Expression of miR-4695-5p in the serum of colorectal cancer patients and the mechanism of its effect on the proliferation and invasion of colorectal cancer cells / 国际外科学杂志

Objective:To explore the expression of microRNA-4695-5p in the serum of colorectal cancer patients and its effect on the proliferation and invasion of colorectal cancer CACO-2 cells.Methods:A total of 43 serum samples of colorectal cancer patients who were admitted to the Department of Gastrointestinal Surgery, Luoyang Central Hospital Affiliated to Zhengzhou University from March 2018 to November 2021 were selected, and serum samples of 43 healthy people who underwent outpatient physical examination were used as controls. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expression levels in the serum of colorectal cancer patients and those of healthy individuals. The miR-4695-5p overexpression plasmid or the negative control plasmid were transfected into CACO-2 cells, namely the miR-4695-5p group and the control group, and the transfection efficiency was verified by qRT-PCR. CCK8 method and Transwell experiment were used to detect the effect of overexpression of miR-4695-5p on the proliferation and invasion of CACO-2 cells. The dual luciferase reporter gene experiment was used to verify the targeted binding relationship between miR-4695-5p and Ras-related C3 botulinum toxin substrate 1 (RAC1). qRT-PCR and Western blot were used to detect the effect of overexpression of miR-4695-5p on the expression of RAC1 gene and Wnt/β-Catenin signaling pathway protein.The software of SPSS28.0 was used to conduct data analysis. The measurement data of normal distribution were espressed by Mean±SD. The t-test was used to compare the means between two groups, and the one-way analysis of variance was used to compare the means of multiple groups. Results:The expression level of miR-4695-5p in the serum of patients with colorectal cancer was significantly lower than that of healthy individuals ( P<0.01). The relative expression levels of miR-4695-5p in the control group and miR-4695-5p group were 1.09 ± 0.65 and 8.83±2.03, respectively. The expression level of miR-4695-5p in CACO-2 cells in the miR-4695-5p group was 8.10 times that of the control group, and CACO-2 cells overexpressing miR-4695-5p were successfully constructed ( P<0.01). Compared with the control group, the proliferation ability of CACO-2 cells in the miR-4695-5p group was significantly reduced ( P<0.05), and the invasion ability of CACO-2 cells was significantly reduced ( P<0.01). Bioinformatics tools and dual luciferase reporter gene experiments confirmed that miR-4695-5p can target and bind RAC1 ( P<0.01). Compared with the control group, the expression of RAC1 gene in the miR-4695-5p group was significantly decreased ( P<0.01), and the expression of Wnt/β-Catenin signaling pathway proteins Wnt3a, β-catenin, and c-MYC decreased significantly. Conclusions:miR-4695-5p is lowly expressed in the serum of colorectal cancer patients. miR-4695-5p inhibits the proliferation and invasion of colorectal cancer CACO-2 cells by targeting the inhibition of RAC1 gene expression and down-regulating the activity of the Wnt/β-Catenin signaling pathway.

Application of grey model and ARIM multiple seasonal model in the prediction of brucellosis in Chengde of Heibei Province / 中华地方病学杂志

Objective The grey model [GM (1,1)] and the ARIMA multiple seasonal model were used to predict the incidence trend of brucellosis in Chengde,and the effects of predictions of the two models is compared.Methods According to the statistical results of epidemiological research results of monthly brucellosis patients in Chengde from 2008 to 2014,we established the ARIMA multiple seasonal model and GM (1,1) model,individually predicted the trend of brucellosis in 2015.Compared with the actual monitoring results,the average relative error was used to verify the reliability of the model.Results GM (1,1) model was (X)0(k) =0.001 6 (X)0 + 23 712.31) exp [0.001 6 (k-t0)],and the optimal model for determining the ARIMA multiple seasonal model of product was ARIMA (0,1,1) × (0,1,0)12.The average relative errors of the two models predicted in 2015 from January to December compared with the actual monitoring data were 114.82％ and 11.66％.Conclusion The product ARIMA multiple seasonal model has better predictive effect on brucellosis,and can be used for short-term forecasting.

