Influence of different levels of heparin and antithrombin Ⅲ on human coagulation factor Ⅸ activity / 中国输血杂志

【Objective】 To study the effect of different concentrations of heparin, ATⅢ or a mixture of heparin and antithrombin Ⅲ (ATⅢ) (1∶1)on the activity of human coagulation factor Ⅸ (FⅨ). 【Methods】 The heparin or heparin/ATⅢ with different concentrations were added into human coagulation Ⅸ products or human prothrombin complex (PCC) to prepare heparin or heparin/ATⅢ samples, containing 0, 0.1, 0.3, 0.5, 0.8, 1, 2 and 4 IU per unit. ATⅢ with different concentrations were added into FⅨ or PCC to prepare ATⅢ samples containing ATⅢ 0, 0.1, 0.5 and 1 IU per unit. The FⅨ activity of the samples prepared was tested by one-stage coagulation method. Then corresponding amount of protamine sulfate were added to neutralize heparin or heparin/ATⅢ to detect the FⅨ activity again. Their influence of heparin, ATⅢ and heparin/ATⅢ with different concentrations on the activity of FⅨ were analyzed. 【Results】 When the content of heparin or heparin/ATⅢ was 0, 0.1, 0.3 and 0.5 IU per unit of FⅨ, the detection results of FⅨ titer in samples were consistent. When the content of heparin or heparin/ATⅢ per unit of FⅨ was 0.8, 1, 2 and 4 IU, the detection results of FⅨ titer were all lower than those of samples without heparin. When the ATⅢ content was 0, 0.1, 0.5 and 1 IU, the FⅨ titer of the samples was consistent. 【Conclution】 When the content of heparin or heparin/ATⅢ in the product is less than or equal to 0.5 IU per IU of FⅨ, the step of protamine sulfate adding could be omitted as it has little effect on FⅨ activity. When >0.5 IU per IU of FⅨ, however, protamine sulfate adding, to neutralize heparin, is necessary before FⅨ activity testing.

Study on separation and purification process of C1 esterase inhibitor / 中国输血杂志

【Objective】 To study the technology of separating and purifying C1 esterase inhibitor (C1-INH) by using the waste washing liquid as raw materia during the preparation of human prothrombin complex (PCC) l. 【Methods】 C1-INH was isolated and purified by a two-step method of polyethylene glycol (PEG) 4000 precipitation and cation chromatography. The pH of raw materials and the concentration of PEG4000 were adjusted to investigate the optimal conditions of PEG4000 precipitation method. After PEG was precipitated and centrifuged, the supernatant is treated as the loading solution for cation exchange chromatography, using Fractogel EMD SE HiCap(M) gel and CM Sepharose FF gel for ion exchange chromatography. The most suitable gel and separation conditions were selected by comparing the C1-INH antigen yield, activity yield and specific activity. 【Results】 Under the condition of pH 6.1, when the mass fraction of PEG4000 was 14%, the recovery rate of C1 esterase inhibitor was close to 70%, and the removal rate of ceruloplasmin was more than 95% after stirring for 10 minutes. As fractogel EMD SE HiCap(M) gel was used for cation exchange chromatography, when the eluent salt concentration was 0.25 M sodium chloride, the activity yield of C1 esterase inhibitor was greater than 80%, and the specific activity was greater than 5 IU/mg. 【Conclusions】 Using the waste washing liquid as the raw material during the preparation of PCC, the C1 esterase inhibitor with high specific activity can be prepared through PEG precipitation and purification by Fractogel EMD SE HiCap(M) ion exchange chromatography.

Functional Analysis of SI10875 in the Regulation of PS I Activity in Synechocystis sp.PCC 6803 / 中国生物化学与分子生物学报

