Clinicopathologic and endoscopic features of sessile serrated lesions and conventional adenomas: a large inpatient population-based study in China.

Objectives: Sessile serrated lesions (SSLs) are precursors of sporadic colorectal cancer (CRC) and have distinct characteristics compared with conventional adenomas (CAs). Several lifestyle and environmental factors may play critical roles in the development of advanced lesions. Our aim is to describe the features of SSLs and CAs and further explore risk factors for advanced lesions. Methods: This is an observational study that collected demographic, endoscopic, and histological data from the China-Japan Friendship Hospital among the inpatient population with pathologically reported as SSL or CA between 2015 and 2022. We analyzed the clinicopathology and endoscopic differences between SSL alone, CA alone, and synchronous SSL+CA groups, and identified risk factors using multiple regression analysis. Results: A total of 9236 polyps from 6598 patients were included in the cohort. Patients with SSL+CA were more likely to be older (p=0.008), while individuals with SSL alone had a higher proportion of early-onset polyps (p<0.001), and SSLs were more common in advanced polyps than CAs (p<0.001). A greater proportion of advanced polyps in the SSL and CA groups were diagnosed as Yamada III, Yamada IV, and laterally spreading tumor (p=0.002, p<0.001, respectively), and multiple SSLs and CAs were more represented in nonadvanced polyps than in advanced polyps. In multiple regression analysis, older patients were more likely to develop advanced SSLs (aOR 1.05, 95% CI 1.02-1.09, p=0.005). Conclusion: SSLs and CAs have diverse demographic, endoscopic, and histological characteristics, and their advanced lesions share different risk factors, which advances the understanding of the etiology and progression of SSLs.

Noninvasive Diagnostic Technique for Nonalcoholic Fatty Liver Disease Based on Features of Tongue Images.

OBJECTIVE: To investigate a new noninvasive diagnostic model for nonalcoholic fatty liver disease (NAFLD) based on features of tongue images. METHODS: Healthy controls and volunteers confirmed to have NAFLD by liver ultrasound were recruited from China-Japan Friendship Hospital between September 2018 and May 2019, then the anthropometric indexes and sampled tongue images were measured. The tongue images were labeled by features, based on a brief protocol, without knowing any other clinical data, after a series of corrections and data cleaning. The algorithm was trained on images using labels and several anthropometric indexes for inputs, utilizing machine learning technology. Finally, a logistic regression algorithm and a decision tree model were constructed as 2 diagnostic models for NAFLD. RESULTS: A total of 720 subjects were enrolled in this study, including 432 patients with NAFLD and 288 healthy volunteers. Of them, 482 were randomly allocated into the training set and 238 into the validation set. The diagnostic model based on logistic regression exhibited excellent performance: in validation set, it achieved an accuracy of 86.98%, sensitivity of 91.43%, and specificity of 80.61%; with an area under the curve (AUC) of 0.93 [95% confidence interval (CI) 0.68-0.98]. The decision tree model achieved an accuracy of 81.09%, sensitivity of 91.43%, and specificity of 66.33%; with an AUC of 0.89 (95% CI 0.66-0.92) in validation set. CONCLUSIONS: The features of tongue images were associated with NAFLD. Both the 2 diagnostic models, which would be convenient, noninvasive, lightweight, rapid, and inexpensive technical references for early screening, can accurately distinguish NAFLD and are worth further study.

