Objective:To discuss the effect of circadion rhythm gene TIMELESS(TIM)silencing on immune escape of the ovarian cancer cells,and to clarify its related mechanism.Methods:The CD8+T lymphocytes were isolated and identified by flow cytometry to detect the proportion of CD3+/CD8+cell subsets.The human ovarian cancer SK-OV-3 cells were cultured in vitro and divided into interference plasmid transfected with TIM small interfering(siRNA)(si-TIM),negative control plasmid(si-NC),programmed death ligand 1(PD-L1)over-expression plasmid(oe-PD-L1),and negative control plasmid(oe-NC)groups.The cells were further divided into blank control group(BC group,non-transfection),si-NC group(transfected with si-NC),si-TIM group(transfected with si-TIM),si-NC+oe-NC group(transfected with si-NC and oe-NC),and si-TIM+oe-PD-L1 group(transfected with si-TIM and oe-PD-L1).Real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods were used to detect the expression levels of TIM mRNA and protein in the SK-OV-3 cells to verify TIM gene silencing.The transfected SK-OV-3 cells were co-cultured with activated CD8+T lymphocytes and divided into BC group(SK-OV-3 cells cultured alone),BC/T group,si-NC/T group,si-TIM/T group,si-NC+oe-NC/T group,and si-TIM+oe-PD-L1/T group.CCK-8 method was used to detect the survival rates of the SK-OV-3 cells in various groups;flow cytometry was used to detect the apoptotic rates of the SK-OV-3 cells and positive expression rate of PD-L1 on surface of the cells in various groups;enzyme-linked immunosorbent assay(ELISA)method was used to detect the levels of interferon-γ(IFN-γ)and tumor necrosis factor-α(TNF-α)in the co-culture supernatant;lactate dehydrogenase(LDH)release assay was used to detect the cytotoxicity of the CD8+T lymphocytes in various groups;RT-qPCR method was used to detect the expression levels of TIM and PD-L1 mRNA in the SK-OV-3 cells in various groups;Western blotting method was used to detect the expression levels of TIM and PD-L1 proteins in the SK-OV-3 cells in various groups.Results:After scparated with immune magnetic bead method,the proportion of CD8+T lymphocyte(CD3+/CD8+)subsets was(96.56%±0.59%),indicating high purity of the extracted CD8+T lymphocytes.Compared with BC group,the expression levels of TIM mRNA and protein in the cells in si-TIM group were significantly decreased(P<0.01),suggesting successful TIM gene silencing in the ovarian cancer SK-OV-3 cells.The CCK-8 results showed that compared with BC group,the survival rate of the SK-OV-3 cells in BC/T group was significantly decreased(P<0.01);compared with BC/T group,the survival rate of the SK-OV-3 cells in si-TIM/T group was significantly decreased(P<0.01).The flow cytometry results showed that compared with BC group,the apoptotic rate of the SK-OV-3 cells in BC/T group was significantly increased(P<0.01);compared with BC/T group,the apoptotic rate of the SK-OV-3 cells in si-TIM/T group was significantly increased(P<0.01);compared with si-TIM/T group,the apoptotic rate of the SK-OV-3 cells in si-TIM+oe-PD-L1/T group was significantly decreased(P<0.01).Compared with BC group,the positive expression rate of PD-L1 on surface of the SK-OV-3 cells in si-TIM group was significantly decreased(P<0.01).The ELISA results showed that compared with BC/T group,the levels of IFN-γ and TNF-α in the culture supernatant in si-TIM/T group were significantly increased(P<0.01);compared with si-TIM/T group,the levels of IFN-γ and TNF-α in the supernatant in si-TIM+oe-PD-L1/T group were significantly decreased(P<0.01).The LDH release assay results showed that compared with BC/T group,the cytotoxicity of the CD8+T lymphocytes in si-TIM/T group was significantly increased(P<0.01);compared with si-TIM/T group,the cytotoxicity of the CD8+T lymphocytes in si-TIM+oe-PD-L1/T group was significantly weakened(P<0.01).The RT-qPCR and Western blotting results showed that compared with BC group,the expression levels of PD-L1 mRNA and protein in the SK-OV-3 cells in si-TIM group were significantly decreased(P<0.01);compared with si-TIM group,the expression level of PD-L1 protein in the cells in si-TIM+oe-PD-L1 group was significantly increased(P<0.01).Conclusion:TIM gene silencing enhances the cytotoxic effect of CD8+T lymphocytes on ovarian cancer SK-OV-3 cells and inhibits immune escape,and its mechanism may be related to the regulation of PD-L1 protein expression.

