Background levels of neomorphic 2-hydroxyglutarate facilitate proliferation of primary fibroblasts.

Each cell types or tissues contain certain "physiological" levels of R-2-hydroxyglutarate (2HG), as well as enzymes for its synthesis and degradation. 2HG accumulates in certain tumors, possessing heterozygous point mutations of isocitrate dehydrogenases IDH1 (cytosolic) or IDH2 (mitochondrial) and contributes to strengthening their malignancy by inhibiting 2-oxoglutarate-dependent dioxygenases. By blocking histone de-methylation and 5-methyl-cytosine hydroxylation, 2HG maintains cancer cells de-differentiated and promotes their proliferation. However, physiological 2HG formation and formation by non-mutant IDH1/2 in cancer cells were neglected. Consequently, low levels of 2HG might play certain physiological roles. We aimed to elucidate this issue and found that compared to highest 2HG levels in hepatocellular carcinoma HepG2 cells and moderate levels in neuroblastoma SH-SY5Y cells, rat primary fibroblast contained low basal 2HG levels at early passages. These levels increased at late passage and likewise 2HG/2OG ratios dropped without growth factors and enormously increased at hypoxia, reaching levels compared to cancer HepG2 cells. Responses in SH-SY5Y cells were opposite. Moreover, external 2HG supplementation enhanced fibroblast growth. Hence, we conclude that low 2HG levels facilitate cell proliferation in primary fibroblasts, acting via hypoxia-induced factor regulations and epigenetic changes.

Antioxidant and regulatory role of mitochondrial uncoupling protein UCP2 in pancreatic beta-cells.

Research on brown adipose tissue and its hallmark protein, mitochondrial uncoupling protein UCP1, has been conducted for half a century and has been traditionally studied in the Institute of Physiology (AS CR, Prague), likewise UCP2 residing in multiple tissues for the last two decades. Our group has significantly contributed to the elucidation of UCP uncoupling mechanism, fully dependent on free fatty acids (FFAs) within the inner mitochondrial membrane. Now we review UCP2 physiological roles emphasizing its roles in pancreatic beta-cells, such as antioxidant role, possible tuning of redox homeostasis (consequently UCP2 participation in redox regulations), and fine regulation of glucose-stimulated insulin secretion (GSIS). For example, NADPH has been firmly established as being a modulator of GSIS and since UCP2 may influence redox homeostasis, it likely affects NADPH levels. We also point out the role of phospholipase iPLA2 isoform gamma in providing FFAs for the UCP2 antioxidant function. Such initiation of mild uncoupling hypothetically precedes lipotoxicity in pancreatic beta-cells until it reaches the pathological threshold, after which the antioxidant role of UCP2 can be no more cell-protective, for example due to oxidative stress-accumulated mutations in mtDNA. These mechanisms, together with impaired autocrine insulin function belong to important causes of Type 2 diabetes etiology.

Assessment of mitochondrial DNA as an indicator of islet quality: an example in Goto Kakizaki rats.

BACKGROUND: Diabetic Goto Kakizaki (GK) rats represent an established model of type 2 diabetes that exhibit an onset of pancreatic islet (PI) pathology characterized by islet hypertrophy with a decreased number of insulin-secreting ß-cells. Among the remaining ß-cells, oxidative phosphorylation (OXPHOS) and consequently glucose-stimulated insulin secretion (GSIS) are impaired, perhaps owing to a deficit in mitochondrial DNA (mtDNA). We sought to identify this abnormality. METHODS: ß-Cells were obtained from Accutase-dissolved PI isolated from GK or Wistar rats and sorted based on the positive Zn(2+) signal of Newport Green. The mtDNA copy number per cell was quantified as the amplicon ratio by polymerase chain reaction using specific primers against the rat ND5 mt gene and UCP2 nuclear gene. RESULTS: The 12-month-old GK rats exhibited drastically reduced copy numbers per remaining ß-cell, from 7,400 ± 600 in 12-month old Wistar rats (100%) to 24 ± 4%; mtDNA content in heart and liver was 70 ± 25% and 60 ± 20%, respectively. Versus age-paired Wistar rats, 6- and 4-month-old GK rats showed reductions to 60 ± 15% and 50 ± 20%, respectively. CONCLUSIONS: OXPHOS of remnant ß-cells in diabetic GK was drastically impaired due to the lack of sufficient mtDNA levels. We suggest the use of mtDNA quantification to quickly assess PI quality before transplantation.

Mitochondrial phospholipase A2 activated by reactive oxygen species in heart mitochondria induces mild uncoupling.

