Clinical implementation of comprehending dentin on a microscopic level

SUMMARY: The aim of the present in vitro study is to visualize dentin to get an in-depth knowledge of the nature of dentin that could provide useful information regarding conditioning dentinal substrate when treating dentinal lesions. Forty-nine extracted human third molars were obtained and prepared to produce artificial dentinal lesions through demineralizing with acetic acid for 7 and 14 days, or lactic acid for 7 days. The teeth were divided into groups and treated with either NaOCl, pepsin, trypsin, or phosphoric acid. To obtain information on the morphology of the treated dentinal surfaces, all samples were visualized under high resolution field emission scanning electron microscope. With high magnification reaching x50000 dentin was clearly visualized together with its constitutes. The effect of various demineralization approaches and various treatment protocols were demonstrated clearly. The relationship between the conditioning procedure steps and the subsequent bond strength was discussed. To our best knowledge, there is no previous clear highly magnified scanning electron microscope images for dentin, and dentinal components and constitutes with and without various treatments. The current in vitro study suggests the complexity nature of dentin as a substrate that should be treated carefully especially with technique sensitive procedures such as adhesive restorations.

El objetivo del presente estudio in vitro fue visualizar la dentina para obtener un conocimiento completo de la naturaleza de ella lo que podría proporcionar información útil sobre el acondicionamiento del sustrato dentinario en el tratamiento de lesiones dentinarias. Se obtuvieron 49 terceros molares humanos extraídos y se prepararon para producir lesiones dentinales artificiales mediante desmineralización con ácido acético por 7 y 14 días, o ácido láctico por 7 días. Los dientes se dividieron en grupos y se trataron con NaOCl, pepsina, tripsina o ácido fosfórico. Para obtener información sobre la morfología de las superficies dentinarias tratadas, todas las muestras se visualizaron bajo un microscopio electrónico de barrido de emisión de campo de alta resolución. Con un gran aumento que alcanzó x50000, la dentina se visualizó claramente junto con sus componentes. Se demostró el efecto de varios enfoques de desmineralización y varios protocolos de tratamiento. Se discutió la relación entre los pasos del procedimiento de acondicionamiento y la subsiguiente fuerza de unión. Hasta donde sabemos, no hay imágenes claras previas de microscopio electrónico de barrido altamente ampliadas para la dentina y los componentes y constituyentes de la dentina con y sin diferentes tratamientos. El estudio in vitro actual sugiere la naturaleza compleja de la dentina como sustrato que debe tratarse con cuidado, especialmente en los procedimientos sensibles a la técnica, tal como las restauraciones adhesivas.

Preliminary analysis of isolation effect of influenza virus by recombinant MDCK cells stably expressing trypsinogen / 中国生物制品学杂志

@#ObjectiveTo investigate the isolation effect of influenza virus by recombinant MDCK cells(MTY6 cells)stably expressing trypsinogen.MethodsAccording to the virus isolation method recommended by the World Health Organization(WHO)Global Influenza Surveillance Network(GISN)and the National Influenza Centers(NICs),a total of 20 throat swab specimens containing positive nucleic acid for H1N1,H3N2 and B influenza virus were isolated simultaneously using MDCK and MTY6 cells. Guinea pig red blood cells and chicken red blood cells were used for agglutination test respectively and the agglutination effects of different types of red blood cells,the positive rate of virus and the titer of hemagglutinin isolated from different cells were statistically compared.ResultsThe agglutination effect of the same virus isolate on the two types of red blood cells was different. The complete agglutination time of guinea pig red blood cells was about 2 times that of chicken red blood cells,and the deposition shape showed a ring shape. The average hemag-glutinin titer was 23. 6 ± 1. 2times that of chicken red blood cells. Under the same conditions,3 samples were negative for both types of cells,11 samples were positive for both types of cells,and the other 6 samples were negative for MDCK cells while positive for MTY6 cells. The positive rate of MTY6 cells was 30% higher than that of MDCK cells. The isolated positive samples included 8 cases of H1N1 subtype and the hemagglutinin titer of virus isolated by MTY6 cells was significantly higher than that by MDCK cells[13. 0(1. 7,23. 0)times on average]. 2 cases of H3N2 and 2 cases of B were isolated,the hemagglutinin titer of each virus isolated by MTY6 cells was 11. 3 and 32. 0 times higher than that by MDCK cells on average respectively.ConclusionIn conclusion,guinea pig red blood cells were superior to chicken red blood cells for influenza virus detection by cell isolation. Under the same conditions,MTY6 cells were more sensitive than MDCK cells for influenza virus isolation,and had the potential to be used as a high-quality cell matrix for influenza virus isolation.