Objective: To uncover the underlying mechanisms of action of the Yinlai decoction on high-calorie diet-induced pneumonia through proteomics analysis. Methods: Based on the Gene Expression Omnibus (GEO) database, lung tissue samples from normal and high-fat diet (HFD) fed mice in the GSE16377 dataset were selected as test cohorts to identify differentially expressed genes and conduct bioinformatics analyses. In the animal experiments, mice were randomly divided into the control (N), high-calorie diet pneumonia (M), and Yinlai decoction treatment (Y) groups. Mice in the M group received high-calorie feed and a 0.5 mg/mL lipopolysaccharide solution spray for 30 min for 3 d. The mice in the Y group were intragastrically administered 2 mL/10 g Yinlai decoction twice daily for 3 d. Pathological evaluation of the lung tissue was performed. Differentially expressed proteins (DEPs) in the lung tissue were identified using quantitative proteomics and bioinformatics analyses. The drug-target relationships between Yinlai decoction and core DEPs in the lung tissue were verified using AutoDock Vina and Molecular Graphics Laboratory (MGL) Tools. DEPs were verified by western blot. Results: GEO data mining showed that an HFD altered oxidative phosphorylation in mouse lung tissue. The Yinlai decoction alleviated pathological damage to lung tissue and pneumonia in mice that were fed a high-calorie diet. A total of 47 DEPs were identified between the Y and M groups. Enrichment analysis revealed their association with energy metabolism pathways such as the tricarboxylic acid cycle (TCA) and oxidative phosphorylation. The protein-protein interaction network revealed that Atp5a1, Pdha1, and Sdha were the target proteins mediating the therapeutic effects of Yinlai decoction. Molecular docking results suggested that the mechanism of the therapeutic effect of Yinlai decoction involves the binding of brassinolide, praeruptorin B, chrysoeriol, and other components in Yinlai decoction to Atp5a1. Conclusion The Yinlai decoction alleviated lung tissue damage and pneumonia in mice that were fed a high-calorie diet by regulating the TCA and oxidative phosphorylation. Our study highlights the importance of a healthy diet for patients with pneumonia and provides a scientific basis for the prevention and treatment of pneumonia through dietary adjustments.

Objective To investigate the changing distribution and antibiotic resistance profiles of clinical isolates of Staphylococcus in hospitals across China from 2015 to 2021.Methods Antimicrobial susceptibility testing was conducted for the clinical isolates of Staphylococcus according to the unified protocol of CHINET(China Antimicrobial Surveillance Network)using disk diffusion method and commercial automated systems.The CHINET antimicrobial resistance surveillance data from 2015 to 2021 were interpreted according to the 2021 CLSI breakpoints and analyzed using WHONET 5.6.Results During the period from 2015 to 2021,a total of 204,771 nonduplicate strains of Staphylococcus were isolated,including 136,731(66.8％)strains of Staphylococcus aureus and 68,040(33.2％)strains of coagulase-negative Staphylococcus(CNS).The proportions of S.aureus isolates and CNS isolates did not show significant change.S.aureus strains were mainly isolated from respiratory specimens(38.9±5.1)％,wound,pus and secretions(33.6±4.2)％,and blood(11.9±1.5)％.The CNS strains were predominantly isolated from blood(73.6±4.2)％,cerebrospinal fluid(12.1±2.5)％,and pleural effusion and ascites(8.4±2.1)％.S.aureus strains were mainly isolated from the patients in ICU(17.0±7.3)％,outpatient and emergency(11.6±1.7)％,and department of surgery(11.2±0.9)％,whereas CNS strains were primarily isolated from the patients in ICU(32.2±9.7)％,outpatient and emergency(12.8±4.7)％,and department of internal medicine(11.2±1.9)％.The prevalence of methicillin-resistant strains was 32.9％in S.aureus(MRSA)and 74.1％in CNS(MRCNS).Over the 7-year period,the prevalence of MRSA decreased from 42.1％to 29.2％,and the prevalence of MRCNS decreased from 82.1％to 68.2％.MRSA showed higher resistance rates to all the antimicrobial agents tested except trimethoprim-sulfamethoxazole than methicillin-susceptible S.aureus(MSSA).Over the 7-year period,MRSA strains showed decreasing resistance rates to gentamicin,rifampicin,and levofloxacin,MRCNS showed decreasing resistance rates to gentamicin,erythromycin,rifampicin,and trimethoprim-sulfamethoxazole,but increasing resistance rate to levofloxacin.No vancomycin-resistant strains were detected.The prevalence of linezolid-resistant MRCNS increased from 0.2％to 2.3％over the 7-year period.Conclusions Staphylococcus remains the major pathogen among gram-positive bacteria.MRSA and MRCNS were still the principal antibiotic-resistant gram-positive bacteria.No S.aureus isolates were found resistant to vancomycin or linezolid,but linezolid-resistant strains have been detected in MRCNS isolates,which is an issue of concern.

