Effect of 6-aminonicotinamide on metabolism of astrocytes and C6-glioma cells

Brain tissue cells have been shown to use two predominant pathways for energy production. The first of these is the pentose phosphate shunt, and the second is glycolysis, followed by the TCA cycle. Inhibition of these pathways can result in a reduction of ATP, and changes in the concentration of various metabolites. In the present study, the acute and chronic effect of 6-aminonicotinamide (6-AN) (0.01, 0.02, and 0.03 mg/ml) was examined on astrocytes and C6-glioma cells. Following this treatment, glucose, lactate, glutamate, ATP, and PCr were assayed according to the procedures of Lowry and Passonneau. Our data indicated that following 15 minutes treatment of astrocytes and C6-glioma with 6AN there was no significant difference in the concentration of metabolites measured. However, following 24 hours treatment there was a significant increase in glucose concentration and significant reduction in the concentration of ATP, PCr, lactate and glutamate in both cell types. Morphological changes appeared later following 48 hours treatment with 6-AN in both cell types. Glucose accumulation can be explained by the fact that it is the precursor to both glycolysis and the pentose phosphate shunt. If these processes are inhibited, glucose will obviously accumulate and products like ATP, PCr, lactate and glutamate will decrease. Additionally, there was significant differences in concentration of glucose and lactate between astrocytes and C6-glioma cells. The significance of these differences has been discussed.     

尿百草枯与动脉血乳酸浓度对急性百草枯中毒的预后评估

目的：观察急性百草枯中毒（APP）患者入院时尿百草枯（PQ）浓度与动脉血乳酸（Lac）浓度变化,探讨APP 患者预后特异度和敏感度较高的监测指标。方法收集我院急诊内科2012年1月－2015年1月 APP 患者142例的临床资料,根据中毒后30 d 是否死亡分为死亡组与存活组,记录入院时尿 PQ 浓度和血 Lac 浓度,通过 ROC 曲线评估其对 APP 预后的诊断价值。结果死亡组患者入院时尿 PQ 浓度和血 Lac 浓度明显高于存活组（P ＜0．05）;两者对 APP 患者死亡预测的 ROC 曲线下面积（AUC）显示,尿 PQ 浓度的 AUC 为0．843,截断值为50．12 mg／L,敏感度为78．6％,特异度为88．5％,Youden 指数0．671;血 Lac 浓度的 AUC 为0．782,截断值为3．76 mmol／L,敏感度为85．4％,特异度为76．7％,Youden 指数0．621。结论入院时尿 PQ 浓度（≥50．12 mg／L）和（或）血 Lac 浓度（≥3．76 mmol／L）可作为预测 APP 患者预后的参考指标。     

DNA Sequence Polymorphism of the Lactate Dehydrogenase Genefrom Iranian Plasmodium vivax and Plasmodium falciparum Isolates

Background:

Parasite lactate dehydrogenase (pLDH) is extensively employed as malaria rapid diagnostic tests (RDTs). Moreover, it is a well-known drug target candidate. However, the genetic diversity of this gene might influence performance of RDT kits and its drug target candidacy. This study aimed to determine polymorphism of pLDH gene from Iranian isolates of P. vivax and P. falciparum.

Methods:

Genomic DNA was extracted from whole blood of microscopically confirmed P. vivax and P. falciparum infected patients. pLDH gene of P. falciparum and P. vivax was amplified using conventional PCR from 43 symptomatic malaria patients from Sistan and Baluchistan Province, Southeast Iran from 2012 to 2013.

Results:

Sequence analysis of 15 P. vivax LDH showed fourteen had 100% identity with P. vivax Sal-1 and Belem strains. Two nucleotide substitutions were detected with only one resulted in amino acid change. Analysis of P. falciparum LDH sequences showed six of the seven sequences had 100% homology with P. falciparum 3D7 and Mzr-1. Moreover, PfLDH displayed three nucleotide changes that resulted in changing only one amino acid. PvLDH and PfLDH showed 75%–76% nucleotide and 90.4%–90.76% amino acid homology.

Conclusion:

pLDH gene from Iranian P. falciparum and P. vivax isolates displayed 98.8–100% homology with 1–3 nucleotide substitutions. This indicated this gene was relatively conserved. Additional studies can be done weather this genetic variation can influence the performance of pLDH based RDTs or not.     