Phylloquinone is a unique cofactor of photosystem I (PS I ) , made up of a redox-active naphthoquinone ring attached to a partially saturated C-20 phytyl side chain.At present, the research on the biosynthesis of phylloquinone in cyanobacteria is mainly focused on the formation of naphthoquinone ring, while there was a shortage of reports in the biosynthesis of phytyl side chain.In this study, a highly homologous protein S110875 was found in Synechocystis sp.PCC 6803 by homologous sequence alignment with VTE6, a kinase involved in phylloquinone biosynthesis by converting phytyl-phosphate into phytyl- diphosphate in Arabidopsis thaliana.The resulting S110875 mutant, called As/Z0875, accumulates none phylloquinone and tocopherol, as well as low amounts of chlorophyll content (P<0.05).The mutant had retarded growth in the absence of added glucose.Chlorophyll fluorescence, P700 absorbance changes, 77 K fluorescence emission spectra and Western blot analyses showed that in As/Z0875, PS I function was impaired and accumulation of the PS I complex was reduced remarkably (P<0.01), indicating that phvlloquinone deficiency affected PS I function, thus hindering the normal growth of cyanobacteria.Our results provide the evidence that the phytol phosphorylation pathway is essential for phylloquinone biosynthesis in cyanobacteria for the first time, and a basis for further investigate the protein synthesis, assembly and stability of PS I complex in cyanobacteria.

PCC0208017, a novel small-molecule inhibitor of MARK3/MARK4, suppresses glioma progression and

Gliomas are the most common primary intracranial neoplasms among all brain malignancies, and the microtubule affinity regulating kinases (MARKs) have become potential drug targets for glioma. Here, we report a novel dual small-molecule inhibitor of MARK3 and MARK4, designated as PCC0208017. PCC0208017 strongly inhibited kinase activity against MARK3 and MARK4, and strongly reduced proliferation in three glioma cell lines. This compound attenuated glioma cell migration, glioma cell invasion, and angiogenesis. Molecular mechanism studies revealed that PCC0208017 decreased the phosphorylation of Tau, disrupted microtubule dynamics, and induced a G2/M phase cell cycle arrest. In an glioma model, PCC0208017 showed robust anti-tumor activity, blood-brain barrier permeability, and a good oral pharmacokinetic profile. Molecular docking studies showed that PCC0208017 exhibited high binding affinity to MARK3 and MARK4. Taken together, our study describes for the first time that PCC0208017, a novel MARK3/MARK4 inhibitor, might be a promising lead compound for treatment of glioma.

Effect of GSH and niacin combination on protein oxidation, ER stress, glycation and aggregation in HLE cells under high glucose condition / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM:To evaluate the effects of reduced glutathione(GSH)and niacin combination on protein oxidative stress, endoplasmic reticulum(ER)stress, glycation, and aggregation of the αβ crystalline in human lens epithelial(HLE)cells treated with high glucose levels. <p>METHODS:HLE cells were cultured and exposed to 25 mmol/L glucose to promote high glucose conditions. Groups of cells were co-treated with three different combinations of dosages: 10 μmol/L GSH+2 μmol/L niacin, 30 μmol/L GSH+25 μmol/L niacin, and 100 μmol/L GSH+25 μmol/L niacin. After 72h incubation, protein carbonyl content(PCC)and glucose reactive protein(GRP78)content were assessed using ELISA examinations. After two-week incubation, advanced glycation end products(AGEs)were also assessed and the expression of αβ crystalline was measured using Western Blot examination. The SPSS 18.0 statistical package was used for all data analyses.<p>RESULTS:PCC and GRP78 levels in the co-treated groups were not significantly reduced compared to control(<i>P</i>>0.05). In contrast, there was a significant decrease of the AGEs levels in all groups co-treated with GSH and niacin when compared with the control group(<i>P</i><0.05). In addition, the αβ crystalline expression increased after high dose glucose administration, but decreased in all groups co-treated with GSH and combinations of GSH and niacin.<p>CONCLUSION:The results suggest that combinations of GSH and niacin inhibit the aggregation of proteins and prevent glycation in hyperglycemic human lens epithelial cells. This study shows that this combination may play an active role in preventing diabetic cataract mainly from the AGEs pathway.

