Effective antimicrobial activity of Cbf-14-2 against penicillin-resistant bacteria in vitro and in vivo / 中国药科大学学报

The antibacterial activity and mechanism of the antimicrobial peptide mutant Cbf-14-2 against NDM-1 carrying recombinant bacteria (E.coli BL21 (DE3)-NDM-1) was investigated in this study.The minimum inhibitory concentration (MIC),minimum bactericidal concentration (MBC) and killing curves (KCs) in vitro were determined by the broth microdilution method.Mice septicemia model was established by interaperitotoneal injection of E.coli BL21 (DE3)-NDM-1 to evaluate the antibacterial activity of this peptide in vivo.Results showed that Cbf-14-2 exhibited a potent antibacterial activity with MIC of 16 μg/mL and killed almost all recombinant bacteria within 120 min.Meanwhile,it significantly improved the survival rate of infected mice up to 70％ with the decreasing of bacterial load in mice lung,liver,spleen and kidney.This powerful clearance ability of Cbf-14-2 against bacteria mainly related to its enhanced membrane penetration ability through neutralizing the negative charges and disrupting the integrity of the bacterial cell membrane.Therefore,Cbf-14-2 is expected to be a potential antimicrobial agent for the treatment of infection induced by multi-drug resistant bacteria,especially for the NDM-1carrying bacteria.

Markerless DNA deletion based on Red recombination and in vivo I-Sec I endonuclease cleavage in Escherichia coli chromosome / 生物工程学报

Red-based recombineering has been widely used in Escherichia coli genome modification through electroporating PCR fragments into electrocompetent cells to replace target sequences. Some mutations in the PCR fragments may be brought into the homologous regions near the target. To solve this problem in markeless gene deletion we developed a novel method characterized with two-step recombination and a donor plasmid. First, generated by PCR a linear DNA cassette which comprises a I-Sec I site-containing marker gene and homologous arms was electroporated into cells for marker-substitution deletion of the target sequence. Second, after a donor plasmid carrying the I-Sec I site-containing fusion homologous arm was chemically transformed into the marker-containing cells, the fusion arms and the marker was simultaneously cleaved by I-Sec I endonuclease and the marker-free deletion was stimulated by double-strand break-mediated intermolecular recombination. Eleven nonessential regions in E. coli DH1 genome were sequentially deleted by our method, resulting in a 10.59% reduced genome size. These precise deletions were also verified by PCR sequencing and genome resequencing. Though no change in the growth rate on the minimal medium, we found the genome-reduced strains have some alteration in the acid resistance and for the synthesis of lycopene.

Early apoptosis leads to decrease of B cells in MRL/lpr mice / 中国免疫学杂志

Objective:To explore the change of B cell numbers in active MRL/lpr lupus mice , and their regulation mechanisms.Methods:B cell cycle and the percent of B cells in spleen lymphocytes of active MRL /lpr lupus mice and normal C 57/B6 mice were analyzed by using flow cytometry .The apoptotic B cells and their subclass were analyzed by Annexin V and PI staining.Further more ,B cells were purified by magnetic sorting , and real-time quantitative PCR was carried out to detect apoptosis-related gene.Results:Compared with the C57/B6 mice,the percent of B cells in active MRL/lpr lupus mice were significantly reduced (P<0.01),while the percent of apoptotic cells were significantly increased (P<0.01).The percent of early apoptotic B cells were sig-nificantly increased ( P <0.01 ) which including the immature and mature B cells , while the late apoptotic B cells were unchanged.Further more,we found that the anti-apoptotic protein BIRC3 was significantly reduced in active lupus B cells (P<0.01), while the pro-apoptotic protein BCL2L1 and BBC3(PUMA) were significantly increased(P<0.01).Conclusion: B cells in active lupus mice were significantly reduced while early apoptotic B cells were increased , which may be attributed to the changed balance between the anti-apoptotic and pro-apoptotic proteins , suggesting the reduction of B cells in SLE patients may be related to their increased early apoptosis .

Immunomodulatory activity of polysaccharide and research progresses of polysaccharide medicine / 中国生化药物杂志

Polysaccharides are a class of natural macromolecules with biological activities which could regulate the immune system by activating immune cells and promoting the secretion of cytokines to exert the anti-tumor and anti-oxidation activities.The article reviews the mechanism of polysaccharides acting on the immune system and its anti-tumor immunotherapy applications.

Application of metabolic regulation in the antibiotic biosynthesis of actinomycetes / 中国药科大学学报

@#The secondary metabolites of actinomycetes are rich and diverse, which have become important sources of antibiotics and their lead compounds. The study of microbial secondary metabolism is focused on metabolic regulation of antibiotic biosynthesis in actinomycetes by genetic manipulation. On the basis of the advances of recent years, this paper summarizes the application progress of metabolic regulation used for improving biosynthesis of antibiotics in actinomycetes, concluding of regulating the expression of regulatory genes, increasing the copy numbers of gene clusters and heterologous expression, over-expressing the resistance genes and transfer gene, improving precursor metabolic flux, and ribosome engineering.