Homeostasis of reactive oxygen species (ROS) in cardiomyocytes is critical for elucidation of normal heart physiology and pathology. Mitochondrial phospholipases A2 (mt-PLA2) have been previously suggested to be activated by ROS. Therefore, we have attempted to elucidate physiological role of such activation. We have found that function of a specific i-isoform of mitochondrial phospholipase A2 (mt-iPLA2) is activated by tert-butylhydroperoxide in isolated rat heart mitochondria. Isoform specificity was judged from the inhibition by bromoenol lactone (BEL), a specific iPLA2 inhibitor. Concomitant uncoupling has been caused by free fatty acids, since it was inhibited by bovine serum albumin. The uncoupling was manifested as a respiration burst accompanied by a slight decrease in mitochondrial inner membrane potential. Since this uncoupling was sensitive to carboxyatractyloside and purine nucleotide di- and tri-phosphates, we conclude that it originated from the onset of fatty acid cycling mediated by the adenine nucleotide translocase (major contribution) and mitochondrial uncoupling protein(s) (minor contribution), respectively. Such a mild uncoupling may provide a feedback downregulation of oxidative stress, since it can further attenuate mitochondrial production of ROS. In conclusion, ROS-induced function of cardiac mt-iPLA2 may stand on a pro-survival side of ischemia-reperfusion injury.

[Relation between alcohol intake and some metabolic and cardiovascular risk factors in healthy men]. / Vztah mezi dietním príjmem alkoholu a nekterymi metabolickými a kardiovaskulárními rizikovými faktory u zdravých muzu.

BACKGROUND: Czech Republic belongs to countries with worldwide highest alcohol per capita intake. Although mild intake of alcohol may exhibit protective effects, its abuse is associated with true deleterious consequences for the whole organism. The aim of our study was to assess relation between alcohol intake and several metabolic and cardiovascular risk factors. METHODS AND RESULTS: In the group of clinically healthy men (n=102, mean age 39.1 +/- 10.8 years) complete clinical examination, biochemical work-up and weekly dietary profile assessment were performed. Data were compared using standard statistical tests and linear regression analyses. Participants were divided into 4 groups according to the regular weekly alcohol intake (group 0: <70 g; group 1: 70-210 g: group 2: 211-420 g; group 3: >420 g alcohol/week). We found close relation between alcohol intake and several risk factors for cardiovascular diseases and metabolic syndrome such as WHR, BMI, total caloric intake, blood pressure, serum lipids, or iron metabolites and markers of oxidative stress (AGEs, AOPP). CONCLUSIONS: This study proves an important relationship between alcohol intake and risk factors for cardiovascular diseases or metabolic syndrome. Subjects abusing alcohol thus seem not to be only at higher risk for alcohol-mediated liver damage, but also for the cardiovascular and common metabolic diseases.

Arachidonic acid and long-chain n-3 polyunsaturated fatty acid contents in meat of selected poultry and fish species in relation to dietary fat sources.

Arachidonic acid (AA) content, long-chain n-3 polyunsaturated fatty acid (PUFA) equivalent [LCE; calculated as 0.15 x linolenic acid (LA) + eicosapentaenoic acid (EPA) + docosahexaenoic acid (DHA)], and PUFA n-6/PUFA n-3 ratio were determined in meat [breast meat (BM), thigh meat (TM), and fillets (F), respectively] within four sets of chickens, five sets of turkeys, one set of common carp, and four sets of rainbow trout, fed either commercial diet or diets with manipulated PUFA n-3 and PUFA n-6 contents. AA content was within the range of 20 mg/100 g (F of rainbow trout fed the diet with linseed oil, LO) to 138 mg/100 g (TM of chickens fed restrictively the diet based on maize to the age of 90 days). AA content in BM of turkeys fed the diet with LO or fish oil (FO) did not differ (P > 0.05) from that of rainbow trout F. LCE was in the range of 16 mg/100 g (BM of turkeys fed a commercial feed mixture) to 681 mg/100 g (F of rainbow trout fed a commercial feed mixture). With regard to BM, only turkeys fed the diet with LO deposited more (P < 0.01) LCE (71 mg/100 g) as compared to all other poultry sets except turkeys fed the diet with FO (123 mg/100 g). Apart from all fish samples, also both BM and TM of turkeys fed the diet with either LO or FO met the recommended value of the PUFA n-6/PUFA n-3 ratio (<4). AA content in the tissue increased significantly (P < 0.001) with increasing dietary LA in both all chicken tissues and all turkey tissues, which is contrary to the suggested strong metabolic regulation of the AA formation. When all tissues within all animal species were taken as a one set, both AA percentage and EPA + DHA percentage in the tissue (Y, %) decreased (P < 0.001) with increasing fat content in the tissue (X, %), according to the equation Y = 4.7 - 0.54X (R (2) = 0.41) and Y = 6.0 - 0.33X (R (2) = 0.35), respectively. AA content in chicken BM, chicken TM, and turkey BM, respectively, decreased linearly (P < 0.01) with increasing live weight reached at the slaughter age.

