LC–MS-based serum metabolomics analysis for the screening and monitoring of colorectal cancer

BackgroundColorectal Cancer (CRC) is a prevalent digestive system tumour with significant mortality and recurrence rates. Serum metabolomics, with its high sensitivity and high throughput, has shown potential as a tool to discover biomarkers for clinical screening and monitoring of the CRC patients.MethodsSerum metabolites of 61 sex and age-matched healthy controls and 62 CRC patients (before and after surgical intervention) were analyzed using a ultra-performance liquid chromatography-high resolution mass spectrometer (UPLC-MS). Statistical methods and pathway enrichment analysis were used to identify potential biomarkers and altered metabolic pathways.ResultsOur analysis revealed a clear distinction in the serum metabolic profile between CRC patients and healthy controls (HCs). Pathway analysis indicated a significant association with arginine biosynthesis, pyrimidine metabolism, pantothenate, and CoA biosynthesis. Univariate and multivariate statistical analysis showed that 9 metabolites had significant diagnostic value for CRC, among them, Guanosine with Area Under the Curve (AUC) values of 0.951 for the training group and0.998 for the validation group. Furthermore, analysis of four specific metabolites (N-Phenylacetylasparticacid, Tyrosyl-Gamma-glutamate, Tyr-Ser and Sphingosine) in serum samples of CRC patients before and after surgery indicated a return to healthy levels after an intervention.ConclusionOur results suggest that serum metabolomics may be a valuable tool for the screening and monitoring of CRC patients

Spreading of Pandemic Vibrio parahaemolyticus O3:K6 and Its Serovariants: A Re-analysis of Strains Isolated from Multiple Studies

In China, V. parahaemolyticus has been a leading cause of foodborne outbreaks and bacterial infectious diarrhea since the 1990s, and most infections have been associated with the pandemic V. parahaemolyticus O3:K6 and its serovariants. However, a comprehensive overview of the sero-prevalence and genetic diversity of the pandemic V. parahaemolyticus clone in China is lacking. To compensate for this deficiency, pandemic isolates in both clinical and environmental Chinese samples collected from multiple studies were analyzed in this study. Surprisingly, as many as 27 clinical pandemic serovariants were identified and were widely distributed across nine coastal provinces and two inland provinces (Beijing and Sichuan). O3:K6, O4:K68, and O1:KUT represented the predominant clinical serovars. Only four environmental pandemic serovariants had previously been reported, and they were spread throughout Shanghai (O1:KUT, O3:K6), Jiangsu (O3:K6, O4:K48), Zhejiang (O3:K6), and Guangdong (O4:K9). Notably, 24 pandemic serovariants were detected within a short time frame (from 2006 to 2012). The pandemic isolates were divided into 15 sequence types (STs), 10 of which fell within clonal complex (CC) 3. Only three STs (ST3, ST192, and ST305) were identified in environmental isolates. Substantial serotypic diversity was mainly observed among isolates within pandemic ST3, which comprised 21 combinations of O/K antigens. The pandemic O3:K6 serotype showed a high level of sequence diversity, which was shared by eight different STs (ST3, ST227, ST431, ST435, ST487, ST489, ST526, and ST672). Antimicrobial susceptibility testing revealed that most isolates shared similar antibiotic susceptibility profiles. They were resistant to ampicillin but sensitive to most other drugs that were tested. In conclusion, the high levels of serotypic and genetic diversity of the pandemic clone suggest that the involved regions are becoming important reservoirs for the emergence of novel pandemic strains. We underscore the need for routine monitoring to prevent pandemic V. parahaemolyticus infection, which includes monitoring antimicrobial responses to avoid excessive misuse of antibiotics. Further investigations are also needed to delineate the specific mechanisms underlying the possible seroconversion of pandemic isolates

Expression and prognostic value of miR-486-5p in patients with gastric adenocarcinoma.

MicroRNA (miR)-486-5p expression is often reduced in human cancers. However, its expression in gastric carcinoma and its relation to clinicopathological features and prognosis are unclear. Tissue microarrays were constructed from 84 patients with gastric adenocarcinoma (GC) who were undergoing radical resection. miR-486-5p expression was detected by miRNA-locked nucleic acid in situ hybridization, and its correlations with clinicopathological features and overall survival were analyzed. Bioinformatic studies predict that fibroblast growth factor 9 (FGF9) is a potential target gene of miR-486-5p. miR-486-5p was mainly located in the cytoplasm of GC cells and neighboring normal tissues. Compared with paracancerous normal tissue, miR-486-5p expression was decreased in 63.1% (53/84) of the GC samples, increased in 32.1% (27/84) and unchanged in 4.8% (4/84). FGF9 expression was decreased in 69.0% (58/84) of GC samples and increased in 31.0% (26/84) compared with normal paracancerous tissues using immunohistochemical analysis. Low or unchanged miR-486-5p expression (P = 0.002), tumor stage (P = 0.001), tumor status (P = 0.001), node status (P = 0.001), tumor size (P = 0.004), and depth of tumor invasion (P = 0.013) were significant negative prognostic predictors for overall survival in patients with GC. After stratification according to American Joint Committee on Cancer (AJCC) stage, low/unchanged miR-486-5p expression remained a significant predictor of poor survival in stage II (P = 0.024) and stage III (P = 0.003). Cox regression analysis identified the following predictors of poor prognosis: tumor status (hazard ratio [HR], 7.19; 95% confidence interval [CI], 1.75-29.6; P = 0.006), stage (HR, 2.62; 95%CI, 1.50-4.59; P = 0.001), lymph node metastasis (HR, 2.52; 95% CI, 1.27-4.99; P = 0.008), low/unchanged miR-486-5p (HR, 2.47; 95% CI, 1.35-4.52; P = 0.003), high level of FGF9 (HR, 2.41; 95% CI, 1.42-4.09; P = 0.001) and tumor size (HR, 2.50; 95% CI, 1.30-4.82; P = 0.006). Low or unchanged expression of miR-486-5p compared with neighboring normal tissues was associated with a poor prognosis, while high expression was associated with a good prognosis in GC. miR-486-5p may thus be useful for evaluating prognosis and may provide a novel target treatment in patients with GC

