Concentraciones séricas de Beta-hidroxibutirato y su asociación con enfermedades posparto en bovinos de leche / Serum Beta-hydroxybutyrate concentrations and its association with postpartum diseases in dairy cows

RESUMEN Objetivo. Determinar la prevalencia de cetosis bovina según las concentraciones séricas (mmol/Lt) de beta-hidroxibutirato (βHB) y estimar su asociación con enfermedades del posparto temprano en vacas lecheras. Materiales y métodos. Se llevó a cabo un estudio epidemiológico de corte transversal con información individual de los niveles sanguíneos de βHB y presentación clínica de enfermedades puerperales de n=1149 animales; además se evaluó la eficiencia de la prueba como predictora de enfermedad posparto. Posteriormente, se realizó X 2 y un modelo de regresión logística final para explorar los factores asociados con hipercetonemia. La relación entre βHB sanguíneo, la presentación de cetosis y la ocurrencia de enfermedades puerperales se calculó analizando Característica Operativa del Receptor (Receiver Operating Characteristic -ROC). Resultados. Según los niveles de -3HB, la prevalencia de cetosis fue de 7.9%, cetosis clínica 0.6% y cetosis subclínica 7.3%. La prueba fue predictora de enfermedad puerperal (LR+ del 13.6). La condición corporal ≥3.5, número de partos igual a 2 y ≥3, la presentación de retención de placenta, fiebre de leche y la enfermedad posparto, son factores de riesgo de cetosis. El análisis de la curva ROC mostró que la medición de βHB (≥1.2 mmol/Lt) en sangre sirve para diagnosticar cetosis (p<0.0001). Conclusiones. La medición de niveles sanguíneos de βB permitió determinar que la presentación de cetosis es baja, es una alerta no solamente para esta enfermedad sino para las enfermedades del posparto temprano. Los resultados de este estudio confirman lo reportado en estudios previos sobre los factores de riesgo de cetosis.

ABSTRACT Objective. Determine the prevalence of bovine ketosis based on serum concentrations (mmol/Lt) of beta-hydroxybutyrate (βHB) and estimate its association with early postpartum diseases in dairy cows. Materials and methods. An epidemiological cross-sectional study was carried out with individual information on blood levels of βHB and clinical presentation of puerperal diseases in 1149 animals; In addition, the efficiency of the test was evaluated as a predictor of postpartum disease. Subsequently, univariate analysis and a final logistic regression model were performed to explore the factors associated with hyperketonemia. The association between blood βHB, the presentation of ketosis and the occurrence of puerperal diseases was calculated by analyzing the Receiver Operating Characteristic -ROC. Results. According to βHB levels, the prevalence of ketosis was 7.9%, clinical ketosis 0.6% and subclinical ketosis 7.3%. The test was a predictor of puerperal disease (LR+ of 13.6). Body condition score ≥3.5, number of births 2 and ≥3, presentation of retained fetal membranes, milk fever and postpartum disease, are risk factors for ketosis. The analysis of the ROC curve showed that the measurement of βHB (≥1.2 mmol/Lt) in blood serves to diagnose ketosis (p<0.0001). Conclusions. The measurement of βHB blood levels allowed to determine that ketosis presentation is low, it is an alert not only for this disease but for early postpartum diseases. The results of this study confirm risk factors observed in previous studies.

Influência da restrição alimentar e do estresse térmico sobre parâmetros fisiológicos em ovinos / [Influence of food restriction and heat stress on physiological parameters in sheep]

