Teaching reform and practice of "food enzymology and enzyme engineering" based on cutting-edge researches / 生物工程学报

Food enzymology and enzyme engineering is an important professional course of food science. The course includes the basic theory of enzymology, enzyme engineering technology and the application of enzymes in food industry. Considering the knowledge gap between the teaching contents and the cutting-edge researches, the team constantly adjusted and optimized the course contents to enable students to keep up with state-of-the-art progress by carefully mining the cutting-edge researches. Taking cutting-edge researches as the breakthrough point, we explored the problem-based learning (PBL) teaching model under the guidance of outcome-based education (OBE) concept, and highlighted the importance of the teacher-student and student-student interactions to improve students' enthusiasm and participation. A diversified assessment system was established to evaluate the performance of students in the learning process. The teaching reform consolidated the basic knowledge and expanded the academic frontiers, and fostered students' ability in analyzing problems, designing solutions and achieving team communication. The course may give new insights into the teaching reform of food enzymology and enzyme engineering and other related courses.

Grandes mujeres científicas: Maud Leonora Menten / Great scientific women: Maud Leonora Menten

Maud Leonora Menten nació en Canadá, tuvo cuatro títulos universitarios: Bachiller en Artes, Master en Fisiología, médica y Doctora en Bioquímica. Trabajó en Estados Unidos, Alemania y Canadá. Trabajó en diferentes áreas: en la distribución de los iones cloruro en el sistema nervioso central, en tumores experimentales y su tratamiento con bromuro de radio, en el equilibrio ácido-base durante la anestesia, en el mecanismo hiperglucemiante de toxinas bacterianas, en el descubrimiento de un mecanismo de acoplamiento en química orgánica y hasta en la electroforesis de las hemoglobinas humanas. Sin embargo, el aporte por el cual es más conocida es su trabajo en el estudio de la cinética enzimática junto a Leonor Michaelis en 1913. El propósito de este trabajo es exponer la vida personal y académica de una científica conocida por la gran mayoría de los profesionales de la salud. La mujer que a principios del siglo XX trabajó con grandes investigadores de Canadá, Estados Unidos y Alemania, cuyos aportes científicos fueron reconocidos muchas décadas después. (AU)

Maud Leonora Menten was born in Canada; she had four university degrees, Bachelor of Arts, Master of Physiology, Physician and Doctor of Biochemistry. She worked in the United States, Germany, and Canada. Maud worked in different areas: the distribution of chloride ions in the central nervous system, experimental tumors and their treatment with radium bromide, the acid-base balance during anesthesia, the hyperglycemic mechanism of bacterial toxins, the discovery of a coupling mechanism in organic chemistry and even the electrophoresis of human hemoglobins. However, the contribution for which she is best known is for her work in the study of enzymatic kinetics with Leonor Michaelis in 1913. The aim of this paper is to expose the personal and academic life of a scientist known to the vast majority of Health professionals. The woman who, at the beginning of the 20th century, worked with great researchers from Canada, the United States and Germany, whose scientific contributions were recognized many decades later. (AU)

Cartílago y gonartrosis / Cartilage and knee osteoarthritis

RESUMEN Fundamento: el cartílago articular es un tejido avascular, aneural y alinfático que desempeña un importante papel en la articulación, su afección más frecuente es la de tipo degenerativa. Objetivo: actualizar los conocimientos sobre el cartílago articular normal, envejecido y con cambios degenerativos. Métodos: la búsqueda y análisis de la información se realizó en un periodo de tres meses (primero de octubre al 31 de diciembre de 2018) y se emplearon las siguientes palabras: cartilage AND osteoarthritis, cartilage AND knee osteoarthritis, cartilage a partir de la información obtenida se realizó una revisión bibliográfica de un total de 164 artículos publicados en las bases de datos PubMed, Hinari, SciELO y Medline mediante el gestor de búsqueda y administrador de referencias EndNote, de ellos se utilizaron 50 citas seleccionadas para realizar la revisión, de ellas 48 de los últimos cinco años. Resultados: se mencionan los aspectos macro y microscópicos del tejido, así como su organización por zonas y áreas. Se describen los cambios que ocurren en el proceso degenerativo a diferentes niveles y en el envejecimiento. Conclusiones: el cartílago articular es la estructura anatómica más afectada en la articulación por el proceso degenerativo. Se encuentra organizado por zonas y áreas, las que se afectan a medida que avanza la enfermedad. El origen de la destrucción del cartílago es enzimático y repercute en las demás estructuras como el tejido sinovial y hueso subcondral. Es importante conocer las diferencias entre el envejecimiento y la afección degenerativa de este tejido.