Effect of age on utilization of selenium by chickens.

Utilization of selenium was examined in slow-growing laying-type chickens (SG) and in fast-growing broiler hybrids (FG) fed ad libitum on a diet with 265 microg of selenium/kg, including 128 microg of selenium added as sodium selenite. To 40 d of age, coefficients of selenium retention increased (P < 0.05) daily in the SG and FG groups by 0.76 and 0.61%, respectively. From 40 to 100 d, the regression coefficients were not significant. Coefficients of selenium retention and retention per unit of body gain were higher in SG chickens. The influence of age on selenium content in BW gain of birds was evident (P < 0.01). From 5 to 40 d, allometric coefficients were 1.444 and 1.070 for SG and FG, respectively, and from d 40 to 100 the corresponding values were 1.282 and 1.081, respectively.

Cholesterol content in meat of some poultry and fish species as influenced by live weight and total lipid content.

Total cholesterol content in 24 fillets (F) of males and females of common carp, 35 samples of male chicken breast meat (BM) and thigh meat (TM), and 48 samples of male turkey BM and TM, respectively, was determined by high-performance liquid chromatography after total lipid (TL) extraction using n-hexane/2-propanol mixture. Cholesterol content in male carp fillets (77.6 mg/100 g) was higher (P < 0.001) in comparison with females (69.4 mg/100 g). Irrespective of the sex differences, cholesterol content increased (P < 0.01) in the sequence chicken BM (53.0 mg/100 g) = turkey BM (53.0) < turkey TM (61.5) < carp F (73.5) < chicken TM (82.9 mg/100 g). Cholesterol content in chicken TM decreased (P < 0.05) with increasing live weight reached at the age of 43 days, but did not change (P > 0.05) in other tested tissues. Cholesterol concentration in TL of all five tested tissues within three animal species decreased sharply (P < 0.001) with increasing TL content reached in a given tissue at the fixed age. It follows from the results of the study that a two hundred gram portion of carp F and chicken TM without skin represents 49 and 55% of the upper limit of daily cholesterol intake, respectively.

[Effect of sodium chloride on elasticity of silicone matrices]. / Vliv chloridu sodného na elastické chování silikonových matic.

An increase in the availability of the active ingredient from silicone matrices can be achieved by using sodium chloride as the filler. An addition of sodium chloride also influences the mechanical properties of silicone systems. The temperature dependence of real Young's model E' of addition silicone systems with sodium chloride was measured. With increasing NaCl concentration in the silicone system, the values of Young's module E are increased. The increase is also influenced by the particle size of NaCl. The values of the module are moderately increased also with increasing temperature, which corresponds to the kinetic theory of caoutchouc elasticity. A significant increase in E takes place at NaCl concentrations higher than 20%. With a 50% NaCl content in the system, E is increased approximately 3.5 times in comparison with an unfilled sample. Various theories of elastic behaviour of composites can be employed for the description of the effect of the filler on the mechanical properties of silicone systems under small deformations. The effect of chloride is best described by the relation proposed by Nielsen. The value of the constant A in Nielsen's relation, dependent on the shape of the filler particles and Poisson's constant of the unfilled matrix, decreases with increasing particle size. With a change in particle size, a certain change in particle shape takes place. Hydrodynamic theory provides good results for sodium chloride particle size of 102.5 microns, but for particle sizes of 20 microns and 60 microns it is less successful than Nielsen's relation.

[Effect of fillers on the strength of silicone systems]. / Vliv plniv na pevnostní chování silikonových systému.

The present paper examines the strength behaviour of addition silicone systems composed of Lukopren A 6200 and three inorganic fillers (Skron SF 600, Cab-O-Sil LM 150, sodium chloride) in different concentrations and particle sizes. The tensile strength sigma (MPa) and the relative lengthening of samples on rupture epsilon (%) were measured. With increasing concentration of the filler in the system relative lengthening of matrices is decreased. This trend corresponds with the theoretical hypotheses and it is understandable also with regard to the fact that the filler, in comparison with silicone rubber, possesses very low ductility. An addition of Sikron increases tensile strength of samples. Dependence of tensile strength on the amount of Sikron in the system is linear. This behaviour is typical of fibrous composites and the composites with high adhesion between the filler and matrix. Concentrations of Cab-O-Sil up to 5% did not exert an effect on tensile strength. An important increase in strength was shown only by the systems containing 10% of Cab-O-Sil. Experimental dependence of tensile strength on the amount of Cab-O-Sil in the system can be described by the equation for composites with Hookean behaviour till destruction. The effect of sodium chloride on tensile strength is more complicated. In the systems with NaCl particles of 20 microns, strength is not practically changed up to a content of 40% of NaCl in the system. In larger size fractions, tensile strength is decreased, and with a change in particle size, shape is probably changed as well.