Humans ; Asthma/drug therapy* ; Biological Therapy

Objective:To investigate the clinical application value and safety of magnetically controlled capsule gastroscopy (MCCG) in gastric and duodenal examination of children in comparison with conventional gastroscopy.Methods:Data of 160 outpatients or inpatients with abdominal pain accompanied by Helicobacter pylori infection aged 8-16 who underwent either MCCG or conventional gastroscopy in Shanghai Children's Hospital from March 2020 to March 2022 were retrospectively analyzed. Children were divided into the MCCG group ( n=80) and the conventional gastroscopy group ( n=80) according to different examination methods. The detection and examination time of lesions in upper gastrointestinal tract, tolerance and safety between the two groups were analyzed. Results:MCCG was successfully performed in 79 children and conventional gastroscopy was successfully performed in 78 children, respectively. The positive detection rates were 1.3% (1/79) and 1.3% (1/78) in the esophagus ( χ2=0.000, P>0.999), 87.3% (69/79) and 91.0% (71/78) in the stomach ( χ2=0.552, P=0.327) , 15.2% (12/79) and 19.2% (15/78) in duodenum ( χ2=0.450, P=0.533) with no significant difference between the two groups. There was no significant difference in the examination time [72.0 (41.0, 109.5) min VS 6.0 (4.3, 7.0) min, U=24, P<0.001] in the MCCG group and the conventional gastroscopy group. No adverse event occurred in either group. Conclusion:There is no significant difference in the detection rate of gastric and duodenal lesions between the MCCG group and the conventional gastroscopy group. MCCG is safe and stable, and can be used as an diagnostic tool for gastric and duodenal diseases in children.

To investigate the diagnostic value of narrow-band imaging (NBI) endoscopy for esophageal polyps in children. Microscopic morphology of various polyps in 35 children with esophageal polyps in Children's Hospital of Shanghai from January 2016 to June 2020 were observed under both traditional white light endoscopy and NBI endoscopy. The sensitivity and specificity of traditional white light endoscopy and NBI endoscopy were compared with the pathological results as the gold standard. A total of 70 esophageal polypoid lesions were found in 35 children, including 27 single polyps. Pathological results indicated that the majority of polyps were non-neoplastic polyps (52.9%, 37/70).The sensitivity of NBI endoscopy in the diagnosis of esophageal neoplastic polyps was significantly higher than that of white light endoscopy [93.9% (31/33) VS 90.9% (30/33), P < 0.001], and the specificity was also higher [89.2% (33/37) VS 78.4% (29/37), P=0.864]. By observing the microscopic structure of esophageal polyps, NBI endoscopy contributes to the clinical prediction of the pathological properties of polyps. Its sensitivity is superior to the white light endoscopy.

ObjectiveTo screen out the mRNAs involved in the resistance of hepatoma cells to anlotinib using ceRNA microarray. MethodsHigh-dose shock combined with low-dose induction was used to culture hepatoma cells resistant to anlotinib, and CCK8 assay was used to verify the difference in the proliferation of drug-resistant hepatoma cells treated by anlotinib. The ceRNA microarray was used to screen out the differentially expressed genes between drug-resistant hepatoma cells and normal hepatoma cells, and real-time PCR was used to verify the differentially expressed genes detected by some microarrays. the independent samples t-test was used for comparison of continuous data between two groups, and the Kaplan-Meier method was used to analyze the overall survival of hepatoma cells samples, and the log-rank test was used to compare survival rates. Fisher’s exact test was used for chip screening. ResultsThere was a significant difference in gene expression between drug-resistant hepatoma cells and normal hepatoma cells, and 10 genes with the greatest difference were screened out for analysis by reducing the range. There were 4 genes associated with drug resistance and tumor growth, i.e., BIRC2, BIRC7, ABCC2, and MAPK8. There were significant reductions in the expression levels of BIRC2, ABCC2, and MAPK8 (P=0001 4, 0001 2, and 0.011 8), and there was a significant increase in the expression of BIRC7 (P＜0.001). The results of real-time PCR were consistent with those of microarray (t=10.74,32.65,18.34, and 2.80; P=0.000 4, 0.000 1, 0.000 1, and 0.044 8). The high expression of BIRC7 and the low expression of MAPK8 were associated with the significant reduction in survival time (P=0.022 0 and 0.005 6). ConclusionBIRC2, BIRC7, ABCC2, and MAPK8 are differentially expressed between anlotinib-resistant hepatoma cells and normal hepatoma cells and may be involved in the resistance of hepatoma cells to anlotinib.