Objetivou-se avaliar as alterações fisiológicas de ovinos submetidos à restrição alimentar, expostos à radiação solar e à sombra. Dez borregas mestiças foram divididas igualmente em dois grupos: grupo radiação solar - GRS, animais expostos ao sol durante o dia; e grupo sombra - GS, animais alocados dentro de um galpão à sombra. Nos seis dias iniciais de experimento, os animais foram submetidos à restrição alimentar, com acesso apenas à água, já expostos ao sol ou à sombra. Nos seis dias subsequentes, foi ofertada dieta composta por ração e feno de alfafa (3%PV). Os parâmetros avaliados foram frequência respiratória (FR), temperatura retal (TR), ß-hidroxibutirato (BHB), pH urinário, ingestão de matéria seca (IMS) e dados bioclimáticos (temperatura, umidade relativa do ar e índice de temperatura e umidade - ITU). A FR foi o parâmetro em que houve maior diferença significativa entre os grupos (p<0,05). O balanço energético negativo foi constatado em todos os animais (BHB>0,8mmol/L; pH<7,0). Ovelhas que sofrem restrição alimentar durante períodos de alta temperatura e umidade têm menor frequência respiratória e temperatura interna, independentemente de estar na sombra ou pleno sol. Porém, quando normalizado a oferta de alimento as ovelhas em pleno sol têm menor ingestão de matéria seca.(AU)

We aimed to evaluate physiological changes in sheep submitted to food restriction, exposed to the sun or shade. Ten ewe lambs were equally divided into two groups: Sun Radiation Group - GRS, exposed to sun during day hours, and Shade Group - GS, with animals placed inside a barn. In the first six days of experiment, animals were submitted to food restriction, with access to fresh water, already exposed to sun and shade. In the following six days alfalfa hay and concentrate were provided to the animals (3%BW). Parameters assessed were respiratory rate (FR), rectal temperature (TR), ß-hydroxybutyrate, urinary pH, dry matter intake (IMS) and climate data (air temperature, humidity, and temperature-humidity index - ITU). The parameter that most differed between groups was FR. All animals presented negative energy balance (BHB>0.8mmol/L; pH<7,0). Ewe lambs submitted to food restriction during periods of high temperature and humidity have lower respiratory rate and intern temperature, independently of being exposed to sun or shade. However, when food was offered animals exposed to sun had lower dry matter intake.(AU)

Efeito da suplementação de colina protegida no perfil metabólico e intervalo entre parto e concepção de vacas leiteiras / Prepartum supplementation of protected choline on metabolic profile and interval between calving and conception of dairy cows

Durante o periparto, as vacas leiteiras são submetidas a uma grande demanda de energia, ao mesmo tempo em que reduzem sua ingestão de matéria seca. O balanço energético negativo, resultante dessa equação, acarreta severos transtornos metabólicos, à produção e, principalmente, à reprodução. O objetivo do presente estudo foi avaliar o efeito da colina protegida sobre os parâmetros metabólicos, o intervalo entre parto e concepção e a produção de leite em vacas no período de transição. Cinquenta e quatro vacas leiteiras foram divididas em três grupos: controle, suplementação com colina por 10 dias pré-parto (T10) e suplementação com colina por 20 dias pré-parto (T20). Após o parto, foram mensurados os teores de frutosamina, colesterol, ácidos graxos não esterificados (AGNE), beta-hidroxibutirato (BHB), aspartato aminotransferase (AST), gamaglutamiltransferase (GGT) e total de oxidantes (TOS), nos dias 10, 20 e 30. Ainda foram avaliadas produção de leite e intervalo entre parto e concepção. Não houve efeito da suplementação com colina sobre os parâmetros sanguíneos e a produção. O intervalo entre parto e concepção foi menor no grupo T20. A colina suplementada por 20 dias durante o pré-parto melhorou a performance reprodutiva de vacas leiteiras(AU)

During the periparturient dairy cows undergo a large energy demand, at the same time reducing their intake of dry matter. The negative energy balance resulting from this equation leads to severe metabolic disorders in production, and mainly in reproduction. The aim of this study was to evaluate the effect of protected choline on metabolic parameters, reproductive performance, and milk production in cows during the transition period. Fifty-four dairy cows were divided into three groups: control, supplementation with choline for 10 days prepartum (T10) and supplementation with choline for 20 days prepartum (T20). After delivery we measured fructosamine levels, cholesterol, non-esterified fatty acids (NEFA), beta-hydroxybutyrate (BHB), aspartate aminotransferase (AST), gamma glutamyltransferase (GGT), and total oxidant (TOS) on days 10, 20 and 30. We also evaluated milk production and interval between calving and conception. There was no effect of supplementation with choline on blood and production parameters. The interval between calving and conception was lower in the T20 group. Choline supplemented by 20 during the antepartum improved reproductive performance of dairy cows, although it did not change the metabolic profile.(AU)

