Dermatitis, Occupational ; diagnosis ; Diagnostic Errors ; Female ; Humans ; Trichloroethylene ; adverse effects ; Young Adult

Objective:To prepare the production of TIGIT-Fc fusion protein using H22 cells stably integrated the gene by lentivirus vector , and to explore the immunoregulatory effect on macrophages by TIGIT-Fc.Methods: TIGIT-Fc fusion gene were constructed by molecular cloning.The fusion gene was then subcloned to plasmids contained the secretion signaling peptide .The secrected TIGIT-Fc fusion gene was inserted into the lentivirus backbone vector.The purified lentivirus vector was the used to infect the murine H22 cell line.TIGIT-Fc protein was purified by protein A column from the ascites of H 22-injected C57BL/6 mice.Macrophages stimulated by lipopolysaccharide ( LPS ) was challenged to TIGIT-Fc treatment or control.Cytokine levels was then detected by ELISA.Results: TIGIT-Fc protein was purified from the ascites of H 22-injected mice.PVR was upregulated in LPS-treated macrophages.IL-10 level was upregulated in TIGIT-Fc treated macrophages.Conclusion: TIGIT-Fc promotes the mature macrophages to secrete anti-inflammatory cytokine IL-10.

To construct a plasmid expressing MDR1 short hairpin RNA (shRNA) mediated by pEGFP-C1/U6 vector, two coding sequences of 19 nucleotides were selected from MDR. Two pairs of oligonucleotides were designed for these two fragments. After annealing the formed double-stranded DNAs were ligated with plasmid pEGFP-C1/U6 (pEGFP-C1 vector with U6 promoter). The plasmids producing MDR1 shRNA were constructed from the inverted motif containing 9 spacers and four Ts. The results showed that the constructed plasmids were named pEGFP-C1/U6/A and pEGFP-C1/U6/B, and the constructs were identified by restriction and sequence analysis, no any base mutation was observed. It is concluded that plasmids of pEGFP-C1/U6/A and pEGFP-C1/U6/B expressing MDR1 shRNA were successfully constructed, providing a highly effective method for reversing the multidrug resistance in clinic.
ATP-Binding Cassette, Sub-Family B, Member 1 ; biosynthesis ; genetics ; DNA-Binding Proteins ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; Humans ; Plasmids ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; biosynthesis ; genetics ; RNA, Small Nuclear ; genetics

Objective To investigate the infectious status,etiological spectrum and epidemiological characteristics of rotavirus (group A/B/C),calicivirus (novovirus Ⅰ/Ⅱ,sapovirus),astrovirus and enteric adenovirus in diarrhea cases below 5 years old from 2008 to 2015 in Henan provinces.Methods Totally 2541 stool samples were collected from cases below 5 years old in four sentinel hospitals.All stool specimens were tested for group A rotavirus by double antibody sandwich ELISA method.G/P genotyping of group A rotavirus was determined by nested multiplex PCR.Viral RNA was extracted from all samples and rotavirus (group B/C),calicivirus,astrovirus and enteric adenovirus were detected by two-step reverse transcription-polymerase chain reation (RT-PCR)/PCR.Results One thousand four hundred and twenty-one out of 2 541 samples were positive with a total positive rate of 55 .9%,among which,102 were mixed infection.The isolation rate of rotavirus was 36.0% (914 samples)(group A:785 cases,group B:36 cases,group C:93 cases),calicivirus was 12.1 % (308 samples)(novovirus Ⅰ:64 cases,novovirusⅡ:193 cases,sapovirus:51 cases),astrovirus was 5 .9% (151 samples),enteric adenovirus was 1 .9%(48 samples).The group A rotavirus gene type combinations were composed mainly of G9P[8],G2P[4], G3P[8 ],G1P [8 ]and most cases were identified from September to November and March to May. Novovirus Ⅱ was predominant in calicivirus and most cases were identifed between March and May. Rotavirus or calicivirus infection was mainly among children aged 4—12 months or 3—5 years, respectively.Clinical manifestations included fever,diarrhea,vomiting,dehydration.Gender and region distributions differed according to the types of pathogen.Conclusions Group A rotavirus and novovirus Ⅱare the major viral pathogen in diarrhea cases younger than 5 years old in Henan province.Different viral infections exhibit extinct epidemiologic and clinical characteristics.

