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921.
干扰素α2b(interferonα2b,IFNα2b)是一种用于病毒性疾病和肿瘤性疾病治疗的多功能细胞因子,因其在体内的半衰期短限制了其在临床上的应用。将IFNα2b连接到人血清白蛋白(human serum albumin,HSA)的C端,构建融合蛋白HSA-IFNα2b。构建含融合蛋白的真核表达质粒p MH3/HSA-IFNa2b,经电转的方法转入中国仓鼠卵巢(Chinese hamster ovary,CHO)细胞中。经G418抗性压力筛选和目的蛋白的表达量筛选,最终获得一株高表达的稳定细胞株(CHO/p MH3/HSA-IFNa2b)。表达的目的蛋白经Western blot验证显示,产物具有IFNα2b和HSA的双抗原性。经悬浮驯化稳定后,通过批次筛选得到一株稳定的高克隆表达株,产量约为65mg/L,进一步选取高表达克隆株在悬浮驯化中不同代数进行批次培养,不同代数之间蛋白质的表达量和生长情况没有明显的差异,获得一株稳定遗传表达的单克隆细胞株,3L摇瓶的流加培养结果显示,最佳发酵时间为15天,蛋白质表达量为121mg/L。经离心获得的发酵液,经两步纯化后获得蛋白质纯度高达96.8%的目的蛋白,总回收率为22.3%。参照《中国药典》2015版对IFNα2b的检测方法,结果显示,CHO表达的HSA-IFNα2b比活性为4.16×106IU/mg。首次将HSA-IFNα2b在哺乳动物细胞CHO中构建表达,表达获得高活性的HSA-IFNα2b融合蛋白。 相似文献
922.
Combined metabolomics and proteomics reveals hypoxia as a cause of lower productivity on scale‐up to a 5000‐liter CHO bioprocess
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Yuanwei Gao Somak Ray Shujia Dai Alexander R. Ivanov Nicholas R. Abu‐Absi Amanda M. Lewis Zhuangrong Huang Zizhuo Xing Michael C. Borys Zheng Jian Li Barry L. Karger 《Biotechnology journal》2016,11(9):1190-1200
Large‐scale bioprocessing is key to the successful manufacturing of a biopharmaceutical. However, cell viability and productivity are often lower in the scale‐up from laboratory to production. In this study, we analyzed CHO cells, which showed lower percent viabilities and productivity in a 5‐KL production scale bioreactor compared to a 20‐L bench‐top scale under seemingly identical process parameters. An increase in copper concentration in the media from 0.02 µM to 0.4 µM led to a doubling of percent viability in the production scale albeit still at a lower level than the bench‐top scale. Combined metabolomics and proteomics revealed the increased copper reduced the presence of reactive oxygen species (ROS) in the 5‐KL scale process. The reduction in oxidative stress was supported by the increased level of glutathione peroxidase in the lower copper level condition. The excess ROS was shown to be due to hypoxia (intermittent), as evidenced by the reduction in fibronectin with increased copper. The 20‐L scale showed much less hypoxia and thus less excess ROS generation, resulting in little to no impact to productivity with the increased copper in the media. The study illustrates the power of 'Omics in aiding in the understanding of biological processes in biopharmaceutical production. 相似文献
923.
