Xenopus Paraxial Protocadherin (PAPC), which was initially identified in a screen for genes present in the Spemann organizer of Xenopus embryos, is required for gastrulation, somitogenesis and otic vesicle formation. In order to investigate its function in various developmental events, an antibody was prepared which could specifically recognize Xenopus PAPC. Glutathione S transferase (GST) expression system was used to express the fusion protein GST-PAPC. Rabbits were immunized with GST-PAPC Western blotting analysis of FL-PAPC transfected HEK 293T cells lysates, which could be specifically blocked by pre-adsorption of prokaryotic expressed GST-PAPC fusion protein. Furthermore, by using immunofluorescence analysis the polyclonal antibody recognized membrane-bound PAPC in FL-PAPC transfected 293T cells and Xenopus animal cap cells. By Western blotting analysis,the endogenous 150 ku PAPC protein was detected in Xenopus embryos using the anti-PAPC antibody. Take together it could be concluded that a polyclonal antibody specifically against Xenopus PAPC was developed.

The association of the serum phospholipid fatty acid components with lipid profiles in patients with newly-diagnosed type 2 diabetes mellitus was investigated.Saturated fatty acid in patients with newly diagnosed type 2 diabetes mellitus was increased(48.79±1.55 vs 42.58±1.96,P<0.01),whereas monounsaturated fatty acid and polyunsaturated fatty acid were decreased(10.72±1.53 vs 12.09±1.32,33.41±2. 16 vs 35.79±2.41.6.08±1.66 vs 9.54±1.54,all P

AIM: To investigate the heterogeneity of basal intracellular free calcium concentration( i) in peritoneal macrophages(PM) and whether it is relative to the reactivity of PM at the single cell level. METHODS: i implicated stimulated were measured by fluorescent microscopic imaging system after loading with fluorescent probe fura-2/AM. Superoxide(O _2)produced by single PM was determined by modified NBT test. RESULTS: The values of basal i determined in 392 PMs of 7 mice showed normal distribution [(54?24) nmol/L, n=392] with wide range(less than 20 nmol/L to more than 100 nmol/L), among which about 50% were in the range of 40-60 nmol/L. When stimulated with PMA or fMLP, i was increased, the peak values were positively correlated with the basal i in one mouse(PMA stimulated cells: r=0.52, P

Objective To investigate the differences of miRNA-21 and miRNA-146a expression between colorectal neoplasms tissues and serum of the patients with colorectal neoplasm,and the clinical significance was analyzed.Methods The endoscopic biopsy tissues and serum samples of 100 colorectal cancer (CRC),80 colorectal adenoma (CRA)patients and 65 healthy controls were collected.The expressions of miRNA-21 and miRNA-146a were detected by quantitative real time polymerase chain reaction.The relationship between the expression and clinicopathological features were analyzed.And then,the diagnostic value of expression difference of serum miRNA in colorectal neoplasm were evaluated. The Mann-WhitneyU test and Kruskal-Wallis test were performed for comparisons between groups,and the correlation of miRNA expression between tissues and serum was analyzed by Spearman test.Results
The expressions of miRNA-21 in the tissues of CRC and CRA were 8.573 ±0.898 and 7.746 ±1 .183, respectively,which were significantly higher than that of healthy controls 6.160 ±0.835 (U =120.129 and 33.230,both P <0.01).The expressions of miRNA-21 in the serum of CRC and CRA were 1 .829± 0.303 and 1 .624 ±0.226,respeotively,which were higher than that of healthy controls 1 .391 ±0.221 (U =40.353 and 15 .512,both P < 0.01 ).The expression of miRNA-21 in the serum and tissues of patients with CRC were both higher than those of patients with CRA (U =11 .384 and 10.189,both P <0.01).The expression of miRNA-21 in patients with CRC was associated with TNM stage and lymph node metastasis.The expression level of miRNA-21 in CRA was correlated with histological types.The results of Spearman analysis indicated that the expression of miRNA-21 in the tissues and serum of patients with CRC was positively correlated (r=0.459,P <0.01).The expression of miRNA-146a in the tissues and serum of CRC patients were 2.556±0.351 and 0.249±0.038,respectively,which were lower than those of healthy controls (3.428 ±0.328 and 0.279 ±0.053)(U =102.134 and 30.111 ,both P <0.01).The expression in the tissues and serum of CRA patients were 3.255 ±0.332 and 0.290±0.036, respectively,which were also lower than those of healthy controls,however the difference was not statistically significant (U = 3.936 and 3.180,both P > 0.05 ).The expression of miRNA-146a in the tissues and serum of CRC patients were both lower than those of CRA patients (U =73.809 and 21 .123, both P <0.01).The degree of decreased expression of miRNA-146a in CRC patients was correlated with TNM staging and tumor differentiation degree,however there was no correlation between the expression in CRA and clinical features.According to receiver operating characteristic (ROC)curve analysis,the AUC of miRNA-21 ,miRNA-146a and a combination of them in CRC and health individuals was 0.889, 0.791 and 0.863,respectively;in CRA and health individuals was 0.784,0.692 and 0.761 ,respectively;in CRC and CRA was 0.705 ,0.820 and 0.713,respectively.Conclusion The different expressions of miRNA-21 and miRNA-146a had potential values in early detection of colorectal cancer.