ABSTRACT Background: articular cartilage is an important avascular, alinphatic and aneural tissue in joints, it is mainly affected by degenerative disease. Objective: to update knowledge about normal, aging and degenerative articular cartilage. Methods: a three months research and analysis were conducted from October 1st, 2018 to December 31th, 2018. Our review included 164 articles published in PubMed, Hinari, SciELO and Medline databases by using EndNote. The words used were cartilage AND osteoarthritis, cartilage AND knee osteoarthritis, cartilage. Fifty citations were selected, 48 of all them within the last five years, were used to write the present paper. Results: macro and microscopic features of articular cartilage were mentioned as well as its organization in zones and areas. Degenerative process was described at different levels, and in aging. Conclusions: articular cartilage is the most affected tissue in osteoarthritis. Cartilage is organized by zones and areas which are affected as degenerative disease progresses. The start point of osteoarthritis is enzymatic and gradually affects synovial tissue and sub-chondral bone. It is important to know the main differences between aging and degenerative cartilages.

Effect of calcium carbonate nanoparticles on alkaline protease production from Streptomyces spororaveus

Aims@#This study aims to detect the effect of nano-sized calcium carbonate as an ingredient in growth medium on the production of alkaline protease by Streptomyces spororaveus. The proportional relationship between highly production of alkaline protease and calcium carbonate nanoparticles emphasizes the unique and super properties of nanotechnology that applied in all field. @*Methodology and results@#The high production of protease from S. spororaveus accompanied with presence of calcium carbonate nanoparticles as one of growth medium's constituents. Both qualitative and quantitative tests for proteolytic activity proved this fact. Agar-well diffusion method revealed that, the proteolytic activity with calcium carbonate nanoparticles (45 mm) is higher than that with calcium carbonate (30 mm). Calcium carbonate nanoparticles led to 150 μg/mL of protease, while calcium carbonate led to 65 μg/mL only. The crude protease was purified by ammonium sulphate precipitation and gel filtration column chromatography using Sephadex G-100. The purified protease was separated by SDS-PAGE as a single band at 30 kDa. The highest proteolytic activity was obtained at pH 8.5 and 45 °C as optimum environmental conditions. The purified protease has inhibited the growth of Candida albicans ATCC-10231 and Aspergillus brasiliensis ATCC-16404 at 8 mm and 10 mm of inhibition zone respectively. @*Conclusion, significance and impact of study@#Calcium carbonate nanoparticles in the composition of starch nitrate broth is good stimulus for highly proteolytic activity of S. spororaveus. Shake-flask fermentation method proved that, the concerned protease is an alkaline and thermostable up to 70 °C. However, the best pH and temperature values are 8.5 and 45 °C, respectively. This study can be applied to manufacture a modified starch nitrate broth medium for highly production of proteases from Streptomyces bacteria.

Función de los confórmeros de ataque cercano en la acilación enantioselectiva del (R,S)-propranolol catalizada por lipasa B de Candida antárctica / Rol of the near attack conformers during enantioselective acylation of (R,S)-propranolol catalysed by Candida antarctica lipase B

RESUMEN La lipasa B de Candida antárctica (CalB) se ha utilizado en la acilación quimio- y enantioselectiva del racemato (R,S)-propranolol. CalB tiene enant¡oselect¡v¡dad moderada (£=63) por el R-propranolol. La enantioselectividad, se origina en la reacción de transferencia del grupo acilo desde la serina catalítica, acilada, al propranolol. La fase inicial de esta reacción involucra la formación de complejos de Michaelis y posteriormente conformaciones de ataque cercano. El análisis de las conformaciones de ataque cercano ha permitido en varios casos explicar el origen de la catálisis o reproducir el efecto catalítico. En este trabajo se profundiza en la comprensión la función de las conformaciones de ataque cercano en la enantioselectividad de la acilación del (R,S)-propranolol catalizada por CalB. Para lo anterior se realizó un estudio detallado de los complejos de Michaelis y de las conformaciones de ataque cercano del paso enantioselectivo de la reacción de acilación del (R,S)-propranolol utilizando un protocolo de dinámica molecular QM/MM (SCCDFTB/CHARMM) utilizando 6 distribuciones de velocidades iniciales y simulaciones de 2,5 ns. Se estudiaron 7 complejos CalB-propranolol. Los enlaces de hidrógeno del sitio activo de CalB acilada relevantes para la actividad catalítica fueron estables en todas las simulaciones. Las poblaciones de los complejos de Michaelis y de las conformaciones de ataque cercano son dependientes de la distribución de las velocidades iniciales de la dinámica molecular. La enantioselectividad moderada de CalB acilada, encontrada experimentalmente, puede ser parcialmente atribuida a la alta población de conformaciones de ataque cercano observada para el S-propranolol.

