Isolation and identification of multidrug-resistant Staphylococcus haemolyticus from a laboratory-breeding mouse

Objective:To analysis and identify a bacterium strain isolated from laboratory breeding mouse far away from a hospital.Methods:Phenotype of the isolate was investigated by conventional microbiological methods,including Gram-staining,colony morphology,tests for haemolysis, catalase,coagulase,and antimicrobial susceptibility test.The meek and 16S rRNA genes were amplified by the polymerase chain reaction(PCR) and sequenced.The base sequence of the PCR product was compared with known 16S rRNA gene sequences in the CenBank database by phylogenetic analysis and multiple sequence alignment.Results:The isolate in this study was a gram positive,coagulase negative,and catalase positive coccus.The isolate was resistant to oxacillin,methicillin,penicillin,ampicillin,cefazolin,cipr of loxacin erythromycin,et al.PCR results indicated that the isolate was meek gene positive and its 16S rRNA was 1465 bp.Phylogenetic analysis of the resultant 16S rRNA indicated the isolate belonged to genus Saphylococcus,and multiple sequence alignment showed that the isolate was Saphylococcus haemolyticus with only one base difference from the corresponding 16S rRNA deposited in the CenBank.Conclusions: 16S rRNA gene sequencing is a suitable technique for non-specialist researchers.Laboratory animals are possible sources of lethal pathogens,and researchers must adapt protective measures when they manipulate animals.

Effect of hepatocyte growth factor signaling pathway activation on Plasmodium berghei infection

Objective:To explore the effect of hepatocyte growth factor signaling pathway activation on Plasmodium berghei infection.Methods:In this study,hepatocyte growth factor was detected by ELISA and Western blotting assay.Hepatocyte injury was detected by FITC-dextran absorption assay,and hepatocyte growth factor expression was shown to be expressed in the same injury cells by immunofluorescence against hepatocyte growth factor.In addition,Activation of hepatocyte growth factor and its receptor signaling pathway was detected with immunoprecipitation and detection of phosphorylation status.Results:It was found that injury of hepatocytes by sporozoite migration induced the secretion of hepatocyte growth factor and it was hepatocyte growth factor that rendered hepatocytes susceptible to Plasmodium sporozoite infection.In addition,hepatocyte infections depended on activation of the hepatocyte growth factor and its receptor signaling pathway.Conclusions:Our results indicate that hepatocyte growth factor and its receptor may possibly be potential targets for new approaches to malaria treatment.

Construction and identification of recombinant plasmid pUIS3-BLC^+

Objective:To construct and identify recombinant plasmid pUIS3-BLC^+.Methods:The cDNA of B-lymphocyte chemoattractant(BLC) was amplified from the total RNA of spleen tissues by PCR method,and were inserted into plasmid of Plasmodium berghei with U1S3 knockout by digestion of restrictive endonuclease and T7 ligation.The recombinant plasmids were screened,and then underwent restriction enzymatic digestion and DNA sequencing.Then the confirmed plasmid was further transfected into COS-1 cells by lipofectamine and the BLC expression was tested by RT-PCR and Western blotting.Results:The cDNA of BLC gene was correctively amplified by RT-PCR and the recombinant plasmid pUIS3-BLC^+ was constructed successfully,which was confirmed by restriction enzymatic digestion and DNA sequencing.RT-PCR and Western blotting also showed the BLC gene expression in COS-1 cells.Conclusions:The recombinant plasmid pUIS3-BLC^+ has BLC expression in COS-1 cells,and is useful for further study on BLC transgene and UIS3 gene knockout in Plasmodium berghei.

Cy5.5-MSA-G250 nanoparticles(CMGNPs)induce M1 polarity of RAW264. 7 macrophage cells via TLR4-dependent manner

Cy5.5-MSA-G250 nanoparticles(CMGNPs)had been proved to have unique advantages for cancer treatment,including excellent photothermal performance,tumor cell-selective cytotoxicity,direct visualization,and good biocompatibility.However,to cellular systems,the CMGNPs are considered as fo reign invaders,and the effect of CMGNPs on immunity system is still unknown.Therefore,more efforts are needed to understand the role of CMGNPs on the immunity system.In this study,we attempted to screen the pro-inflammatory responses on RAW264.7 macrophages after treated with the CMGNPs.In vitro experiments clearly showed that CMGNPs not only enhances phagocytosis capacity of RAW264.7 cells,but also promotes Ml polarization,associated with changes in cell morphology and increased expression of inflammatory cytokines.This ability to induce Ml polarization may be beneficial to CMGNPs to achieve better anticancer effects in clinical trials.Moreover,the observed Ml macrophages’ polarization triggered by CMGNPs can be abolished after adding TLR4 inhibitor,CLI095,suggesting that TLR4 is involved in CMGNP-induced inflammation.

A novel therapeutic vaccine based on graphene oxide nanocomposite for tumor immunotherapy

With an intensive understanding of the mechanism of immune system,developing a therapeutic tumor vaccine is one of the most perspective strategy of cancer immunotherapy.In this study,we report a facile approach to prepare graphene oxide(GO)-based therapeutic cancer-nanovaccine.The model antigen(ovalbumin,OVA)and adjuvant(CpG ODN),are conjugated with GO-PEI nanosheet through electrostatic interaction.The addition of PEG can improve biocompatibility and prevent nanoparticle aggregation.The prepared GO-based nanovaccine,GO-PEI-OVA-PEG-CpG,exhibits good biocompatibility and low toxicity both in vivo and in vitro.More importantly,it can efficiently induce the maturation of dendritic cells(DCs),the enhancement of antigen cross-presentation ability,and the amplification of cytokine production of immune cells.Impressively,this nanovaccine shows a remarkable therapeutic effect against preestablished B16-OVA-melanoma tumors,which can significantly inhibit tumor growth and prolong the survival time of the OVA-expressed tumor-bearing mice.Moreover,combining GO-PEI-OVA-PEG-CpG with NLG919,an IDO-1(indoleamine-2,3-dioxygenase)inhibitor which can regulate the tumor microenvironment,displays a synergistic therapeutic effect.These findings indicate the GO-PEI-OVA-PEG-CpG nanovaccine actively induces an antigen-specific antitumor immune response and it combined with NLG919 could achieve better therapeutic outcomes.