Objective To establish HPLC fingerprints of soaking Euodiae fructus with water decoction of Radix glycyrrhizae by Zhangbang method(hereinafter referred to as"soaking Euodiae fructus"),and to determine the content of evodiamine,rutaecarpine and evocarpine,so as to provide the basis for the quality control and standard improvement of soaking Euodiae fructus.Methods The fingerprint of soaking Euodiae fructus was established based on wavelength switching technology,and the similarity evaluation was conducted.Taking the rutaecarpine as the internal reference,the relative correction factors of evodiamine and evocarpine were calculated by slope correction method,and the differences in measurement results between quantitative analysis of multi-components by single-marker(QAMS)method and external standard method were compared.Results The established fingerprint of soaking Euodiae fructus had a total of 20 common peaks,the similarity between it and the control fingerprint spectrum was 0.970 to 0.998,and 7 common peaks of them were identified.The contents of evodiamine and evocarpine determined by QAMS method were 1.754-7.542 mg/g and 1.281-2.455 mg/g in 10 batches of samples,and the results obtained by QAMS method and external standard method were similar.Conclusion The established HPLC method is simple,reliable,with good separation and repeatability,and can be used for quality evaluation of processed Euodiae fructus.

OBJECTIVE To investigate the effect of vitamin D receptor(VDR) genetic polymorphism on the concentration of tacrolimus in renal transplant recipients at early stage after transplantation.METHODS The 360 cases of renal transplant recipients who received tacrolimus, mycophenolic acid, and glucocorticoid were recruited. CYP3A5(rs776746) and VDR(VDR ApaI rs7975232, VDR BsmI rs1544410, VDR FokI rs2228570 and VDR TaqI rs731236) genotypes were determined. The differences of concentration(C), dose(D) and the ratio of concentration to dose(C/D) of tacrolimus were compared among all of the genotype groups at the seventh day after renal transplantation. RESULTS The C and C/D of tacrolimus in CYP3A5 non-expresser(GG genotype) were all significantly higher than CYP3A5 expresser(AA and AG genotype)(P<0.05). When taking the different CYP3A5 genotypes in consideration, it was found that the C/D in patients with VDR ApaI rs7975232 AA genotype was significantly lower than those with AC and CC genotypes for CYP3A5 non-expresser(P<0.05). However, VDR ApaI rs7975232 gene polymorphism had no influence on C and C/D of tacrolimus in CYP3A5 expresser. Besides, no matter in CYP3A5 expresser or in non-expresser, VDR BsmI rs1544410, VDR TaqI rs731236 and VDR FokI rs2228570 had no effect on C, D and C/D of tacrolimus. CONCLUSION During the early stage of renal transplantation, the polymorphism of VDR ApaI rs7975232 show significant relevance with tacrolimus concentration in CYP3A5 non-expresser. The detection of the genotype might be helpful to guide individual therapy.

Objective: To explore the effect of high mobility group box-1 protein (HMGB1) on the balance of Th17/Treg in patients with immune thrombocytopenia (ITP). Methods: A total of 30 patients who were first diagnosed as ITP in the Fifth People's Hospital of Datong from July 2017 to April 2018 were selected as the case group, and another 30 healthy volunteers in the corresponding period were taken as the control group. The proportion of Th17 and Treg cells was detected by using flow cytometry, and the concentration of HMGB1, interleukin (IL)-17 and transforming growth factor β (TGF-β) in plasma was tested by using enzyme-linked immunosorbent assay (ELISA). Isolated peripheral blood mononuclear cells (PBMC) were cultured in vitro. After the treatment with recombinant human HMGB1 (rhHMGB1), real-time polymerase chain reaction (RT-PCR) was applied to detect the mRNA expression changes in Treg cell transcription factor intracellular forkhead helix transcription factor 3 (Foxp3) and Th17 cell transcription factor retinoid related orphan receptor γt (RORγt). The differences of indicators in Treg cell transcription factor peripheral blood between the case group and the control group were compared, and the balance correlation between HMGB1 and Th17/Treg was analyzed. Results: Compared with the healthy control group, the proportion of Th17 cells and the expression level of HMGB1 and IL-17 in peripheral blood of ITP patients were increased (all P < 0.01), while the proportion of Treg cells and the level of TGF-β were decreased (all P < 0.01). The proportion of Th17 cells and the expression level of IL-17 and HMGB1 in peripheral blood of ITP patients were positively correlated with the concentration of HMGB1 (all P < 0.01); the proportion of Treg cells and the level of TGF-β were negatively correlated with the expression level of HMGB1 (all P < 0.01). In vitro experiments, the expression of intracellular RORγt mRNA was increased compared with the negative control group (1.50±0.24 vs. 0.93±0.22, t = 9.612, P < 0.01), and the expression of Foxp3 mRNA was decreased compared with the negative control group after the stimulation of PBMC by rhHMGB1 (0.72±0.19 vs. 1.08±0.18, t = 7.387, P < 0.01). Conclusion The high level of HMGB1 in the peripheral blood of ITP patients induces Th17/Treg imbalance and aggravates inflammatory reactions, which may be an important cause of ITP.