Trichostrongylus colubriformis infection in Santa Inês lambs: impact on feed digestibility, blood markers, and nitrogen balance / Infecção por Trichostrongylus colubriformis em cordeiros Santa Inês: impacto na digestibilidade dos alimentos, marcadores sanguíneos e balanço de nitrogênio

Abstract This study aimed to evaluate the effects of Trichostrongylus colubriformis infection on the hemato-biochemical parameters, feed digestibility, and nitrogen balance in Santa Inês lambs. Eighteen three-month-old Santa Ines castrated male lambs (16.9 ± 1.43 kg of body weight) were randomly distributed in two experimental treatments: infected with T. colubriformis (I, n = 9) and uninfected (U, n = 9). The I group received a total of 45,000 L3 larvae of T. colubriformis (5,000 infective larvae, three times per week, for three weeks). During the experimental period, blood, feed digestibility, and nitrogen balance were evaluated. The I lambs showed a reduction in erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, and total proteins, as well as an increase in platelets and eosinophils compared to those in the U group (p < 0.05). With the exception of total protein content, these values were within the normal range for the species. Furthermore, lower dry matter and organic matter digestibility were observed in the I lambs (p = 0.08). The present findings highlight that T. colubriformis infection has the potential to impair some hemato-biochemical parameters as well as feed digestibility in lambs, which could affect their productivity.

Resumo O objetivo deste estudo foi avaliar os efeitos da infecção por Trichostrongylus colubriformis nos parâmetros hemato-bioquímicos, digestibilidade aparente dos nutrientes e balanço de nitrogênio de cordeiros Santa Inês. Dezoito cordeiros Santa Inês, de três meses de idade e castrados (16,9 ± 1,43 kg de peso corporal), foram distribuídos aleatoriamente em dois tratamentos experimentais: infectados com T. colubriformis (I, n = 9) e não infectados (U, n = 9). O grupo I recebeu um total de 45.000 larvas L3 de T. colubriformis (5.000 larvas infectantes, três vezes por semana, durante três semanas). Durante o período experimental, foram avaliadas as variáveis sanguíneas, digestibilidade e balanço de nitrogênio. Os cordeiros I apresentaram redução de eritrócitos, hemoglobina, hematócrito, volume corpuscular médio e proteínas totais, além de aumento de plaquetas e eosinófilos, quando comparados ao grupo U (p< 0,05). Contudo, exceto para proteínas totais, os valores estavam dentro do intervalo normal para a espécie. Além disso, a digestibilidade da matéria seca e da matéria orgânica foi menor no grupo I de cordeiros (p = 0,08). Os presentes achados destacam que a infecção por T. colubriformis teve potencial para prejudicar alguns parâmetros hemato-bioquímicos, bem como a digestibilidade aparente da dieta, o que poderia afetar a produtividade dos cordeiros.

Inhibitory effect of ketogenic diet on neuroblastoma in BALB/c-nu mouse models / 南方医科大学学报

OBJECTIVE@#To investigate the inhibitory effect of ketogenic diet (KD) on growth of neuroblastoma in mice.@*METHODS@#BALB/c-nu mouse models bearing neuroblastoma xenografts were established by subcutaneous injection of human neuroblastoma cell line (SH-SY5Y). When the tumor volume reached 250 mm3, the mice were randomized into SD group with standard diet and PBS treatment, KD group with ketogenic diet and PBS treatment, and CP+KD group with ketogenic diet and cyclophosphamide (60 mg·kg·day) treatment, =8. The tumor volume, body weight, blood glucose, ketone body (β-Hydroxybutyrate) levels, and hepatic steatosis in the mice were assessed. The expressions of caspase-3 and caspase-8 were detected by Western blotting, and Ki67 expresison was detected using immunohistochemistry (IHC). Transmission electron microscopy (TEM) was employed for the autophagosomes, and the autophagic protein Beclin1, LC3A/B and P62 were detected by IHC and Western blotting.@*RESULTS@#On day 28 post tumor cell injection, the mice in KD and CP+KD groups could prolong the overall survival rates than that in SD group ( < 0.001). On day 22 post the injection, the tumor volume in KD group was smaller than that in SD group ( < 0.05); on 16, 19, and 22 day post the injection, the tumor volume in CP+KD group was smaller than that in SD group ( < 0.01). The mice in SD group showed greater body weight on day 19 and higher blood glucose level on day 13 post the injection than those in the other two groups ( < 0.05). Blood ketone level and hepatic steatosis score were higher and glucose ketone index (GKI) was lower in KD and CP+KD groups than those in SD group (all < 0.05). The expressions of Ki67 and apoptotic proteins were detected in the tumor tissues of all groups. TEM revealed more autophagosomes in the tumor tissues of KD group than that of SD group. P62 expression was lowered ( < 0.01) and Beclin1 and LC3A/B expressions were up-regulated in the tumor tissues of KD group ( < 0.05), which is consisitent with IHC.@*CONCLUSIONS@#KD has a strong anti-tumor effect in the xenograft mouse model possibly by regulating cell autophagy.

