Preparation, characterization and performance of gamma-polyglutamic acid/carboxymethyl chitosan-calcium phosphate cement / 中国组织工程研究

BACKGROUND: Nowadays complex bone defects have become a great challenge to orthopedists. A synergistic contribution of various growth factors and a crosstalk between their signaling pathways have been suggested as determinatives for the overall osteogenic outcome.OBJECTIVE: To develop calcium phosphate cement (CPC) incorporated with γ-polyglutamic acid/carboxymethyl chitosan (PGA/CMCS), and to evaluate its physical and chemical properties and sustained-release function. METHODS: The γ-PGA/CMCS polymer composites were prepared by graft copolymerization and spray freeze drying methods, and then loaded with recombinant human bone morphogenetic protein 2 (rhBMP-2) growth factor. CPC served as control group, and γ-PGA/CMCS-CPC containing different contents of rhBMP-2 as experimental groups. A γ-PGA/CMCS-CPC scaffold with regular blade-like crystalline structure was fabricated by injection compression molding. Before mixed with the liquid phase, the solid additives were properly mixed by wet method of CPC solid and the γ-PGA/CMCS carrier, then the pre-blended mix was freeze-dried. The setting time and compressive strength of bone cement in each group were detected, and the microstructure of the material surface was observed under scanning electron microscopy. In vitro release of rhBMP-2 was investigated. The effect of bone cement extracts on cell proliferation was determined through MTS assay.RESULTS AND CONCLUSION: γ-PGA/CMCS-CPC had the same physicochemical properties to the CPC. Initial and final setting time, compressive strength of bone cement had no significant differences among groups. The scanning electron microscope results showed that the γ-PGA/CMCS-CPC scaffold was covered by regular blade-like crystalline structure and the γ-PGA/CMCS particles were uniformly dispersed in the CPC crystals. A sustained release of rhBMP-2 was observed from the γ-PGA/CMCS-CPC. The cell experiments exhibited that the samples with regular blade-like crystalline structure had better cell response compared to CPC control groups with irregular crystalline structure. These findings indicate that γ-PGA/CMCS-CPC can maintain good physicochemical properties, and release growth factor or drug to promote bone formation.

Codon usage bias in the straw mushroom Volvariella volvacea / 生物工程学报

We analyzed the whole genome coding sequence of Volvariella volvacea to study the pattern utilization of codons by Codon W 1.4.2. As results, 24 optimal codons were identified. Moreover, the frequency of codons usage was calculated by CUSP program. We compared the frequency of codons usage of V. volvacea with other organisms including 6 modal value species (Homo sapiens, Saccharomys cerevisiae, Arabidopsis thalian, Mus musculus, Danio rerio and Drosophila melanogaster) and 4 edible fungi (Coprinopsis cinerea, Agaricus bisporus, Lentinula edodes and Pleurotus ostreatus). We found that there were less differences in 3 edible fungi (excluding Pleurotus ostreatus) than 6 modal value species, comparing with the frequency of codons usage of V. volvacea. With software SPSS16.0, cluster analysis which showed differences in the size of codon bias, reflects the evolutionary relationships between species, which can be used as a reference of evolutionary relationships of species. This was the first time for analysis the codon preference among the whole coding sequences of edible fungi, serving as theoretical basis to apply genetic engineering of V. volvacea.

Effects of cholesterol overload on visfatin secretion from adipocytes and the mechanism / 中华内分泌代谢杂志

3T3-L1 adipocytes were cultured and differentiated into mature adipocytes in vitro. The adipocytes were intervened by ACAT inhibitor( 2 μg/ml) and ox-LDL with various concentrations (0,25,50,75,and 100 μg/ml)for 48 h,ACAT inhibitor( 2 μg/ml) and ox-LDL( 50 μg/ml) at the 0,6,18,36,and 48 h,or ACAT inhibitor( 2 μg/ml),ox-LDL( 50 μg/ml),and TUDCA with various concentrations(0,100,200,and 400 μ mol/L)for 48 h,respectively.The levels of visfatin in supernatant were examined by ELISA and the expressions of protein GRP78 and CHOP in adipocytes were detected by Western blot.After the adipocytes were treated with ACAT inhibitor and ox-LDL at different concentrations for 48 h,the cholesterol concentration and the expressions of GRP78 and CHOP protein in adipocytes and the visfatin levels in the supernatant fluid were increased with the increase of the ox-LDL concentration.The differences had statistical significance in the experimental groups compared with blank control group( all P＜0.05 ).After the intervention with ACAT inhibitor and ox-LDL for different durations,the expressions of GRP78 and CHOP protein in adipocytes and the visfatin levels in the supernatant fluid were up-regulated in a timedependent manner.The differences between experimental groups and blank control had statistical significance( all P＜0.05 ).After the intervention with ACAT inhibitor,ox-LDL,and different concentrations of TUDCA for 48 hours,the expressions of GRP78 and CHOP protein in adipocytes and visfatin levels in the supernatant fluid were down-regulated in a dose-dependent manner and as compared with blank control group the difference were statistically significant( all P＜ 0.05 ).The increase of cholesterol load in adipocyte may promote the visfatin secretion,denoting that the mechanism might be due to the enhancement of endoplasmic reticulum stress in aidpocytes.

Cloning and sequence analysis of V_H and V_L gene of anti-CD3 McAb / 中国免疫学杂志

Objective:To amplify and sequence of the variable region genes of anti CD3 McAb.Methods:The V H?V L genes were amplified by RT PCR from total RNA that were extracted from WuT3 hybridoma.Recombinant cloning vector was constructed and sequenced after the enzyme digestion.Results:It showed that V H gene consisted of 363 bp encoding amine acid residues,belongs to mouse heavy chain subgroup IIB;V L gene consisted of 330 bp encoding amine acid residues,belongs to mouse ? light chain subgroup III.Comparing with Kabat database,the V H?V L genes were in agreement with the characterization of DNA sequences present in the mouse Ig V H?V L regions respectively.Conclusion:The success of cloning of the V H?V L genes of WuT3 McAb lay a good foundation for the construction and expression of chimeric antibody.