Leukocyte immunoglobulin-like receptor B2 overexpression as a promising therapeutic target and noninvasive screening biomarker for colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) has become the second most deadly malignancy in the world, and the exploration of screening markers and precise therapeutic targets is urgent. Our previous research identified leukocyte immunoglobulin-like receptor B2 (LILRB2) protein as a characteristic protein of CRC, but the association between LILRB2 expression and clinicopathological features, the internal mechanism related to CRC progression, and screening diagnostic efficacy are not clear. Therefore, we hypothesized that LILRB2 is significantly highly expressed in CRC tissues, correlated with advanced stage and a poor prognosis, and could be used as a therapeutic target and potential screening biomarker for CRC. AIM: To explore whether LILRB2 can be used as a potential therapeutic target and noninvasive screening biomarker for CRC. METHODS: Patients who underwent radical surgery for CRC at China-Japan Friendship Hospital between February 2021 and October 2022 were included. Cancer and paracancerous tissues were collected to verify LILRB2 expression, and the association between LILRB2 expression and clinicopathological features was analysed. Serum was collected from CRC patients, adenoma patients and healthy controls during the same period to assess the diagnostic value of LILRB2 as a noninvasive screening biomarker, and its diagnostic value was further compared with that of the traditional markers carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9). RESULTS: A total of 58 CRC patients were included, and LILRB2 protein was significantly overexpressed in cancer tissues compared with paracancerous tissues (P < 0.001). Angiopoietin-like protein 2 (ANGPTL2) protein, as the ligand of LILRB2, was synergistically overexpressed in CRC tissues (P < 0.001), and overexpression of LILRB2 and ANGPTL2 protein was significantly correlated with poor to moderate differentiation, vascular involvement, lymph node metastasis, distant metastasis, advanced tumor-node-metastasis stage and a poor prognosis (P < 0.05), which suggested that LILRB2 and ANGPTL2 are closely associated with CRC progression. In addition, serum LILRB2 concentrations increased stepwise in healthy individuals, adenoma patients and CRC patients with statistically significant differences. The sensitivity of serum LILRB2 for the diagnosis of CRC was 89.74%, the specificity was 88.89%, the area under the curve was 0.95, and the diagnostic efficacy was better than that of conventional CEA and CA19-9. CONCLUSION: LILRB2 protein can be used as a potential novel therapeutic target and noninvasive screening biomarker for CRC, which is beneficial for early screening and precise treatment.

Proteomics-based identification of proteins in tumor-derived exosomes as candidate biomarkers for colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the second leading cause of cancer-related death, with high morbidity worldwide. There is an urgent need to find reliable diagnostic biomarkers of CRC and explore the underlying molecular mechanisms. Exosomes are involved in intercellular communication and participate in multiple pathological processes, serving as an important part of the tumor microenvironment. AIM: To investigate the proteomic characteristics of CRC tumor-derived exosomes and to identify candidate exosomal protein markers for CRC. METHODS: In this study, 10 patients over 50 years old who were diagnosed with moderately differentiated adenocarcinoma were recruited. We paired CRC tissues and adjacent normal intestinal tissues (> 5 cm) to form the experimental and control groups. Purified exosomes were extracted separately from each tissue sample. Data-independent acquisition mass spectrometry was implemented in 8 matched samples of exosomes to explore the proteomic expression profiles, and differentially expressed proteins (DEPs) were screened by bioinformatics analysis. Promising exosomal proteins were verified using parallel reaction monitoring (PRM) analysis in 10 matched exosome samples. RESULTS: A total of 1393 proteins were identified in the CRC tissue group, 1304 proteins were identified in the adjacent tissue group, and 283 proteins were significantly differentially expressed between them. Enrichment analysis revealed that DEPs were involved in multiple biological processes related to cytoskeleton construction, cell movement and migration, immune response, tumor growth and telomere metabolism, as well as ECM-receptor interaction, focal adhesion and mTOR signaling pathways. Six differentially expressed exosomal proteins (NHP2, OLFM4, TOP1, SAMP, TAGL and TRIM28) were validated by PRM analysis and evaluated by receiver operating characteristic curve (ROC) analysis. The area under the ROC curve was 0.93, 0.96, 0.97, 0.78, 0.75, and 0.88 (P < 0.05) for NHP2, OLFM4, TOP1, SAMP, TAGL, and TRIM28, respectively, indicating their good ability to distinguish CRC tissues from adjacent intestinal tissues. CONCLUSION: In our study, comprehensive proteomic profiles were obtained for CRC tissue exosomes. Six exosomal proteins, NHP2, OLFM4, TOP1, SAMP, TAGL and TRIM28, may be promising diagnostic markers and effective therapeutic targets for CRC, but further experimental investigation is needed.

microRNA-627-5p inhibits colorectal cancer cell proliferation, migration and invasion by targeting Wnt2.

