Objective:To study the trinucleotide repeats of GCN (GCA, GCT, GCC, GCG) encoding Alanine in exon 3 of the PHOX2B gene among healthy individuals from southwest China and two patients with Congenital central hypoventilation syndrome (CCHS). Methods:The number and sequence of the GCN repeats of the PHOX2B gene were analyzed by capillary electrophoresis, Sanger sequencing and cloning sequencing of 518 healthy individuals and two newborns with CCHS, respectively. Results:Among the 1036 alleles of the 518 healthy individuals, five alleles were identified, including (GCN) 7, (GCN) 13, (GCN) 14, (GCN) 15 and (GCN) 20. The frequency of the (GCN) 20 allele was the highest (94.79%). And five genotypes were identified, which included (GCN) 7/(GCN) 20, (GCN) 13/(GCN) 20, (GCN) 14/(GCN) 20, (GCN) 15/(GCN) 20, (GCN) 20/(GCN) 20. The homozygous genotypes were all (GCN) 20/(GCN) 20, and the carrier rate was 89.58%. Four GCN sequences of the (GCN) 20 homozygous genotypes were identified among the 464 healthy individuals. The GCN repeat numbers in the exon 3 of the PHOX2B gene showed no significant difference between the expected and observed values, and had fulfilled the, Hardy-Weinberg equilibrium. The genotypes of the two CCHS patients were (GCN) 20/(GCN) 25 and (GCN) 20/(GCN) 30, respectively. Conclusion:It is important to determine the GCN repeats and genotypic data of the exon 3 of the PHOX2B gene among the healthy individuals. The number of GCN repeats in 518 healthy individuals was all below 20. The selection of appropriate methods can accurately detect the polyalanine repeat mutations (PARMs) of the PHOX2B gene, which is conducive to the early diagnosis, intervention and treatment of CCHS.

Objective To investigate the molecular typing,virulence,and drug resistance features of bacterial strains in a simultane-ous outbreak of paratyphoid fever A and B,and then provide evidence for the prevention and treatment of the simultaneous transmission of different types of paratyphoid fever.Methods The clinical data of 31 patients confirmed as paratyphoid fever in the Hospital of Xin-jiang Production and Construction Corps from September 2018 to November 2018 were retrospectively analyzed.The isolated strains were performed serotyping and drug sensitivity tests.The molecular typing and the detection of virulence and drug resistance genes were carried out by multiplex PCR,pulsed-field gel electrophoresis(PFGE),and multilocus sequence typing(MLST).Results A total of 32 strains of Salmonella paratyphi were isolated from 31 patients,with 19 strains classified as type A and 13 as type B.The intermedi-ate rates of all strains against ciprofloxacin were 100%.The molecular typing and serotyping results of 11 representative strains were consistent.The PFGE fingerprints of Salmonella paratyphi A and B were also consistent.The MLST of Salmonella paratyphi A was ST85,and that of Salmonella paratyphi B was ST86.All strains carried virulence island SPI1-SPI5 representative genes such as invA,sitC,sseL,sifA,mgtC,siiE,and sopB,and regulatory gene phoP.Salmonella paratyphi A also carried cytolethal distending toxin(CDT)genes with trimeric structure such as cdtB,pltA,and pltB.The virulence plasmid genes such as pefA,prot6E,and spvB were all negative.Conclusion The simultaneous transmission of Salmonella paratyphi A and B has the characteristics of high pathogenicity and poor sensitivity to ciprofloxacin,which should be highly concerned by clinical and laboratory personnel.

Objective To investigate the effect of human colorectal fibroblast(CCD-18Co)-conditioned medium(CCD18-Co-CM)on biological behaviors of colorectal cancer(CRC)cells and explore the possible molecular mechanisms.Methods Real-time cellular analysis(RTCA),clone formation assay and wound healing assay were used to analyze the changes in proliferation,clone formation,and migration abilities of CRC cell lines HCT116 and Caco-2 treated with CCD18-Co-CM.Western blotting was used to detect the changes in ATK,ERK and STAT3 signaling pathways in the CRC cells activated by CCD18-Co-CM.The effect of CCD18-Co-CM on spheroidization ability of the cells was assessed with sphere-formation assay,and the changes in expressions of CRC stemness markers were detected using RT-PCR.Results CCD-18Co-CM significantly promoted proliferation,colony formation,and migration of HCT116 and Caco-2 cells,enhanced sphere-forming ability and expressions of CRC stemness markers,and increased ERK phosphorylation in the cells.Treatment with SCH772984 effectively inhibited CCD-18Co-CM-induced ERK signaling pathway activation,suppressed the malignant phenotype,and lowered the sphere-forming ability and expression of stemness markers of the two CRC cells.Conclusion Colorectal fibroblasts promote malignant phenotype of CRC cells by activating the ERK signaling pathway.