Construction of T7 RNA polymerase gene expression system in Anabaena sp. PCC 7120 for the expression of hG-CSF / 生物工程学报

The low expression rate of exogenous genes in cyanobacteria is one of the bottlenecks of cyanobacteria genetic engineering. The T7 RNA polymerase expression system has achieved the efficient expression of exogenous genes in Escherichia coli. Cyanobacteria and E. coli are both Gram-negative bacteria with high genetic homology. The construction of T7 RNA polymerase expression system in cyanobacteria may improve the expression of foreign genes. In order to construct the T7 RNA polymerase expression system in Anabaena sp. PCC 7120, methods such as overlapping extension PCR and digestion-ligation technique were used to construct a site-specific integration vector pEASY-T1-F1-TacT7RNAPCmR-F2 and a shuttle expression vector pRL-T7-hG-CSF. The site-specific integration vector is capable of expressing T7 RNA polymerase, and the shuttle expression vector expresses hG-CSF driven by the T7 promoter. Then we introduced the site-specific integration vector into the wild type cyanobacteria by electroporation and transferred the shuttle expression vector into the site-integrated transgenic cyanobacteria by triparental conjugative transfer. In the end, we identified the presence of foreign genes in cyanobacteria by PCR, tested the transcription level of foreign genes in cyanobacteria by RT-PCR, and detected the protein expression of foreign genes in cyanobacteria by Western blotting. The two vectors were successfully constructed, the T7 RNA polymerase gene and hG-CSF gene were transferred into cyanobacteria well, and both genes were also expressed in cyanobacteria. In summary, the T7 RNA polymerase expression system was successfully constructed in cyanobacteria, and the expression rate of hG-CSF gene was doubled than the traditional cyanobacteria expression systems. This expression system will provide a better tool for the application of cyanobacteria genetic engineering and will promote the development of cyanobacteria as a chassis cell in the fields of synthetic biology in the future.

Panorama sobre a regulamentação do sistema APPCC em estabelecimentos de carne de aves sob inspeção federal / The regulation of the HACCP system in poultry establishments under federal inspection

Desde 2006, o Ministério da Agricultura, Pecuária e Abastecimento (MAPA) vem desenvolvendo regulamentação específica para aplicação do sistema de Análise de Perigos e Pontos Críticos de Controle (APPCC) nos matadouros de aves sob inspeção federal, incluindo diretrizes para a implantação desse programa de qualidade nas indústrias. Este artigo apresenta um panorama sobre a evolução dessa regulamentação, atualmente estagnada, e os métodos vigentes para fiscalização da aplicação do sistema APPCC em estabelecimentos de carne de aves. Dessa forma, os autores buscam facilitar a compreensão do processo regulatório por pesquisadores e técnicos da área e, com isso, incentivar esses profissionais a contribuir para o aperfeiçoamento das diretrizes governamentais.

Effects of light quality on cell growth and psbA promoter of engineered Synechococcus sp. PCC7002 / 生物工程学报

Light quality can regulate both psbA genes and vector promoter psbA of the engineered Synechococcus. Through light regulation, we tried to improve yield of the recombinant protein for vp28 gene-expressed Synechococcus sp. PCC7002. To drive photon-capturing efficiently, three limiting factors (irradiance, temperature and pH) were optimized by measuring net photosynthesis. High cell density cultures were performed with variant ratios of white, red and blue light in a 5-L photo-bioreactor. Yields of biomass, expressions of vp28 and transcription levels of psbA were compared. High ratio blue light-induced vp28 transcription had tripled and the relative accumulation of VP28 protein was doubled. The relative expressions of psbAII and psbAIII had positive correlations with higher ratio of blue light, not the red light. With high ratio red light inducing, dry biomass reached 1.5 g/L in three days. Therefore, we speculated that red light accelerated biomass accumulation of the transgenic strain and blue light promoted transcription for PpsbA and psbA. These results provided useful information for mass production of cyanobacteria and its secondary metabolites.

The correlation study between early postsurgery phonetic acquisition of speech stop and speech outcomes of the children aged 30 months after cleft palate repair / 实用口腔医学杂志

Objective:To study the correlation between early postsurgery phonetic acquisition of speech stop and speech outcomes of young children.Methods:28 children with cleft palate were included in the study.An one-stage palatal repair procedure was per-formed by one surgeon for the children before the age of 1 8 months.Naming tests were used in the speech therapy room when the chil-dren were aged 23 months and 30 months.Stop consonant inventory number,percent correct consonants(PCC),percent correct man-ners(PCM)and percent correct places(PCP)were analyzed.Results:Number of stop consonant was significantly correlated with PCC,PCM and PCP at 24 and 30 months of age in the children.Coefficient of determination between stop consonant number and PCC was 0.535.Conclusion:Speech stop may be used as the “sensitive sound”for the analysis of speech development of the chinese children aged 2-3 years after cleft palate repair and as the individuation guideline to determine the best assessment and therapy time.