BACKGROUND: microRNA-627-5p (miR-627-5p) dysregulation has been observed in several cancer types, such as hepatocellular carcinoma, oral squamous cell carcinoma, glioblastoma multiforme, and gastric cancer. The biological function of miR-627-5p in colorectal cancer (CRC) growth and metastasis is yet unclear. AIM: To investigate the effects of miR-627-5p on the malignant biological properties of colorectal malignant tumour cells by targeting Wnt2. METHODS: The levels of miR-627-5p in colorectal tumour tissues were assessed in Gene Expression Omnibus datasets. In order to identify Wnt2 transcript expression in CRC tissues, quantitative real-time polymerase chain reaction (qRT-PCR) analysis was used. Luciferase reporter tests were used to explore whether miR-627-5p might potentially target Wnt2. Wnt2 transcript and protein levels were detected in CRC cells with high miR-627-5p expression. To learn more about how miR-627-5p affects CRC development, migration, apoptosis, and invasion, functional experiments were conducted. Cotransfection with the overexpression vector of Wnt2 and miR-627-5p mimics was utilized to verify whether overexpression of Wnt2 could cancel the impact of miR-627-5p in CRC. Western blot and qRT-PCR were conducted to investigate the effects of miR-627-5p on the Wnt/ß-catenin signalling pathway. RESULTS: miR-627-5p was notably decreased in colorectal tumour tissues, while the gene level of Wnt2 was notably upregulated. A dual luciferase reporter assay revealed that miR-627-5p specifically targets the 3'-untranslated regions of Wnt2 and miR-627-5p upregulation markedly reduced the protein and gene expression of Wnt2 in CRC cells. In vitro gain-of-function assays displayed that miR-627-5p overexpression decreased CRC cells' capabilities to invade, move, and remain viable while increasing apoptosis. Wnt2 overexpression could reverse the suppressive functions of miR-627-5p. Moreover, upregulation of miR-627-5p suppressed the transcript and protein levels of the downstream target factors in the canonical Wnt/ß-catenin signalling, such as c-myc, CD44, ß-catenin, and cyclinD1. CONCLUSION: miR-627-5p acts as a critical inhibitory factor in CRC, possibly by directly targeting Wnt2 and negatively modulating the Wnt/ß-catenin signalling, revealing that miR-627-5p could be a possible treatment target for CRC.

Atypical Whipple's disease with special endoscopic manifestations: A case report.

BACKGROUND: Whipple's disease is a rare systemic infection caused by Tropheryma whipplei. Most patients present with nonspecific symptoms, and routine laboratory and imaging examination results also lack specificity. The diagnosis often relies on invasive manipulation, pathological examination, and molecular techniques. These difficulties in diagnosing Whipple's disease often result in misdiagnosis and inappropriate treatments. CASE SUMMARY: This paper reports on the case of a 58-year-old male patient who complained of fatigue and decreased exercise capacity. The results of routine blood tests indicated hypochromic microcytic anemia. Results of gastroscopy and capsule endoscopy showed multiple polypoid bulges distributed in the duodenal and proximal jejunum. A diagnosis of small intestinal adenomatosis was initially considered; hence, the Whipple procedure, a pylorus-preserving pancreaticoduodenectomy, was performed. Pathological manifestations showed many periodic acid-Schiff-positive macrophages aggregated in the intestinal mucosa of the duodenum, upper jejunum, and surrounding lymph nodes. Based on comprehensive analysis of symptoms, laboratory findings, and pathological manifestations, the patient was finally diagnosed with Whipple's disease. After receiving 1 mo of antibiotic treatment, the fatigue and anemia were significantly improved. CONCLUSION: This case presented with atypical gastrointestinal manifestations and small intestinal polypoid bulges, which provided new insight on the diagnosis of Whipple's disease.