Evaluation of different sites for blood collection for ß-hydroxybutyrate assessment in dairy cows using hand-held meter / Avaliação de diferentes locais de coleta de sangue para avaliação de ß-hidroxibutirato em vacas de leite usando aparelho portátil automático

Cetose subclínica é uma grande preocupação em rebanhos leiteiros, e seu diagnóstico e prevenção podem ter um grande impacto na saúde animal. Este estudo comparou quatro locais diferentes para a coleta de amostras de sangue (jugular, veias mamárias e coccígeas e ponta da cauda) para medição de ß-hidroxibutirato (BHBA), usando-se um medidor portátil automático. Foram utilizadas seis vacas Holandesas, e a coleta de sangue foi feita no segundo, quinto, 10º, 15º e 21º dias pós-parto. Os resultados do medidor portátil foram semelhantes aos resultados do laboratório e apresentaram uma correlação forte de 0,83. As concentrações séricas de BHBA nas amostras de sangue coletadas na ponta da cauda, na jugular e na coccígea foram semelhantes. No entanto, o sangue retirado da veia mamária tinha uma concentração mais baixa. Portanto, as amostras de sangue para aferição de BHBA podem ser recolhidas nas veias jugular e coccígeas e na ponta da cauda, sendo as duas últimas as opções mais fáceis para monitorar o BHBA em rebanho leiteiro.(AU)

Comparação de métodos para avaliação de beta-hidroxibutirato em ovelhas / Comparison of methods for evaluation of beta-hydroxybutyrate in ewes

Atualmente o uso de sensores portáteis para mensuração de corpos cetônicos está padronizado e difundido na rotina clínica, contudo estudos em ovinos são escassos. Assim, a presente pesquisa objetivou avaliar a acurácia dos sensores portáteis de uso humano e de uso veterinário para a determinação de beta-hidroxibutirato (BHB) em ovelhas no final da gestação e no pós-parto recente. Foram utilizadas 37 amostras de sangue provenientes de nove ovelhas mestiças Corriedale. A determinação bioquímica de BHB no soro, considerada como o padrão-ouro, foi realizada utilizando-se metodologia enzimática colorimétrica. A média obtida na bioquímica sérica foi de 0,497mmol/L; no sensor de uso humano, a média foi igual a 0,537mmol/L, enquanto no sensor de uso veterinário foi de 0,751mmol/L. Foi verificada alta correlação entre o dosímetro de uso humano e o padrão-ouro (r=0,93, P<0,001). A média do aparelho de uso veterinário diferiu das demais (51%; P<0,05), superestimando os resultados em ovelhas. As medições obtidas no aparelho veterinário também apresentaram menor precisão e veracidade. Concluiu-se que o sensor portátil de uso humano é mais acurado e mais preciso no diagnóstico precoce de toxemia da gestação em ovelhas.

