Aim To explore the mechanism of E.coli heat-labile enterotoxin B subunit(LTB)as adjuvant by analysis of cellular proteins interacting with LTB. Methods Whole cell proteins were purified from RAW 264.7 cell after treated with LTB or NaCl 12 h, respectively.The cellular proteins were interacted with LTB and the interacting proteins were purified by pull-down assay and identified by mass spectrography.The LTB interaction proteins were conformed with Western blot and immunofluorescence assay.Results 25 LTB interaction proteins were found,and their interaction network was mapped;four proteins (Jup,Dsp,Ddx5 and Vimentin)were indicated to be related with LTB adjuvant activity;immunofluorescence assay indicated that GM130 interacted with LTB,however,Vimentin had no interaction with LTB in vivo.After treated by LTB,the expression of β-actin was upregulated obvi-ously in RAW 264.7 cell,whereras,Hspd1 did not show any change.Conclusions LTB exerts adjuvant activity through binding to GM1 of immune cells,cau-sing endocytosis and transporting to the Golgi apparatus by vesicles.Then LTB might bind to Jup and affect TCF/LEF activity,regulating the expression of Bcl 2, IL-6,and Runx3.The result is promoted T cell and B cell proliferation,differentiation and activation by se-cretion of cytokines and immunoglobulins.

BACKGROUND:Classical drug for Parkinson’s disease is levodopa, but long-term application of levodopa can induce complications such as dyskinesias.
OBJECTIVE:To observe the effects of levodopa on learning and memory capacities of Parkinson’s disease rats and to study its mechanisms.
METHODS:The rat models of Parkinson’s disease were established using 6-hydroxydopamine. The 228 model rats were randomly divided into control group and experimental group. Rats in the experimental group were intraperitoneal y injected with 10, 20 and 30 mg/(kg?d) levodopa for 28 consecutive days. At 1, 3, 5, 7, 14 and 28 days after intraperitoneal injection, we observed the rats’ learning and memory capacities and tested plasma concentration of homocysteine and folic acid. Acetylcholinesterase activities in the rat hippocampus were measured. Hippocampal neurofibril ary tangles were observed using Bielschowsky staining.
RESULTS AND CONCLUSION:Increased dose of levodopa and prolonged application time obviously decreased learning and memory capacities in rats (P<0.001), increased plasma homocysteine levels, reduced folic acid levels (P<0.001), diminished acetylcholine esterase activities in the rat hippocampus (P<0.001), and increased neurofibril ary tangles in the rat hippocampus (P=0.000). Results suggested that a large dose of levodopa could significantly decrease the learning and memory capacities, and disease acetylcholine esterase activities, and increase neurofibril ary tangles in hippocampus. Its mechanism possibly associated with the increased plasma concentration of homocysteine.

[ ABSTRACT] AIM:To investigate the effect of DEK downregulation on the apoptosis of gastric carcinoma SGC-7901 cells, and to explore its associations with NF-κB signaling pathway and apoptosis related proteins.METHODS:SGC-7901 cells with different treatments were divided into 3 groups including untreated group, control siRNA group and DEK siRNA group.The expression of DEK at mRNA and protein levels in the SGC-7901 cells was detected by real-time PCR and Western blot.The cell apoptosis was examined by flow cytometry.Furthermore, the activities of caspase-3 and caspase-9 in the SGC-7901 cells were investigated by Caspase-Glo?-3/9 kit.Finally, the expression of key regulatory pro-tein p65 of NF-κB signaling pathway and apoptosis-related proteins Bcl-2 and Bax in the SGC-7901 cells was investigated by Western blot.RESULTS:Compared with untreated group and control siRNA group, the expression of DEK at mRNA and protein levels was significantly downregulated in DEK siRNA group (P<0.05).In addition, the ratios of early phase apoptosis and total apoptosis in DEK siRNA group were markedly higher than those in untreated group and control siRNA group (P<0.05).Most notably, the decrease in p65 and Bcl-2 proteins, increase in Bax protein and the increases of caspase-3 and caspase-9 activities were observed in DEK siRNA group.CONCLUSION:Downregulation of DEK mediates cell apoptosis of gastric carcinoma may be tightly associated with NF-κB signaling pathway.