ABSTRACT Candida antarctica lipase B (CalB) has been used for chemo- and enantioselective acylation of racemic (R,S)-propranolol, with moderate enantioselectivity (£=63) for R-propranolol. The enantioselective step in this reaction is the transfer of an acyl group from the catalytic acylated serine to propranolol. The initial phase of this reaction involves the formation of Michaelis complexes, followed by the formation of near-attack complexes. The analysis of the near-attack complexes has in several cases permitted to explain the origin of the catalysis or to reproduce the catalytic effect. The aim of this study was improve the understanding of the role of the near-attack complexes for the enantioselectivity of the acylation of (R,S)-propranolol, catalyzed by CalB. To this purpose a detailed investigation of the Michaelis and near-attack complexes of the enantioselective step of the acylation of (R,S)-propranolol using QM/MM molecular dynamics was performed. Several simulations (each 2,5 ns) with different initial velocity distributions were performed. In total seven CalB-propranolol complexes were studied. The hydrogen bonds in the active site of CalB, which are relevant for the catalytic activity, are stable in all simulations. The lifetime of the Michaelis complexes is considerably shorter than the simulation time. Conclusions: The populations of the Michaelis and near-attack complexes depend on the initial velocity distribution in the molecular dynamics simulations. The experimentally observed moderate enantioselectivity may be partially attributed to the high population of near-attack conformations of S-propranolol.

Enzymology of snails under treatment of molluscicides / 中国血吸虫病防治杂志

Studies on the enzymology of snails are important in the study of molluscicidal mechanism.The alteration of activi-ties of enzymes after molluscicidal treatment was reported in large numbers of papers.This paper reviews the progress of studies on the enzymology of snails under the treatment of molluscicides.

Bioinformatic analysis and characterization of myxobacteria laccase-like multicopper oxidases / 生物工程学报

Laccase is a widely-used environment-friendly copper-containing oxidase found in many plants, insects and fungi. Recently, more and more laccases are also found in bacteria. Myxobacteria are an important bacteria resource. However, myxobacteria are much more difficult to isolate and purify than other bacteria. We used bioinformatic approach to screen myxobacteria proteomes available in NCBI. Based on conserved sequences of four copper binding sites in multicopper oxidase, 30 potential laccase sequences were obtained. Among them, nine genes were synthesized and expressed in Escherichia coli BL21 (DE3). Seven proteins showed laccase activity when tested with traditional laccase substrates. One protein, named rSC-2, was chosen for further research because it exhibited the highest activity towards 2,6-dimethyl phenol (DMP). The molecular weight of rSC-2 was 57 kDa. Its specific activity to DMP was 0.27 U/mg. The optimal temperature and the optimal pH were 60 ℃ and 7.0, respectively. About 50% of the original activity was retained after incubation at 60 ℃ and pH 7.0-8.0 for 1 h. Metals showed different effects on rSC-2. rSC-2 activity was enhanced by several metalsat concentration of 1 mmol/L, such as Ca²⁺ and Mn²⁺. With a higher concentration of 5 mmol/L, the activity of rSC-2 was apparently inhibited. This is the first report of bioinformatics screening myxobacteria laccases in combination with expression in E. coli.

CONSUMO DE ÁLCOOL E A INFLUÊNCIA DO EXERCÍCIO FÍSICO NA ATIVIDADE ENZIMÁTICA DE RATOS WISTAR / ALCOHOL CONSUMPTION AND THE INFLUENCE OF PHYSICAL EXERCISE ON ENZYME ACTIVITY OF WISTAR RATS / CONSUMO DE ALCOHOL Y LA INFLUENCIA DEL EJERCICIO FÍSICO EN LA ACTIVIDAD DE LA ENZIMA DE RATAS WISTAR

RESUMO Introdução: Biomarcadores vem sendo utilizados para monitorar o uso do álcool e, atualmente, os mais sensíveis e específicos são enzimas hepáticas, por exemplo, gama glutamiltransferase (GGT), alanina aminotransferase (ALT), aspartato aminotransferase (AST) e fosfatase alcalina (ALP). Objetivo: Verificar, a partir da experimentação animal, as alterações provocadas pelo uso de álcool e pela prática de atividade física nas enzimas hepáticas e pancreáticas. Métodos: Vinte e quatro ratos da linhagem Wistar foram distribuídos aleatoriamente em grupos experimentais, alojados em gaiolas com ambiente controlado, divididos de acordo com os tratamentos recebidos. No tratamento inicial, foi administrado álcool aos grupos álcool sedentário (AS) e álcool exercitado (AE) e, ao final da quarta semana, iniciou-se o programa de treinamento físico em esteira com os grupos AE e controle exercitado (CE). A coleta de sangue foi realizada por punção cardíaca ao final de cada experimento. Na análise estatística, utilizou-se teste de análise de variância (ANOVA) seguido de teste de Tukey e teste de Kruskal-Wallis. Resultados: O grupo AS apresentou valores significativamente mais elevados de ALT e ALP quando comparado aos grupos CE e AE, respectivamente. Não foram observadas diferenças significativas entre os quatro grupos estudados para os parâmetros AST, GGT e amilase. Conclusão: A associação entre consumo de álcool e sedentarismo aumentou a liberação das enzimas ALT e ALP em ratos Wistar; a prática de exercício físico aeróbico após abstinência alcoólica evitou o aumento da liberação de ALP no plasma desses animais.

