12E7 Antigen ; Antigens, CD ; metabolism ; CD57 Antigens ; metabolism ; Cell Adhesion Molecules ; metabolism ; Female ; Humans ; Immunohistochemistry ; Neuroectodermal Tumors, Primitive, Peripheral ; metabolism ; pathology ; therapy ; Pancreas ; chemistry ; pathology ; surgery ; Pancreatectomy ; methods ; Pancreatic Neoplasms ; metabolism ; pathology ; surgery ; Phosphopyruvate Hydratase ; metabolism ; Young Adult

Graft vs Host Disease ; metabolism ; surgery ; Graft vs Tumor Effect ; HLA Antigens ; genetics ; Hematopoietic Stem Cell Transplantation ; Humans ; Receptors, KIR ; genetics ; metabolism

AIM:To construct eukaryotic expression vector expressing double shRNA sections targeting Survivin gene.METHODS: Eukaryotic expression vector expressing double shRNA sections targeting Survivin gene were designed and chemically synthesized.They were directionally inserted into plasmid pGenesil-1 with respectively U6 promoter and termination code,the common green fluorescence protein(EGFP) gene and Neo gene. In this way,the vector of pGenesil-1 shRNA containing 2 sections of Survivin shRNA were constructed and they were transfected into the pancreatic cancer cell Bx-PC3.Transfection was detected by fluorescence microscope.The inhibition expression of Survivin mRNA was measured by RT-PCR.RESULTS: HE1 and HE2 plasmids were identified by the biocatalyst cut which confirmed the exactitude and were analyzed by the sequence analysis which verified the perfect clone plasmid inserted by them.CONCLUSION: A eukaryotic expression vector of double short hairpin RNA for Survivin gene is successfully constructed.The pancreatic cancer cells Bx-PC3 succeed to be transfected and expression of Survivin mRNA is inhibited obviously.

Pediatrics internship is a bridge that brings medical theory into clinical reality.Cultivate systematic clinical thinking during internship is the key to the growth of clinical doctors.Good outcome has been achieved among these eight-year medical students by using teaching methods such as case discussions,lectures,teacher instructions,which can cultivate proper clinical thinking and make students more active.

Objective By detecting the mutations spectrum of phenylalanine hydroxylase(PAH)gene in phe-nylketonuria(PKU)patients and their parents. The researchers analyzed the gene mutation features and high - frequency mutations and determined the relationship between the genotype and the phenotype,which would provide a theoretical basis for the early diagnosis and genetic consultation of PKU children in the region. Methods In this study,13 exons and their flanking introns of the PAH gene in 32 PKU patients and their parents from Wuxi and Suqian in Jiangsu province were sequenced by using the next - generation sequencing(NGS)technology. Results Sixty - one mutant sites and 32 mutant genes were detected in 32 PKU patients,and the mutation detection rate was 95. 31%(61 / 64 cases). The variants at c. 721C ﹥ T,c. 1068C ﹥ A,c. 611A ﹥ G,c. 1197A ﹥ T,c. 728G ﹥ A,c. 331C ﹥ T and c. 442 -1G ﹥ A were common mutations in the region with mutation frequency over 5% . What's more,4 novel variants of c. 699C ﹥ G,c. 265C ﹥ T,c. 722G ﹥ A and c. 1194A ﹥ G were found. Of those,c. 699C ﹥ G was not recorded in the PAH variant database and HGMD database and. c. 265C ﹥ T,c. 722G ﹥ A,and c. 1194A ﹥ G were first reported in the Chi-nese population. Genotype - accurate biochemical phenotype correlation by using the Guldberg AV system revealed con-sistency rate of 38. 0%(8 / 21 cases),which the consistency rate between accurate biochemical phenotype and predic-tive phenotype of moderate to severe genotype was 92. 3%(12 / 13 cases),and mild genotype was 50. 0%(4 / 8 cases). Conclusions The PAH gene variants of PKU patients in Jiangsu province are distributed mainly in exons 7,of which the highest frequency gene mutation is c. 721 c ﹥ T. Moreover,one novel variant c. 699C ﹥ G was reported for the first time. The PKU children inherit the PAH mutation gene mainly from both parents. There are definite correlation between the genotypes and phenotypes.

Objective To explore the changes of CD40 and CD40 ligand(CD40L) levels and investigate their significances in children with graft-versus-host disease(GVHD)after related-donor human leukocyte antigen(HLA) matching allogeneic hematopoietic stem cell transplantation(HSCT).Methods Nineteen patients with ?-thalassemia major and 1 patient with congenital inherent hemolytic anemia accepted umbilical cord blood transplantation(UCBT) and allogeneic peripheral blood stem cell transplantation(allo-PBSCT),respectively,and all cases were received successfully related-donor human leukocyte antigen matching allogeneic HSCT.Peripheral blood samples were obtained before and after transplantation,the time when GVHD happened,the expressions of CD40 and CD40L were measured by using immunofluresence asssy.Results Four UCBT children and 3 allo-PBSCT children had no acute GVHD.Thirteen children had acute GVHD(degreeⅠ-Ⅳ),the expressions of CD40L on CD4+ and CD8+ T cells in the patients with acute GVHD increased,especially in allo-PBSCT.Five cases of UCBT and 12 cases of allo-PBSCT patients had chronic GVHD,the expressions of CD40L+,CD25+ and CD69+ on CD4+ and CD8+ T cells in patients with chronic GVHD increased obviously.The expression of CD19+CD40+ was lower than normal within 3 months after transplantation.Conclusions The high expression of CD40L+T cells in periferal blood after HSCT was related to the activation and proliferation of T cells in the development of GVHD in HSCT.