Currently the use of portable sensors for measuring ketone bodies is standardized and diffused in the clinical routine, however, studies in sheep are scarce. Therefore, the present study aimed to evaluate the accuracy of the human portable sensor and the veterinary portable sensor for the determination of beta-hydroxybutyrate (BHB) in sheep at the end of gestation and postpartum. We used 37 samples of blood from nine crossbred Corriedale sheep. Biochemical determination of serum BHB, considered gold standard, was performed using colorimetric enzymatic methodology. The mean serum biochemistry was 0.497mmol/L, in the human sensor the mean was 0.537mmol/L, while in the veterinary sensor it was 0.751mmol/L. A high correlation was verified between the dosimeter for human use and the gold standard (r= 0.93, P< 0.001). The mean of the veterinary apparatus differed from the others, being 51% (P< 0,05), higher than the standard, that is, it was less accurate and had lower veracity, overestimating the results in sheep. It was concluded that the portable sensor for human use is more accurate and accurate in the early diagnosis of toxemia of pregnancy in sheep.

Effect of reducing the activity of respiratory chain on biosynthesis of poly(3-hydroxybutyrate-co-lactate) in Escherichia coli / 生物工程学报

Poly(3-hydroxybutyrate-co-lactate) [P(3HB-co-LA)] belongs to the polyhydroxyalkanoates (PHA) family and possesses promising properties including biocompatibility and biodegradability. In this study, we directly synthesized P(3HB-co-LA) with glucose by introducing the β-ketothiolase and acetoacetyl-CoA reductase from Ralstonia eutropha, the engineered propionate CoA transferase from Clostridium propionicum and the engineered polyhydroxyalkanoate synthase from Pseudomonas fluorescens strain 2P24 into Escherichia coli. The polymer content was 83.9% (W/W), and the molar percentage of lactate reached 1.6%. On this basis, in order to accumulate lactate, we reduced the activity of respiratory chain by deleting the ubiX gene, which is involved in the synthesis of coenzyme Q8. Moreover, we removed the dld gene to avoid the conversion of lactate to pyruvate during the fermentation. With these manipulations, the molar percentage of lactate in the polymer was improved to 14.1%, with an 81.7% (W/W) of polymer content. The test results indicated that the strategy of reducing the activity of respiratory chain effectively increased the lactate units in the polymer, and it contributed a new approach to change the content of monomer components in the polymer.

Microbial production of poly (glycolate-co-lactate-co-3-hydroxybutyrate) from glucose and xylose by Escherichia coli / 生物工程学报

Escherichia coli was metabolically engineered to produce poly(glycolate-co-lactate-co-3-hydroxybutyrate) using glucose and xylose as carbon sources. The combinatorial biosynthetic route was constructed by the overexpression of a series of enzymes including D-tagatose 3-epimerase, L-fuculokinase, L-fuculose-phosphate aldolase, aldehyde dehydrogenase, propionyl-CoA transferase, β-ketothiolase, acetoacetyl-CoA reductase, and polyhydroxyalkanoate synthase. Overexpression of polyhydroxyalkanoate granule associated protein significantly improved biopolymer synthesis, and the recombinant strain reached 3.73 g/L cell dry weight with 38.72% (W/W) biopolymer content. A co-culture engineering strategy was developed to produce biopolymer from a mixture of glucose and xylose, achieving 4.01 g/L cell dry weight containing 21.54% (W/W) biopolymer. The results of this work offer an approach for simultaneously utilizing glucose and xylose and indicate the potential for future biopolymer production from lignocellulosic biomass.

Cardioprotective Potential of an SGLT2 Inhibitor Against Doxorubicin-Induced Heart Failure

BACKGROUND AND OBJECTIVES: Recent studies have shown that sodium-glucose co-transporter 2 (SGLT2) inhibitors reduce the risk of heart failure (HF)-associated hospitalization and mortality in patients with diabetes. However, it is not clear whether SGLT2 inhibitors have a cardiovascular benefit in patients without diabetes. We aimed to determine whether empagliflozin (EMPA), an SGLT2 inhibitor, has a protective role in HF without diabetes. METHODS: Cardiomyopathy was induced in C57BL/6J mice using intraperitoneal injection of doxorubicin (Dox). Mice with HF were fed a normal chow diet (NCD) or an NCD containing 0.03% EMPA. Then we analyzed their phenotypes and performed in vitro experiments to reveal underlying mechanisms of the EMPA's effects. RESULTS: Mice fed NCD with EMPA showed improved heart function and reduced fibrosis. In vitro studies showed similar results. Phloridzin, a non-specific SGLT inhibitor, did not show any protective effect against Dox toxicity in H9C2 cells. SGLT2 inhibitor can cause increase in blood ketone levels. Beta hydroxybutyrate (βOHB), which is well known ketone body associated with SGLT2 inhibitor, showed a protective effect against Dox in H9C2 cells and in Dox-treated mice. These results suggest elevating βOHB might be a convincing mechanism for the protective effects of SGLT2 inhibitor. CONCLUSIONS: SGLT2 inhibitors have a protective effect in Dox-induced HF in mice. This implied that SGLT2 inhibitor therapy could be a good treatment strategy even in HF patients without diabetes.