Construction of a Single Chain Variable Fragment Antibody Against Sulfonamides and Its Interaction with Sulfathiazole / 分析化学

The interaction between the antibody and the corresponding target molecule determines the characteristics of immunoassay. In this study, a single chain variable fragment antibody (scFv4C7) derived from the hybridoma strain 4C7 were prepared via genetic engineering technique. The recognition properties of scFv4 C7 was determined and compared to those of the parent monoclonal antibody by indirect competitive enzyme-linked immunosorbent assay(ic-ELISA). Three dimensional structure of the scFv4C7 was presented by Swiss-Model, and sulfathiazole ( STZ) was docked to the scFv4C7 model to obtain the structure of the binding complex. The results from the ic-ELISA showed that the binding properties of scFv4C7 were comparable with the parent monoclonal antibody and STZ was almost completely buried in a deep binding pocket formed by the heavy chain and light chain of the antibody. The interaction between STZ and scFv4C7 was more closely related to the heavy chain and the complementarity-determining region ( CDR ) H3 loop played more important role than other CDR loops. The study preliminary provided the necessary structural information for the preparation of antibody with broader specificity and higher affinity.

Performance verification of portable Celercare M1 analyzer in CRP determination / 国际检验医学杂志

Objective To verify the analytical performance of the portable Celercare M1 analyzer in the C-reactive protein(CRP) quantitative determination.Methods According to the standard program of the American Clinical and Laboratory Standards Insti-tute (CLSI),The portable Celercare M1 analyzer was performed the verification on the CRP detection in the aspects of the linearity range,precision,accuracy,interference test and method comparison.Results The CRP detection showed good linearity in the range of 0.5-200 mg/L and the regression equation was Y =0.979X +0.456.The within-run coefficients of variation(CV)of samples with low and high CRP levels were 5.9% and 2.0%,respectively.The between-day CV of samples with low and high CRP levels were 6.3% and 2.2%,respectively.The accuracy of the assay was admirable,and bias of CRP results at low and high levels were 2.3% and 0.7%,respectively.No significance interference was observed when serum bilirubin ≤340 μmol/L,triglyceride ≤10 mmol/L and hemoglobin ≤8 g/L in this method.The regression equation of Celercare M1 analyzer(Y )and the IMMAGE 800 (X) for detecting CRP was Y =0.944X +0.206,R 2 =0.996(P <0.05).The relative bias of CRP results in three medical decision levels (3,10,100 mg/L)was 1 .0%,4.0% and 5.0%,repectively,which was lower than 1/2 of total error allowance.Conclusion The linearity,precision,accuracy and interference of the Celercare M1 analyzer in CRP determination are admirable and can be accepted in clinical practice.

Comparative analysis of common biochemical assay results for venous blood and peripheral blood / 国际检验医学杂志

Objective To investigate the comparability of the results of common biochemical items detected by Celercare M1 an‐alyzer in the peripheral blood and the venous whole blood .Methods The samples of peripheral blood and venous whole blood were collected from subjects .The biochemical items including Mg2+ ,Cl- ,tCO2 ,K+ ,Na+ ,Ca2+ ,α‐HBDH ,LDH ,AST ,CK ,CK‐MB ,TP , ALB ,TBIL ,ALT ,GGT ,ALP ,UREA ,GLU ,UA ,CHOL ,and HDL‐C were determined by Celercare M1 analyzer ,and the results were compared .Results The tCO2 results of venous blood was significantly higher than that of peripheral blood (P<0 .05) .How‐ever ,the results of α‐HBDH ,LDH ,CK and CK‐MB of venous blood samples were significantly lower than those of peripheral blood samples ,and the difference was statistically significant (P<0 .05) .Conclusion The peripheral blood can replace venous blood for biochemical analysis on Celercare M1 analyzer ,except for the electrolyte test items and cardiac enzyme items such as α‐HBDH , LDH ,CK and CK‐MB .