Objective To study the mutations of Env sequence of SIVmac239 after infection of Chinese rhesus monkeys, and compare the differences and characteristics of Gp120 sequences of enterotropic and neurotropic SIV strains. Methods Six strains of simian immunodeficiency virus were analyzed in this study: four separated from peripheral blood mononuclear cells of SIVmac239-infected monkeys and two neurotropic SIVmac251 strains.Isolated and cultured monoclonal virus was obtained by limiting dilution assay.Gp120 sequences were amplified after the RNA extraction and phylogenetic analysis was processed thereafter.So did the Gp120 amino acid sequence and N-glycosylation sites analysis of the enterotropic and neurotropic strains.Results SIVmac239 had different mutations in four rhesus monkeys.The diversity in amino acid sequences of the enterotropic and neurotropic strains concentrated in the V1 and V4 regions of Gp120.The enterotropic strains had an addition of glycosylation site in V4 but the glycosylation site changes of neurotropic strains were located in the conservative regions of C1, C2 and C3.Conclusions The addition of one glycosylation site in V4 region of GP120 and loss of one glycosylation site in C1 region are associated with enhanced enterotropism and neurotropism.The differences between the enterotropic and neurotropic strains are not dipicted in Gp120 V3 region which is closely related with the tropism of strains.

Objective Salidroside is a major active component of integripetal rhodiola herbal medicine, which has a significant activity against hypoxia and ischemia.This study was to investigate the effects of side chain carbon losssalidroside analogues (N04) on the expressions of HIF-1α-related factors in the hypoxia-injured EAhy926 human vascular endothelial cells.Methods EAhy926 human umbilical vein endothelial cells in the logarithmic growth phase were randomly divided into a normal control, a hypoxia model control, a salidroside, a high-dose N04, a medium-dose N04, and a low-dose N04 group.The hypoxia model was established by depriving the culture medium of sugar and serum and culturing the EAhy926 cells in an environment of 95%N2+5%CO2 for 2 hours, followed by intervention with salidroside at 1×10-6 mol/L and N04 at 1×10-6, 1×10-7, and 1×10-8 mol/L, respectively.Then, the activity of the cells was detected by MTT assay, their LDH activity examined by spectrophotometry, the mRNA expressions of HIF-1α and VEGF measured by RT-PCR, the protein expressions of HIF-1α, VEGF and pVHL determined by Western blot, and the activity of eNOS measured by ELISA.Results Compared with the normal control group, the hypoxia model cells showed significantly reduced activity (0.51±0.05 vs 0.27±0.02, P<0.01), an elevated LDH level ([6.65±1.43] vs [78.82±2.33] U/L, P<0.01), and decreased eNOS activity ([1.56±0.23] vs [1.16±0.20] U/100 mL, P<0.01).In comparison with the hypoxia model group, the cells treated with high-, medium-, and low-dose N04 exhibited remarkably increased activity (0.27±0.02 vs 0.0.42±0.05, 0.40±0.03 and 0.37±0.04, P<0.01), a reduced LDH level ([78.82±2.33] vs [53.05±3.90], [58.42±4.45] and [62.73±3.63] U/L, P<0.01), and increased eNOS activity ([1.16±0.20] vs [3.01±0.47], [2.60±0.26] and [2.32±0.29] U/100 mL, P<0.01).The activity of eNOS was also increased in the salidroside group ([2.32±0.29] U/100 mL, P<0.01).The cell activity in the high-and medium-dose N04 groups was markedly higher than that in the salidroside group (P<0.05), and so was the eNOS activity in the high-dose N04 group and the LDH level in the medium-and low-dose N04 groups (P<0.05).In comparison with the normal control group, the expressions of HIF-1α mRNA, HIF-1α protein and VEGF protein were significantly up-regulated in the hypoxia model group (P<0.01) while that of the pVHL protein markedly down-regulated (P<0.01).Compared with the hypoxia model group, the expressions of HIF-1α mRNA, HIF-1α protein and VEGF protein were remarkably reduced (P<0.05), while that of the pVHL protein markedly elevated (P<0.05).Both the expressions of VEGF mRNA and HIF-1α protein were significantly lower in the medium-and low-dose N04 groups than in the salidroside group (P<0.05).Conclusion N04 can protect vascular endothelial cells against hypoxia-induced injury by regulating the expression of HIF-1α-related factors and eNOS.

Objective To investigate the anti-inflammatory and analgesic activities of different extracts from X. mongolicum compared with that of X. sibiricum. Methods Seventy male Kunming mices were randomly divided into seven groups:control group,aspirin group,the ethanol extracts of X. sibiricum group and X. mongolicum group,the extracts from X. mongolicum with ether,ethyl acetate,and n-butyl alcohol group(n=10 each). The mice were administrated with 500 mg·kg-1 ( p. o. ) of different extracts. The hot-plate tests and the xylene-induced ear edema in mice were performed to observe the analgesic and anti-inflammatory activities,respectively. Results In the hot-plate tests,the pain threshold from the extracts of X. sibiricum and the different extracts from X. mongolicum at 60,120,180 min were prolonged,and there was no statistically significant differences(P>0. 05) between the ethanol extracts from X. sibiricum and X. mongolicum. The best analgesic activity was the ether and butyl alcohol extracts from X. mongolicum. Meanwile,the ear edema from the extracts of X. sibiricum and the different extracts of X. mongolicum were inhibited significantly(P<0. 05). There was also no significant differences between the ethanol extracts of X. sibiricum and X. mongolicum(P>0. 05). The ether and butyl alcohol extracts from of X. mongolicum showed the best anti-inflammatory effect. Conclusion X. mongolicum exhibites significant anti-inflammatory and analgesic activities comparable to that of X. sibiricum. The best anti-inflammation and analgesic activities were from ether and butyl alcohol extracts from X. mongolicum.