Preparation of the first batch of national standard of recombinant trypsin / 中国生物制品学杂志

@#Objective To prepare the 1st batch of national standard of recombinant trypsin in order to standardize and improve the quality of recombinant trypsin.Methods Enzyme-substrate identification,HPLC identification,N-terminal sequencing and TOF-MS were used to confirm the property and structure of recombinant trypsin;the purity was determined by HPLC;according to the methods in Chinese Pharmacopoeia(Volume Ⅲ 3603,2020 edition),the specific activity of the candidate standard was determined,and the stability and uniformity were investigated.Results The structure of recombinant trypsin was confirmed,and the specific activity of the 1 st batch of national standard of recombinant trypsin was 5 169 U/mg,containing 80% β-trypsin and 9% α-trypsin.The RSD of purity of α,β-trypsin and retention time of 12 candidate standards were all less than 2.0%.The purity of α,β-trypsin showed no obvious decrease stored at 25 ℃ and relative humidity(RH) 80% for 10 d,while the purity of β-trypsin decreased slightly and the purity of α-trypsin increased slightly stored at 40℃ and RH 80% for 10 d.The purity of β-trypsin decreased slightly when exposed to light(4 000 lx) for 10 d.Conclusion The national standard of recombinant trypsin with accurate structure and high purity was prepared,which can be used for system suitability test of the purity determination.

Systemic therapy with bromelain-trypsin-rutoside combination in inflammation: A narrative review of the pharmacodynamics

Inflammation and wound healing are attended by interactions of numerous cell types, mediators, cytokines, and the vascular system. Any imbalance in these interactions can set off a cascade of processes, often working in different directions, and which continue till homeostasis is restored. Systemic enzyme therapy with trypsin-bromelain-rutoside combination has long been used as dietary supplements or pharmaceuticals commercially to counter various inflammatory conditions. The clinical benefits are believed to be due to the multipronged action of the combination, which include anti-kinin, antioxidant, antiplatelet, anti-chemokine, pro-fibrinolytic, and pro-fibrotic activities; with more recent evidence indicating inhibition of pro-inflammatory signal transduction. The primary modes of action of the individual agents on different components and processes involved in inflammation have been evaluated and elucidated, using various in vitro, animal model, and clinical studies. A narrative review of the evidence collected over the years has been provided.

LC-MS/MS method for the quantitation of serum tocilizumab in rheumatoid arthritis patients using rapid tryptic digestion without IgG purification / 药物分析学报

The quantitation of serum tocilizumab using liquid chromatography tandem-mass spectrometry(LC-MS/MS)method has not been widely applied in clinical settings because of its time-consuming and costly sample pretreatments.The present study aimed to develop a validated LC-MS/MS method for detecting serum tocilizumab by utilizing immobilized trypsin without an immunoglobulin G purification step and evaluate its applicability in the treatment of rheumatoid arthritis(RA)patients administered intrave-nously or subcutaneously with tocilizumab.The tocilizumab-derived signature peptide was deciphered using a nano-LC system coupled to a hybrid quadrupole-orbitrap mass spectrometer.The serum tocili-zumab was rapidly digested by immobilized trypsin for 30 min.The chromatographic peak of the signature peptide and that of the internal standard were separated from the serum digests for a total run time of 15 min.The calibration curve of serum tocilizumab concentration was linear with a range of 2-200 μg/mL.The intra-and inter-day accuracy and relative standard deviation(RSD)were 90.7％-109.4％and＜10％,respectively.The serum tocilizumab concentrations in the RA patients receiving intravenous and subcutaneous injections were 5.8-28.9 and 2.4-63.5 pg/mL,respectively.The serum tocilizumab concentrations using the current method positively correlated with those using the enzyme-linked immunosorbent assay,although a systematic error was observed between these methods.In conclu-sion,a validated LC-MS/MS method with minimal sample pretreatments for monitoring serum tocili-zumab concentrations in RA patients was developed.

Energy values and protein digestibility of soybean milk by-product in pigs based on in vitro assays / Valores energéticos y digestibilidad proteica del subproducto de la leche de soja en cerdos con base en ensayos in vitro / Valores energéticos e digestibilidade protéica do subproduto do leite de soja em suínos com base em ensaios in vitro

Abstract Background: Soybean milk by-product (SMBP) is a potential alternative feed ingredient in swine diets due to its high protein content. However, information on energy and nutritional values of SMBP used as swine feed ingredient is limited. Objective: To estimate energy values and protein digestibility of SMBP in pigs based on in vitro assays. Methods: Four SMBP samples were obtained from 3 soybean milk-producing facilities. In vitro total tract disappearance (IVTTD) and in vitro ileal disappearance (IVID) of dry matter (DM) in the SMBP samples were determined. In vitro ileal disappearance of crude protein was determined by analyzing crude protein content in undigested residues after determining IVID of DM. Digestible and metabolizable energy of SMBP were estimated using gross energy, IVTTD of DM, and prediction equations. Results: Sample 4 had greater IVTTD of DM than that of sample 3 (97.7 vs. 94.4%, p<0.05), whereas IVID of DM in sample 4 was lower compared with sample 1 (53.5 vs. 65.0%, p<0.05). In vitro ileal disappearance of crude protein in sample 2 was greater than that in sample 1 and 3 (92.6 vs. 90.6 and 90.1%; p<0.05). The estimated metabolizable energy of SMBP ranged from 4,311 to 4,619 kcal/kg as-is basis and the value of sample 3 was the least (p<0.05) among SMBP samples. Conclusion: Energy values and protein digestibility should be determined before using SMBP in swine diets.