ABSTRACT Introduction: Biomarkers have been used to monitor the use of alcohol and currently the most sensitive and specific are the liver enzymes, for example, gamma-glutamyl transferase (GGT), alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP). Objective: To determine through animal experiments the changes caused by alcohol and the physical activity in liver and pancreatic enzymes. Methods: Twenty-four Wistar rats were randomly assigned into experimental groups, housed in cages with controlled environment, divided according to the treatment received. In the initial treatment, alcohol was administered to sedentary alcohol (SA) and exercised alcohol (EA) groups and at the end of the fourth week the program of physical training on a treadmill began with the AE and control exercised (CE) groups. Blood collection was performed by cardiac puncture at the end of each experiment. For the statistical analysis we used analysis of variance (ANOVA) followed by Tukey test and Kruskal-Wallis test. Results: The AS group had significantly higher values of ALT and ALP when compared to CE and EA groups, respectively. No significant differences were observed between the four groups for the parameters AST, GGT and amylase. Conclusion: The association between alcohol consumption and physical inactivity increased the release of the enzymes ALT and ALP in Wistar rats; the practice of aerobic exercise after alcohol withdrawal prevented the increased release of ALP in the plasma of these animals.

RESUMEN Introducción: Los biomarcadores se han utilizado para controlar el consumo de alcohol y en la actualidad las enzimas hepáticas son las más sensibles y específicas, por ejemplo, gamma glutamil transferasa (GGT), alanina aminotransferasa (ALT), aspartato aminotransferasa (AST) y fosfatasa alcalina (ALP). Objetivo: Determinar, a partir de experimentos con animales, los cambios provocados por el alcohol y la actividad física en las enzimas hepáticas y pancreáticas. Métodos: Veinticuatro ratones Wistar fueron asignados al azar a los grupos experimentales, fueron alojados en jaulas con ambiente controlado, divididos de acuerdo a los tratamientos recibidos. En el tratamiento inicial, el alcohol se administró a los grupos alcohol sedentario (AS) y alcohol ejercicio (AE) y el final de la cuarta semana, se inició el programa de entrenamiento físico en una caminadora con los grupos AE y el control ejercitado (CE). La recogida de sangre se realizó mediante punción cardiaca al final de cada experimento. En el análisis estadístico se utilizó el análisis de varianza (ANOVA), seguido por la prueba de Tukey y la prueba de Kruskal-Wallis. Resultados: El grupo AS tuvo valores significativamente más elevados de ALT y ALP en comparación con los grupos CE y AE, respectivamente. No se observaron diferencias significativas entre los cuatro grupos para los parámetros de AST, GGT y amilasa. Conclusión: La asociación entre el consumo de alcohol y la falta de actividad física aumenta la liberación de las enzimas ALT y ALP en ratones Wistar; la práctica de ejercicio aeróbico después de la retirada del alcohol impidió el aumento de la liberación de ALP en el plasma de estos animales.

The significance of neutrophil alkaline phosphatase activity in the differential diagnosis of polycythemia / 国际检验医学杂志

Objective To discuss clinical significance of the neutrophil alkaline phosphatase(NAP) activity in polycythemia pa‐tients .Methods 35 cases of patients with secondary polycythemia(secondary group) and 22 patients with polycythemia vera (poly‐cythemia vera group) were enrolled in the study ,whose peripheral blood smears were stained by using chemical method and the pos‐itive rate and scores of NAP were assessed .In addition to that ,30 cases of healthy people who had underwent physical examination were recruited as control group .Results NAP positive rate and scores of the polycythemia vera group were significantly higher than those of secondary and control group(P0 .05) .Conclusion NAP score is a good methord to diagnose polycythemia ,which could facilitate rapid ,accurate diagno‐sis and be used as auxiliary indicators in the differential diagnosis of polycythemia .

