin affinity chromatography and high-performance liquid chromatography. The purification method showed good reproducibility and was easy to operate. Purified recombinant hTα1 of high homogeneity was characterized and found to be of high purity (over 99%), as determined by high-voltage electrophoresis and high-performance liquid chromatography analysis. Isoelectric focusing analysis indicated a pI of approximately 4.0, and full wavelength screening showed an optimal absorbance wavelength at around 214nm.

Downregulation of AP-1 gene expression is an initial event in the oridonin-mediated inhibition of colorectal cancer: studies in vitro and in vivo.

BACKGROUND AND AIM: Oridonin is the active ingredient isolated from the Chinese herb Rabdosia rubescens. We used both in vivo and in vitro approaches to elucidate the underlying mechanism of the oridonin-mediated inhibition of colorectal cancer. METHODS: Two colorectal cell lines, Lovo and SW480, were treated with oridonin in solution. The effect of this treatment on the inhibition of the cell proliferation rate was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. The changes in gene expression that occurred in both cell lines in response to treatment with oridonin were determined via an illumine expression sensor. Additionally, a colorectal cancer colostomy implantation model was established. Animals were injected intraperitoneally with an oridonin solution. RESULTS: The treatment of Lovo and SW480 cells with oridonin inhibited cell proliferation in a dose-dependent manner. Furthermore, the rate of inhibition increased with prolonged treatment. The growth rate of the colorectal cancer colostomy implantation model was significantly lower than control cells when treated with oridonin (P<0.001), which meant that oridonin treatment had a significant effect on the tumor growth rate. In the tumor model, activator protein-1 (AP-1) was the only gene found to be downregulated after oridonin treatment by the gene expression sensor. After 4 weeks of treatment, AP-1, nuclear factor-κB (NF-κB) and P38 were all found to be downregulated. CONCLUSIONS: Our study confirmed the inhibitory effects of oridonin on colorectal cancer. These results indicate that the downregulation of AP-1 might be an initial response to treatment by oridonin. This regulation could, in turn, affect the expression of the NF-κB and mitogen-activated protein kinase pathways, thereby inhibiting tumor growth.

[Expression pattern and clinical significance of hairy enhancer of split 1 in colorectal cancer].

OBJECTIVE: To explore the roles of hairy enhancer of split 1 (Hes1) gene in colorectal cancer and analyze its clinical significance. METHODS: A total of 146 cases with colorectal cancer at our hospital were collected prospectively and followed up. There were 84 males and 62 females with an average age of (61 ± 11) years old. Tissue microarray was prepared and the expression of Notch signal genes were detected by immunohistochemical staining. RESULTS: The differential expressions of Hes1 were significant among various types [negative or weakly positive: 8% (12/146), positive(+): 77% (112/146), positive(++): 15% (22/146)]. Among all, 134 were followed up successfully for an average duration of (42 ± 13) months. According to the Kaplan-Meier life curve, the overall 1, 3 and 5-year survival rates were 93% (136/146), 74% (108/146) and 67% (98/146) respectively. The long-term survival rate was correlated with TNM stage and pathological types (all P = 0.000), but not with Hes1 (P = 0.267). CONCLUSION: The expression of Hes1 is correlated with pathological types and differentiation types. However the long-term survival rate is not correlated with its expression.

The Value of Multi-targeted Fecal DNA Methylation Detection for Colorectal Cancer Screening in a Chinese Population.

Objective: To design a multi-targeted fecal DNA methylation kit and explore its value for clinical application among Chinese people. Methods: Based on previous research, a multi-targeted fecal DNA methylation detection kit, using four genes, was designed and clinically validated. Results: The methylation PCR from 279 patients met the requirements for the detection criteria. When all four molecular markers were negative, the negative predictive value (NPV) for colorectal cancer was 100% and the NPV for colorectal polyps was 84.21%. When one molecular marker was positive, the sensitivity (Se) for colorectal cancer was 76.4%-90.3%, the specificity (Sp) was 68.3-93.4%, and the positive predictive value (PPV) for colorectal cancer was 54.5-85.5%, and the NPV was 87.0-95.0%. For colorectal polyps, the Se was 41.0-52.5%, Sp 69.5-91.5%, and the PPV for colorectal polyps was 41.0-70.3%, the NPV was 75.2-79.3%. When two molecular markers were positive, the Se for colorectal cancer was 52.6-73.7%, the Sp was 93.2-98.3%, the PPV for colorectal cancer was 84.6-96.2%, the NPV was 76.0-85.3%. For colorectal polyps, the Se was 25.9-40.7%, Sp was 93.2-98.3%, PPV for screening of colorectal polyps was 63.6-90.0%, and the NPV was 73.3-78.1%. When three molecular markers were positive, the Se for colorectal cancer was 31.6-52.6%, the Sp was 98.3-100.0%, the PPV for colorectal cancer was 94.4-100.0%, the NPV was 73.4-76.6%. For colorectal polyps, the Se was 14.8-25.9%, and Sp was 98.3-100.0%, the PPV for colorectal polyps was 85.7-100.0%, the NPV was 72.0-74.7%. When four molecular markers were positive, the Se for colorectal cancer was 31.6%, the Sp was 100.0%, and the colorectal cancer PPV was 100.0% and the NPV was 69.4%. For polyps, the Se was 14.8%, Sp was 100.0%, and PPV was 100.0% and the NPV was 72.0%. Conclusion: The multi-targeted fecal DNA methylation detection kit for colorectal cancer and polyps had the sensitivity and specificity to meet the requirements for screening of colorectal tumors, which is easy to operate, has stable results and important clinical value. Among the four molecular markers studied, when one marker was positive for DNA methylation, colonoscopy was required; as the number of positive methylation markers increased, the specificity for the diagnosis gradually increased as well.