Resumen Antecedentes: El subproducto de la leche de soja (SMBP) es un ingrediente alimenticio alternativo con uso potencial en dietas porcinas dado su alto contenido de proteína. Sin embargo, la información sobre sus valores energéticos y nutricionales para alimentación de cerdos es muy limitada. Objetivo: Estimar los valores de energía y la digestibilidad de la proteína del SMBP en cerdos con base en ensayos in vitro. Métodos: Se obtuvieron cuatro muestras de SMBP de tres empresas productoras de leche de soja. Se determinaron la desaparición de tracto total in vitro (IVTTD) y la desaparición ileal in vitro (IVID) de la materia seca (DM) en las muestras de SMBP. La desaparición ileal in vitro de proteína cruda se determinó analizando el contenido de proteína cruda en residuos no digeridos después de determinar la IVID de la DM. La energía digestible y metabolizable de SMBP se estimó utilizando la energía bruta, IVTTD de la DM y ecuaciones de predicción. Resultados: La muestra 4 tuvo una mayor IVTTD de la DM que la muestra 3 (97,7 vs. 94,4%, p<0,05), mientras que la IVID de la DM en la muestra 4 fue menor en comparación con la muestra 1 (53,5 vs. 65,0%, p<0,05). La desaparición ileal in vitro de la proteína cruda en la muestra 2 fue mayor que la de las muestras 1 y 3 (92,6 vs. 90,6 y 90,1%; p<0,05). La energía metabolizable estimada de SMBP varió de 4.311 a 4.619 kcal/kg (en base húmeda) y el valor de la muestra 3 fue el menor (p<0.05) entre las muestras de SMBP. Conclusión: Los valores de energía y la digestibilidad de la proteína deben determinarse antes de usar el SMBP en dietas porcinas.

Resumo Antecedentes: O subproduto do leite de soja (SMBP) é um potencial ingrediente alternativo na dieta de suínos, considerando seu alto teor de proteínas. No entanto, as informações sobre os valores energéticos e nutricionais do SMBP usado como ingrediente alimentar para suínos são limitadas. Objetivo: Estimar valores energéticos e digestibilidade protéica do SMBP em suínos com base em ensaios in vitro. Métodos: Foram obtidas quatro amostras de SMBP de três instalações produtores de leite de soja. Foram determinados o desaparecimento total do trato in vitro (IVTTD) e o desaparecimento ileal in vitro (IVID) da matéria seca (DM) nas amostras de SMBP. O desaparecimento ileal in vitro da proteína bruta foi determinado pela análise do conteúdo de proteína bruta em resíduos não digeridos após a determinação da IVID do DM. A energia digerível e metabolizável do SMBP foi estimada usando energia bruta, IVTTD do DM e equações de predição. Resultados: a amostra 4 apresentou maior IVTTD de DM do que a amostra 3 (97,7 vs. 94,4%, p<0,05) enquanto a IVID do DM na amostra 4 foi menor em comparação com a amostra 1 (53,5 vs. 65,0%, p<0,05). O desaparecimento ileal in vitro da proteína bruta na amostra 2 foi superior ao da amostra 1 e 3 (92,6 vs. 90,6 e 90,1%; p<0,05). A energia metabolizável estimada do SMBP variou de 4.311 a 4.619 kcal/kg no estado em que se encontra e o valor da amostra 3 foi o menor (p<0,05) entre as amostras do SMBP. Conclusão: os valores energéticos e a digestibilidade das proteínas devem ser determinados antes do uso do SMBP nas dietas suínas.

Effects of different doses of ulinastatin on pulmonary function in patients undergoing total aortic arch replacement / 中华麻醉学杂志

Objective:To evaluate the effects of different doses of ulinastatin on lung function in patients undergoing total aortic arch replacement.Methods:One hundred and thirty five patients with acute Stanford type A aortic dissection of both sexes, aged 20-70 yr, with body mass index of 16.2-33.3 kg/m 2, of American Society of Anesthesiologist physical status Ⅳ, were divided into 3 groups ( n=45 each) using a random number table method: high-dose ulinastatin group (group H with total dose of 30 000 U/kg), low-dose ulinastatin group (group L with total dose of 20 000 U/kg) and control group (group C). In group H and group L, half of the total dose of ulinastatin was given after induction of anesthesia, the rest of the total dose was primed after being added to cardiopulmonary bypass (CPB) circuit, while normal saline 100 ml was given at the same time point in group C. After induction of anesthesia (T 0), and at 3, 6, 12, 24 and 48 h after the beginning of CPB (T 1-5), blood samples from the central vein were collected for determination of plasma concentrations of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). The oxygenation index (OI) and alveolar-arterial partial pressure of oxygen difference (P A-aO 2) at T 0 and T 2-T 5, intraoperative blood loss and blood transfusion, postoperative mechanical ventilation time, length of intensive care unit (ICU) stay and the incidence of complications within 30 days after surgery were collected. Results:Compared with group C, the plasma concentrations of TNF-α and IL-6 were significantly at T 1-T 5, OI was increased, and P A-aO 2 was decreased at T 2, 3 in H and L groups ( P<0.05). There was no significant difference in the mechanical ventilation time, length of ICU stay and incidence of complications within 30 days after surgery among the 3 groups ( P>0.05). Conclusion:Ulinastatin can inhibit inflammatory responses and improve lung function in patients undergoing total aortic arch replacement, but it has no value for clinical outcomes.