Atherogenic index of plasma combined with waist circumference and body mass index to predict metabolic-associated fatty liver disease.

BACKGROUND: Early identification of metabolic-associated fatty liver disease (MAFLD) is urgent. Atherogenic index of plasma (AIP) is a reference predictor of obesity-related diseases, but its predictive value for MAFLD remains unclear. No studies have reported whether its combination with waist circumference (WC) and body mass index (BMI) can improve the predictive performance for MAFLD. AIM: To systematically explore the relationship between AIP and MAFLD and evaluate its predictive value for MAFLD and to pioneer a novel noninvasive predictive model combining AIP, WC, and BMI while validating its predictive performance for MAFLD. METHODS: This cross-sectional study consecutively enrolled 864 participants. Multivariate logistic regression analysis and receiver operating characteristic curve were used to evaluate the relationship between AIP and MAFLD and its predictive power for MAFLD. The novel prediction model A-W-B combining AIP, WC, and BMI to predict MAFLD was established, and internal verification was completed by magnetic resonance imaging diagnosis. RESULTS: Subjects with higher AIP exhibited a significantly increased risk of MAFLD, with an odds ratio of 12.420 (6.008-25.675) for AIP after adjusting for various confounding factors. The area under receiver operating characteristic curve of the A-W-B model was 0.833 (0.807-0.858), which was significantly higher than that of AIP, WC, and BMI (all P < 0.05). Subgroup analysis illustrated that the A-W-B model had significantly higher area under receiver operating characteristic curves in female, young and nonobese subgroups (all P < 0.05). The best cutoff values for the A-W-B model to predict MAFLD in males and females were 0.5932 and 0.4105, respectively. Additionally, in the validation set, the area under receiver operating characteristic curve of the A-W-B model to predict MAFLD was 0.862 (0.791-0.916). The A-W-B level was strongly and positively associated with the liver proton density fat fraction (r = 0.630, P < 0.001) and significantly increased with the severity of MAFLD (P < 0.05). CONCLUSION: AIP was strongly and positively associated with the risk of MAFLD and can be a reference predictor for MAFLD. The novel prediction model A-W-B combining AIP, WC, and BMI can significantly improve the predictive ability of MAFLD and provide better services for clinical prediction and screening of MAFLD.

Comprehensive proteomic signature and identification of CDKN2A as a promising prognostic biomarker and therapeutic target of colorectal cancer.

BACKGROUND: The carcinogenesis of colorectal cancer (CRC) involves many different molecules and multiple pathways, and the specific mechanism has not been elucidated until now. Existing studies on the proteomic signature profiles of CRC are relatively limited. Therefore, we herein aimed to provide a more comprehensive proteomic signature profile and discover new prognostic markers and therapeutic targets by performing proteomic analysis of CRC and paired normal tissues. AIM: To investigate the proteomic signature and identify novel protein prognostic biomarkers of CRC. METHODS: Cancer tissues and paired normal tissues were collected from 48 patients who underwent surgical removal at the China-Japan Friendship Hospital from January 2020 to June 2021. Data independent acquisition (DIA) quantitative proteomic analysis was performed using high-performance liquid chromatography-mass spectrometry/mass spectrometry (nano-UHPLC-MS/MS) to identify differentially expressed proteins, among which those with a P adj value (t test, BH correction) < 0.05 and an absolute fold change (|log2FC|) > 2 were identified as potential markers. Differentially expressed proteins were selected by bioinformatics analysis and validated by immunohistochemical tissue microarrays, and their association with prognosis was further analyzed with the Gene Expression Profiling Interactive Analysis database to identify prognostic protein biomarkers of CRC. RESULTS: Significantly differential protein expression was observed between cancer tissues and normal tissues. Compared with normal tissues, 1115 proteins were upregulated and 705 proteins were downregulated in CRC based on P adj < 0.05 and |log2FC| > 2, and bioinformatics analysis revealed that the differentially expressed proteins were involved in multiple biological processes associated with tumorigenesis, including ribosome biogenesis in eukaryotes, focal adhesion, extracellular matrix-receptor interactions and other tumor metabolism processes. Moreover, cyclin-dependent kinase inhibitor 2A (CDKN2A) expression was markedly upregulated in CRC, as validated by immunohistochemistry (0.228 vs 0.364, P = 0.0044), and was significantly enriched in tumor proliferation and signal transduction pathways such as the cell cycle and p53 signaling pathways. High CDKN2A expression was significantly correlated with poor prognosis (P = 0.021). These results demonstrated that CDKN2A functions as a driver of CRC. CONCLUSION: Our study provides a comprehensive proteomic signature of CRC and highlights CDKN2A as a potential powerful prognostic marker and precision therapeutic target.