Objective To investigate the effect of human colorectal fibroblast(CCD-18Co)-conditioned medium(CCD18-Co-CM)on biological behaviors of colorectal cancer(CRC)cells and explore the possible molecular mechanisms.Methods Real-time cellular analysis(RTCA),clone formation assay and wound healing assay were used to analyze the changes in proliferation,clone formation,and migration abilities of CRC cell lines HCT116 and Caco-2 treated with CCD18-Co-CM.Western blotting was used to detect the changes in ATK,ERK and STAT3 signaling pathways in the CRC cells activated by CCD18-Co-CM.The effect of CCD18-Co-CM on spheroidization ability of the cells was assessed with sphere-formation assay,and the changes in expressions of CRC stemness markers were detected using RT-PCR.Results CCD-18Co-CM significantly promoted proliferation,colony formation,and migration of HCT116 and Caco-2 cells,enhanced sphere-forming ability and expressions of CRC stemness markers,and increased ERK phosphorylation in the cells.Treatment with SCH772984 effectively inhibited CCD-18Co-CM-induced ERK signaling pathway activation,suppressed the malignant phenotype,and lowered the sphere-forming ability and expression of stemness markers of the two CRC cells.Conclusion Colorectal fibroblasts promote malignant phenotype of CRC cells by activating the ERK signaling pathway.

Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)is the main pathogen that causes COVID-19,which is fast-mutating and highly transmissible.The infection has led to a global epidemic.As the main preventive and control measure,vaccination plays a critical role in fighting a-gainst COVID-19.Although a large number of epitope-based and mucosal vaccines have been stud-ied,few peptide epitope vaccines targeting the mucosa and their functional evaluation have been re-ported.In this study,we used SARS-CoV-2 structural protein M peptide epitope predicted by the IEDB database as an antigenic target to design the MS-3S gene containing 3 050 and 1 229 signal peptides and DCpep optimized for insertion into MS2 phage coat proteins.The expression plasmid pSIP:MS-3S was constructed by cloning the PCR fragments seamlessly and was transformed into Lactiplantibacillus plantarum 18 to obtain the recombinant bacterium LP18:MS-3S.Expression conditions such as induction time,inducer concentration,rotational speed and initial pH were opti-mized.The intranasal immunization experiments were performed to examine the vaccine efficacy.The results showed that the 916 bp-long target gene MS-3S modified and optimized was amplified and used to successfully construct the recombinant bacterial strain LP18:MS-3S.The optimal con-ditions for recombinant protein expression were obtained and verified by Western blot,flow cy-tometry,immunofluorescence and other detection methods.The optimal expression conditions were determined as follows:induction time was 4 h with 100 pg/L of SppIP as the optimal induction concentration.Antibody-specific for the epitope was verified by ELISA experiments in serum,alve-olar lavage fluid and fecal dilutions of mice.In summary,a recombinant bacterial strain expressing the epitope antigen of the SARS-CoV-2 M protein peptide was constructed.The obtained protein can induce the body to produce humoral and mucosal immunity,which lays the foundation for the development of a vaccine candidate for the mucosal immunity of COVID-19.

Background/Aims: Low educational attainment is a well-established risk factor for nonalcoholic fatty liver disease (NAFLD) in developed areas. However, the association between educational attainment and the risk of NAFLD is less clear in China. Methods: A cross-sectional study including over 200,000 Chinese adults across mainland China was conducted. Information on education level and lifestyle factors were obtained through standard questionnaires, while NAFLD and advanced fibrosis were diagnosed using validated formulas. Outcomes included the risk of NAFLD in the general population and high probability of fibrosis among patients with NAFLD. Logistic regression analysis was employed to estimate the risk of NAFLD and fibrosis across education levels. A causal mediation model was used to explore the potential mediators. Results: Comparing with those receiving primary school education, the multi-adjusted odds ratios (95% confidence intervals) for NAFLD were 1.28 (1.16 to 1.41) for men and 0.94 (0.89 to 0.99) for women with college education after accounting for body mass index. When considering waist circumference, the odds ratios (95% CIs) were 0.94 (0.86 to 1.04) for men and 0.88 (0.80 to 0.97) for women, respectively. The proportions mediated by general and central obesity were 51.00% and 68.04% for men, while for women the proportions were 48.58% and 32.58%, respectively. Furthermore, NAFLD patients with lower educational attainment showed an incremental increased risk of advanced fibrosis in both genders. Conclusions In China, a low education level was associated with a higher risk of prevalent NAFLD in women, as well as high probability of fibrosis in both genders.