Effect of ulinastatin on hyperoxia-induced acute lung injury in infantile rats: relationship with Wnt/β-catenin signaling pathway / 中华麻醉学杂志

Objective:To evaluate the effect of ulinastatin on hyperoxia-induced acute lung injury (ALI) and its relationship with Wnt/β-catenin signaling pathway in infantile rats.Methods:A total of 36 clean-grade Sprague-Dawley rats, aged 14 days, weighing 40-50 g, were divided into 3 groups ( n=12 each): control group (C group), hyperoxia-induced ALI group (ALI group) and ulinastatin group (UTI group). Hyperoxia-induced ALI was induced by inhaling oxygen at concentration greater than 90% for 72 h. At 1 day after the model was established successfully, ulinastatin 50 000 U/kg was injected intraperitoneally daily at the same time for 3 consecutive days in group UTI, while the equal volume of normal saline was injected intraperitoneally at the same time point in C and ALI groups.The animals were sacrificed at 4 days after the model was established successfully, the lung tissues were taken for determination of the wet/dry weight ratio (W/D ratio), for microscopic examination of the pathological changes which were scored, for measurement of interleukin-6 (IL-6) IL-1β and tumor necrosis factor-alpha (TNF-α) (by enzyme-linked immunosorbent assay) and for detection of the expression of phosphorylated glycogen synthase kinase (p-GSK-3β), Wnt3a and β-catenin by Western blot, and ultrastructure was examined with with an electron microscope. Results:Compared with C group, W/D ratio and lung injury score were significantly increased, the contents of IL-6, IL-1β and TNF-α were increased, and the expression of p-GSK-3β, Wnt3a and β-catenin were up-regulated in lung tissues in group ALI ( P<0.05). Compared with group ALI, W/D ratio and lung injury score were significantly decreased, the contents of IL-6, IL-1β and TNF-α were decreased, and the expression of p-GSK-3β, Wnt3a and β-catenin were down-regulated in lung tissues in group UTI ( P<0.05). The ultrastructure injury in group UTI was reduced as compared with group ALI. Conclusion:The mechanism by which ulinastatin can alleviate hyperoxia-induced ALI is related to inhibiting the activation of Wnt/β-catenin signaling pathway and decreasing inflammatory response in infantile rats.

Relationship between mechanism of ulinastatin reducing perioperative myocardial injury and ferroptosis in peripheral blood mononuclear cells in pediatric patients undergoing heart surgery under cardiopulmonary bypass / 中华麻醉学杂志

Objective:To investigate the relationship between the mechanism of ulinastatin reducing perioperative myocardial injury and ferroptosis in peripheral blood mononuclear cells (PBMCs) in pediatric patients undergoing heart surgery under cardiopulmonary bypass (CPB).Methods:A total of 60 pediatric patients of either sex, aged 4-8 yr, of American Association of Anesthesiologists physical status Ⅱ or Ⅲ, undergoing elective repair of ventricular septal defect under CPB, were divided into 2 groups by a random number table method: control group (C group) and ulinastatin group (UTI group), with 30 cases in each group.Combined intravenous-inhalational anesthesia was used.In UTI group, ulinastatin 20 000 U/kg was diluted to 100 ml in normal saline, 50 ml was infused through the central vein over 15 min starting from 20 min before skin incision, and the remaining 50 ml was instilled through the CPB pipeline over 15 min starting from 10 min of CPB.The equal volume of normal saline was given instead in C group.Blood samples from the internal jugular vein were collected after anesthesia induction and before skin incision (T 1), at 30 min after start of CPB (T 2), immediately after termination of CPB (T 3) and at 24 h after termination of CPB (T 4) for determination of the levels of amino-terminal B-type pro-brain natriuretic peptide (NT-proBNP), cardiac troponin I (cTnI) and creatine kinase isoenzymes (CK-MB) in plasma by enzyme-linked immunosorbent assay.PBMCs were extracted by modified Ficoll density gradient centrifugation method for determination of the concentrations of Fe 2+ and malondialdehyde (MDA) and activity of superoxide dismutase (SOD) in PBMCs (by colorimetric method) and expression of long-chain acyl-CoA synthase 4 (ACSL4) and glutathione peroxidase 4 (GPX4) in PBMCs (by Western blot). Results:Compared with the baseline at T 1, the levels of NT-proBNP, cTnI and CK-MB in plasma were significantly increased, the concentrations of Fe 2+ and MDA in PBMCs were increased, the expression of ACSL4 in PBMCs was up-regulated, and the activity of SOD was decreased, and the expression of GPX4 was down-regulated at T 2-4 in two groups ( P<0.05). Compared with C group, the plasma levels of NT-proBNP, cTnI and CK-MB were significantly decreased, the concentrations of Fe 2+ and MDA in PBMCs were decreased, the expression of ACSL4 in PBMCs was down-regulated, the activity of SOD was increased, and the expression of GPX4 was up-regulated at T 2-4 in UTI group ( P<0.05). Conclusion:The mechanism by which ulinastatin reduces perioperative myocardial injury may be related to inhibition of ferroptosis in PBMCs in the pediatric patients undergoing open heart surgery under CPB.