Quartz strip resonators as a temperature sensor.

The miniature frequency-temperature sensor in the form of a small quartz strip vibrating in the thickness-shear mode with the resonant frequency in the range from 4 to 8 MHz is considered. Y-cut strips rotated in the range from 0 degrees to 27 degrees are the subject of the study. The temperature coefficients of the resonance frequency for the different rotation angles are given. The different temperature dependence of the resonant frequency of the quartz temperature sensors vibrating on the fundamental and harmonic frequency of the thickness-shear modes is pointed out.

Influence of electrodes arrangement on the quartz resonator equivalent circuit.

The strip electrodes deposited on the main surfaces of the rotated Y-cut quartz plate in the Z' direction and overlapping only in the central part of the plate excite a non-negligible electric field parallel to the surface of the plate. The influence of this field on the parameters of the electrical equivalent circuit and frequency-temperature characteristic of the AT-cut quartz plates is examined. The influence of this electric field parallel to the surface depends on the orientation of electrodes in the rectangular coordinate system of quartz.

Dynamics and distribution of radiocaesium in broiler chicken.

The distribution and biological half-life of radiocaesium (137Cs) in broiler chickens after three oral applications (in course of 1 day at the age of 14 days) of artificially contaminated feed mixture were studied. There was a rapid uptake of the orally administered 137Cs (within a few hours) and also a rapid loss of 137Cs which varied in the different organs (the initial biological half-life was: liver 0.6 day, intestine 0.6 day, breast meat 2 days, leg meat 1.2 days). More than one-half of the total administered 137Cs activity (55%) was excreted from the body within the 1st day after dosage, and after 14 days more than 90% had been excreted. The highest accumulation of 137Cs occurred in meat (50%-90%), and the proportion of total activity in breast and leg meat varied during decontamination. The transfer of radiocaesium from feed into the chicken body (measured as ratios of the 137Cs activity concentrations in the organ to the 137Cs activity concentration in the applied dose) 1 day after application was: 0.0220, 0.0294, 0.0216 and 0.0195 for breast meat, leg meat, intestine and liver, respectively. Significant differences between the values were demonstrated (P < 0.05) except between those of breast meat and intestine. For the first 3 days there was a higher proportion of 137Cs activity in leg meat, whereas from the 4th day a greater part of total activity was found in breast meat. The latter results were confirmed in a subsequent study. Data from this study suggest that if broiler chickens are contaminated by radiocaesium to a level of 5 kBq/chicken in the course of 1 day at the age of 14 days, then immediate feeding with uncontaminated feed mixture for 18 days should be effective in decontaminating the chicken's meat below the intervention levels for radiocaesium in animal products, i.e. below 1000 Bq. kg-1.

Effects of sex, age and food intake upon metabolisable energy values in broiler chickens.

1. Nitrogen-corrected apparent metabolisable energy (AME[n]) values of 2 diets with different energy:protein ratio were estimated in an experiment with 8 groups of female and 8 of male chickens in 15 sequential 3-day balance periods from the 12th to the 56th d of age. 2. The effect of sex on AME(n) values was not significant. 3. The effect of age was highly significant. AME(n) values of the mixture with the narrower energy:protein ratio increased with age. The dependence of AME(n) of the diet with the wider energy:protein ratio on age was parabolic, AME(n) values increasing only until the 37th day of life. 4. Under conditions of ad libitum feeding, AME(n) of the mixture with the wider energy:protein ratio given to female chickens decreased significantly with increasing food intake.

Minimal forces to move patients.

OBJECTIVES: Health care workers who have patient transfer duties are at risk for back injury. Transferring patients between beds and gurneys is a rigorous pulling task that requires back, leg, and arm strength. This study analyzed the efficiency of commercially available transfer devices, namely a patient roller, patient shifter, and draw sheet. METHOD: With the use of one or two force scales attached to each transfer device, the amount of force needed to transfer 15 participants, weighing from 101 lb to 240 lb, back and forth between a bed and a gurney was measured. Ten transfers per device per participant were performed. RESULTS: The patient roller was superior to the patient shifter and draw sheet in reducing transfer forces. Additionally, gurney-to-bed transfers tended to be more demanding for all transfer devices and for heavier participants. CONCLUSIONS: The patient roller was the most efficient transfer device in moving participants compared with the draw sheet and patient shifter. Transfer forces can be estimated with the use of linear equations, with patient weight, direction of transfer, and transfer device as the independent variables. These estimated forces can assist occupational therapists in the returning their injured health care coworkers to patient transfer duties. The results further indicated that high forces are required to transfer patients; therefore, patient-transfer personnel should obtain assistance when moving patients.