Health reform in grass-roots units of China according to the monitoring data of public opinion during NPC and PCC / 中华医学图书情报杂志

In order to show the social situation and public opinions for the decision-makers of medical reform, the viewpoints of main media, NPC and PCC members, and Internet users were analyzed according to the real-time monitoring data of public opinions on health-related topics ( new rural cooperative medical system, basic public health service, and rural medical workers) during the NPC and PCC, which showed the public awareness of health reform in grass-roots units of China.

Study on the inlfuening faltors of altivity detemination of four coagulation factors in human prothrombin complex concentrates / 中国生化药物杂志

Objective To study the inlfuence factors on detection of activity of four coagulation factors in prothrombin complex concentrates (PCC) by several factors. Methods Using Pharmacopoeia of the People’s Republic of China (2010) as reference, the activity of four coagulation factors in PCC were investigated by choosing different pre-treatments, different diluents, different salt concentration, different standard human plasma and different company reagents. Results The activity of FII, FVII, FX were decreased and FIX was increased in the condition of adding protamine sulfate, and there were no differences of four coagulation factors whether warm bath in 37 for 15 min or not. However, the differences of four coagulation factors were significant by using deficient plasma, saline, distilled water and commercial dilution buffer(P<0.05). The activity of coagulation factor II, X in 1 mol/L salt concentration of PCC were significantly lower than in 0.25 mol/L(P<0.05), while coagulation factor VII, IX were not. The activity of FII, FVII, FIX, and FX were different by using different standard human plasma to make standard curve. The activity of four coagulation factors existed significant difference(P=0.00) by using SIEMENS company reagents and domestic reagents. Conclusion Choosing different pre-treatments, different dilution buffers, salt concentration, standard human plasma and commercial kits will inlfuence the detection result of coagulation factors.

Utilidad diagnóstica del anticuerpo antipéptidocíclico citrulinado como prueba diagnóstica enpacientes con artritis reumatoide / Diagnostic utility of antibody to cyclic citrullinated peptide as a diagnostic test for rheumatoid arthritis patients

Introducción: La artritis reumatoide es una enfermedad autoinmune sistémica, inflamatoria y crónica. En el diagnóstico confirmatorio la presencia del anticuerpo contra el péptido cíclico citrulinado parece tener mejor especificidad que el factor reumatoide. Objetivo: Evaluar las características operativas de los anticuerpos contra el péptido cíclico citrulinado y factor reumatoide en pacientes con artritis reumatoide. Métodos: Estudio retrospectivo convencional de pruebas diagnósticas, en 68 pacientes con artritis reumatoide, y 68 con patologías ortopédicas. El diagnóstico de la artritis se hizo a través de los criterios del Colegio Americano de Reumatología. La determinación del anticuerpo contra el péptido cíclico citrulinado se realizó por electroquimioluminiscencia y factor reumatoide por inmunoturbidimetría. Para el análisis de datos se usó el software R. A través de tabulaciones cruzadas se determinaron las características operacionales del método. Se construyó la gráfica de Receiver Operating Curve. Resultados: La sensibilidad, especificidad, valor predictivo positivo y negativo del anticuerpo contra el péptido cíclico citrulinado fueron 81%, 92%, 90% y 82%, respectivamente. Para el factor reumatoide los resultados fueron similares, excepto en la especificidad (90%). Con cualquiera de las dos pruebas positivas la sensibilidad alcanza 89% y la especificidad 86%, con razón de verosimilitud positiva de 6,35. La positividad simultánea de ambas pruebas disminuye la sensibilidad a 67% y la especificidad aumenta 100% con razón de verosimilitud negativa de 0,33. Conclusión: Por la sensibilidad y especificidad del anticuerpo contra el péptido cíclico citrulinado es considerado como una prueba útil para el diagnóstico temprano. El uso combinado de las dos pruebas aumenta significativamente la sensibilidad, especificidad y razón de verosimilitud positiva.