Objective:To evaluate the diagnostic performance of probe-based confocal laser endomicroscopy (pCLE) for indeterminate biliary strictures.Methods:Twelve patients with indeterminate biliary strictures who underwent pCLE and brush cytology from April 1, 2013 to December 30, 2016 were enrolled. Clinical data, the results of endoscopic retrograde cholangiopancreatography, pCLE examination and brush cytology were collected. Compared with post-operative pathology and follow-up over 12 months, sensitivity, specificity, positive predictive value(PPV), negative predictive value(NPV), and accuracy of pCLE and brush cytology of the diagnosis of malignant biliary strictures were analyzed.Results:The final diagnosis were 9 malignant and 3 benign. The sensitivity, specificity, PPV, NPV and accuracy of brush cytology were 3/9, 3/3, 3/3, 3/9 and 50.0%(6/12), respectively. The corresponding indicators of pCLE were 9/9, 2/3, 9/10, 2/2, and 91.7%(11/12), respectively.Conclusion:pCLE can be used for differential diagnosis of indeterminate biliary stricture.

Objective To analyze the clinical application of HLA donor specific antibodies (DSAs) detected by Luminex single antigen beads,and to discuss the impact of early intervention on renal function.Method In 64 cases of living-relative renal transplantation,DSA was detected using a Luminex single antigen assay before and after transplantation.The positive recipients were given large doses of intravenous irnmunoglobulin (IVIG) and increased doses of mycophenolate mofetil (MMF).The relationship between DSA and renal function was analyzed.Result DSA was negative in all recipients before transplantation.Ten cases of DSA positive recipients were found in HLA mismatch after transplantation.After the intervention,two cases of DSA positive recipients became negative,immunofluorescence intensity was decreased by more than 50％ in 6 cases,and no significant reduction was found in the other two cases.Antibody-mediated rejection (AMR) occurred in two cases of intervention ineffective recipients after 3 to 6 months and the renal function was impaired.Conclusion Dynamic monitoring of DSA using Luminex single antigen beads may timely predict changes of renal function.Early application of large doses of IVIG and increasing doses of MMF can reduce the incidence of AMR.

Objective To analyze the distribution and drug resistance of pathogens isolated from blood culture specimens in a grade 3A hospital so as to provide a basis for the prevention and control of bloodstream infection ,and rational antibacterial drugs use .Methods A total of 3 241 blood culture results in our center from June 2014 to September 2016 were analyzed retrospectively . The blood samples were cultivated with the BacT/ALERT 3D and VersaTrek instruments .The bacterial identification and antibiotic susceptibility tests were conducted with VITEK‐2 Compact and ARIS 2X instruments .Results Among 3 241 blood culture speci‐mens ,99 strains of pathogenic bateria were isolated with the positive rate of 3 .1% ,including 42(42 .4% ) strains of Gram‐negative bacteria ,54(54 .5% ) strains of Gram‐positive bacteria and 3(3 .0% ) strains of fungus .The top three of Gram‐negative bacteria were in turn Eescherichia coli ,Klebsiella pneumonia and Pseudomonas aeruginosa ;the top three of Gram‐positive bacteria were in turn coagulase‐negative staphylococcus (CNS ) ,Staphylococcus aureus and streptococcus pyogenes .Enterobacteriaceae remained 100 .0% sensitivity to imipenem ,meropenem ,piperacillin/tazobactam and amikacin .The resistance rate of Gram‐positive bacterium to vancomycin ,linezolid ,teicoplanin and gentamycin was 0 .0% ,the rest had different degrees of drug resistance .Conclusion Ee‐scherichia coli ,CNS and Staphylococcus aureus are the most frequent pathogenic bacteria of the blood stream infection in our hospi‐tal .Pathogens show different resistance to different kinds of antibacterial agents .Clinic should rationally select the antibacterial a‐gents according to the drug susceptibility test results so as to slow down the generation of drug resistance bacterial strains .