Performance Evaluation of the Autokit Total Ketone Bodies

In this study, the Autokit Total Ketone Bodies kit (Wako Pure Chemical, Japan), a total ketone measurement assay using an enzymatic method, was evaluated using a Roche Cobas e702 instrument (Roche Diagnostics, Germany). Precision, linearity, carryover, and reference range verification were evaluated with reference to Clinical Laboratory Standards Institute guidelines. Standard materials provided by the manufacturer and patient samples were used for the evaluation. The precision and carryover of the evaluation result satisfied the acceptance criteria. Linearity was also acceptable at more than 0.99. The quantitative Autokit Total Ketone Bodies kit is precise, and can be widely used in clinical laboratories.

Preparation and characterization of polyhydroxyalkanoate bioplastics with antibacterial activity / 生物工程学报

Polyhydroxyalkanoates (PHAs), as a novel class of biopolymer, are attracting more attention due to their diverse material properties and environment-independent biodegradability. Here we report the preparation of PHA exhibiting efficient antibacterial activity by embedding Nisin, a food additive generally recognized as safe, into poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx), a type of PHA with high biocompatibility. We first prepared Nisin-containing PHBHHx films using solvent casting method. Confocal laser scanning microscopy analysis showed that a well-mixed integrated structure of the films with an even distribution of the Nisin particles in the PHBHHx matrices. Then the antimicrobial activity of PHBHHx/Nisin films against Micrococcus luteus was quantified on agar plate by measuring the size of inhibition zone. Cultivation in liquid media further confirmed the releasing of Nisin from the films and the long-time antibacterial activity. Results showed that the threshold of Nisin concentration for long-time and effective inhibition against bacteria growth is 25 μg/g. These results altogether establish a technological foundation for the application of PHA in biomedicine and food industry.

Biomanufactured polyhydroxyalkanoates (PHA) modification: a review / 生物工程学报

In this review, we presented the industrial status of biomanufactured polyhydroxyalkanoates (PHA), including poly (3-hydroxybutyrate) (PHB), poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), poly (3-hydroxybutyrate-co-4-hydroxybutyrate) (P3/4HB)), and poly (3-hydroxybutyrate-3-hydroxycaproate) (PHBH). A lot of modification studies, aimed at solving problems of poor thermal stability, narrow processing window and other drawbacks of PHA, are discussed. The properties of PHA can be optimized by using proper modification method, in order to expand its applications.

Expression and characterization of a novel halohydrin dehalogenase from Tistrella mobilis KA081020-065 / 生物工程学报

Halohydrin dehalogenase is of great significance for biodegradation of the chlorinated pollutants, and also serves as an important biocatalyst in the synthesis of chiral pharmaceutical intermediates. A putative halohydrin dehalogenase (HheTM) gene from Tistrella mobilis KA081020-065 was cloned and over-expressed in Escherichia coli BL21 (DE3). The recombinant enzyme was purified by Ni-NTA column and characterized. Gel filtration and SDS-PAGE analysis showed that the native form of HheTM was a tetramer. It exhibited the highest activity at 50 degrees C. The nature and pH of the buffer had a great effect on its activity. The enzyme maintained high stability under the alkaline conditions and below 30 degrees C. HheTM catalyzed the transformation of ethyl(S)-4-chloro-3-hydroxybutyrate in the presence of cyanide, to give ethyl (R)-4-cyano-3-hydroxybutyrate, a key intermediate for the synthesis of atorvastatin.

