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OBJECTIVE@#To investigate the effect of prostaglandin E recoptor 4 antagonist (EPA) on the self-renewal ability of human CD34 cells and its mechamism.@*METHODS@#The peripheral blood hematopoietic stem cell of 20 healthy donors received the G-CSF-mobilization were collected, then the human CD34 cells were sorted out by MACS microbead kit. The human CD34 cells were treated with DMSO (control group), EPA (EPA group) and EPA+EPA antagonist (EPA+EPA group) for 72 hours. The differential genes and pathways related with CD34 cell stemness were detected by Thermogram and Pathway enrichment analysis. and then the expression levels of protein and gene (β-catenin, Nanog, Oct4, Sox2, Stat3, AKT, P38) were detected by qRT-PCR and Western blot respectively.@*RESULTS@#EPA could elevate the mRNA and protein expression of β-catenin, Nanog, Oct4, Sox2, in comparison with control group, however, mRNA and protein expression of STAT3, AKT, P38 were not changed. When human CD34 cell were cultured with EPA+XAV939 it was found that the mRNA and protein expression of β-catenin was downregulated, moreover the mRNA and protein expression of Nanog, Oct4, Sox2 were reduced.@*CONCLUSION@#EPA can upregulate stemness factors-β-catenin, Nanog, Oct4 and Sox2 in human CD34 cell in vitro, but not STAT3, AKT and P38.
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Objective:To obtain a high specificity and high affinity anti-PcrV protein monoclonal antibody which can be used for the treatment of Pseudomonas aeruginosa infected.Methods: The PcrV gene was amplified by PCR using P.aeruginosa PAO1 genome DNA as the template.The expression vector(pET-28a-PcrV) was constructed and transformed into E.coli BL21(DE3).The re-combinant PcrV protein was expressed by IPTG induction and purified by Ni2+affinity chromatography.The specific binders of PcrV were screened by phage display.The genes encoding VH and VL were amplified respectively by PCR using the plasmid of positive clone as the template.Then the recombinant expression vectors were constructed and transfected into 293E cells.Monoclonal antibody were purified by the Protein A affinity resin from the culture supernatants.The affinity of antibody was detected by ELISA and the function of YG5 was verified in murine pneumonia model caused by P.aeruginosa.Results: Recombinant PcrV protein was expressed and purified.A full human monoclonal antibody(named as YG5) against PcrV was obtained by phage display.The results of ELISA showed that YG5 had a high affinity with EC50=61 ng/ml.Furthermore,it was found that YG5 could protect mice from infection caused by P.aeruginosa.Conclusion:Our findings present a novel human monoclonal antibody YG5 against PcrV,which inhibits the infection casued by P.aeruginosa and may be a potential drug for treatment of P.aeruginosa infection.
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<p><b>OBJECTIVE</b>To investigate the potential signaling pathway that regulates the proliferation of human CD34cells stimulated by prostaglandin E2 receptor 4 agonist (EP4A) in vitro.</p><p><b>METHODS</b>Twenty samples of peripheral blood containing stem cells were collected from the G-CSF mobilized healthy donors in our department of hematology. Human CD34cells were isolated by magnetic activated cell sorting (MACS) microbeads kit. The Cell Counting Kit-8 (CCK8) assay was used to determine the optimal concentration and time of EP4A to promote human CD34cell proliferation in vitro. Under the optimal condition, quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect mRNA level of β-catenin, and Western blot was used to assay protein expression of β-catenin and P-GSK-3β in human CD34cells treated with EP4A.</p><p><b>RESULTS</b>Culturing with 10 µmol/L EP4A for 72 h, it was found that EP4A promoted human CD34cell proliferation significantly, and the proliferation rate of human CD34cells was 1.36 times higher than that of the control(P=0.002). Under the optimal condition, it was also found that EP4A enhanced the β-catenin expression at both mRNA and protein levels, and up-regulated phosphorylation of GSK-3β in human CD34cells, but these effects could be inhibited by the EP4A antagonist EP4AA.</p><p><b>CONCLUSION</b>EP4A can enhance human CD34cell proliferation in vitro by activating Wnt/β-catenin signaling pathway.</p>
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Objective To evaluate the quality of life (QOL)among patients with anti -TB treatment in Zhoushan city and to analyze the influence factors.Methods Our self -designed questionnaire and WHOQOL -BREF scale were used to investigate the general information and the quality of life among 187 tuberculosis patients.Scores of life quality under different conditions were compared.Results The scores of physiological domain,psychological domain,social relations and environmental domains were 61.13 ±13.12,53.24 ±16.59,59.91 ± 12.08 and 60.12 ± 15.68 respectively. The scores of all domains excluding the environmental relations were significantly lower than the normal level (P <0.05). The scores of males were significantly higher than females in psychological domain (P <0.05),and the score differences among age groups were of statistical significance both in physiological domain and psychological domain (both P <0.05). Those with high education levels scored higher than those with lower education level in psychological and social domains (both P <0.05).The scores of different economic income in 4 domains showed significant differences (all P <0.05). The first time treated,hospitalization time less than 1 month and effectively treated patients scored higher than those not (all P <0.05).Conclusion The quality of life among tuberculosis patients could be influenced by sex,age,income, education,and treatment factors,so appropriate health care such as psychological support and health education should be provided.
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A series of novel self-assembled polymeric micelles based on carboxymethyl chitosan bearing long chain alkyl chains (N-octyl-O, N-carboxymethyl chitosan, OCC) was synthesized. PTX loaded OCC polymeric micelles (PTX-OCC) were prepared by dialysis method. The effects of the degree of substitutions (DS) of octyl groups on the solubilizing abilities of OCC for paclitaxel were studied. The PTX-OCC were characterized using drug loading content, drug encapsulation efficiency, dynamic light scattering, zeta potential and transmission electron microscopy (TEM). Take PTX injection (PTX-INJ) as control, the safety of PTX-OCC including hemolysis, hypersensitiveness in guinea pigs and acute toxicity in mice were also evaluated. OCC showed excellent loading capacities for paclitaxel with the DS of octyl groups in the range of 37.9% - 58.6%. Drug loading contents were up to 24.9% - 34.4% with drug encapsulation efficiency 56.3% - 89.3%, which both increased with the increasing of DS of octyl groups. The mean size of PTX-OCC was 186.4 - 201.1 nm which decreased with the increasing of DS of octyl groups. The zeta potential was -47.5 to -50.9 mV, which had no obvious relation with the DS of octyl groups. The TEM images showed a spherical shape. No burst release phenomena were observed and drug cumulative release was in the range of 60% -95% in 15 days. PTX-OCC with higher DS of octyl groups showed stronger sustained releasing ability. In terms of the induction of membrane damage and hypersensitiveness, PTX-OCC was superior to PTX-INJ. The LD50 and its 95% confidence interval of PTX-OCC were 134.4 (125.0 - 144.6) mg x kg(-1), which was 2.7 fold of PTX-INJ. The present PTX-OCC could be potentially useful as safety carriers for intravenous delivery.