Objective To develop a novel skin microdialysis technology in vivo,and to determine the pharmacokinetic of ba-icalin after transdermal administration in rats. Methods An HPLC-MS/MS method used for the determination of baicalin in skin mi-crodialysis samples was established ,SD rats were pretreated with skin microdialysis operation under anesthesia , and then the baicalin gel was applied to the skin surface of probe in vivo.The baicalin concentration of skin microdialysates was determined , the time curve of baicalin concentration was drawn and the topical pharmacokinetics parameters of percutaneous absorption was calculated. Results Baicalin was optimized at the transitions m/z 447.3→271.2.The linearity correlation was good and the assay exhibited good precision and accuracy.The subcutaneous probe recovery of baicalin in vivo was(24.40 ±0.91)%and was stable over the 240 min study peri-od.Baicalin could be detected in the microdialysis samples after transdermal administration , and its concentration continued to rise in 8 h.AUC0-t in skin tissue was(50.04 ±34.17) mg· min· L-1. Conclusion The method of skin microdialysis in vivo could be used in the local pharmacokinetic research of baicalin.

Objective In the present study, we investigated the effects of advanced oxidation protein products(AOPP) on reactive oxygen species(ROS) production in murine osteoblastic MC3T3-E1 cells by NADPH oxidase enzymes pathway. Methods Experiments were divided into three groups, including control group, rats albumin(RSA) group, and AOPP group. Different concentrations of AOPP were added to the osteoblastic MC3T3-E1 cells culture medium. The production of ROS in MC3T3-E1 cells was measured by the fluorescence intensity of intracellular fluoroprobe ( DCFD ) . In order to verify the effect of enzyme of the production of ROS, the specific inhibitors of corresponding enzymes were added in the MC3T3-E1 cells which were cultured in the medium with AOPP. Finally, western blot and immunofluorescence were used to observe the changes of NADPH oxidase enzymes subunits. Results Different concentrations of AOPP (50,100,200μg/ml) induced MC3T3-E1 cells to produce different amount of ROS. The higher concentrations of AOPP were added, the more ROS were produced. Furthermore,200μg/ml AOPP induced the maximum amount of ROS production(P<0. 05). Meanwhile, AOPP induced MC3T3-E1 cells to produce different amount of ROS with a time-dependent manner. The peak amount of ROS production in MC3T3-E1 cells was observed in 3h when AOPP were added (P<0. 05). In addition, when specific inhibitors of corresponding enzymes were added in the MC3T3-E1 cells, the production of ROS were significantly suppressed by C-SOD, DPI, and apocynin(P<0. 05). On the other hand, AOPP can up-regulate the expression of Nox4 protein of the MC3T3-E1 cells, which is one of the subunits of NADPH oxidase enzymes. Meanwhile, AOPP can also induce the membrane migration of p47phox subunit. Conclusion AOPP induces osteoblastic MC3T3-E1 cells to produce ROS by NADPH oxidase enzymes pathway, and which may be one of the pathogenesis of AOPP involved in osteoporosis.

Objective To clone and identify the differentially expressed genes of rat intestinal epithelial cell line (IEC 6) before and after exposure to high dose radiation so as to provide proof for the investigation of the molecular mechanisms in the repair of radiation damage of intestinal epithelial cells. Methods A subtractive cDNA library for differentially expressed genes was constructed by suppression subtractive hybridization (SSH) and T/A cloning technique after IEC 6 cells were exposed to radiation at the dose of 35 Gy ? ray. The expressed sequence tag (EST) library was screened by reverse Northern hybridization. Positive clones were sequenced and the similarity was searched against the DNA database in GenBank. Limited clones were identified by Northern hybridization. Results More than 2 000 white clones were harvested after the library amplification. Ninety six of them were randomly picked out for PCR amplification, and 15 positive clones which corresponded to 12 individual genes were identified by reverse Northern hybridization. These genes were involved in cell skeleton, cell stress, cell cycle control, and signal transduction, etc. In addition, a novel cDNA sequence was also obtained. Conclusion A subtractive cDNA library for differentially expressed genes in IEC 6 cells exposed to the radiation at the dose of 35 Gy ? ray has been successfully constructed with SSH and T/A clone techniques. Several positive ESTs which correspond to genes involving in cell skeleton, cell stress, cell cycle control, and signal transduction are identified. These genes may play important roles in the process of the damage and repair of the intestinal epithelial cells exposed to radiation.