Culture effects of mouse induced pluripotent stem cells by three-dimensional films in vitro / 中华心血管病杂志

<p><b>OBJECTIVE</b>To observe the proliferation and differentiation capacities of mouse induced pluripotent stem cells (miPSCs) cultured in 3-hydroxybutyrate-co-3-hydroxyhexanoate(PHBHHx) three-dimensional films three-dimension films for the purpose of finding a suitable polymeric biomaterials for forming myocardial patches.</p><p><b>METHODS</b>miPSCs were recovered, passaged, cultured and identified, then miPSCs divided into the experimental and control groups. MiPSCs in the experimental group were cultured with PHBHHx three-dimension films. MiPSCs in the control group were cultured with traditional culture dish. Stem cell culture medium or differential medium were added to miPSCs to detecte cell vitality by CCK-8 after 72 hours or to measure the cTnT expression of miPSCs through immunofluorescence or the cTnT expression quantity through flow cytometry after 15 days.</p><p><b>RESULTS</b>Cell activity assay showed that the absorbance values were 0.836 ± 0.038 in the experimental group, 0.312 ± 0.004 in the control group (P<0.05). Scanning electron microscope (SEM) observation showed that miPSCs grew well on the PHBHHx dimensional films with normal shape. Immunofluorescence results demonstrated positive cTnT expression in both groups and flow cytometry measured cTnT expression was (60.32 ± 1.76)% in the experimental group and (47.54 ± 1.46)% in the control group (P<0.05).</p><p><b>CONCLUSIONS</b>miPSCs can survive, proliferate and differentiate on PHBHHx dimensional films. miPSCs proliferation and differentiation capacities are significantly higher in PHBHHx three-dimensional films culture compared with the traditional cell culture.</p>

Determining the Cause of Natural Death: A Case of Previously Unknown Sturge-Weber Syndrome / 대한법의학회지

In some cases, it is difficult to determine a single cause of death even after conducting full autopsy and additional tests. A 49-year-old man, reportedly having diabetes mellitus, was found unconscious by his mother and revealed to be dead. He had several contusions all over his body, including the right periocular area, but they did not appear fatal. A focal area of polymicrogyria and cortical dysplasia was found on the right preoccipital notch, accompanied with dystrophic calcification and leptomeningeal angiomatosis. These findings were considered indicative of Sturge-Weber syndrome, a rare neurocutaneous disorder, of atypical type without facial lesions. Blood level of beta-hydroxybutyrate was 859 microg/mL, implying that he also had diabetic ketoacidosis. His ketoacidosis may not have been corrected appropriately because of status epilepticus in association with brain lesion, resulting in his death, but neither direct evidence nor statement was obtained. In cases with several apparent causes of death, the examiner's assumption should be based not on imagination but on evidence, and logic should not be overlooked. It is more helpful for the investigators or the bereaved to obtain more detailed information rather than come to a hasty conclusion.

Establishment of blood β-hydroxybutyrate threshold for diagnosis of type 2 diabetes ketoacidosis / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish of blood beta hydroxybutyrate (βOHB) threshold for diagnosing type 2 diabetes ketoacidosis (DKA) and explore the relationship between βOHB levels and the severity of DKA.</p><p><b>METHODS</b>Correlation analysis was performed between serum βOHB and [HCO(3)] in type 2 diabetic patients admitted in the emergency department in the past year. Regression equation was used to calculate the concentration of βOHB corresponding to a [HCO(3)] level of 18.0, 15, and 10.0 mmol/L, and βOHB concentration corresponding to a [HCO(3)] level of 18.0 mmol/l was used as the DKA diagnostic threshold.</p><p><b>RESULTS</b>The serum βOHB level and [HCO3] concentration showed a good correlation (R²=0.7023, P<0.001). βOHB concentrations that corresponded to a [HCO(3)] level of 18.0, 15, and 10.0 mmol/L were 3.0, 4.70, and 7.5 mmol/L, respectively, in accordance with the severity of DKA. Combined with the blood glucose concentration ≥ 13.9 mmol/L, a blood βOHB≥3.0 mmol/L showed a sensitivity of 99%, specificity of 86%, and total effectiveness of 92.81% for diagnosing DKA.</p><p><b>CONCLUSION</b>A serum βOHB level above 3.0 mmol/L can be used as the diagnostic threshold of DKA. βOHB can serve as an index for assessing the severity of DKA.</p>