P-cymene prevent high-fat diet-associated colorectal cancer by improving the structure of intestinal flora.

Objective: To investigate the preventing effect of P-cymene on high fat diet-related colorectal cancer and its mechanism. Methods: Forty Wistar rats were randomly divided into G1 group (high-fat diet), G2 group (high-fat diet + DMH), G3 group (high-fat diet + P-cymene), and G4 group (high-fat diet + DMH + P-cymene).G2 and G4 groups were subcutaneously injected with dimethylhydrazine (DMH), and G3 and G4 groups were intragastrically administered with P-cymene to investigate the effects of P-cymene on tumor formation, inflammatory factors, glucose, lipid metabolism and gut microbes. Results: No tumors were formed in the high-fat diet group (G1) or the high-fat diet + P-cymone group (G3). 7 rats (70%) of the high-fat diet + DMH group (G2) developed 8 cancerous nodules, including 6 adenocarcinomas and 2 signet ring cell carcinomas; 4 rats (40%) in the high-fat diet + DMH + P-cymene group (G4) group formed 4 cancerous nodules, all of which were adenocarcinoma. There was no significant difference in the changes of glucose and lipid metabolism in each group. After the use of P-cymene, IL-1 decreased, IL-6 increased, and LEP decreased in the G4 group.The difference was statistically significant.The contents of Candida and Unclassified Bacteria in the G3 group rats were significantly lower than those in the G1 group.At the species level comparison, compared with the G2 group, the content of Clostridium XlVa in the intestinal tract of the G2 group rats was significantly increased compared to the G1 group. Conclusion: In this study, it was found that p-cymenen can prevent the occurrence of colorectal cancer related to high-fat and high-calorie diet. The mechanism may be is reducing the expression of inflammatory factors such as IL-1 and LEP, increasing the expression of inflammatory factors of IL-6, and promoting the growth of probiotics such as bifidobacteria, isobacteria and clostridium IV in the intestinal tract.

The Anti-oxidant Monoterpene p-Cymene Reduced the Occurrence of Colorectal Cancer in a Hyperlipidemia Rat Model by Reducing Oxidative Stress and Expression of Inflammatory Cytokines.

BACKGROUND/AIM: Overexpression of inflammatory cytokines and oxidative stress increase the risk of colorectal cancer (CRC) in obesity and hyperlipidemia. The aim of this study was to investigate whether the monoterpene antioxidant p-cymene would reduce the incidence of CRC in a rat model of hyperlipidemia. MATERIALS AND METHODS: The hyperlipidemic CRC rat model was established by a high-fat diet and dimethyl hydrazine (DMH) induction. All rats received 30 mg/kg DMH to induce CRC, and were then assigned to groups with a normal diet or high-fat diet with/without 30 mg/kg/day p-cymene orally during the entire experimental period. Tumor incidence in each group, and the level of serum inflammatory cytokines and oxidative stress-related markers in intestinal tissues were measured. RESULTS: p-Cymene significantly inhibited CRC occurrence in hyperlipemic rats (p=0.024) by reducing the expression of serum inflammatory cytokines (interleukin-1 by 54.5%; interleukin-6 by 28.3%; adiponectin by 26.3%; cyclo-oxygenase-2 by 48.4%) and intestinal oxidative-stress cytokines (total antioxidant capacity by 30.4%; superoxide dismutase by 30.3%; malondialdehyde by 47.1%). CONCLUSION: p-Cymene has clinical potential to reduce the incidence of CRC in hyperlipemia.

The study of effect of tea polyphenols on microsatellite instability colorectal cancer and its molecular mechanism.