Objective: To study the incidence and clinicopathological features of intermediate fibroblastic/myofibroblastic tumors(IF/MFT) in infants and the young children. Methods: All available cases with soft tissue tumors in infants and children were retrieved from the files of Women and Children′s Hospital Affiliated to Qingdao University, from January 2012 to December 2017.The incidence rate of IF/MFT was observed.Cases of IF/MFT were identified and investigated by light microscopy and immunohistochemistry by reviewing the related literature. Results: Among 290 soft tissue tumors, 15 cases(5.2%) were IF/MFT, accounted for 88.2%(15/17 cases) of borderline soft tissue tumors.Twelve cases were male, 3 cases were female, the median age was 8 months, and 4 cases were congenital.Clinically, 11 cases were presented with slow-growing painless masses located in the trunk or extremities.According to histopathology, 9 cases(60.0%) were categorized as infantile fibromatosis(IFM), including 5 cases(33.3%) desmoid-type and 4 cases(26.7%) diffuse-type; 3 cases(20.0%) as lipofibromatosis(LFM); 2 cases(13.3%) as infantile fibrosarcoma(IFS) and 1 case(6.7%) as giant cell fibroblastoma(GCF). All 15 tumors were characterized by the presence of spindle fibroblasts and myofibroblasts with infiltration of the surrounding structures.Immunohistochemically, all the 15 cases were diffusely positive for Vimentin(Vim), but negative for Myogenin, MyoD1, Desmin and S-100.Smooth muscle actin(SMA), β-catenin and Bcl-2 were positive in some cases to a certain degree.The Ki-67 proliferation index was higher in diffuse-type IFM and IFS, the former was 5.0%-20.0%, and the latter was about 20.0%, however, the other cases showed Ki-67 <5.0%.The main clinical treatment was complete or extensive excision. Conclusions IF/MFT accounts for a high proportion of intermediate soft tissue tumors in infants and young children, mostly seen in male children, and IFM and LFM are the main types.The clinical signs and symptoms associated with these tumors are often nonspecific, and their histopathologic manifestations may overlap.The final diagnosis of IF/MFT must depend on the characteristics of age, location, histopathologic changes and immunohistoche-mical findings.

Research based learning (RBL) is a brand new teaching mode implicated by Shanghai Jiaotong University School of Medicine specifically designed for clinical undergraduates.The nature of RBL is about exploring unknown knowledge and designing and executing comprehensive experiments.RBL aims at creating an open,active student participation,and close student-mentor interaction teaching mode.IBL includes literature study,research aim selection,experiment design and implementation,statistical analysis,results and conclusions,final report writing and defense of the report.Our experience indicates that RBL can substantially improve students' scientific logic and critical thinking and experimental skills,and develop the spirit of a team-player.This scientific training enables students to receive more comprehensive scientific training,facilitate their subsequent clinical research and practice,and assist them to participate in more international scientific exchanges.We propose that the RBL mode should be adopted for clinical student education in other universities.

Objective To investigate the expression of PES1 in ovarian cancer and its relationship with the vascular endothelial growth factor(VEGF) expression .Methods The expression of PES1 in ovarian cancer tissues and corresponding pericancerous tis-sues was detected with Western blot .Results The expression of PES1 in ovarian cancer tissues was obviously higher than that in the pericancerous tissues .The secretion amounts of VEGF in cell culture fluid of CAOV-3 and ES-2 with transfection of FLAG-PES1 were elevated from (178 .0 ± 11 .8) pg/mL and (309 .5 ± 18 .5) pg/mL to (375 .0 ± 18 .3) pg/mL and (633 .2 ± 25 .7)pg/mL respectively (P<0 .01) ,the secretion amounts of VEGF were also increased(P<0 .01) .The relative VEGF mRNA levels were also raised by 1 .8 times and 2 times respectively (P< 0 .01);Western blot simultaneously showed that the over expression of PES1 could increase the expression of HIF-1αin these two kinds of cells .The luciferase report gene transcriptional activation activity de-tection reveald that PES1 had no direct effect on the VEGF transcription .After cotransfecting HIF-1αSiRNA and FLAG-PES1 to CAOV-3 and ES-2 cells ,Western blot demonstrated that HIF-1αSiRNA could obviously inhibit the increase of HIF-1αexpression induced by PES1 ,at the same time the secretion amounts of VEGF and mRNA level were also supressed .Conclusion The expres-sion of PES1 is obviously up-regulated in ovarian cancer tissues .