The Role of Individual Disulfide Bonds of μ-Conotoxin GIIIA in the Inhibition of NaV1.4

μ-Conotoxin GIIIA, a peptide toxin isolated from Conus geographus, preferentially blocks the skeletal muscle sodium channel NaV1.4. GIIIA folds compactly to a pyramidal structure stabilized by three disulfide bonds. To assess the contributions of individual disulfide bonds of GIIIA to the blockade of NaV1.4, seven disulfide-deficient analogues were prepared and characterized, each with one, two, or three pairs of disulfide-bonded Cys residues replaced with Ala. The inhibitory potency of the analogues against NaV1.4 was assayed by whole cell patch-clamp on rNaV1.4, heterologously expressed in HEK293 cells. The corresponding IC50 values were 0.069 ± 0.005 μM for GIIIA, 2.1 ± 0.3 μM for GIIIA-1, 3.3 ± 0.2 μM for GIIIA-2, and 15.8 ± 0.8 μM for GIIIA-3 (-1, -2 and -3 represent the removal of disulfide bridges Cys3–Cys15, Cys4–Cys20 and Cys10–Cys21, respectively). Other analogues were not active enough for IC50 measurement. Our results indicate that all three disulfide bonds of GIIIA are required to produce effective inhibition of NaV1.4, and the removal of any one significantly lowers its sodium channel binding affinity. Cys10–Cys21 is the most important for the NaV1.4 potency

Population Structure of Clinical <i>Vibrio parahaemolyticus</i> from 17 Coastal Countries, Determined through Multilocus Sequence Analysis

<div><p><i>Vibrio parahaemolyticus</i> is a leading cause of food-borne gastroenteritis worldwide. Although this bacterium has been the subject of much research, the population structure of clinical strains from worldwide collections remains largely undescribed, and the recorded outbreaks of <i>V. parahaemolyticus</i> gastroenteritis highlight the need for the subtyping of this species. We present a broad phylogenetic analysis of 490 clinical <i>V. parahaemolyticus</i> isolates from 17 coastal countries through multilocus sequence analysis (MLST). The 490 tested isolates fell into 161 sequence types (STs). The eBURST algorithm revealed that the 161 clinically relevant STs belonged to 8 clonal complexes, 11 doublets, and 94 singletons, showing a high level of genetic diversity. CC3 was found to be a global epidemic clone of <i>V. parahaemolyticus</i>, and ST-3 was the only ST with an international distribution. <i>recA</i> was observed to be evolving more rapidly, exhibiting the highest degree of nucleotide diversity (0.028) and the largest number of polymorphic nucleotide sites (177). We also found that the high variability of <i>recA</i> was an important cause of differences between the results of the eBURST and ME tree analyses, suggesting that <i>recA</i> has a much greater influence on the apparent evolutionary classification of <i>V. parahaemolyticus</i> based on the current MLST scheme. In conclusion, it is evident that a high degree of genetic diversity within the <i>V. parahaemolyticus</i> population and multiple sequence types are contributing to the burden of disease around the world. MLST, with a fully extractable database, is a powerful system for analysis of the clonal relationships of strains at a global scale. With the addition of more strains, the pubMLST database will provide more detailed and accurate information, which will be conducive to our future research on the population structure of <i>V. parahaemolyticus</i>.</p></div

An ME tree was constructed using the concatenated sequences of the seven loci of each of the 161 STs obtained in this study.

<p>Squares, circles, and triangles with different shading represent the eight clonal complexes observed via eBURST. The scale represents the evolutionary distance, and bootstrap values over 50% are shown on the branches.</p

Genetic Diversity of the Seven Loci in 490 <i>Vibrio parahaemolyticus</i> Isolates.

<p>Genetic Diversity of the Seven Loci in 490 <i>Vibrio parahaemolyticus</i> Isolates.</p

Survival curves in patients with GC according to miRNA-486–5p levels.

<p><b>a</b> Overall survival curves in patients with GC according to miRNA-486–5p levels (<i>P</i> = 0.002). <b>b</b> Survival curves in stage-II GC according to miRNA-486–5p levels (<i>P</i> = 0.024). <b>c</b> Survival curves in stage-III GC according to miRNA-486–5p levels (<i>P</i> = 0.003).</p

miR-486-5p expression and clinicopathological features in patients with gastric adenocarcinoma.

<p>miR-486-5p expression and clinicopathological features in patients with gastric adenocarcinoma.</p