Pyruvate carboxylase (PC) is a mitochondrial, biotin-containing enzyme catalyzing the ATP-dependent synthesis of oxaloacetate from pyruvate and bicarbonate, with a critical anaplerotic role in sustaining the brain metabolism. Based on the studies performed on animal models, PC expression was assigned to be glia-specific. To study PC distribution among human neural cells, we probed the cultured human astrocytes and brain sections with antibodies against PC. Additionally, we tested the importance of PC for the viability of cultured human astrocytes by applying the PC inhibitor 3-chloropropane-1,2-diol (CPD). Our results establish the expression of PC in mitochondria of human astrocytes in culture and brain tissue and also into a subpopulation of the neurons in situ. CPD negatively affected the viability of astrocytes in culture, which could be partially reversed by supplementing media with malate, 2-oxoglutarate, citrate, or pyruvate. The provided data estimates PC expression in human astrocytes and neurons in human brain parenchyma. Furthermore, the enzymatic activity of PC is vital for sustaining the viability of cultured astrocytes.

• MeSH
• astrocyty * metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• lidé MeSH
• mozek metabolismus   MeSH
• neurony metabolismus   MeSH
• pyruvátkarboxylasa * metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kyselina pyrohroznová MeSH
• pyruvátkarboxylasa * MeSH

Pseudomonas putida KT2440 is an attractive bacterial host for biotechnological production of valuable chemicals from renewable lignocellulosic feedstocks as it can valorize lignin-derived aromatics or glucose obtainable from cellulose. P. putida EM42, a genome-reduced variant of strain KT2440 endowed with advantageous physiological properties, was recently engineered for growth on cellobiose, a major cellooligosaccharide product of enzymatic cellulose hydrolysis. Co-utilization of cellobiose and glucose was achieved in a mutant lacking periplasmic glucose dehydrogenase Gcd (PP_1444). However, the cause of the co-utilization phenotype remained to be understood and the Δgcd strain had a significant growth defect. In this study, we investigated the basis of the simultaneous uptake of the two sugars and accelerated the growth of P. putida EM42 Δgcd mutant for the bioproduction of valuable compounds from glucose and cellobiose. We show that the gcd deletion lifted the inhibition of the exogenous β-glucosidase BglC from Thermobifida fusca exerted by the intermediates of the periplasmic glucose oxidation pathway. The additional deletion of hexR gene, which encodes a repressor of the upper glycolysis genes, failed to restore rapid growth on glucose. The reduced growth rate of the Δgcd mutant was partially compensated by the implantation of heterologous glucose and cellobiose transporters (Glf from Zymomonas mobilis and LacY from Escherichia coli, respectively). Remarkably, this intervention resulted in the accumulation of pyruvate in aerobic P. putida cultures. We demonstrated that the excess of this key metabolic intermediate can be redirected to the enhanced biosynthesis of ethanol and lactate. The pyruvate overproduction phenotype was then unveiled by an upgraded genome-scale metabolic model constrained with proteomic and kinetic data. The model pointed to the saturation of glucose catabolism enzymes due to unregulated substrate uptake and it predicted improved bioproduction of pyruvate-derived chemicals by the engineered strain. This work sheds light on the co-metabolism of cellulosic sugars in an attractive biotechnological host and introduces a novel strategy for pyruvate overproduction in bacterial cultures under aerobic conditions.

• Klíčová slova
• Cellobiose, Co-utilization of sugars, Glucose, Metabolic engineering, Metabolic model, Pseudomonas putida, Pyruvate,
• MeSH
• celobiosa metabolismus   MeSH
• celulosa metabolismus   MeSH
• Escherichia coli metabolismus   MeSH
• glukosa metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• metabolické inženýrství MeSH
• proteiny přenášející monosacharidy genetika   metabolismus   MeSH
• proteiny z Escherichia coli * genetika   MeSH
• proteomika MeSH
• Pseudomonas putida * genetika   metabolismus   MeSH
• symportéry * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• celobiosa MeSH
• celulosa MeSH
• glukosa MeSH
• kyselina pyrohroznová MeSH
• LacY protein, E coli MeSH Prohlížeč
• proteiny přenášející monosacharidy MeSH
• proteiny z Escherichia coli * MeSH
• symportéry * MeSH