Circulating miR-627-5p and miR-199a-5p are promising diagnostic biomarkers of colorectal neoplasia.

BACKGROUND: Early detection of colorectal neoplasms, including colorectal cancers (CRCs) and advanced colorectal adenomas (AAs), is crucial to improve patient survival. Circulating microRNAs (miRNAs) in peripheral blood are emerging as noninvasive diagnostic markers for multiple cancers, but their potential for screening colorectal neoplasms remains ambiguous. AIM: To identify candidate circulating cell-free miRNAs as diagnostic biomarkers in patients with colorectal neoplasms. METHODS: The study was divided into three phases: (1) Candidate miRNAs were selected from three public miRNA datasets using differential gene expression analysis methods; (2) an independent set of serum samples from 60 CRC patients, 60 AA patients and 30 healthy controls (HCs) was included and analyzed by quantitative real-time polymerase chain reaction for miRNAs, and their diagnostic power was detected by receiver operating characteristic (ROC) analysis; and (3) the origin and function of miRNAs in cancer patients were investigated in cancer cell lines and tumor tissues. RESULTS: Based on bioinformatics analysis, miR-627-5p and miR-199a-5p were differentially expressed in both the serum and tissues of patients with colorectal neoplasms and HCs and were selected for further study. Further validation in an independent cohort revealed that both circulating miR-627-5p and miR-199a-5p were sequentially increased from HCs and AAs to CRCs. The diagnostic power of miR-672-5p yielded an area under the curve (AUC) value of 0.90, and miR-199a-5p had an AUC of 0.83 in discriminating colorectal neoplasms from HCs. A logistic integrated model combining miR-199a-5p and miR-627-5p exhibited a higher diagnostic performance than either miRNA. Additionally, the levels of serum miR-627-5p and miR-199a-5p in CRC patients were significantly lower after surgery than before surgery and the expression of both miRNAs was increased with culture time in the culture media of several CRC cell lines, suggesting that the upregulated serum expression of both miRNAs in CRC might be tumor derived. Furthermore, in vitro experiments revealed that miR-627-5p and miR-199a-5p acted as tumor suppressors in CRC cells. CONCLUSION: Serum levels of miR-199a-5p and miR-627-5p were markedly increased in patients with colorectal neoplasms and showed strong potential as minimally invasive biomarkers for the early screening of colorectal neoplasms.