OBJECTIVE: To investigate the mechanism by which conditioned medium of colorectal cancer cells promotes the formation of cancer-associated fibroblasts (CAFs). METHODS: Normal human colorectal fibroblasts (CCD-18Co cells) in logarithmic growth phase were treated with the conditioned media of colorectal cancer HCT116 cells (HCT116-CM) or Caco-2 cells (Caco-2-CM) alone or in combination with 300 nmol/L ERK inhibitor SCH772984. The expression levels of CAFs-related molecular markers were detected in the treated cells with real-time quantitative PCR (RT- qPCR) and immunofluorescence assay, and the changes in cell proliferation, colony formation and migration were assessed with RTCA, colony formation and wound healing assays; Western blotting was performed to detect the activated signaling pathways in the fibroblasts and the changes in CAFs formation after blocking of the signaling pathway. RESULTS: HCT116-CM and Caco-2-CM significantly upregulated mRNA expression levels of CAFs markers (including α-SMA, FAP, FN and TGF-β) in CCD-18Co cells, and strongly promoted fibroblast transformation into CAFs (P < 0.05). The two conditioned media also promoted the proliferation, colony formation and migration of CCD-18Co cells (P < 0.05) and significantly increased the levels of α-SMA protein and ERK phosphorylation in the cells (P < 0.05). The ERK inhibitor SCH772984 obviously inhibited the expression of α-SMA and the transformation of CCD-18Co cells into CAFs induced by the conditioned medium of colorectal cancer cells (P < 0.05). CONCLUSION Colorectal cancer cells may induce the formation of colorectal CAFs by activating the ERK pathway in the fibroblasts.
Humans ; Cancer-Associated Fibroblasts/metabolism* ; Culture Media, Conditioned/pharmacology* ; MAP Kinase Signaling System ; Caco-2 Cells ; Fibroblasts ; Signal Transduction ; Cell Proliferation ; Cell Line, Tumor ; Colorectal Neoplasms/genetics* ; Cell Movement

Acute-on-chronic liver failure (ACLF) is a clinical syndrome with short-term rapid deterioration, multiple organ failure, and high mortality and has become a research hotspot in the field of severe liver diseases. Early, accurate, and quick judgment of clinical prognosis and timely intervention of disease progression are of great significance for improving disease outcome. This article summarizes the inflammation-related prognostic markers for HBV-related ACLF found in recent years, so as to improve the existing prognostic indicators and provide more basis for evaluating the prognosis of ACLF patients by clinicians.

Objective:To explore the clinical significance of different metabolic parameters measured by 18F-fluorodeoxyglucose (FDG) PET/CT in predicting the effectiveness of chemotherapy in advanced lung adenocarcinoma patients. Methods:A set of metabolic parameters of PET/CT and clinical characteristics which were detected from 127 patients (70 males, 57 females, age (56.8±10.1) years) with advanced lung adenocarcinoma treated with at least two cycles of chemotherapy in Hainan Cancer Hospital between August 2017 and June 2019 were retrospectively analyzed. The effects of those parameters on patients′ survival were analyzed by receiver operating characteristic (ROC) curve, Kaplan-Meier method (log-rank test) and Cox proportional hazards model.Results:Maximum standardized uptake value (SUV max), metabolic tumor volume 30% (MTV 30), and total lesion glycolysis 30% (TLG 30) had larger areas under the curve (0.581, 0.606 and 0.693 respectively) compared with other imaging parameters, and the optimal cut-off values were 10.12, 20.21 cm 3 and 81.25 g respectively. Kaplan-Meier univariate and Cox analyses synergistically showed that clinical stage (hazard ratio ( HR)=0.293(95% CI: 0.190-0.451), P<0.001), smoking ( HR=0.732(95% CI: 0.605-0.885), P=0.001), and MTV 30 ( HR=1.555(95% CI: 1.078-2.242), P=0.018) had significant predictive value for progression-free survival (PFS). Stratified analysis showed that smoking and MTV 30>20.21 cm 3 were independent prognostic factors for poor PFS in patients with advanced lung adenocarcinoma receiving chemotherapy ( HR=0.738(95% CI: 0.611-0.893), P=0.002; HR=1.502(95% CI: 1.037-2.177), P=0.032). Conclusions:Clinical stage, smoking and MTV 30 are independent prognostic factors of PFS in patients with advanced lung adenocarcinoma receiving chemotherapy. MTV 30≤20.21 cm 3 is expected to be an image biomarker for predicting survival and selecting patients with advanced lung adenocarcinoma who are more likely to benefit from chemotherapy.