Production of Herbal Protein Isolates with the Enzymatic Hydrolysis Technology

The article aims to develop the technology of the production of herbal protein isolates with 41% proteinconcentration by enzymatic hydrolysis. The technology includes preparing a hydro module from shredded lupineseeds and water in proportion 1:10, hydrolysis of starch with alpha-amylase and glucoamylase, centrifugation,autoclaving of the obtained centrifugate at the temperature of 120-130º С and the pressure of 6х105 Pa for 5-6hours, cooling it to the temperature of 36º С and hydrolyzing it with trypsin solution in phosphate buffer solutionat pH 7.5 for 50-60 minutes, centrifugation, heating and drying at the temperature of 95-100 ºС to get the dryresidue concentration of 45% in the protein preparation. Before adding trypsin, it is intensified by blue spectrumlight with the luminous flux of 35 µW/cm2. Based on the research, there are regulated quality indicators of proteinpreparation, storage requirements, and retention periods: retention period of 9 months at the temperature of 0-4°С with relative humidity not more than 75%.

A carbon nanoparticle-peptide fluorescent sensor custom-made for simple and sensitive detection of trypsin / 药物分析学报

Herein, we report a novel sensor to detect trypsin using a purpose-designed fluorescein-labelled peptide with negatively charged carbon nanoparticles (CNPs) modified by acid oxidation. The fluorescence of the fluorescein-labelled peptide was quenched by CNPs. The sensor reacted with trypsin to cleave the peptide, resulting in the release of the dye moiety and a substantial increase in fluorescence intensity, which was dose-and time-dependent, and trypsin could be quantified accordingly. Correspondingly, the biosensor has led to the development of a convenient and efficient fluorescent method to measure trypsin activity, with a detection limit of 0.7μg/mL. The method allows rapid determination of trypsin activity in the normal and acute pancreatitis range, suitable for point-of-care testing. Furthermore, the applicability of the method has been demonstrated by detecting trypsin in spiked urine samples.

Research progress on role of mast cells in irritable bowel syndrome / 中国药理学通报

Irritable bowel syndrome (IBS) is one kind of functional gastrointestinal tract disorders with high incidence in the world. IBS is induced by avariety of factors,such as visceral hypersensitivity .abnormal gastrointestinal motility, intestinal barrier dysfunction, intestinal inflammation, neuro-immunity disorder, etc. The pathogenesis of IBS is complex and diverse. Mast cells (MCs) play an important role in the occurrence and development of IBS. The present article reviews the role of MCs both in the pathogenesis and in the treatment of IBS.

Construction of acellular porcine bladder scaffolds / 中国组织工程研究

BACKGROUND: Bladder repair is currently one of the main treatments for bladder defects. Homologous tissue is less affected by various factors. Tissue engineered acellular bladder matrix has become an increasing area of interest. Porcine bladder acellular matrix has a wide range of sources and has a natural extracellular scaffold structure, which has become a hot topic in tissue engineering bladder substitute materials. OBJECTIVE: To explore the feasibility of acellular porcine bladder as a tissue engineering scaffold material. METHODS: The cell-free matrix of pig bladder was prepared by liquid nitrogen freezing and thawing, dodecyl sodium sulfate and trypsin decellularization method. According to different decellularization methods, pig bladders were divided into normal control group (without any treatment), experimental group (treated with 0.6% trypsin and 5% sodium lauryl sulfate (pH 8.0)) and acellular control group (treated with 0.75% trypsin (pH 8.0), 1% trypsin (pH 8.0), 5% sodium lauryl sulfate (pH 7.6) or 10% sodium dodecyl sulfate (pH 7.6)). The decellularization effect of pig bladder was observed by hematoxylin-eosin staining, van Gieson staining, DNA quantification, and α-Gal antigen detection. RESULTS AND CONCLUSION: Hematoxylin-eosin staining revealed that in the experimental group, the components of the bladder cells of the pigs were basically removed. van Gieson staining revealed that the DNA residues and α-Gal antigen residues in the cells were significantly lower than those in the control group (P < 0.05). These results suggest that treatment of pig bladder with 0.6% trypsin and 5% sodium dodecyl sulfate can effectively remove its cellular components while retaining the extracellular matrix of porcine bladder tissue. This provides a reference value for constructing accelular porcine bladder scaffolds.