Hepatitis B virus replication and viral gene expressions do not affect CDC37 level in hepatocytes in vitro / 南方医科大学学报

<p><b>OBJECTIVE</b>To study the influence of hepatitis B virus (HBV) replication and expressions of different viral genes on CDC37 level in hepatocytes.</p><p><b>METHODS</b>We amplified and cloned 6 HBV genes (P, preS1, preS2, S, C and X) into pCMV expression vectors, which were transfected in Huh7 and HepG2 hepatoma cell lines, and CDC37 expression level in the cells was detected using Western blotting. Wealso cloned the promoter sequence of CDC37 into pGL3 vector, and co-transfected pGL3 with pCMV recombinant plasmids into Huh7 and HepG2 cells and the fluorescent signals were detected. To study the influence of HBV replication on CDC37 expression, we constructed 1.28-copy overlength genomes of HBV genotypes B, C, D and CD recombinant. The overlength HBV genomes were transformed into Adeasier-1 cells for recombination and into 293 cells for packaging. Huh7 and HepG2 cell lines infected with the packaged HBV recombinant adenoviruses were examined for CDC37 expression with Western blotting.</p><p><b>RESULTS</b>Western blotting showed that the expression of different HBV genes did not obviously affect the protein level of CDC37 in the hepatocytes. The protein expression of HBV genes had no effect on the activity of CDC37 promoter. Huh7 and HepG2 cells infected with 1.28-copy HBV replicon showed no significant changes in the expression level of CDC37.</p><p><b>CONCLUSION</b>HBV replication and its gene expression have no effect on the level of CDC37 in hepatocytes in vitro.</p>

Construction of plasmids expressing Sars-CoV encoding proteins and their effects on transcription of hfgl2 prothrombinase / 华中科技大学学报（医学）（英德文版）

SARS coronavirus (SARS-CoV) is the etiologic agent of severe acute respiratory syndrome. The aim of this study was to construct Sars-CoV membrane (M), nucleocapsid (N) and spike 2 (S2) gene eukaryotic expression plasmids, and identify their expression in vitro. Gene fragments encoding N protein, M protein and S2 protein of SARS-CoV were amplified by PCR using cDNA obtained from lung samples of SARS patients as template, and subcloned into pcDNA3.1 vector to form eukaryotic expression plasmids. SARS-CoV protein eukaryotic expression plasmids were transfected respectively into CHO cells. Immunohistochemistry was employed to detect the expression of the structural proteins of SARS-CoV in transfected cells. SARS-CoV protein eukaryotic expression plasmids were successfully constructed by identification with digestion of restriction enzymes and sequencing. M, N and S2 proteins of SARS-CoV were detected in the cytoplasm of transfected CHO cells. It was concluded that these recombinant eukaryotic expression plasmids were constructed successfully, and SARS-CoV encoding proteins could activate transcription and expression of hfgl2 gene.

Identification of two newly discovered subgenotypes of hepatitis B virus genotype C / 中华传染病杂志

Objective To analyze the genotypic characteristics of hepatitis B virus (HBV)C/D recombinant of two types of newly discovered HBV genotypea found in Western China.Methods The whole genomes of 17 HBV strains isolated from Western China were amplified by polymerase chain reaction (PCR). Bioinformatic softwares were used for the analysis of full genome structure,genetic distances and recombination points.Results The heterogenicity of the HBV C/D recombinant was more than 8% compared with genotype A,B,D,E or F,but 3.8% 0A-8.O% compared with genotype C Based on phylogenetie analysis, a11 C/D recombinant strains clustered within genotype C.but were rouped into two other clusters within the genotype C independently from C1-C5 subgenotypes,which were two kinds of new HBV/C genotypea.Condusion The HBV C/D recombinant could he considered as tWO kinds of new subgenotypea of HBV genotype C which are different from subgenotype C1-C5 based on the genetic distances analysis.

Treatment of Stye by Auricular Point Blood-letting Therapy: A Report of 85 Cases / 针灸推拿医学（英文版）

Eighty-five cases of stye were treated by pricking and extruding some blood in auricular points Eye 1 and Eye 2. After 5 treatments, 63 cases were cured, 17 cases obtained improvement, 5 cases obtained no effects, and the total effective rate is 94.1%.