Association of endothelial nitric oxide synthase gene polymorphisms and avascular necrosis of femoral head / 中国医师杂志

Objective To investigate the association of endothelial nitric oxide synthase (eNOS) polymorphisms and avascular necrosis of femoral head (ANFH),to explore possible relationship between the ANFH incidence and 27-bp repeat polymorphism in intron 4 and G894T polymorphism in exon 7.Methods Totally 125 atraumatic ANFH patients and 126 healthy controls without hip trauma history were enrolled.Gene polymorphisms in 27-bp repeat polymorphism in intron 4 and G894T polymorphism in exon 7 were determined.Medical history was collected for etiology analysis.Results In between-group comparison,the frequency of b/a genotype intron 4 in ANFH group was significantly higher than that in healthy control group [19/125 (15.2％) vs 6/126 (4.8％),P =0.0001,OR =4.501],and the result of G/T genotype exon 7 in the ANFH group also indicated a statistical significance with the healthy control group [30/125 (24.0％) vs 17/126 (13.5％),P =0.0094,OR =3.804].In subgroup analysis,genotype b/a and G/T were especially higher in the idiopathic group than that in the healthy control group.Conclusions eNOS gene polymorphisms might be a risk factor of ANFH,there is underlying relevance of eNOS and the disease.Both 27-bp repeat polymorphism in intron 4 and G894T polymorphism in exon 7 are associated with ANFH incidence.

Change of serum metalloproteinases and metalloproteinases inhibitors in exfoliation syndrome in Uyghur population / 中华实验眼科杂志

Background Exfoliation syndrome is a systemic disease with abnormal accumulation of extracellular matrix.Oxidative stress and imbalance between matrix metalloproteinases(MMPs) and tissue inhibitor of matrix metalloproteinases(TIMPs)may play an important role in the pathogenesis of exfoliation syndrome.Researches showed that the incidence of exfoliation syndrome is higher in Uyghur nationality than that in Han nationality.However,whether the imbalance of serum MMPs and TIMPs is associated with pathogenesis of different ethnic groups is unclear.Objective The aim of the study was to discuss the change of serum MMP-9 and TIMP-2 in Uyghur patients.Methods This study protocol was approved by Ethic Committee of Xinjiang Medical University and followed Declaration of Helsinki.A prospective cohort study was performed.Forty Uyghur nationality (46 eyes) with exfoliation syndrome were collected from March 2012 to May 2013 in Affiliated First Hospital of Xinjiang Medical University and First People Hospital of Kashi.Forty cases(40 eyes)age-and gender-matched normal volunteers were included in the same duration.The peripheral blood was collected under the informed consent.Serum MMP-9 and TIMP-2 levels were detected by ELISA,and the results between the two groups were compared using independent samples t test.Results Slit-lamp examination found that part of pigmentation was depigmented and white dandrufflike substance attached in the pupil margin in all the patients,and stripping white dandruff-like substance was deposited in the front surface of the lens capsule which distributed in 3 zones.The pupils were disk-shaped pupil,and the surrounding area was the ring granular and the middle was transparent area without ablative material after dilation.Serum MMP-9 levels were (57.88±18.63)μg/L in the exfoliation syndrome group and (9.35±2.78)μg/L in the normal control group;serum TIMP-2 levels were (17.36±4.66) μg/L in the exfoliation syndrome group and (25.73±3.59) μg/L in the control group.The ratios of MMP-9/TIMP-2 were 3.57± 1.45 in the exfoliation syndrome group and 0.37±0.11 in the control group,with statistically significant differences in serum MMP-9 and TIMP-2 levels as well as ratio of MMP-9/TIMP-2 between the two groups(t=11.52,-6.36,9.87,all at P=0.00).Conclusions The upregulation of serum MMP-9 and downregulation of serum TIMP-2 are found in Uyghur patients with exfoliation syndrome.It is verified that the imbalance of serum MMP-9 and TIMP-2 contributes to the pathogenesis of exfoliation syndrome in Uyghur patients.

