The clinical characteristics and survival study of patients with hepatogenic diabetes / 重庆医学

Objective To investigate the clinical characteristics and living conditions of hepatic diabetes (HD) patients and to analyze the risk factors of the prognosis of HD patient .Methods One hundred and sixty‐eight cases of cirrhosis cases were divided into liver cirrhosis group(n=74) ,the combined group cases(n=48)and combined HD type 2 diabetes (T2MD) group(n=46)se‐lected from January 2011 to December 2012 .Death was end point .The clinical features and prognosis of patients were compared . The risk factors for liver cirrhosis were analyzed by the simple factor analysis and Logistic regression analysis .Results The pa‐tients with history of diabetes in T2MD group were more than those in the HD group and the cirrhosis group (P0 .05) .The total bilirubin (TBil) of HD group was higher than those of the T2MD group and liver cirrhosis group(P<0 .05) .The levels of fasting glucose levels(FPG) ,insulin resistance index (HOMA‐IR) ,total cholesterol (TG) ,C‐peptide of liver cirrhosis group were lower than HD group and T2MD group (P<0 .05) .The levels of FPG ,C peptide of HD group were higher than that of T2MD group (P<0 .05) .The levels of TG of T2MD group were higher than that of HD group (P<0 .05) .Logistic multivariate analysis showed that age ,alcohol consump‐tion ,diabetes ,Child‐Pugh score were independent risk factors for death in patients with liver cirrhosis .Conclusion The morbidity and mortality of HD patients may be higher than patients with liver cirrhosis ,and HD could increase the risk of death in patients with liver cirrhosis .

Purification of recombinant human antithrombin III expressed in a goat mammary bioreactor / 生物工程学报

Antithrombin III (AT III) is the most important anti-clotting substance. Recombinant human antithrombin III (rhAT III) expressed in transgenic goat milk attracts more and more attention. Develop an effective purification route for rhAT III is vital to its industrial production. An efficient purification method was developed for the rapid purification of rhAT III by isoelectric precipitation and heparin affinity chromatography. First, casein was effectively removed by isoelectric precipitation. rhAT III was further purified by heparin affinity chromatography. In the process of heparin affinity chromatography, the effects of pH and temperature on the stability of rhAT III were studied, and the effects of operating conditions, elution gradient, flow rate and sample loaded, on the purification efficiency were also studied. Under the optimized conditions, the protein recovery of rhAT III was about 90% with purity over 99%, while its activity recovery was about 50%. Such a purification process is very simple and effective, and it would provide a valuable reference for the further scaling-up of industrial production.

Purification and characterization of recombinant human lactoferrin expressed in a cattle mammary bioreactor / 生物工程学报

Novel ion exchange adsorbents were synthesized by immobilizing sulfopropyl derivative onto homemade highly cross-linked agarose beads. The effects of different ligand densities (from 0.05 to 0.24 mol/L) on static and dynamic adsorption of the adsorbents were investigated using lysozyme as a model protein. Based on these results, rHLF was purified from the transgenic milk by our SP media. 1 mL high density (0.24 mol/L) adsorbent could handle 50 mL rHLF-containing milk. The mass recovery of rHLF was 86.5% and the purity was 98.5%. CD spectra demonstrated that the native structure of rHLF was not affected in the purification process. The biological functions of the purified rHLF, including iron binding, releasing and antimicrobial activities were then investigated. The results showed that rHLF had comparable iron binding and releasing activity to that of native HLF. 5 g/L concentration of rHLF significantly inhibited the growth of Escherichia coli. These studies lay a solid foundation for the wide application of our self-prepared ion exchange adsorbents in protein purification.

Effect of affinity medium and solution conditions on endotoxin removal from protein solutions / 生物工程学报

Endotoxin removal is essential for the safety of biological products. To remove endotoxin efficiently, we used polymyxin B (PMB) affinity adsorbent to remove endotoxin from protein solutions by static adsorption. We studied the effects of spacer length and ligand density of the affinity adsorbent, pH, salt type and concentration, protein type and concentration, endotoxin concentration, and additive on endotoxin removal and protein recovery. Endotoxin content and protein concentration were determined by test and Lowry assay respectively. The results showed that PMB affinity adsorbent had high capacity, high adsorption speed, high removal efficiency and good reusability. In addition, ligand density, pH, salt concentration and the isoelectric point and hydrophobicity of protein all had remarkable influence on the endotoxin removal. Under the optimal conditions, the recoveries of hemoglobin, human serum albumin and lysozyme were 87.2%, 73.4% and 97.3%, respectively, and the corresponding endotoxin removal rates 99.8%, 97.9% and 99.7%, respectively. This study illustrated the effects of solution conditions on the efficiency of endotoxin removal and protein recovery, and would provide useful reference for the efficient removal of endotoxin from biological products.

Effects of solvent environment on the structure of hepatitis B surface antigen (HBsAg) / 生物工程学报

As a virus-like particle, hepatitis B surface antigen (HBsAg) was the primary component of hepatitis B vaccine. HBsAg was maintained by the non-covalent interaction of proteins and lipids. The intact structure of HBsAg particle was vital to its function. However, there was no report about the effects of solvent environment on HBsAg structure. In this paper, we studied the effects of temperature, pH, ionic type and salt concentration on HBsAg structure. The results showed that HBsAg was stable at normal temperature, but began to denature above 60 degrees C. The aggregation of HBsAg at pH 3.0 and 4.0 was nearly irreversible, but partly reversible at pH 5.0. The influence of ionic type on HBsAg was generally in accordance with Hofmeister sequence, except that SO4(2-) caused more aggregation than F-. HBsAg aggregates started to be visible in 0.4 mol/L (NH4)2SO4, and the extent of aggregation increased with the salt concentration. Therefore, caution must be taken when using (NH4)2SO4 in the hydrophobic chromatography purification of HBsAg.

Effect of solution environment on the purification of pertussis toxin / 生物工程学报

The low recovery of pertussis toxin (PT) and the low resolving efficiency of chromatography, due to the instability of PT in low salt condition, are the main challenges for its purification. We aplied 2 mol/L urea to prevent the aggregation and disassociation of PT during the purification by ion-exchange chromatography (IEC) and gel filtration chromatography (GFC). The effect of urea on the purification of PT was studied by ELISA assay and non-reduced SDS-PAGE. The activity recoveries of PT and filamentous hemagglutinin (FHA) in IEC and GFC, the resolution efficiency in GFC and the purities of PT and FHA were improved obviously by adding 2 mol/L urea in the mobile phase. The results highlight the potential application of urea in the acellular pertussis vaccine (APV) manufacture procedure.