Objective: To investigate the expressionof proline-rich protein 11 (PRR11) in esophageal carcinoma (EC) tissues and to study it’s effect on the proliferation and metastasis of human EC TE-2 cells in vitro. Methods: Eighty patients were pathologically diagnosed with EC the Department of Thoracic Surgery of the Second Affiliated Hospital of Zhengzhou University from October 2016 to October 2018, and their surgically resected cancer tissues and corresponding para-cancerous tissues were collected for this study. qPCR was used to detect the expression of PRR11 mRNAin tissues or cells. Log-rank Test was used to analyzethe relationship between the expression of PRR11 in EC tissues and general data, histological type, lymphatic metastasis, depth of invasion and TNM stageof the EC patients. Kaplan-Meierplot was used to analyze the association between PRR11 mRNA and patients’prognosis. TE-2 cells were transfected with lentivirus shRNA to construct cell line with PRR11 knockout and corresponding control cell lines, as shPRR11#1, shPRR11#2 and control group. qPCR and WB assays were used to verify the mRNA and protein expressions of PRR11 in cell lines respectively. MTT was used to examine the proliferation of transfected cells, and Transwell experiments were used to detect cell invasion and migration. Results: The expression of PRR11 mRNA in EC was higher than that in para-cancer tissues (P<0.05). There was significant correlation between PRR11 over-expression and histological type, lymphatic metastasis, depth of invasion and TNM stage(all P <0.05), and high PRR11 expression was significantly related with the poor prognosis of EC patients (P<0.05). The mRNA and protein expressions of PRR11 in cells of shPRR11#1 and shPRR11#2 groups were significantly lower than those in control group (all P <0.05). MTT assay showed that the proliferation of cells in shPRR11#1 and shPRR11#2 groups was significantly lower than that in the control group (P<0.05 or P<0.01). The results of Transwell invasion and migration assays showed that the average number of cells with in each field of viewin shPRR11#1 and shPRR11#2 groups was significantly lower than that in the control group (P<0.01). Conclusion: PRR11 is over-expressed in EC tissues and PRR11 over-expression is closely related to the occurrence, progression and prognosis of esophageal cancer. In vitro experiments have also demonstrated that knockdown of PRR11 can inhibit the proliferation, invasion and migration of EC. PRR11 can be used as a potential molecule marker and drug targets for EC. ··

Objective:To explore the effect of quality management circulation (plan-do-check-act, PDCA) mode combined with clinical pathway nursing on leakage rate and nursing satisfaction in patients with ascites due to cirrhosis.Methods:A total of 418 patients with cirrhotic ascites in hospital infectious department were selected and divided into control group and 209 patients in study group according to random digital table method. The leakage rate, quality of life score and total nursing satisfaction were compared between the two groups.Results:The local leakage rates were 17.22% (36/209), 14.83% (31/209), 12.92% (27/209), 8.61% (18/209), respectively, at 24 hours, 48 hours, 72 hours and 1 week after abdominal tube placement. The control group was 22.01% (46/209), 20.10% (42/209), 23.92% (50/209), 25.84% (54/209). The difference between 72 hours and 1 week was statistically significant ( χ2 values were 8.421, 1.746, P<0.01). Quality of life scores were (3.15±1.15), (1.25±1.10), (1.15±1.08), (2.40±1.45), (2.35±1.50) in the study group and (3.75±1.05), (3.25±1.05), (3.60±1.25), (4.35±1.25), (3.15±1.25) in the control group, and the difference was statistically significant ( t values were 5.570-21.441, P<0.05). The total satisfaction of the study group was 96.65% (202/209), which was significantly higher than that of the control group 85.65% (179/209), and the difference was statistically significant ( χ2 value was 15.686, P<0.01). Conclusion:PDCA mode combined with clinical pathway nursing can reduce the leakage rate of abdominal administration, improve the quality of life score and nursing satisfaction, and improve the quality of nursing, which is a safe and effective nursing mode.

Objective:To discuss the effect of downregulating the proline-rich protein 11(PRR11)expression on drug resistance of the esophageal cancer drug resistant cells,and to clarify the related mechanism.Methods:The drug resistant cells EC9706/cisplatin(DDP)were established by incrementally stimulating the human esophageal cancer EC9706 cells with the increasing concentrations of DDP.The drug sensitivity of the EC9706/DDP cells was detected by MTT assay;the expression levels of PRR11 mRNA and protein in the EC9706/DDP cells and their parent EC9706 cells were detected by real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods.The EC9706/DDP cells were divided into control group,sh-NC group(infected with sh-NC),sh-PRR11 group(infected with sh-PRR11),sh-NC+DDP group(infected with sh-NC and treated with 4 mg·L-1 DDP),and sh-PRR11+DDP group(infected with sh-PRR11 and treated with 4 mg·L-1 DDP).The expression levels of PRR11 mRNA in the cells in various groups were detected by RT-qPCR method;the expression levels of PRR11,phosphoinositide 3-kinase(PI3K)p110α,protein kinase B(AKT),phosphorylated AKT(p-AKT),P-glycoprotein(P-gp),and multidrug resistance-associated protein 1(MRP1)proteins in the cells in various groups were detected by Western blotting method;the apoptotic rates of the cells in various groups were detected by flow cytometry.Results:The DDP-resistant cell line EC9706/DDP was successfully obtained,and the drug resistance index was 7.23±0.86.Compared with the EC9706 cells,the expression levels of PRR11 mRNA and protein in the EC9706/DDP cells were increased(P<0.05).Compared with control and sh-NC groups,the expression levels of PRR11 mRNA and protein in the cells in sh-PRR11 group were decreased(P<0.05),and the 50%inhibitory concentration(IC50)of DDP was decreased(P<0.05).Compared with sh-NC group,the expression levels of PI3K p110α,p-AKT,P-gp,and MRP1 proteins in the cells in sh-NC+DDP and sh-PRR11 groups were decreased(P<0.05),and the apoptotic rate of the cells was increased(P<0.05).Compared with sh-NC+DDP group and sh-PRR11 group,the expression levels of PI3K p110α,p-AKT,P-gp,and MRP1 proteins in the cells in sh-PRR11+ DDP group were increased(P<0.05),and the apoptotic rate of the cells was increased(P<0.05).Conclusion:Downregulating the expression of PRR11 gene in the drug resistant EC9706/DDP cells can inhibit the expressions of drug resistance-related proteins,reverse the resistance to DDP,and induce the apoptosis;its mechanism may be related to the inhibition of activation of the PI3K/AKT signaling pathway.