ObjectiveTo explore the efficacy and safety of percutaneous antegrade stenting in the treatment of ureteral obstruction after renal transplantation.MethodsWe retrospectively reviewed 11patients with renal graft ureteral obstruction (2 cases of acute obstruction and 9 cases of chronic obstruction) from March 2009 to March 2011.The etiology of the obstruction was renal graft-ureter-bladder anastomotic stricture in 5 cases,stone obstruction in 2 cases,and undetermined in 4 cases.Renal graft and collecting system were examined by ultrasonography preoperatively to select suitable puncture position,and then ureteropyelography was performed under X-ray guidance.When the obstruction location was clear,the urology guidewire was implanted to the bladder by needle,and then guidewire was released by cystoscopy.Ureteral stent was implanted along the guidewire,and upper ureteral stents was observed under X-ray. After removal of guidewire,the stent location was confirmed once again.The renal pelvis fistula drainage lasted for 1-2 weeks,and ureteral stent to 6 months to one year.Ultrasound and renal function were tested after 1week,1month,3 months and 6 months,and then every six months.ResultsOperation was done successfully in 10 patients,and failed in one case due to a long segment of ureteral stenosis.The operating time of ureteral stent implantation was 54±27 min.Serum creatinine of patients was reduced from preoperative 326±147 to postoperative 89±49 μmol/L.During a follow-up period of 6 to 27 months,no complications occurred.ConclusionPercutaneous antegrade stenting in the treatment of ureteral obstruction after renal transplantation is safe and effective.

Objective To evaluate the safety and feasibility of simplified fast track surgery in gastric surgery.Methods The study enrolled consecutive cases of elective gastric surgery from March 2010 to December 2011.The patients were randomized into two groups:simplified fast track surgery group (n =149) and traditional clinical pathway group (n =154).Demographic,intra-operative and post-operative data between two groups were compared.Results The outcome of demographic and intra-operative data between simplified fast track surgery group and traditional clinical pathway group were not different statistically (P ＞ 0.05).The time of post-operative recovery and discharge of simplified fast track surgery group was significantly shorter than traditional clinical pathway group (P ＜ 0.05).Difference of post-operative complications of two groups was not significant (P ＞ 0.05).Conclusion The programme of simplified fast track surgery in gastric surgery is safe and feasible.

Knockout is an important method for gene function study, while vector is the core of gene knockout. In order to obtain an effective vector for rapid construction of mutant and essentiality identification of the corresponding gene, we constructed a recombinant plasmid named plDM-T based on the temperature-sensitive and replication- defective plasmid plDM1 by inserting an Xcm I adapter into the EcoR I and Pst I sites of pIDM1. Digesting with Xcm I, pIDM-T can be prepared as a linear T-vector for PCR products cloning and be used to knockout the corresponding gene in the genome with insertion duplication mutagenesis. After the verification of temperature sensitivity of the replication of the plasmid, we cloned two Salmonella pullorum genes eno and ybdr into the constructed pIDM-T, and two recombinant plasmids pIDM-T_eno and pIDM-T_ybdr were identified. The recombinant plasmids were then transformed into S. pullorum strain CVCC527 and the IPC (Integration rate per cell) values were calculated. As a result, we identified the eno gene as an essential gene and the ybdr as a non-essential gene in CVCC527. We verified the correctness of recombination site in ybdr recombinant 527 clones (Sal delta ybdr) by PCR and sequencing. The pIDM-T vector can be used for gene knockout in S. pullorum, as well as the identification of essentiality of the corresponding genes, which offers an effective and rapid tool for gene function study in Salmonella.
Cloning, Molecular ; Gene Knockout Techniques ; Genetic Vectors ; genetics ; metabolism ; Plasmids ; genetics ; Recombinant Proteins ; genetics ; metabolism ; Recombination, Genetic ; Salmonella ; classification ; genetics

BACKGROUND: Vasorelaxation plays an important role In the occurrence of cyclosporine A (CsA)-induced nephrotoxlcity.OBJECTIVE: To observe the alleviative effects of green tea polyphenols (GTP) on CsA-induced inhibition of vasorelaxation and the underlying mechanisms.METHODS: Sprague-Dawley rats were randomly and evenly divided into four groups: CsA, control, CsA + GTP, and GTP. After 5 weeks of drug treatment, blood urea nitrogen (BUN) and creatinine (Cre) levels were determined. Then the thoracic aorta rings were mounted on a bath system, and acetylcholine was used to induce vasorelaxation. The effects of L-NAME and indomethacin and the denuded vasorelaxation were evaluated.RESULTS AND CONCLUSION: The BUN and Cre levels in the CsA group were significantly higher than those in the control group (P < 0.05). The maximal response (Emax%) for acetylcholine-induced vasorelaxation in the CsA group was significantly lower than that in the control and GTP groups. After pretreatment with L-NAME, vasorelaxation was significantly lower in the CsA,CsA+GTP and GTP groups than in the control group. After pretreatment with indomethacin, vasorelaxation was significantly higher in the control, CsA +GTP, and GTP groups than in the CsA group. The level of nitric oxide metabolites in the vascular tissue in the CsA group was significantly lower compared with other groups. The results demonstrated that CsA can decrease nitric oxide levels in vascular tissues and induce abnormal endothelium-dependent vasorelaxation, which is mediated by nitric oxide pathway.