Identification of circ_0000375 and circ_0011536 as novel diagnostic biomarkers of colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) imposes a tremendous burden on human health, with high morbidity and mortality. Circular ribonucleic acids (circRNAs), a new type of noncoding RNA, are considered to participate in cancer pathogenesis as microRNA (miRNA) sponges. However, the dysregulation and biological functions of circRNAs in CRC remain to be explored. AIM: To identify potential circRNA biomarkers of CRC and explore their functions in CRC carcinogenesis. METHODS: CircRNAs and miRNAs differentially expressed in CRC tissues were identified by analyzing expression profiles from the Gene Expression Omnibus (GEO) database. Circ_0000375 and circ_0011536 were selected as CRC biomarker candidates. Quantitative real-time polymerase chain reaction was utilized to evaluate the expression of these 2 circRNAs in CRC tissues, serums and cell lines. Receiver operating characteristic curves were generated to assess the diagnostic performances of these 2 circRNAs. Then, functional experiments, including cell counting kit-8, wound healing and Transwell invasion assays, were performed after the overexpression of circ_0000375 and circ_0011536 in CRC cell lines. Furthermore, candidate target miRNAs of circ_0000375 and circ_0011536 were predicted via bioinformatics analysis. The expression levels of these miRNAs were explored in CRC cell lines and tissues from GEO datasets. A luciferase reporter assay was developed to examine the interactions between circRNAs and miRNAs. Based on the target miRNAs and downstream genes, functional enrichment analyses were applied to reveal the critical signaling pathways involved in CRC carcinogenesis. RESULTS: Downregulated circ_0000375 and circ_0011536 expression was observed in CRC tissues in GSE126095, clinical CRC tissue and serum samples and CRC cell lines. The areas under the curve for circ_0000375 and circ_0011536 were 0.911 and 0.885 in CRC tissue and 0.976 and 0.982 in CRC serum, respectively. Moreover, the serum levels of these 2 circRNAs were higher in patients at 30 d postsurgery than in patients before surgery, suggesting that the serum expression of circ_0000375 and circ_0011536 is related to CRC tumorigenesis. Circ_0000375 and circ_0011536 overexpression inhibited the proliferation, migration and invasion of CRC cells. Furthermore, miR-1182 and miR-1246, which were overexpressed in CRC tissues in GSE41655, GSE49246 and GSE115513, were verified as target miRNAs of circ_0000375 and circ_0011536, respectively, by luciferase reporter assays. The downstream genes of miR-1182 and miR-1246 were enriched in some CRC-associated pathways, such as the Wnt signaling pathway. CONCLUSION: Circ_0000375 and circ_0011536 may function as tumor suppressors in CRC progression, serving as novel biomarkers for CRC diagnosis and as promising candidates for therapeutic exploration.

Identification of the circRNA-miRNA-mRNA regulatory network and its prognostic effect in colorectal cancer.

BACKGROUND: The high morbidity and mortality of colorectal cancer (CRC) have posed great threats to human health. Circular RNA (CircRNA) and microRNA (miRNA), acting as competing endogenous RNAs (ceRNAs), have been found to play vital roles in carcinogenesis. However, the biological function of ceRNAs in CRC pathogenesis and prognosis remains largely unexplored. AIM: To identify the CRC-specific circRNA-miRNA-mRNA regulatory network and uncover the subnetwork associated with its prognosis. METHODS: CircRNAs, miRNAs and mRNAs differentially expressed (DE) in CRC tissues were selected by expression file analysis in the Gene Expression Omnibus (GEO) database, and the downstream target molecules of circRNAs and miRNAs were predicted. Then, the intersection of differentially expressed RNA molecules with the predicted targets was determined to obtain a ceRNA network. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted to elucidate the possible mechanism of pathogenesis. A survival analysis using the gene profiles and clinical information in The Cancer Genome Atlas (TCGA) database was performed to identify the mRNAs associated with the clinical outcome of CRC patients and construct a prognostic subnetwork. RESULTS: We downloaded three datasets (GSE126095, GSE41655 and GSE41657) of large-scale CRC samples from the GEO database. There were 55 DEcircRNAs, 114 DEmiRNAs and 267 DEmRNAs in CRC tissues compared with normal tissues. After intersecting these molecules with predicted targets, 19 circRNAs, 13 miRNAs and 28 mRNAs were chosen to develop a circRNA-miRNA-mRNA network. GO and KEGG functional enrichment analyses indicated that the retinol metabolic process, leukocyte chemotaxis, extracellular matrix remodeling, endoplasmic reticulum stress, alcohol dehydrogenase activity, gastric acid secretion, nitrogen metabolism and NOD-like receptor signaling pathway might participate in the tumorigenesis of CRC. After verifying the identified mRNA effect in the TCGA database, we finally recognized 3 mRNAs (CA2, ITLN1 and LRRC19) that were significantly associated with the overall survival of CRC patients and constructed a ceRNA subnetwork including 5 circRNAs (hsa_circ_0080210, hsa_circ_0007158, hsa_circ_0000375, hsa_circ_0018909 and hsa_circ_0011536) and 3 miRNAs (hsa-miR-601, hsa-miR-671-5p and hsa-miR-765), which could contain innovative and noninvasive indicators for the early screening and prognostic prediction of CRC. CONCLUSION: We proposed a circRNA-miRNA-mRNA regulatory network closely associated with the progression and clinical outcome of CRC that might include promising biomarkers for carcinogenesis and therapeutic targets.