YMDD mutations of HBV in HBeAg positive and HBeAg negative chronic hepatitis B / 中华传染病杂志

Objective To compare the occurrence of YMDD mutations in patients with HBeAg positve and HBeAg negative chronic hepatitis. Methods Two hundred and forty seven chronic hepatitis B patients who received lamivudine treatment were followed up.Liver function tests and hepatitis B virological tests including YMDD mutant analysis,HBV DNA quantitation and HBV serological marker analysis were performed regularly. Results YMDD mutants were detected in forty two patients.The cumulative rate of occurrence of YMDD mutation increased with the duration of lamivudine treatment.The mutation rate was significantly higher in pretreatment HBeAg positive patients than that in HBeAg negative patients.The ALT flare was more evident after emergence of YMDD mutants in pretreatment HBeAg positive patients than that in HBeAg negative patients.Conclusions The cumulative rate of occurrence of YMDD mutation was higher in pretreatment HBeAg positive patients than that in pretreatment HBeAg negative patients.

Endoscopic band ligation without cauterization: a good technique for excision of upper gastrointestinal small leiomyomas / 中华消化内镜杂志

Objective To evaluate the method of endoscopic band ligation in resecting upper gastrointestinal leiomyoma . Methods We selected 59 patients with 64 small upper gastrointestinal tract leiomyo-mas by endoscopy , EUS and EUS guided fine needle aspiration. Of the 64 leiomyomas, their distribution were in esophagus 50, stomach 12 and duodenum 2. When the endoscope with transparent cap was introduced to reach the lesion, it was sucked into the cap, then released the band to ligate it. Two weeks after operation, endoscopic monitoring of the lesion was performed weekly until the complete healing of the wound. Results In the 64 lesions, 50 esophageal and 2 duodenal leiomyomas were thoroughly resected and the mean concrescence time is 3. 1 weeks and 4. 5 weeks respectively. Nine of 12 gastric leiomyomas were resected thoroughly and the other 3 were partially ligated and resulted in incomplete excision with the mean concrescence time of 4. 5weeks. No perforation occurred. Conclusions Endoscopic band ligation is an ideal method in resecting the small upper gastrointestinal leiomyomas.

Differential diagnosis of tumors in upper GI muscularis propria by EUS guided fine needle aspiration and immunohistochemical evaluation / 中华消化内镜杂志

Objective It is difficult to differentiate tumors in upper GI muscularis proporia, such as GI stromal tumors (GISTs) , leiomyomas and schwannomas. We performed EUS guided fine needle aspirations (FNA) and immunohistochemical evaluation of these tumors in upper GI muscularis proporia. Methods We selected 35 Patients with lesions in upper GI muscularis proporia by EGD and EUS. We assessed the shape , size, and position of the lesions and the status of metastasis. After excluding blood vessel between the lesions and the fine needle, EUS FNA were carried out and immunohistochemical staining for CD-117 (c-kit) , CD34 and smooth muscle actin ( SMA) was performed. Results All patients were performed EUS FNA and enough specimens were obtained in 31 patients. The other 4 failed, although we tried it for several times. Among the 31 patients, Cytology and immunohistochemisty demonstrated 21 GISTs and 10 leiomyomas. All the results were evaluated by surgery. The sensitivity is 88. 6% , and the speciality is 100%. No complication occurred. Conclusion EUS-guided FNA and immunohistochemical evaluation is an accurate method for diferential diagnosis of tumors in upper GI muscularis propria.

Study on the cloning, expression and identification of the renal cell carcinoma-associated antigen G250/MN/CA Ⅸ / 中华泌尿外科杂志

Objective To report the cloning, expression and identification of the tumor-associated antigen G250/MN/CA Ⅸ. Methods The total RNA was extracted from renal cell carcinoma tissue samples from 54 male patients. Gene fragments encoding G250 was obtained by RT-PCR,and was cloned into prokaryotic expression vector pET22b(+) and expressed in E.coli BL21(DE3);and the results were examined by SDS-PAGE gel electrophoresis. The recombinant protein was studied by Western blot test. Results DNA sequence analysis showed that the obtained sequence was the same as that showed in GenBank.Gene of G250 was expressed in E.coli BL21 successfully. Western blot analysis showed that the recombinant protein could be specially recognized by monoclonal antibody. It had better antigenicity and specificity. Conclusions This study provides experimental basis for the purification of G250/MN/CA Ⅸ protein and the further study of G250/MN/CA Ⅸ function and preparation of antibody.