Synergistic inhibition of Lactobacillus acidophilus and Clostridium butyricum against Shigella flexneri in vitro / 中国药科大学学报

@#An agar diffusion method was used to investigate the antimicrobial activities of fermentation broth of Lactobacillus acidophilus(L. acidophilus), and Clostridium butyricum(C. butyricum)against Shigella flexneri(S. flexneri)infection in vitro. It was found that cell-free culture supernatants(CFCSs)of L. acidophilus and C. butyricum possessed remarkable synergistic anti-S. flexneri activity in vitro and the antimicrobial activity of the mixed culture was 17. 2% and 22. 4% greater, than single strains alone, respectively. Meanwhile, there was a symbiotic relationship between L. acidophilus and C. butyricum. The result showed that the biomass accumulation of the mixed culture reached 4. 27 g/L(DCW)and increased by 6. 0% and 30. 6% compared to the L. acidophilus and C. butyricum, respectively. Moreover, L. acidophilus and C. butyricum clearly inhibited S. flexneri adhesion to Caco-2 cells by 75. 8% and 81. 2%, and the combination of these two probiotic strains demonstrated the highest inhibition rate, reaching 84. 2%, while the viability of Caco-2 cells treated with L. acidophilus, C. butyricum or their combination was increased by 11. 5%, 12. 5% and 22. 9%, respectively. The synergistic effect of L. acidophilus and C. butyricum provided better protection against Shigella flexneri infection, which represents a promising alternative therapy for shigellosis.

JMJD3 participates in activation and apoptosis of IFN-αand TLR7-induced B cells / 中国免疫学杂志

Objective:To explore the effect of histone demethylase JMJD3 on B cell activation and apoptosis.Methods:B cells were sorted and purified from the peripheral blood of healthy people and SLE patients by using magnetic bead.After B cells were treated with IFN-αor R848 or IFN-α+R848,the percentages of CD86+B cells,CD69+B cells,CD86+Annexin V+B cells and CD69+Annexin V+B cells were detected by flow cytometry.The expression of JMJD3 was detected by Real Time PCR and Western blot.Results:The purity of sorted B cells was up to 95%.IFN-αenhanced both the activation and apoptosis and the JMJD3 expression of TLR7-activated B cells.The expression of JMJD3 was dependent on MAPK signal pathway,but not the NF-κB signaling pathway.Moreover,JMJD3 was highly expressed in B cells of peripheral blood from SLE patients compared to those from healthy people.Furthermore,JMJD3 inhibitors could inhibit the activation and apoptosis of IFN-αand R848 activated B cells.Conclusion:JMJD3 participated in the activation and apoptosis of IFN-αand TLR7-induced B cells, suggesting JMJD3 inhibitors may possess therapeutic effect for alleviating symptom of SLE.

High RNA accumulation in Candida tropicalis is affected by specific growth rate and different medium composition / 中国药科大学学报

Batch and continuous fermentation were adopted to investigate the effect of specific growth rate and amino acid components on RNA accumulation in Candida tropicalis ATCC 20408 in fermentation medium ( FM), yeast peptone dextrose medium (YPD), molasses fermentation medium ( MFM) and FM without corn steep liquor. The data showed that obvious differences in intracellular RNA accumulation were observed at different cell growth phases in bath fermentation prosess, and RNA level reached 11. 8% (g-RNA /g-DCW) during exponential phase, and only 6.9% during stationary phases. It was also found that intracellular RNA accumulation increased with the increase of specific growth rate in continue fermentation prosess, and the highest RNA level reached 15. 6% with the glucose conversion rate of 42. 8% at the dilution rate of 0. 5 h-1. Furthermore, the data showed that RNA lever was notably increased in batch fermentation process when amino acids or peptone was added into the fermentation medium containing no corn steep liquor. Taken together, it was reported for the first time that specific growth rate and amino acid components plays a leading role on the intracellular RNA accumulation in C. tropica lis, and specific growth rate is more important.

Progress in the research of therapeutic enzyme / 生物工程学报

With the development of the research on biotechnology and modern pharmacy, the application of enzyme drugs have grown rapidly and enzyme drugs have become an important branch of biopharmaceutics. In this article, some new varieties of therapeutic enzymes, enzyme targets, mechanisms and new technologies of application in therapeutic enzymes were reviewed, and the direction of development of therapeutic enzymes were discussed.

Histocompatibility evaluation of nano hydroxyapatite-40% zirconia composite bioceramic / 中国矫形外科杂志

[Objective] To evaluate the histocompatibility of nano hydroxyapatite-zirconia composite bioceramic.[Methods]According to the standard of ISO 10993-1,cytotoxicity experiment,acute toxicity test,hemolysis test and in vivo implantation(90 days)test were conducted to evaluate the histocompatibility of nano hydroxyapatite-zirconia composite bioceramic.[Results]The score of cytotoxicity experiment was lower than grade I,and there was no significant inhibition of cell growth,no acute toxic reaction or hemolytic reaction.And the in vivo implantation met the requirements of the biological evaluation of implant materials.[Conclusion]Nano hydroxyzpatie-zirconia composite bioceramic showed a good histocompatibility.It has a broad prospect as a biomaterial scaffold in the bone tissue engineering.

STUDIES ON THE SUBMERGED-CULTURE TECHNOLOGY FOR COPRINUS COMATUS FR. CC-8810 / 微生物学通报

Suitable conditions for polysaccharide production in submerged culture by Coprinus comatus Ft. CC-8810 were: glucose 40g, Soybean cake powder 30g, yeast extract powder 5.0g, bran 30g, KH_2PO_4 1.0g, MgSO_4?7H_2O 0.5g, tap water 1L, pH5—6, 28℃, 160r/min. Seed culture grown in shake flask for 4 days were inoculated into 1.5L fermentor. After 5—6 days the fermentation ended up with decreases in pH to 4.9—5.0 and in reducing sugar to 1.2%. 45.1g(dry Wt.)/L hyphae and 3.6g/L polysaccharide were obtained.