Objective: To search for specific metabolites in the lungs of pneumonia rats fed with a high-calorie diet, as well as explore the changes in the lung metabolites of young rats treated with Yinlai Decoction (YD) and its effects on inflammation-related metabolic pathways.Methods: Lipopolysaccharides (LPS) and a special high-calorie diet were used to induce Sprague Dawley (SD) rats to simulate the intestinal state of infant pneumonia. Liquid chromatography-mass spectrometry technology (LC-MS/MS) was used to detect metabolites in each group. Supervised orthogonal partial least squares discriminant analysis (OPLS-DA) model values were used for the detection results to find the differential metabolites. The metabolic pathways that are involved with the differential metabolites were clarified through enrichment analysis and topological analysis. Finally, the T cell receptor signaling pathway (TCR) signal conversion was analyzed by the network pharmacology method. Results: In the high-calorie diet combined with pneumonia group (M3), a total of 55 metabolites were determined to be different from the normal group (N). A total of 36 metabolites were determined to be different from those in the lung metabolites of the YD treatment group (T1). YD had a regulatory effect on glutathione metabolism, arginine and proline metabolism, ascorbic acid and aldehyde metabolism and phenylalanine metabolism. And the small molecule metabolites could act on the FYN and lymphocyte-specific protein tyrosine kinase (LCK) target proteins in the TCR signaling pathway, thereby affecting the immune function of the lungs. Conclusion: A high-calorie diet can cause abnormal sphingolipid metabolism in the lungs of young rats, thereby creating chronic lung inflammation in young rats. YD has a beneficial effect when used to treat young rats with LPS-induced pneumonia fed on high-calorie diets. Its mechanisms of action may affect the body's immune pathways by regulating the oxidative stress pathway affected by glutathione metabolism.

The COVID-19 outbreak may have some impact on the use of biologics in psoriatic patients because immunosuppressive effects of biologics may potentially alter the susceptibility of patients to the virus, deteriorate the condition of infected patients or even change the prognosis of infection. According to currently available recommendations from international psoriasis academic organizations and specialists, as well as specific situation in China, the authors provide some guidance on the use of biologics for psoriatic patients undergoing or planning to undergo treatment with biologics, those with low or high risk of infection, and for those with or without COVID-19 infection, so as to provide references for clinical practice.

Objective To observe and explore the differentiation value of serum hepcidin level in iron defi-ciency anemia(IDA) and anemia of chronic disease(ACD) .Methods 40 cases of IDA in the hospital from Oc-tober 2015 to February 2017 and 40 cases of ACD were selected as the ACD group and in the same period 50 healthy people undergoing physical examination served as the control group .The ELISA method was adopted to detect serum hepcidin level in each group .The levels of serum iron and total iron binding capacity (TIBC) were detected by using the ferrous benzoxazines colorimetric method .Then the serum hepcidin levels were performed the contrastive analysis and statistical processing .Results The serum hepcidin level in the IDA group was lower than that in the control group ,the difference was statistically significant (P<0 .05) ,and ser-um hepcidin level in the ACD group was higher than that in control group ,the difference was statistically sig-nificant (P<0 .05) .The areas of under the receiver operating characteristic (ROC) curve for hepcidin ,serum iron ,TIBC in the differentiation diagnosis of ACD and IDA were 0 .98 ,0 .67 and 0 .86 respectively .Conclusion Serum hepcidin level is a simple and easy method for the differentiation diagnosis of IDA and ACD ,and its accuracy is superior to that of serum iron and TIBC levels .