Fengjuan Zhang Donghai Peng Chunsheng Cheng Wei Zhou Shouyong Ju Danfeng Wan Ziquan Yu Jianwei Shi Yaoyao Deng Fenshan Wang Xiaobo Ye Zhenfei Hu Jian Lin Lifang Ruan Ming Sun 《PLoS pathogens》2016,12(1)
Cell death plays an important role in host-pathogen interactions. Crystal proteins (toxins) are essential components of Bacillus thuringiensis (Bt) biological pesticides because of their specific toxicity against insects and nematodes. However, the mode of action by which crystal toxins to induce cell death is not completely understood. Here we show that crystal toxin triggers cell death by necrosis signaling pathway using crystal toxin Cry6Aa-Caenorhabditis elegans toxin-host interaction system, which involves an increase in concentrations of cytoplasmic calcium, lysosomal lyses, uptake of propidium iodide, and burst of death fluorescence. We find that a deficiency in the necrosis pathway confers tolerance to Cry6Aa toxin. Intriguingly, the necrosis pathway is specifically triggered by Cry6Aa, not by Cry5Ba, whose amino acid sequence is different from that of Cry6Aa. Furthermore, Cry6Aa-induced necrosis pathway requires aspartic protease (ASP-1). In addition, ASP-1 protects Cry6Aa from over-degradation in C. elegans. This is the first demonstration that deficiency in necrosis pathway confers tolerance to Bt crystal protein, and that Cry6A triggers necrosis represents a newly added necrosis paradigm in the C. elegans. Understanding this model could lead to new strategies for nematode control. 相似文献
924.
925.
Xinqing Lee Yimin Huang Daikuan Huang Lu Hu Zhaodong Feng Jianzhong Cheng Bing Wang Jian Ni Tserenpil Shurkhuu 《PloS one》2016,11(3)
Variation of soil organic carbon (SOC) and its major constraints in large spatial scale are critical for estimating global SOC inventory and projecting its future at environmental changes. By analyzing SOC and its environment at 210 sites in uncultivated land along a 3020km latitudinal transect in East Central Asia, we examined the effect of environmental factors on the dynamics of SOC. We found that SOC changes dramatically with the difference as high as 5 times in north China and 17 times in Mongolia. Regardless, C:N remains consistent about 12. Path analysis indicated that temperature is the dominant factor in the variation of SOC with a direct effect much higher than the indirect one, the former breaks SOC down the year round while the latter results in its growth mainly via precipitation in the winter half year. Precipitation helps accumulate SOC, a large part of the effect, however, is taken via temperature. NH4+-N and topography also affect SOC, their roles are played primarily via climatic factors. pH correlates significantly with SOC, the effect, however, is taken only in the winter months, contributing to the decay of SOC primarily via temperature. These factors explained as much as 79% of SOC variations, especially in the summer months, representing the major constraints on the SOC stock. Soil texture gets increasingly fine southward, it does not, however, constitute an apparent factor. Our results suggested that recent global warming should have been adversely affecting SOC stock in the mid-latitude as temperature dominates other factors as the constraint. 相似文献
926.
Wei Zhu Zanlin Mai Jing Qin Xiaolu Duan Yang Liu Zhijian Zhao Jian Yuan Shaw P. Wan Guohua Zeng 《PloS one》2016,11(2)
Background
Diabetic patients are more likely to develop kidney stones than the general population. The underlying mechanisms for this disparity remain to be elucidated. Little is known about the relationship between urine composition and diabetes mellitus in non-stone-forming individuals. We sought to examine the differences in the 24-hour (24-h) urine composition between diabetic and non-diabetic adults who were not stone formers.Methods
A convenience sample of 538 individuals without a history of nephrolithiasis, gout, hyperparathyroidism, or gastroenteric diseases participated in this study. The 24-h urine profiles of 115 diabetic adults were compared with those of 423 non-diabetic adults. Diabetes was defined by self-reported physician diagnosis or medication use. All participants were non-stone formers confirmed by urinary tract ultrasonography. Participants provided a fasting blood sample and a single 24-h urine collection for stone risk analysis. Student’s t-test was used to compare mean urinary values. Linear regression models were adjusted for age, gender, body mass index, hypertension, fasting serum glucose, serum total cholesterol, estimated creatinine clearance rate and urinary factors.Results
Univariable analysis showed that the diabetic participants had significantly higher 24-h urine volumes and lower urine calcium and magnesium excretions than non-diabetic participants (all P < 0.05). After multivariate adjustment, no significant differences in 24-h urine composition were observed between diabetic and non-diabetic participants except for a slightly increased 24-h urine volume in diabetic participants (all P > 0.05). The main limitation of this study is that the convenience samples and self-reported data may have been sources of bias.Conclusion
Our data showed that there were no differences in 24-h urine composition between diabetic and non-diabetic adults who are not stone formers. The reason for it might be the improved glycemic control in diabetic individuals in our study. Therefore, a tighter glycemic control might reduce stone formation in diabetic adults. 相似文献927.