Evaluation of the T-KB-H and 3-HB Kits for the Measurement of Serum Ketone and beta-Hydroxybutyric Acid

BACKGROUND: Diabetes mellitus and alcohol consumption are the most common causes of ketoacidosis in adults. Recently, beta-hydroxybutyric acid (betaHBA) was reported to be a potential serum biomarker in the diagnosis and monitoring of ketoacidosis. We evaluated the performance of T-KB-H and 3-HB kits for the measurement of ketone bodies [acetoacetate (AcAc)+betaHBA] and betaHBA, respectively. METHODS: Quantitative enzymatic assays were performed using the T-KB-H and 3-HB kits (Nittobo Medical Co., Japan) and the Architect ci16200 Integrated System (Abbott Laboratories, USA). Simultaneously, the ketone body levels in these serum samples were determined by gas chromatography-mas spectrometry (GC-MS). We evaluated precision and linearity of these kits and correlation with GC-MS, and established reference intervals in children and adults. RESULTS: The coefficients of variation for the T-KB-H and 3-HB kits were less than 4.0% at analyte levels of 50, 100, and 400 micromol/L. Linearity was observed for AcAc and betaHBA over a 0-1,000 micromol/L range (R2<0.99). Results from the T-KB-H and 3-HB kits were in good agreement with those from the GC-MS analysis, with correlation coefficients of 0.94 for AcAc and 0.96 for betaHBA. Reference intervals determined for the T-KB-H kit were 9.8-270.1 micromol/L and 18.5-531.8 micromol/L in children and adults, respectively. For the 3-HB kit, the reference intervals were 6.4-234.0 micromol/L and 16.0-437.2 micromol/L in children and adults, respectively. CONCLUSIONS: The T-KB-H and 3-HB kits displayed good precision, clinically acceptable linearity, and reliable correlation with an established assay. This indicates that the kits can be used clinically for measuring serum ketone bodies.

Accuracy of capillary blood 3-beta-hydroxybutyrate determination for the detection and treatment of canine diabetic ketoacidosis

In human medicine, diagnosis of diabetic ketoacidosis (DKA) is usually based on measurement of capillary 3-beta-hydroxybutyrate (3-HB) with a hand held ketone sensor. This study was conducted to determine if measurement of capillary 3-HB could be useful for the diagnosis and monitoring of canine DKA. Fifteen dogs with diabetic ketosis and 10 with DKA were evaluated. Paired measurements of 3-HB of capillary and venous blood samples were analysed by the electrochemical sensor and reference method. Use of capillary 3-HB measurement during DKA management was then evaluated through simultaneous measurements of capillary 3-HB, urinary AcAc and venous blood gas analysis. Good agreement between capillary and venous 3-HB measurement was detected by the electrochemical sensor and reference method. Monitoring treatment of DKA revealed a significant correlation between capillary 3-HB and acidosis markers, while no significant correlation was observed between AcAc and acidosis markers. A cut-off value of capillary blood 3-HB >3.8 mmol/L for diagnosis of DKA resulted in 70% and 92% sensitivity and specificity. The electrochemical sensor accurately measures 3-HB concentration in both capillary and venous blood samples, is accurate in diagnosing canine DKA, and appears to reflect the patient's metabolic status during DKA treatment.

novel chemical method for measuring ketone bodies in subclinical ketosis

Subclinical ketosis can cause greater economic loss due to a lack of clinical symptoms. The present study was aimed to design a chemical method for measuring serum ketone bodies in the affected subclinical cows. Acetoacetate concentrations were measured using a nitroprusside reaction and beta-hydroxy butyric acid [BHBA] which was oxidized to acetoacetate using nanosilver particles to determine its concentration. Recovery tests were done for different concentrations of betahydroxy butyrate in bovine pooled serum. Actoacetate levels were in range of 0.1-6 mM, and the values for BHBA were found to be in the range of 0.125-3 mM. It was concluded that this technique can be considered as a simple method for measuring ketone bodies in biological fluids