Alteration of fecal tryptophan metabolism correlates with shifted microbiota and may be involved in pathogenesis of colorectal cancer.

BACKGROUND: Gut tryptophan (Trp) metabolites are produced by microbiota and/or host metabolism. Some of them have been proven to promote or inhibit colorectal cancer (CRC) in vitro and animal models. We hypothesized that there is an alteration of gut Trp metabolism mediated by microbiota and that it might be involved in the pathogenesis of cancer in patients with CRC. AIM: To investigate the features of Trp metabolism in CRC and the correlation between fecal Trp metabolites and gut microbiota. METHODS: Seventy-nine patients with colorectal neoplastic lesions (33 with colon adenoma and 46 with sporadic CRC) and 38 healthy controls (HCs) meeting the inclusion and exclusion criteria were included in the study. Their demographic and clinical features were collected. Fecal Trp, kynurenine (KYN), and indoles (metabolites of Trp metabolized by gut microbiota) were examined by ultraperformance liquid chromatography coupled to tandem mass spectrometry. Gut barrier marker and indoleamine 2,3-dioxygenase 1 (IDO1) mRNA were analyzed by quantitative real-time polymerase chain reaction. Zonula occludens-1 (ZO-1) protein expression was analyzed by immunohistochemistry. The gut microbiota was detected by 16S ribosomal RNA gene sequencing. Correlations between fecal metabolites and other parameters were examined in all patients. RESULTS: The absolute concentration of KYN [1.51 (0.70, 3.46) nmol/g vs 0.81 (0.64, 1.57) nmol/g, P = 0.036] and the ratio of KYN to Trp [7.39 (4.12, 11.72) × 10-3 vs 5.23 (1.86, 7.99) × 10-3, P = 0.032] were increased in the feces of patients with CRC compared to HCs, while the indoles to Trp ratio was decreased [1.34 (0.70, 2.63) vs 2.46 (1.25, 4.10), P = 0.029]. The relative ZO-1 mRNA levels in patients with CRC (0.27 ± 0.24) were significantly lower than those in HCs (1.00 ± 0.31) (P < 0.001), and the relative IDO1 mRNA levels in patients with CRC [1.65 (0.47-2.46)] were increased (P = 0.035). IDO1 mRNA levels were positively associated with the KYN/Trp ratio (r = 0.327, P = 0.003). ZO-1 mRNA and protein levels were positively correlated with the indoles/Trp ratio (P = 0.035 and P = 0.009, respectively). In addition, the genera Asaccharobacter (Actinobacteria) and Parabacteroides (Bacteroidetes), and members of the phylum Firmicutes (Clostridium XlVb, Fusicatenibacter, Anaerofilum, and Anaerostipes) decreased in CRC and exhibited a positive correlation with indoles in all subjects. CONCLUSION: Alteration of fecal Trp metabolism mediated by microbiota is associated with intestinal barrier function and tissue Trp metabolism, and may be involved in the pathogenesis of CRC.