Benzene ; poisoning ; Hazardous Substances ; Humans ; Occupational Exposure ; prevention & control

Child ; China ; epidemiology ; Cytochrome P-450 CYP2E1 ; biosynthesis ; genetics ; Female ; Glutathione Transferase ; biosynthesis ; genetics ; Humans ; Leukemia, Myeloid, Acute ; epidemiology ; genetics ; metabolism ; Male ; NAD(P)H Dehydrogenase (Quinone) ; biosynthesis ; genetics ; Point Mutation ; Polymorphism, Genetic ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; epidemiology ; genetics ; metabolism ; Risk Factors

OBJECTIVETo construct a short hairpin RNA (shRNA) eukaryotic expression vector specific to MDR1 gene in multidrug resistance (MDR) human leukemia cell line K562/A02 to observe its silencing effect on MDR1 and P-glycprotein (P-gp) expression.

METHODSThe shRNA expression vector was constructed by gene recombination, then transfected into the cultured K562/A02 cells. The transcription of MDR1 gene was detected by semi-quantitative RT-PCR and the expression level of P-gp was determined by Western blot. 50% inhibition concentration (IC50) of ADM in K562/A02 cells was determined by MTT method. The intracellular doxorubicin (ADM) concentration was determined by HPLC.

RESULTSThe introduction of pEGFP-C1/U6/MDR1-A or pEGFP-C1/U6/MDR1-B expression vector was shown to efficiently and specifically inhibit the expression of P-gp according to results of Western blot, with an inhibitory rate of 50.67%. Semi-quantitative RT-PCR showed that mRNA transcription of MDR1 gene was reduced by (48.2 +/- 2.5)%. On the contrast, the control plasmid did not exhibit inhibitory effect on the protein expression and mRNA transcription of MDR1. The relative efficiency of K562/A02 to ADM was 40.8% or 62.4%, respectively, and the intracellular accumulation of ADM increased after shRNA treatment.

CONCLUSIONThe shRNA expression vector targeting MDR1 gene showed dramatic inhibition on RNA transcription and protein expression. It could effectively restore the sensitivity of K562/A02 cells to conventional chemotherapeutic agents.

ATP Binding Cassette Transporter, Sub-Family B ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Antibiotics, Antineoplastic ; metabolism ; pharmacology ; Blotting, Western ; Cell Survival ; drug effects ; Dose-Response Relationship, Drug ; Doxorubicin ; metabolism ; pharmacology ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Gene Silencing ; Genetic Vectors ; genetics ; Humans ; K562 Cells ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

Objective To study the inhibitory effect of pterin acid (PTA) against ricin and recombinant ricin A chain protein. Methods Luciferase protein synthesis inhibition assay in a cell-free system and in vitro cytotoxicity experiments were performed to assess the biological activity of ricin and rRTA treated with PTA.Results The result showed that PTA could significantly inhibit the activity of ricin and rRTA in a dose-dependent manner.Conclusion PTA might be used as a small molecular probe to develop an evaluating system for ricin/RTA small molecular inhibitor in vitro. The cell-free system adopted in the current study could also serve as a necessary basis for screening some novel small molecular compounds against ricin and RTA in the future.

Objectives To synthesize hepatitis B virus e gene which was suitable for yeast protein expression and express hepatitis B virus e antigen (HBeAg) in Pichia pastoris.Methods Using synonymous codons preferred by yeast usage on protein expression to replace some native codons of wild-type HBV e gene,the synthetic e gene (syneAg-gene) was achieved by a recursive PCR (rPCR).The syneAg- gene was inserted into the yeast expression vector pPICZ?A.The recombinant plasmid was transformed into GS115 yeast by electroporation.The yeast transformant induced by methanol expressed the HBeAg.Results The restriction analysis and DNA sequencing confirmed that the syneAg-gene was inserted to yeast pPICZ?A in correct orientation.SDS-PAGE,immunoblot and ELISA indicated that the secreted form of HBeAg was expressed by the yeast transformant.The expression level of HBeAg in the strain containing syneAg-gene was 63 mg/L.The titer of the recombinant HBeAg in culture superuatant was 1 :81 920 and the maximum OD (optical density) value of absorbance was 2.8 by ELISA.No cross reactivity between Pichia pastoris-derived HBeAg and anti-HBc antibody was found.Conclusion The recombinant HBeAg with high degree of specificity and immunoreactivity is expressed efficiently in Pichia pastoris.

No abstract available.
Drug Therapy* ; Ovarian Neoplasms*