Importance of exogenous substrates for the energy production of adult rat heart myocytes

The ability of freshly isolated Ca²⁺ tolerant heart myocytes to derive its energy requirements from combinations of exogenously provided glucose, lactate and palmitate was studied. When the substrates were added individually to the incubation medium, myocytes oxidized 5 mm glucose (2.3 ± 0.2 nmol min^?1 mg^?1 protein), 1.0 mm lactate (6.20 ± 0.5), or 0.4 mm palmitate (0.55 ± 0.07) at rates which were several fold higher than previously reported. Palmitate or lactate decreased glucose oxidation by 25% and 50%, respectively, while together they inhibited glucose oxidation rates by 70%. The rate of lactate oxidation was decreased by 50% in the presence of glucose or palmitate. Palmitate oxidation was least affected by the alternative substrates studied. Oxygen consumption was substrate dependent, ranging from 17.9 ± 0.9 nmol min^?1 mg^?1 in the presence of glucose alone, to 29.5 ± 1.3 when both lactate and palmitate were present in the medium. The high rates of exogenous substrate oxidation observed in this study accounted for a major portion of myocyte oxygen consumption, but the combined oxidation of multiple substrates was necessary to supply all their energy requirements.     

High energy phosphate compounds and mitochondrial function in ischemic myocardium of swine with advanced coronary atherosclerosis

High energy compounds (adenosine triphosphate and creatine phosphate), lactate and mitochondrial function and morphology were investigated in swine suffering from persistent myocardial ischemia. The myocardial ischemia was secondary to severe diffuse coronary artery atherosclerosis induced by hypercholesterolemia enhanced by X-irradiation to the heart.In the ischemic myocardia, the high energy phosphate compounds were decreased up to 50% and lactate was increased threefold. The ischemia apparently had also damaged myocardial mitochondria; QO₂ and ACR were lower in mitochondrial preparations from the ischemic tissue. In addition, electron microscopy showed more swollen and broken forms in mitochondria preparations from the ischemic myocardium. Hypercholesterolemia alone or irradiation alone to other groups of swine did not induce alterations in QO₂ or ACR, although the percentage recovery of mitochondria was decreased by X-irradiation to the heart.We cannot yet relate the findings in this study to the ventricular fibrillation and sudden death that these swine are prone to develop. A possibility that might be investigated is that the decrease in high energy compounds impairs function of ion transport systems (e.g. Na-K-ATPase). The resultant disturbances in ion transport may enhance the development of fibrillation. Despite the abnormalities of mitochondria in the myocardia of the ischemic swine, there were no manifestations of congestive heart failure in this group.     

Biochemistry tests in hospitalized COVID-19 patients: Experience from a Canadian tertiary care centre

BackgroundCoronavirus Disease 2019 (COVID-19) has variable clinical presentation, from asymptomatic to severe disease leading to death. Biochemical markers may help with management and prognostication of COVID-19 patients; however, their utility is still under investigation.MethodsA retrospective study was conducted to evaluate alanine aminotransferase, C-reactive protein (CRP), ferritin, lactate, and high sensitivity troponin T (TnT) levels in 67 patients who were admitted to a Canadian tertiary care centre for management of COVID-19. Logistic, cause-specific Cox proportional-hazards, and accelerated failure time regression modelling were performed to assess the associations of initial analyte concentrations with in-hospital death and length of stay in hospital; joint modelling was performed to assess the associations of the concentrations over the course of the hospital stay with in-hospital death.ResultsInitial TnT and CRP concentrations were associated with length of stay in hospital. Eighteen patients died (27%), and the median initial TnT concentration was higher in patients who died (55 ng/L) than those who lived (16 ng/L; P < 0.0001). There were no survivors with an initial TnT concentration > 64 ng/L. While the initial TnT concentration was predictive of death, later measurements were not. Only CRP had prognostic value with both the initial and subsequent measurements: a 20% increase in the initial CRP concentration was associated with a 14% (95% confidence interval (CI): 1–29%) increase in the odds of death, and the hazard of death increased 14% (95% CI: 5–25%) for each 20% increase in the current CRP value. While the initial lactate concentration was not predictive of death, subsequent measurements were.ConclusionCRP, lactate and TnT were associated with poorer outcomes and appear to be useful biochemical markers for monitoring COVID-19 patients.     

Oxidation of palmitate and lactate by beating myocytes isolated from adult dog heart.