Introduction: Rheumatoid arthritis is a systemic autoimmune, inflammatory and chronic disease. In the confirmatory diagnosis, the presence of anti-cyclic citrullinated peptide antibody seems to have better specificity than rheumatoid factor. Objective: To evaluate the operating characteristics of anti cyclic citrullinated peptide antibody and rheumatoid factor in patients with rheumatoid arthritis. Methods: Retrospective conventional diagnostic test study in 68 patients with rheumatoid arthritis and 68 with orthopedic conditions.The diagnostic was made using the American College of Rheumatology criteria. Serum levels of antibody to cyclic citrullinated peptide were determined by electrochemiluminescence and rheumatoid factor by inmunoturbidimetry. Data analysis was used software R. Through cross tabulations, operational characteristics of the method where determined. Receiver Operating Curve grahp was constructed. Results: The sensitivity, specificity, positive and negative predictive value of antibody to cyclic citrullinated peptide were 81%, 92%, 90% and 82% respectively. Similar results for rheumatoid factor except in specificity (90%). With either of the two positive tests the sensitivity reached 89 and specificity 86% and the positive likelihood ratio for these data was 6.3. The simultaneous positivity of both tests decreases the sensitivity to 67% and specificity increases to 100%, and negative likelihood ratio of 0.33. Conclusion: For the sensitivity and specificity of the antibody against cyclic citrullinated peptide test is considered useful for early diagnosis. The combined use of both tests significantly increases the sensitivity, specificity and positive likelihood ratio.

Caracterização dos usuários com esquizofrenia e outros transtornos psicóticos dos Centros de Atenção Psicossocial. / Characterization of the users with schizophrenia in Psychosocial Care Centers.

A rede de Saúde Mental de base comunitária no Brasil ampliou a oferta de serviços substitutivos para a população com doença mental grave. O objetivo deste estudo foi caracterizar o perfil do usuário com transtornos psicóticos nos Centros de Atenção Psicossocial do Estado de Sergipe (CAPS-SE). Foi conduzido um estudo descritivo, com análise quantitativa dos dados sociodemográficos e número de internação. Foram estudados 1.444 prontuários com diagnóstico de transtorno psicótico e encontraram-se 75,3% deles com diagnóstico de esquizofrenia e 24,7% com outros transtornos psicóticos. Os usuários com menos de 24 anos representavam 33% dos usuários, enquanto 54% estavam igualmente distribuídos nas faixas etárias de 25-35 e 35-46 anos. Dos sujeitos com diagnóstico de esquizofrenia, 79,9% eram do gênero masculino e 68%, feminino, mostrando relação significativa entre gênero e diagnóstico. A maioria absoluta dos sujeitos não tem profissão. Quanto ao número de internações, a diferença entre sujeitos diagnosticados como esquizofrênicos para os portadores de outros diagnósticos aponta predominância significativa do primeiro grupo na faixa com mais de seis internações. Conclui-se que entre os usuários dos CAPS é predominante o gênero masculino, com comprometimento importante também na dimensão profissional.

Community based network of Mental Health in Brazil increased the supply of substitute services for the population with grave mental disorders. The objective of this study was to characterize the profile of the user with psychotic disorders in Psychosocial Care Centers (PCC) in the state of Sergipe, Brazil. A descriptive study was carried out by means of a quantitative analysis of the sociodemographic and number of committal data. In this study, 1,444 patients? records with psychotic disorder diagnosis were analysed. It was revealed that 75.3% were diagnosed with schizophrenia and whereas 24.7% with other psychotic disorders. Users with less than 24 years represented 33% of the subjects, while 54% were equally distributed in the age groups 25-35 and 35-46 years. It was shown that among the users diagnosed with schizophrenia, 79.9% belonged to the male gender and 68.7% to the feminine gender. The absolute majority of the subjects have no occupation. Concerning the number of hospitalizations, the difference between subjects diagnosed with schizophrenia for patients with other diagnoses indicates significant predominance more than first group in the range of six admissions. It was concluded that among the users of the PPC with schizophrenia the male gender is prevalent, which also signals impairment towards the professional dimension.