Assessment of hemagglutination activity of porcine deltacoronavirus

Serum concentrations of α2-macroglobulin, α1-antitrypsin, and C-reactive protein in dogs with suspected acute pancreatitis / 대한수의학회지

In canine acute pancreatitis (AP), inappropriate release and activation of zymogen proteases within the pancreas results in the consumption of serum antiproteases. The aim of this study was to examine whether the serum concentrations of α₂-macroglobulin (A2MG), α₁-antitrypsin (A1AT), and C-reactive protein (CRP) differ between dogs with AP and healthy dogs. Twenty healthy dogs and 20 dogs with AP were included in this study. Concentrations of A2MG, A1AT, and CRP were measured in the sera of healthy dogs and dogs diagnosed with AP. Serum A2MG and A1AT concentrations were significantly lower in dogs with AP than in healthy dogs, whereas the serum CRP concentration was significantly higher. In addition, the concentrations of A2MG and A1AT were significantly higher in AP survivors than in AP non-survivors, while the CRP concentration was significantly lower. However, in both AP survivors and non-survivors, the CRP concentrations showed a negative correlation with A2MG concentrations but not with A1AT. These findings indicate that serum antiproteases and CRP concentrations might be associated with the mortality rate of AP in dogs.

Effect of ulinastatin on expression of metabotropic glutamate receptor Ⅱ during cerebral ischemia-reperfusion in rats / 中华麻醉学杂志

Objective To evaluate the effect of ulinastatin on the expression of metabotropic glutamate receptor Ⅱ ( mGluRⅡ) during cerebral ischemia-reperfusion ( I∕R) in rats. Methods Forty-eight male Sprague-Dawley rats, aged 6-8 weeks, weighing 230-280 g, were divided into 3 groups ( n=16 each) by a random number table method: sham operation group (S group), cerebral I∕R group (I∕R group) and ulinastatin group ( U group) . The model of cerebral I∕R injury was established by occluding the right middle cerebral artery using a nylon thread with a rounded tip inserted into internal carotid artery and advanced cranially until resistance was met. Middle cerebral artery occlusion was maintained for 2 h followed by 24-h reperfusion. Ulinastatin 100000 U∕kg was injected via the tail vein immediately after onset of reperfusion in group U. The neurological deficit score ( NDS) was assessed after 24 h of reperfusion. The rats were then sacrificed, and brain tissues were obtained for determination of brain infarction ( by TTC staining) , expression of IκB-α in cerebral ischemic penumbra ( by Western blot) and expression of mGluRⅡ in cerebral ischemic penumbra ( by immunofluorescent staining) . The percentage of cerebral infarct vol-ume was calculated. Results Compared with S group, the NDS and percentage of cerebral infarct volume were significantly increased, the expression of mGluRⅡ in cerebral ischemic penumbra was up-regulated, and the expression of IκB-α in cerebral ischemic penumbra was down-regulated in I∕R and U groups ( P<0. 05). Compared with I∕R group, the NDS and percentage of cerebral infarct volume were significantly de-creased, the expression of mGluRⅡ in cerebral ischemic penumbra was down-regulated, and the expres-sion of IκB-α in cerebral ischemic penumbra was up-regulated in U group ( P<0. 05) . Conclusion The mechanism by which ulinastatin mitigates cerebral I∕R injury may be related to inhibiting the expression of mGluR Ⅱ in cerebral ischemic penumbra of rats.

Effect of ulinastatin on programmed necrosis in hippocampal neurons in a rat model of global cerebral ischemia-reperfusion / 中华麻醉学杂志

Objective To evaluate the effect of ulinastatin on programmed necrosis in hippocampal neurons in a rat model of global cerebral ischemia-reperfusion (I/R).Methods Forty-eight clean-grade healthy adult male Sprague-Dawley rats,aged 8 weeks,weighing 280-320 g,were divided into 3 groups (n=16 each) using a random number table method:sham operation group (Sham group),global cerebral I/R group (I/R group) and ulinastatin group (UT group).Global cerebral I/R was produced by 4-vessel occlusion method in chloral hydrate-anesthetized rats in I/R and UT groups.Ulinastatin 100 000 U/kg was injected via the tail vein at the onset of ischemia in group UT,and the equal volume of normal saline was given instead in Sham and I/R groups.Neurological deficit score (NDS) was estimated at 6,12 and 24 h of reperfusion.Animals were sacrificed at 24 h of reperfusion,brains were removed and the hippocampi were obtained for examination of pathological changes (with a light microscope) and for determination of the malondialdehyde (MDA) content and superoxide dismutase (SOD) activity (by spectrophotometry),and expression of receptor-interacting protein kinase 1 (RIPK1),RIPK3,and mixed lineage kinase domain-like protein (MLKL) in hippocampal tissues (by Western blot).Results Compared with Sham group,the NDS was significantly increased at each time point,the MDA content was increased,the SOD activity was decreased,and the expression of RIPK1,RIPK3 and MLKL was up-regulated in I/R and UT groups (P＜ 0.05).Compared with I/R group,the NDS was significantly decreased at each time point,the MDA content was decreased,the SOD activity was increased,and the expression of RIPK1,RIPK3 and MLKL was down-regulated in UT group (P＜0.05).The pathological changes of hippocampi were significantly attenuated in UT group when compared with I/R group.Conclusion The mechanism by which ulinastatin ameliorates global cerebral I/R injury is related to inhibiting programmed necrosis in hippocampal neurons of rats.