Myocytes were isolated from hearts of adult dogs by repeated digestion with collagenase and hyaluronidase in Ca²⁺-free phosphate buffer followed by centrifugation in the presence of Ficoll. The isolated myocytes were free of other tissue elements and were contracting autorhythmically. The oxidation of palmitate and lactate were studied by incubating cell preparation with [1-¹⁴C]palmitate or [U-¹⁴C]lactate either in the presence or absence of CaCl₂ for 30 min at 37°C. [¹⁴C]O₂ was collected by KOH-soaked filter paper. Cardiac myocytes oxidized 0.1 to 0.18 nmol/mg protein min of palmitate and 1.5 to 2.2 nmol/mg protein min of lactate. The oxidation was linear for 75 min. Maximal rate of oxidation was obtained at substrate concentrations of 0.2 mm palmitate or 5.0 mm lactate. Adenine or pyridine nucleotides had no effect on the oxidation of these substrates. Palmitate oxidation was enhanced by carnitine and also by an increase in the molar ratio of palmitate/albumin, but was depressed (15 to 50%) by CaCl₂ (0.025 to 1.0 mm). CaCl₂ (0.05 to 1.0 mm) increased the rate of lactate oxidation by 50 to 68%. Lactate oxidation was also enhanced (68 to 87%) in the presence of palmitate (0.4 to 0.8 mm). These findings demonstrate the suitability of isolated adult heart myocytes for the study of cardiac metabolism.     

日本血吸虫乳酸脱氢酶(SjLDH)结构与功能的生物信息学分析

目的 应用生物信息学分析软件预测日本血吸虫乳酸脱氢酶（SjLDH）氨基酸序列的结构与功能。 方法 应用NCBI、Expasy等在线生物信息学网站及Vector NTI、PCgene等软件包分析SjLDH与其他物种的同源序列，进行多序列同源比对，分析相同的保守位点及催化活性位点，构建分子进化树；预测二级结构、拓扑结构；同源模建预测三级结构；预测主要抗原表位等。 结果 SjLDH与其他物种的同源序列含相同的保守位点及催化活性位点，与华支睾吸虫LDH（CsLDH）同源性最高为75％，与阴道毛滴虫LDH（TvLDH）同源性最低为17%，与人LDH（HsLDH-A，-B，-C）的同源性为58%~60％； SjLDH与果蝇（DmLDH）的进化距离较秀丽隐杆线虫（CeLDH）为近，3种人LDH中与HsLDH-B、-C的进化距离较HsLDH-A为近；该蛋白具有3个跨膜区域，3个高亲水性区域，主要的线性表位98aa~106aa位于膜外，与原虫LDH相同区域差异显著，而与其他LDH有1~3个氨基酸的差异，关键催化位点之一及底物丙酮酸结合区域位于该区域，蛋白质同源模建分析表明该区域位于蛋白表面形成环状结构，3个关键催化位点位于该区域或在其附近。 结论 生物信息学预测结果提示LDH是研究物种分子进化理想的分子；SjLDH可能是免疫诊断、药物作用及疫苗潜在的靶分子。     

潜在致病性的弗氏枸橼酸杆菌的筛查

目的 筛查具有潜在致病性的弗氏枸橼酸杆菌。方法 对从河南省睢县分离到的36株弗氏枸橼酸杆菌进行黏附HEp-2细胞的检测,以及与HEp-2细胞(MOI: 100)作用10 h后的乳酸脱氢酶(LDH)释放情况分析。同时比较弗氏枸橼酸杆菌CF74(Citrobacter freundii 74)、CF72(Citrobacter freundii 72)、肠聚集性大肠埃希菌(Enteroaggregative E. coli,EAEC)、肠致病性大肠埃希菌(Enteropathogenic E. coli,EPEC)2348/69和HB101的黏附和细胞毒性差异。结果 16株弗氏枸橼酸杆菌几乎没有黏附性,黏附指数1;18株弗氏枸橼酸杆菌具有中等强度的黏附性,黏附指数100;2株弗氏枸橼酸杆菌CF74和CF65(Citrobacter freundii 65)具有强的黏附性。与HEp-2细胞作用10 h后,除了CF74(24.4%),其余的35株弗氏枸橼酸杆菌具有低的LDH释放量,LDH释放量与阴性对照HB101(5.02%)相似。说明除了CF74,其余的弗氏枸橼酸杆菌不具有细胞毒性。CF74具有和EAEC042一样的聚集性黏附类型,并且 CF74引起的LDH释放率明显高于CF72、2348/69和HB101(P0.01)而低于EAEC17-2。结论 作为潜在的致病菌,CF74具有强的黏附性和细胞毒性。