Cloning and High Expression Anabaena Class-II Fructose-1, 6-bisphosphate Aldolase Gene in Escherichia coli / 中国生物工程杂志

The complete genomes of more cyanobacterial strains have been completed for sequence, and genetic engineering of cyanobacteria has evolved in the post-genome era. Since Kaneko and colleagues had completed the sequence for the complete genome of Anabaena sp. PCC 7120 in 2001, functions of some genes in this genome, including fructose-1,6-bisphosphate aldolase (FBA) gene, have been predicated using the method of bioinformatics. However, little information is available regarding whether this gene can encode FBA and its product characteristics of related enzyme. Here, to explore this information, the predicted II-FBA gene-encoding region in Cyanobase database was cloned by PCR method and then ligated into pET-32a to generate the expression vector, pET-FBA-II. The results of SDS-PAGE indicated that the expression level of the expected target polypeptide was approximate 23.4 percent as compared to total protein and the molecular weight is about 40 kDa as compared to the protein molecular marker. The results of enzyme activity analysis showed that the activity of II-FBA was ~11.8 U per mg protein and owned a standard activity of II-FBA. To sum up, the results not only prove the functional prediction of this II-FBA gene from the Cyanobase database, but also provide the important conditions for further studying its physiological and biochemical characteristics and functions of the gene expression product.

Effect of Synechococcus sp.PCC7942 with trans-mutated hCu,Zn-SOD-gene on antioxidation activities in mice / 中国海洋药物

Objective To study the biological activities of Synechococcus sp.PCC7942 with trans-mutated hCu,Zn-SOD-gene.Methods Synechococcus sp.PCC7942 with trans-mutated hCu,Zn-SOD were administered orally for 20d to mice,then the activities of glutathione peroxidase(GSH-Px),catalase(CAT),superoxide dismutase(SOD) and the content of malondialdehyde(MDA) were determined.Results The activities of GSHPx in serum and the activities of CAT in blood increased obviously;the activity of SOD in serum and liver increased markedly;the content of MDA in serum and liver decreased obviously.Conclusion Synechococcus sp.PCC7942 with trans-mutated hCu,Zn-SOD-gene had obvious antioxidant effect in vivo.

Preliminary Studies on the Growth Potential of Transgenic Synechococcus sp.PCC 7002 with Mouse Metallothionein-ⅠGene / 微生物学通报

The growth predominance and culturing potential of transgenic Synechococcus sp.PCC 7002 with mouse metallothionein-Igene are characterized and expatiated by room temperature absorption spectra,photosynthetic oxygen evolution rate,growth kinetic parameters.The results show that the spectral absorption,maximum net photosynthetic rate and saturated light intensity of transgenic Synechococcus sp.PCC 7002 are higher than wild strain,whereas its the respiration rate and compensating light intensity are lower than wild strain.The transgenic cells exhibit a higher growth rate,and the maximum cell concentration is 1.74 times higher than wild ones when cultured in shaking flask.The air lift photobioreactor is suitable for bringing its growth potential into play.

Identification of a Pair of Toxin-antitoxin (TA) Gene in the Chromosome of Cyanobacteria Synechocystis sp. PCC6803 / 微生物学通报

Chromosomally encoded toxin–antitoxin (TA) systems are thought to result in growth arrest and eventual cell death upon exposure to environmental stress in E. coli. In the chromosome of cyanobacteria Synechocystis sp. PCC6803, the genetic organization of a 360 bp open reading frame (ORF), slr0664, and another small ORF of 256 bp, ssr1114, is similar to that of TA system. The predicted protein encoded by slr0664 is homologous to RelE, but neither homologue of ssr1114 nor ssr1114-encoding protein was found in TA system. To see whether slr0664 encodes a toxin protein, ssr1114 encodes an antitoxin, an expressing plasmid containing promoter Plac and PBAD, was constructed. In this construct, Both slr0664 and ssr1114 were controlled by Plac and PBAD, respectively. Expression of slr0664 in Escherichia coli results in the inhi-bition of bacterial growth, the expression of ssr1114 neutralize the toxicity of slr0664 expression. These re-sults show that slr0664 is toxin gene and ssr1114 is antitoxin gene, both ssr1114 and slr0664 constitute achromosomal TA system in Synechocystis sp. PCC6803.