Optimization of the amount of TPCK trypsin adding to MDCK cell suspension for culturing H7N9 avian influenza virus / 中华微生物学和免疫学杂志

Objective@#To investigate the best amount of TPCK trypsin in Madin Darby canine kidney (MDCK) cell suspension for the culture of H7N9 avian influenza virus.@*Methods@#Different concentrations of TPCK trypsin were added during the periods of cell growth and virus production. Their effects on cell growth, viability, glucose and lactate metabolism, and hemagglutination titer were monitored every 12 h. Inter-batch differences were analyzed. The amount of trypsin added in the cell growth phase was 0, 1 μg/ml, 2 μg/ml, 4 μg/ml, 6 μg/ml, 8 μg/ml, 10 μg/ml and 15 μg/ml. The amount of trypsin added during the virus production period was 0, 0.5 μg/ml, 1 μg/ml, 1.5 μg/ml, 2 μg/ml and 2.5 μg/ml. When the hemagglutination titers were same, the adding amount was further optimized at different multiplicity of infection (MOI) of 0.001, 0.005, 0.025 and 0.05.@*Results@#No significant linear effects of TPCK trypsin concentration on cell number, viability, and glucose and lactate metabolism were observed. No toxicity to cell growth was observed when TPCK trypsin concentration reached 15 μg/ml. After the inoculation of H7N9 avian influenza virus, the hemagglutination titers in the 1 μg/ml, 1.5 μg/ml, 2 μg/ml and 2.5 μg/ml TPCK trypsin groups reached the peaks at 48 h, which were 1∶26.5. At 60 h, the hemagglutination titers of the latter two groups decreased faster than those of the former two groups. When the MOI was 0.005, the hemagglutination titer of the 1.5 μg/ml group at 48 h was 26.5 higher than 26 in the 1 μg/ml group under the same condition. There were differences between different batches of TPCK trypsin.@*Conclusions@#Adding 1 μg/ml and 1.5 μg/ml of trypsin could better promote the proliferation of H7N9 avian influenza virus, and 1.5 μg/ml of trypsin had a wider range of MOI applicability.

Electrochemical analysis of the corrosion inhibition properties of L-leucine and trypsin complex admixture on high carbon steel in 1 M H 2 SO 4 solution / Análisis electroquímico de las propiedades de inhibición de la corrosión de la mezcla de L-leucina y complejo de tripsina sobre acero de alto contenido de carbono en solución 1 M de H 2 SO 4 / Análise eletroquímica das propriedades de inibição da corrosão da mistura de L-leucina e complexo de tripsina em aço de alto carbono em solução 1 M de H 2 SO 4

Abstract Corrosion inhibition of biodegradable chemical compounds (L-leucine and trypsin complex) on high carbon steel in 1 M H2SO4 acid media was evaluated with potentiodynamic polarization technique, weight loss analysis, open circuit potential measurement, optical microscopy, and ATR-FTIR spectroscopy. Data obtained showed the mixture has a maximum inhibition efficiency of 82.4% and 90.08% from the electrochemical tests with mixed type inhibition properties. The addition of the mixture shifts significantly the corrosion potential of the steel to passivation values from open circuit potential measurement. Results from thermodynamic calculations indicated chemisorption adsorption mechanism according to Langmuir, Freundlich, and Frumkin isotherms coupled with correlation coefficients of 0.9994, 0.9651 and 0.8834. Statistical analysis showed exposure time to be the most significant variable responsible for corrosion inhibition. Identified functional groups of the compound from ATF-FTIR spectroscopy were adsorbed completely on the carbon steel surface from observation of the decreased peak intensity. Optical microscopy images of the inhibited and uninhibited steel surfaces contrast each other with due to the presence of macro-pits and porous oxide on the uninhibited steel.