Expression of DNA methyltransferases 1,3a,and 3b in retinoblastoma / 中华眼底病杂志

Objective To observe the expressions of DNA methyltransferases (DNMTs) 1,3a and 3b in retinoblastoma (RB).Methods Sixty-two RB samples and six normal retinas were studied,including 17 poorly differentiated and 45 well differentiated samples; 16 invasive and 46 non-invasive samples.The expressions of DNMT1,3a,and 3b,and Ki-67 were detected using immunohistochemical analysis.Brown staining of nuclei was considered to represent the positive stain for DNMT1,3a and 3b,and ki-67,blue staining as negative.The level of high expression of nuclear staining was,positive cells in DNMT1≥65 ％,in DNMT3a≥60％ and in DNMT3b≥40％.The correlations of DNMT1,3a and 3b expression in RB samples,and MIB-1 labeling index were analyzed.Results Viewed under the light microscope,negative expressions of DNMT1,3a and 3b were demonstrated in normal retinas,however,positive expression was observed in RB samples,with 100％ in DNMT1,98％ in DNMT3a and 92％ in DNMT3b.Comparing well differentiated RB samples with poorly differentiated samples,significant differences were found in high expression of DNMT1 (x2 =12.57,P＜0.05) and DNMT3a (x2 =10.54,P＜0.05) ; also in the positive cells of DNMT1 (U=179,P＜0.05) and DNMT3a (U=198,P＜0.05).No significant difference was found comparing high expression (x2=1.5,P＞0.05) and positive cells (U=307,P＞0.05) of DNMT3b.When comparing invasive tumor tissues with non-invasive tumors,significant differences were shown between high expression (x2 =4.72,P＜0.05) and positive cells comparing DNMT1 (U=236,P＜0.05).No significant difference was shown in high expression (x2=3.53,0.84; P＞0.05) in DNMT3a and DNMT3b,or in comparison with positive cells (U=338,257; P＞0.05).The expression of DNMTs was positively correlated with the MIB-1 labeling index in RB tissues (R2=0.554,0.376,0.219; P＜0.05).Conclusion There are high expressions of DNMT1,3a,and 3b in RB.

Protein expressions and clinical significance of β-catenin and glycogen synthase kinase 3 beta in the gastric carcinomas / 中国医师杂志

ObjectiveTo investigate the expressions ofβ-catenin and glycogen synthase kinase 3 beta(GSK-3β) proteins in the gastric carcinomas and to elucidate their clinical significance.Methods The expressions of β-catenin and GSK3-β proteins were analyzed with immunohistochemistry staining of gastric tissue array in the normal gastric mucosa (n =48),paracancerous tissues ( n =24),and primary cancer tissues ( n =48).ResultsThe positive expression rate ofβ-catenin and GSK-3β in gastric carcinomas was 66.7％ and 35.4％,respectively,and β-catenin protein expression in carcinomas was higher than normal and paracancerous tissues ( x2 =65.455,P ＜ 0.05 ).The expression of β-catenin was correlated with cell differential degree,pTNM stage,histological type,lymph node memtastasis,and nerve invasion,respectively ( x2 =4.713,8.242,13.662,11.658,4.550,P ＜ 0.05 or P ＜ 0.01 ).The positive expression rate of GSK3βin carcinomas was lower than normal and paracancerous tissues ( x2 =26.968,P ＜ 0.05).The expression of GSK-3β was correlated with cell differential degree,histological type,and lymph node metastasis,respectively (x2 =3.868,9.665,9.872,P ＜ 0.05 or P ＜ 0.01 ).The expression of β-catenin was negatively correlated with the expression of GSK-3βprotein in the gastric carcinoma tissues ( r =-0.493,P =0.001 ).ConclusionsGSK3-β and β-catenin might play important roles in the progression,differentiation,infiltration,lymph node metastasis of gastric carcinoma,and might be the indicators for diagnosis of a gastric carcinoma.

Characteristics of fiver enzyme abnormalities in acute graft-versus-host disease after bone marrow transplantation / 中华内科杂志

Objective To investigate the characteristics of liver enzyme between acute graft-versushost disease (aGVHD) and non-aGVHD groups after allogeneic hematopoietie stem cell transplantation (allo-HSCT). Methods The liver enzyme data of patients with or without aGVHD was analyzed. Results Among the 371 patients, 158 developed aGVHD(41.6%) with a median time of 29. 5 d. In non-aGVHD group, the median elevating times of ALT, AST, GGT, LDH were all ≤ 20 d after transplantation, which were significantly earlier than those of aGVHD group. The peak value of AST was much higher in aGVHD group, but the remaining liver enzyme showed no significant difference. In aGVHD group, the duration of AST, GGT and LDH was significantly longer than the non-occurrence of aGVHD group, but ALT and ALP showed no difference in duration. In ALT, AST, ALP and LDH elevating group, the occurrence rate of aGVHD was significantly higher, but only ALT and LDH elevation could enter logistic regression model. The sensitivity and veracity of ALT or LDH elevating only were not very good for the diagnosis of aGVHD, but if ALT and LDH beth elevated after day 24 and persisted more than 22 d, the sensitivity and veracity were better. Conclusions The change of liver enzyme is common in allo-HSCT and associates with the occurrence of aGVHD, and the liver enzyme elevating only may not be a good diagnostic index of aGVHD. If the dynamic characteristics of liver enzyme is taken into account and the effect of drug-induced liver injury could be excluded, the elevation of liver enzyme still have the diagnostic value of aGVHD.