Objective:To study the difference in the extraction efficiency of the novel coronavirus (2019-nCoV) nucleic acid by using magnetic beads method, centrifugal column method and one-step method.Methods:On March 5, 2021, 10 throat swabs were collected from the staff working in the nucleic acid sampling room in Department of Clinical Laboratory, Affiliated Taikang Xianlin Drum Tower Hospital, Medical School of Nanjing University. The positive quality control samples were mixed into the swabs and used as mock positive samples. The RNA was extracted from simulated positive samples and their diluted samples by using magnetic beads method, centrifugation column method and one-step method. The purity ( A260/ A280 ratio) and concentration of the nucleic acid obtained were measured by micro-uv photometry, and fluorescence quantitative PCR was performed to compare the CT value and extraction efficiency. The three methods were used to extract the simulated weak positive specimens and to compare the difference of CT values after amplification. The measurement data that followed normal distribution were expressed by xˉ±s, the t test was used for comparing in the same group, and single factor analysis of variance was used for comparing among multiple groups. A P value smaller than 0.05 indicated a significant difference. Results:2019-nCoV nucleic acid extracted by magnetic bead method, centrifugal column method and one-step method could amplify positive results. There was no significant difference between the CT value of RNA amplification extracted by magnetic bead method and one-step method ( t=? 0.995 , P=0.376). The CT values of orf1ab gene amplified by centrifugal column method, magnetic bead method and one-step method were 29.28±0.06, 30.82±0.14 and 29.79±0.01 respectively ( F=11.196 , P=0.041). The CT values of E gene were 28.52±0.40, 27.33±0.78 and 27.38±0.13 respectively ( F=3.407, P=0.169). The CT values of N gene were 28.61±1.02, 27.24±0.20 and 27.25±0.47, respectively ( F=2.880 , P=0.020). The CT values of human genes extracted by centrifugal column method, magnetic bead method and one-step method were 19.68±0.36, 20.14±0.06 and 20.58±0.49 respectively, which was statistically significant ( F=4.904, P=0.048). The CT value of amplified human gene was affected by the dilution of human samples twice. The CT value of undiluted samples was smaller than that of diluted samples twice, with a difference of 2.95±0.22, which was statistically significant ( t=?3.025, P=0.039). The extraction time of one-step method, magnetic bead method and centrifugal column method were (15.00±1.50), (20.00±1.50) and (40.00±5.5) min respectively, and the difference was statistically significant ( F=688 , P=0.027). Conclusions:Magnetic bead method, centrifugal column method and one-step method can be used to extract 2019-nCoV nucleic acid, for the centrifugal column method has a higher extraction efficiency than the magnetic bead method and the one-step method. The one-step method is the fastest, followed by the magnetic bead method and the centrifugal column method. A large number of clinical samples can be processed using the magnetic bead method and one-step method. One-step rapid nucleic acid test can also be performed on samples from emergency and fever clinics. It is not recommended to dilute specimens for testing. In order to improve the detection rate, extracting RNA from highly suspected samples with negative initial nucleic acid test by centrifugal column method is suggested.

Objective:To develop a novel, flexible, dual-arm, master-slave digestive endoscopic minimally invasive surgical robot system named dual-arm robotic endoscopic assistant for minimally invasive surgery (DREAMS) and to evaluate its feasibility for endoscopic submucosal dissection (ESD) by using ex vivo porcine stomachs.Methods:A novel endoscopic robot (DREAMS) system was developed which was composed of a flexible two-channel endoscope, two flexible robotic manipulators, a master controller, a robotic arm, and a control system. A total of 10 artificial round-like lesions with diameters ranging from 15 to 25 mm were created (5 in gastric antrum and 5 in gastric body) by using fresh peeled stomach of healthy pigs as the model. Submucosal dissection was performed with the assistance of the DREAMS system by two operators. The main outcome was submucosal dissection speed, and the secondary outcomes included muscular injury rate, perforation rate, and grasping efficiency of the robot.Results:All 10 lesions were successfully dissected en bloc by using the DREAMS system. The diameter of the artificial lesions was 22.34±2.39 mm, dissection time was 15.00±8.90 min, submucosal dissection speed was 141.79±79.12 mm 2/min, and the number of tractions required by each ESD was 4.2 times. Muscular injury occurred in 4/10 cases of ESD. No perforation occurred. Conclusion:The initial animal experiment shows the DREAMS system is safe and effective.