Resumen Se evaluó la inhibición de la corrosión por parte de compuestos químicos biodegradables (complejo de L-leucina y tripsina) sobre acero de alto contenido de carbono en H2SO4 1 M a través de técnica de polarización potenciodinámica, análisis de pérdida de peso, medición de potencial de circuito abierto, microscopía óptica y espectroscopia ATR-FTIR. Los datos obtenidos mostraron que la mezcla tiene una eficacia de inhibición máxima de 82,4% y 90,08%, a partir de las pruebas electroquímicas con propiedades de inhibición de tipo mixto. La adición de la mezcla cambia significativamente el potencial de corrosión del acero a los valores de pasivación de la medición del potencial de circuito abierto. Los resultados de los cálculos termodinámicos indicaron un mecanismo de adsorción por quimisorción de acuerdo con las isotermas Langmuir, Freundlich y Frumkin, acopladas con coeficientes de correlación de 0,9994; 0,9651 y 0,8834, respectivamente. El análisis estadístico mostró que el tiempo de exposición es la variable más importante en la inhibición de la corrosión. Los grupos funcionales de la mezcla, identificados mediante espectroscopía ATF-FTIR, fueron completamente adsorbidos en la superficie de acero al carbono; esto se dedujo a partir de la observación de la disminución de la intensidad de pico. Las imágenes de microscopía óptica de las superficies de acero inhibidas y desinhibidas contrastan entre sí, debido a la presencia de macro-pozos y óxido poroso en el acero desinhibido.

Resumo Foram feitos estudos de inibição da corrosão de compostos químicos biodegradáveis ccomplexo de L-leucina e tripsina) em aço de alto conteúdo de carbono em meio de H2SO4 1 M, estes foram avaliados com técnica de polarização potenciodinâmica, análise de perda de peso, medição de potencial de circuito aberto, microscopia óptica e espectroscopia ATR-FTIR. Os dados obtidos mostram que a mistura tem uma eficiência de inibição máxima de 82,4% e 90,08% dos testes eletroquímicos com propriedades de inibição do tipo misto. A adição da mistura desloca significativamente o potencial de corrosão do aço para valores de passivação da medição de potencial de circuito aberto. Os resultados dos cálculos termodinâmicos indicam o mecanismo de adsorção de quimisorção de acordo com as isotermas de Langmuir, Freundlich e Frumkin, juntamente com coeficientes de correlação de 0,9994; 0,9651 e 0,8834 respectivamente. A análise estatística mostrou que o tempo de exposição é a variável mais significativa da inibição da corrosão. Os grupos funcionais identificados pela espectroscopia ATF-FTIR foram completamente adsorvidos na superfície do aço ao carbono, de acordo com a observação da intensidade de pico diminuída. As imagens de microscopia óptica das superfícies de aço inibidas e desinibidas contrastam entre si devido à presença de macro-poços e óxido poroso no aço desinibido.

Characterization of digestive enzymes from captive Brazilian flounder Paralichthys orbignyanus / Caracterização de enzimas digestivas do linguado brasileiro de cativeiro Paralichthys orbignyanus

Abstract Knowledge of specific enzyme activity, along with animal habits and digestive capacity is essential in formulating an appropriate diet for any species. In this study, we evaluated and characterized the activity of digestive enzymes present in the liver, intestine, and stomach of Paralichthys orbignyanus. The effects of pH and temperature on enzyme activity were also evaluated via the use of specific substrates. The use of specific substrates and inhibitors showed strong evidence of the presence of trypsin (BApNA= 0.51 ± 0.2 mU mg-1), chimotrypsin (SApNA= 2.62 ± 1.8 mU mg-1), and aminopeptidases (Leu-p-Nan =0.9709 ± 0.83 mU mg-1) in the intestine. Optimum pH for the activity of trypsin, chemotrypsin, leucino aminopeptidase, amilase, and pepsin were 9.5, 9.0, 8.0, 7.5, and 3.5, respectively, while optimum temperatures were 50, 50, 50, 40, and 45 °C, respectively. These results provide additional information regarding the biology of Brazilian flounder and can be used as a basis for further studies regarding fish feeding physiology.

Resumo O conhecimento da atividade enzimática é essencial para formular uma correta dieta específica para espécie, além de estarem correlacionadas com o hábito da alimentação e capacidade digestive. Neste estudo determinamos e caracterizamos a atividade enzimática presente no intestino, estômago e fígado do linguado Paralichthys orbignyanus. Os efeitos da temperatura e pH sobre a atividade enzimática também foram avaliados utilizando substratos específicos. O uso de substratos e inibidores específicos mostrou uma forte evidência da presença da tripsina (BApNA = 0,51 ± 0,2 mU mg-1), quimotripsina (SAPNA = 2,62 ± 1,8 mU mg-1), e as aminopeptidases (Leu-p-Nan = 0,97 ± 0,83 mU mg-1) no intestino. O pH ótimo observado para a atividade de tripsina, quimotripsina, leucino aminopeptidase, amilase e pepsina foi 9,5, 9,0, 8,0, 7,5 e 3,5, respectivamente. A temperatura ótima observada foi 50, 50, 50, 40 e 45 °C, respectivamente. Estes resultados fornecem informações adicionais sobre a biologia do linguado brasileiro e pode ser usado como base para novos estudos sobre fisiologia alimentar.