The evaluation of biochemical parameters in diagnosis of liver disease / 中国基层医药

ObjectiveTo investigate the changes of biochemical parameters in liver disease. MethodsSernm contents of NAG, AFU, LAP, ASTm, GLDH,ADA, and CHE were determined with auto- biochemical analyzer and analyzed in 300 hepatitis patients and 30 healthy subjects. ResultsThe mean values of LAP, ASTm, GLDH,ADA and AFU in acute hepatitis patients were significantly higher than control, positive rate were 93.0% 、81.0%、76. 0% 、82. 0% and 83.2% respectively;The mean values of ADA、AFU and NAG in liver cirrhosis patients were higher than control significantly, positive rate of were 88.5% 、38.2% and 94. 2% respectively;The mean values of ADA and NAG in severe hepatitis patients were higher than control significantly,positive rate are 64. 9% and 95.7% respectively; The mean values of LAP and NAG in liver cancer patients were higher than control significantly, positive rate were75.0% and 88.0% respectively. ConclusionThe index of LAP and ASTm are valuable markers for diagnosing of acute hepatitis;The index of ADA, ASTm and CHE are valuable marker for diagnosing of liver cirrhosis;The index of ASTm, CHE, AFU and GLDH are valuable marker for diagnosing of severe hepatitis, and AFU and LAP are valuable markers for diagnosing of hepatocellular carcinoma.

Endopeptidases of Bacillus subtilis IBTC-3 and B. alcalophilus PB92 in synthesis of precursors of biologically active peptides.

Two endopeptidases (from Bacillus subtilis IBTC-3 and from B. alcalophilus PB92-commercial preparation) efficiently synthesized amino acid esters (NAc-Tyr-OEt and NAc-Phe-OEt) and dipeptides (NAc-Tyr-Gly-NH2 and NAc-Tyr-Arg-NH2) in organic solvent/water systems. The rate of NAc-Tyr-OEt synthesis mediated by the native subtilisin IBTC-3 was maximum (0.23 Umg-1) in ethanol/5-7% w/v water system, while the highest activity of the freeze-dried enzyme (0.18 Umg-1) was achieved, when water content was 9-10% w/v. The preferred system for dipeptide synthesis (using NAc-Tyr-OEt as acyl donor) by both the enzymes was acetonitrile/4% w/v water. In this system, the maximum yield of NAc-Tyr-GlyNH2 was 71 and 80% and that of NAc-Tyr-Arg-NH2 was 53 and 40% for subtilisin IBTC-3 and peptidase PB92, respectively. In contrast to the peptidase PB92, the subtilisin efficiently catalyzed esterification of NAc-Tyr with 1-butanol and isopropanol.

Intracellular amyloid beta interacts with SOD1 and impairs the enzymatic activity of SOD1: implications for the pathogenesis of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease caused by the degeneration of motor neurons. Mutations in Cu/Zn superoxide dismutase (SOD1), including G93A, were reportedly linked to familial ALS. SOD1 is a key antioxidant enzyme, and is also one of the major targets for oxidative damage in the brains of patients suffering from Alzheimer's disease (AD). Several lines of evidence suggest that intracellular amyloid beta (Abeta) is associated with the pathogenesis of AD. In this report we demonstrate that intracellular Abeta directly interacts with SOD1, and that this interaction decreases the enzymatic activity of the enzyme. We observed Abeta-SOD1 aggregates in the perinuclear region of H4 cells, and mapped the SOD1 binding region to Abeta amino acids 26-42. Interestingly, intracellular Abeta binds to the SOD1 G93A mutant with greater affinity than to wild-type SOD1. This resulted in considerably less mutant enzymatic activity. Our study implicates a potential role for Abeta in the development of ALS by interacting with the SOD1 G93A mutant.

An NH(2)-terminal truncated cytochrome P450 CYP3A4 showing catalytic activity is present in the cytoplasm of human liver cells

Cytochrome P450 3A4 (CYP3A4), is the dominant human liver hemoprotein enzyme localized in the endoplasmic reticulum (ER), and is responsible for the metabolism of more than 50% of clinically relevant drugs. While we were studying CYP3A4 expression and activity in human liver, we found that anti-CYP3A4 antibody cross-reacted with a lower band in liver cytoplasmic fraction. We assessed the activities of CYP3A4 and its truncated form in the microsomal and cytoplasmic fraction, respectively. In the cytoplasmic fraction, truncated CYP3A4 showed catalytic activity when reconstituted with NADPH-cytochrome P-450 reductase and cytochrome b5. In order to determine which site was deleted in the truncated form in vitro, we transfected cells with N-terminal tagged or C-terminal tagged human CYP3A4 cDNA. The truncated CYP3A4 is the N-terminal deleted form and was present in the soluble cytoplasmic fraction. Our result shows, for the first time, that N-terminal truncated, catalytically active CYP3A4 is present principally in the cytoplasm of human liver cells.