INTRODUCTION: Tea polyphenol has been shown to have anti-colorectal cancer and anti-gene mutation effects, although the mechanism of inhibition of microsatellite instability (MSI) colorectal cancer is not known. MATERIALS AND METHODS: Using LoVo, HCT-116, HT-29, and SW480 cells treated with an aqueous solution of tea polyphenol, cell proliferation was detected by the methyl thiazolyl tetrazolium method, changes in microsatellite sequences by the Genescan method and changes in the gene expression of LoVo cells using Illumina expression arrays. RESULTS: The proliferation inhibition rate of LoVo, HCT-116, HT-29, and SW480 cells treated with tea polyphenol increased with increasing drug concentration and showed an increasing tendency with time. The proliferation inhibition rate of LoVo and HCT-116 cells with tea polyphenols was higher than that of HT-29 and SW480 cells, and there was a significant difference in the proliferation inhibition rate at 24, 72 h and 1 week. The microsatellite sequence of LoVo cells treated with tea polyphenols remained stable. DISCUSSION: The gene expression arrays and quantitative RT-PCR suggested that tea polyphenol inhibited the gene expression of metallothionein 2A (MT2A), transcription factor (MAFA), hairy and enhancer of split 1 (HES1), and jagged1 (JAG1) nearly twofold over controls. It was also found that tea polyphenol inhibited the BAX and p38 genes with a more than twofold difference but did not significantly inhibited the NFκB pathway. CONCLUSION: Tea polyphenol significantly inhibited the proliferation of MSI colorectal cancer signals maintained stable at the microsatellite state in MSI colorectal cancer. Tea polyphenol inhibited the gene expression of HES1, JAG1, MT2A, and MAFA but upregulated the gene expression of BAX and downregulated that of (P)38. Further research is required to investigate how these pathways are interrelated.

[Impact of polyamidoamine dendrimer liposome on the cellular uptake and cytotoxicity of colonic cancer cells].

OBJECTIVE: To evaluate the effects of polyamidoamine dendrimer (PAMAM) liposome as gene carriers on the cellular uptake and its cytotoxicity in colonic cancer cell. METHODS: The liposome modified PAMAM was synthesized with liposome and polyamidoamine dendrimer. Plasmid PEGFP-N1 was mixed with the liposome-modified PAMAM or unmodified PAMAM to form nanoparticle complexes. The shape and size of the nanoparticle complexes were observed by transmission electron microscope and the zeta potential was measured by analytical tool. The encapsulating efficiency was determined by ultraviolet spectrophotometer in centrifuging method. After the cell lines SW620 (colonic cancer cell), MCF-7 (breast cancer cell), ECV304 (vascular endothelial cell) were transfected by the two kinds of PAMAM nanoparticle complexes, the flow cytometry was used to determine the uptake of enhanced green fluorescent protein (EGFP) gene. The cytotoxicity of PAMAM liposome nanoparticles and PAMAM nanoparticles was evaluated by MTT assay. RESULTS: The diameter of liposome modified PAMAM complex was (192 ± 16) nm, and that of PAMAM complex was (189 ± 19) nm (P > 0.05); and the zeta potential of liposome modified PAMAM complex was higher than that of PAMAM complex [(42 ± 7) mV vs. (32 ± 7) mV, P < 0.05]. There was no significant difference in envelopment rate between the two groups [(82 ± 7)% vs. (84 ± 6)%, P > 0.05]. After the colonic cancer cell line SW620 was transfected with the two kinds of PAMAM nanoparticle complexes, the cellular uptake of the cells with the liposome-modified PAMAM complex was significantly higher than that of the cell with PAMAM complex (P < 0.05). The cellular survival rate of the cell lines with liposome-modified PAMAM complex was significantly higher than that of cell lines with PAMAM complex (P < 0.05). CONCLUSION: The liposome modified PAMAM can improve gene transfection efficiency and suppress its cytotoxicity.

[Tea polyphenol inhibits colorectal cancer with microsatellite instability by regulating the expressions of HES1, JAG1, MT2A and MAFA].

OBJECTIVE: To investigate the mechanism of tea polyphenol in inhibiting microsatellite instability (MSI) of colorectal cancer. METHODS: Using LoVo cells and SW480 cells treated with aqueous solution of tea polyphenol, cell proliferation was detected by methyl thiazolyl tetrazolium (MTT) method, changes in microsatellite sequences were detected by genescan method and changes in gene expression of LoVo cells were detected by illumina expression arrays and quantitative real-time polymerase chain reaction (PCR). RESULTS: The proliferation inhibition rates of LoVo and SW480 cells treated with tea polyphenol increased with the increasing of drug concentration and showed an increasing tendency with time. The proliferation inhibition rate of LoVo cells with tea polyphenol was higher than that of SW480 cells, and there was a significant difference in the proliferation inhibition rates at 24 h, 72 h and one week. The microsatellite sequence of LoVo cells treated with tea polyphenol remained stable. The gene expression arrays and quantitative real-time PCR suggested that tea polyphenol inhibited the gene expressions of MT2A, MAFA, HES1 and JAG1 nearly two-fold over controls. It was also found that tea polyphenol inhibited the BAX and p38 genes with a more than two-fold difference but did not significantly inhibit the nuclear factor-κB pathway. CONCLUSION: Tea polyphenol significantly inhibited the proliferation of MSI colorectal cancer cells and stably maintained the microsatellite state in MSI colorectal cancer. Tea polyphenol inhibited the gene expressions of HES1, JAG1, MT2A and MAFA, up-regulated the gene expression of BAX and down-regulated that of P38. Further research is required to investigate how these pathways are interrelated.