Untargeted metabolomics characteristics of nonobese nonalcoholic fatty liver disease induced by high-temperature-processed feed in Sprague-Dawley rats.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) has become one of the most common chronic liver diseases in the world. In our early clinical data and questionnaire analysis of NAFLD, it was found that the body mass index of some patients did not meet the diagnostic criteria for overweight or obesity. The consumption of high-temperature-processed foods such as fried food, hot pot and barbecue is closely related to the occurrence of nonobese NAFLD. Reducing the intake of this kind of food can reduce disease severity and improve prognosis. AIM: To explore the untargeted metabolomics characteristics of nonobese nonalcoholic fatty liver disease in Sprague-Dawley rats induced by high-temperature-processed feed. METHODS: Fifty-four male Sprague-Dawley rats were divided into three groups: The control group received a standard diet; the nonfried soybeans (NDFS) group received 60% NDFS and 40% basic feed and the dry-fried soybeans (DFS) group received 60% DFS and 40% basic feed. Six rats were sacrificed at week 4, 8, and 12 in each group. The food intake, body weight, Lee's index, liver index, serological index and hepatic histopathology were assessed. Untargeted metabolomics characteristics were used to analyze the changes in liver metabolites of rats at week 12. Correlations between metabolites and pathology scores between the DFS and control groups and between the DFS and NDFS groups were analyzed. We selected some of the metabolites, both within the pathway and outside of the pathway, to explain preliminarily the difference in liver pathology in the three groups of rats. RESULTS: There were no statistically significant differences in the food intake, body weight, Lee's index or serological index between the DFS group and the control group (P > 0.05). At week 8 and week 12, the steatosis scores in the DFS group were significantly higher than those in the other two groups (P < 0.05). At week 12, the liver index of the DFS group was the lowest (NDFS group vs DFS group, P < 0.05). The fibrosis score in the DFS group was significantly higher than those in the other two groups (P < 0.05). The correlation analysis of the liver pathology score and differential metabolites in the DFS and NDFS groups showed that there were 10 strongly correlated substances: Five positively correlated substances and five negatively correlated substances. The positively correlated substances included taurochenodeoxycholate-3-sulfate, acetylcarnitine, 20a,22b-dihydroxycholesterol, 13E-tetranor-16-carboxy-LTE4 and taurocholic acid. The negatively correlated substances included choline, cholesterane-3,7,12,25-tetrol-3-glucuronide, nicotinamide adenine dinucleotide phosphate, lysoPC [16:1 (9Z)] and glycerol 3-phosphate. The correlation analysis of the liver pathology score and differential metabolites in the DFS and control groups showed that there were 13 strongly correlated substances: Four positively correlated substances and 9 negatively correlated substances. The positively correlated substances included 4-hydroxy-6-eicosanone, 3-phosphoglyceric acid, 13-hydroxy-9-methoxy-10-oxo-11-octadecenoic acid and taurochenodeoxycholate-3-sulfate. The negatively correlated substances included lysoPC [16:1(9Z)], S-(9-hydroxy-PGA1)-glutathione, lysoPC [20:5 (5Z, 8Z, 11Z, 14Z, 17Z)], SM (d18:1/14:0), nicotinamide adenine dinucleotide phosphate, 5,10-methylene-THF, folinic acid, N-lactoyl-glycine and 6-hydroxy-5-methoxyindole glucuronide. CONCLUSION: We successfully induced liver damage in rats by using a specially prepared high-temperature-processed feed and explored the untargeted metabolomics characteristics.

A novel RT-LAMP assay for rapid and simple detection of classical swine fever virus.

A simple and rapid assay for the detection of Classical swine fever virus (CSFV) was established using reverse transcription loop-mediated isothermal amplification (RT-LAMP). This study describes the amplification of the genomic RNA of CSFV under isothermal conditions (63 °C) within one hour, using a set of six primers (two outer primers, two inner primers and two loop primers). This RT-LAMP assay showed 100-fold higher sensitivity than the standard RT-PCR method and identified eighteen additional positive cases that were negative when tested by RT-PCR. This RT-LAMP was able to detect all the 13 strains of CSFV but not the BVDV. PRRSV. SIV. PRV-PCV, thus showed a good specificity. Products amplified by RT-LAMP can be visualized by agarose gel electrophoresis and in addition, either as a white precipitate at the bottom of the tube after a pulse spin or as a color change when dyed with SYBR Green I which are visible to the naked eye. Because RT-LAMP is low-cost and produces rapid results, it has the potential to be an excellent tool for CSFV surveillance in the field, especially in developing countries.