Estudo dos polimorfismos das paraoxonases 1 e 2 em pacientes portadores do vírus da imunodeficiência humana e avaliação do potencial de peroxidação lipídica / Frequency of the paraoxonase 1 and 2 genetic polymorphisms and analysis of the lipid peroxidation in plasma of HIV positive patients

A paraoxonase sérica humana (PON) vem sendo amplamente estudada. Além da capacidade de PON1 em hidrolisar compostos organofosfatados, sabe-se, atualmente, que toda a família PON (composta por PON1, PON2 e PON3) promove a proteção de lípides, incluindo-se a lipoproteína de baixa densidade (LDL) contra a oxidação. O gene da PON1 sérica apresenta dois sítios polimórficos bem determinados: a troca Gln192Arg (Q/R) e Met55Leu, os quais estão associados com diferenças na atividade e concentrações séricas da enzima, respectivamente. Também o polimorfismo Cys311Ser parece contribuir sinergisticamente com o alelo PON1-192R quanto ao risco cardiovascular em algumas populações. Já foi demonstrado, por sua vez, que pacientes infectados pelo vírus HIV podem desenvolver dislipidemia e que tanto a atividade como a concentração de PON1 podem ser influenciadas por esta infecção. O objetivo deste estudo foi determinar as freqüências alélicas dos polimorfismos genéticos PON1-192QR, PON1-55LM, PON2-311SC e PON2-148AG, bem como avaliar a atividade de PON1 e a peroxidação lipídica no plasma de indivíduos portadores de HIV. Materiais e Métodos após aprovação pela comissão de ética e da aplicação do termo de consentimento pós-esclarecido, 350 (264 homens e 86 mulheres) pacientes infectados pelo HIV foram incluídos no estudo. Foi avaliado ainda um grupo de 32 (23 homens e 9 mulheres) indivíduos recentemente diagnosticados como portadores do vírus. Uma população saudável composta por 179 doadores de sangue, todos de sexo masculino, foi avaliada como controle. Após a extração do DNA, procedeu-se à genotipagem para os polimorfismos de PON1 e PON2 através de PCR-RFLP. A atividade paraoxonase de PON1 foi avaliada por espectrofotometria empregando-se paraoxon como substrato. O colesterol total, VLDL-colesterol, HDL-colesterol e triglicérides foram determinados por métodos padrão. A fração LDL-colesterol foi calculada pela fórmula de Friedwald. Resultados As frequências alélicas...

Human serum paraoxonase (PON) has been the subject of a number of studies. Beside the capacity of PON1 in hydrolyzing organophosphate compounds, it is known now that the entire PON family (which comprises PON1, PON2 and PON3) protects lipids, including low-density lipoprotein (LDL), from oxidation. Serum PON1 gene presents two well-determined genetic polymorphic sites: a Gln192 Arg (Q/R) and Met55 Leu, which are associated with differences in enzymatic activity and serum concentrations, respectively. Moreover, PON2 Cys311 Ser polymorphism seems to contribute synergistically with PON-192R allele to cardiovascular risk in some populations. It has been shown that HIV infected patients may develop dyslipidemia and that PON1 activity and concentration may be influenced by this infection. The aim of this study was to determine allelic frequencies of PON1-192QR, PON1-55LM, PON2-311SC and PON2-148AG genetic polymorphisms, evaluate PON1 activity and lipid peroxidation in plasma of HIV patients. Methods and Subjects after ethical committee approval and written consent, 350 (264 men and 86 women) HIV infected patients were included in the study. It was also evaluated a group of 32 recently diagnosed HIV individuals (23 men and 9 women). As controls, a healthy population formed by 179 men, blood donors, was studied. After DNA extraction PON1 and PON2 genotyping were performed by PCR-RFLP. Paraoxonase activity of PON1 was evaluated spectrofotometrically using paraoxon as substrate. Serum cholesterol, VLDL-cholesterol, HDL-cholesterol and triglycerides were analyzed by standard methods. LDL-cholesterol was calculated by Friedewald formula. Results: Allelic frequencies for PON1 polymorphisms in patients were: 36,43% for PON1-192R, 57,86% for PON1-55L, 65,57% for PON2-311S and 76,43% for PON2-148A. In recently diagnosed individuals these frequencies were 37,50%, 51,56%, 81,25% and 68,75% respectively. In controls, PON1-192R allelic frequency was 43,02%, PON1-55L...