Yilin Tang Jingjie Ge Fengtao Liu Ping Wu Sisi Guo Zhenyang Liu Yixuan Wang Ying Wang Zhengtong Ding Jianjun Wu Chuantao Zuo Jian Wang 《PloS one》2016,11(4)
PurposeTo characterize cerebral glucose metabolism associated with different cognitive states in Parkinson’s disease (PD) using 18F-fluorodeoxyglucose (FDG) and Positron Emission Tomography (PET).MethodsThree groups of patients were recruited in this study including PD patients with dementia (PDD; n = 10), with mild cognitive impairment (PD-MCI; n = 20), and with no cognitive impairment (PD-NC; n = 30). The groups were matched for age, sex, education, disease duration, motor disability, levodopa equivalent dose and Geriatric Depression Rating Scale (GDS) score. All subjects underwent a FDG-PET study. Maps of regional metabolism in the three groups were compared using statistical parametric mapping (SPM5).ResultsPD-MCI patients exhibited limited areas of hypometabolism in the frontal, temporal and parahippocampal gyrus compared with the PD-NC patients (p < 0.01). PDD patients had bilateral areas of hypometabolism in the frontal and posterior parietal-occipital lobes compared with PD-MCI patients (p < 0.01), and exhibited greater metabolic reductions in comparison with PD-NC patients (p < 0.01).ConclusionsCompared with PD-NC patients, hypometabolism was much higher in the PDD patients than in PD-MCI patients, mainly in the posterior cortical areas. The result might suggest an association between posterior cortical hypometabolism and more severe cognitive impairment. PD-MCI might be important for early targeted therapeutic intervention and disease modification. 相似文献
928.
929.
PurposeDevelop and evaluate an electrochemical method to identify healthy individuals, malignant hematopathic patients and solid tumor patients by detecting the leukocytes in whole-blood.MethodsA total of 114 individual blood samples obtained from our affiliated hospital in China (June 2015- August 2015) were divided into three groups: healthy individuals (n = 35), hematologic malignancies (n = 41) and solid tumors (n = 38). An electrochemical workstation system was used to measure differential pulse voltammetry due to the different electrochemical behaviors of leukocytes in blood samples. Then, one-way analysis of variance (ANOVA) was applied to analyze the scanning curves and to compare the peak potential and peak current.ResultsThe scanning curve demonstrated the specific electrochemical behaviors of the blank potassium ferricyanide solution and that mixed with blood samples in different groups. Significant differences in mean peak potentials of mixture and shifts (ΔEp (mV)) were observed of the three groups (P< = 0.001). 106.00±9.00 and 3.14±7.48 for Group healthy individuals, 120.90±11.18 and 18.10±8.81 for Group hematologic malignancies, 136.84±11.53 and 32.89±10.50 for Group solid tumors, respectively. In contrast, there were no significant differences in the peak currents and shifts.ConclusionsThe newly developed method to apply the electrochemical workstation system to identify hematologic malignancies and solid tumors with good sensitivity and specificity might be effective, suggesting a potential utility in clinical application. 相似文献
930.
Jih-Kai Yeh Yuan-Chuan Hsiao Cian-Ruei Jian Chao-Hung Wang Ming-Shien Wen Chi-Tai Kuo Feng-Chun Tsai Victor Chien-Chia Wu Tien-Hsing Chen 《PloS one》2016,11(1)