In evolution, genes survived that could code for metabolic pathways, promoting long term survival during famines or fasting when suffering from trauma, disease or during physiological growth. This requires utilization of substrates, already present in some form in the body. Carbohydrate stores are limited and to survive long, their utilization is restricted to survival pathways, by inhibiting glucose oxidation and glycogen synthesis. This leads to insulin resistance and spares muscle protein, because being the main supplier of carbon for new glucose production. In these survival pathways, part of the glucose is degraded in glycolysis in peripheral (muscle) tissues to pyruvate and lactate (Warburg effect), which are partly reutilized for glucose formation in liver and kidney, completing the Cori-cycle. Another part of the glucose taken up by muscle contributes, together with muscle derived amino acids, to the production of substrates consisting of a complete amino acid mix but extra non-essential amino acids like glutamine, alanine, glycine and proline. These support cell proliferation, matrix deposition and redox regulation in tissues, specifically active in host response and during growth. In these tissues, also glucose is taken up delivering glycolytic intermediates, that branch off and act as building blocks and produce reducing equivalents. Lactate is also produced and released in the circulation, adding to the lactate released by muscle in the Cori-cycle and completing secondary glucose cycles. Increased fluxes through these cycles lead to modest hyperglycemia and hyperlactatemia in states of healthy growth and disease and are often misinterpreted as induced by hypoxia.

• Klíčová slova
• Cori-cycle, Growth, Inflammation, Insulin resistance, Trauma/Disease, Warburg effect,
• MeSH
• glukosa metabolismus   MeSH
• glykolýza fyziologie   MeSH
• játra metabolismus   MeSH
• kosterní svaly metabolismus   MeSH
• kyselina mléčná metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• ledviny metabolismus   MeSH
• lidé MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• glukosa MeSH
• kyselina mléčná MeSH
• kyselina pyrohroznová MeSH

Introduced about a century ago, suramin remains a frontline drug for the management of early-stage East African trypanosomiasis (sleeping sickness). Cellular entry into the causative agent, the protozoan parasite Trypanosoma brucei, occurs through receptor-mediated endocytosis involving the parasite's invariant surface glycoprotein 75 (ISG75), followed by transport into the cytosol via a lysosomal transporter. The molecular basis of the trypanocidal activity of suramin remains unclear, but some evidence suggests broad, but specific, impacts on trypanosome metabolism (i.e. polypharmacology). Here we observed that suramin is rapidly accumulated in trypanosome cells proportionally to ISG75 abundance. Although we found little evidence that suramin disrupts glycolytic or glycosomal pathways, we noted increased mitochondrial ATP production, but a net decrease in cellular ATP levels. Metabolomics highlighted additional impacts on mitochondrial metabolism, including partial Krebs' cycle activation and significant accumulation of pyruvate, corroborated by increased expression of mitochondrial enzymes and transporters. Significantly, the vast majority of suramin-induced proteins were normally more abundant in the insect forms compared with the blood stage of the parasite, including several proteins associated with differentiation. We conclude that suramin has multiple and complex effects on trypanosomes, but unexpectedly partially activates mitochondrial ATP-generating activity. We propose that despite apparent compensatory mechanisms in drug-challenged cells, the suramin-induced collapse of cellular ATP ultimately leads to trypanosome cell death.

• Klíčová slova
• Trypanosoma brucei, differentiation, drug action, drug mechanisms, energy homeostasis, glycosomes, metabolomics, parasite metabolism, polypharmacology, proteomics, sleeping sickness, suramin, trypanosome,
• MeSH
• adenosintrifosfát metabolismus   MeSH
• energetický metabolismus účinky léků   MeSH
• flagella účinky léků   metabolismus   ultrastruktura   MeSH
• glykolýza účinky léků   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• metabolom účinky léků   MeSH
• mikrotělíska účinky léků   metabolismus   ultrastruktura   MeSH
• mitochondrie účinky léků   metabolismus   ultrastruktura   MeSH
• molekulární modely MeSH
• prolin metabolismus   MeSH
• proteom metabolismus   MeSH
• protonové ATPasy metabolismus   MeSH
• protozoální proteiny metabolismus   MeSH
• suramin farmakologie   MeSH
• Trypanosoma brucei brucei metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adenosintrifosfát MeSH
• kyselina pyrohroznová MeSH
• prolin MeSH
• proteom MeSH
• protonové ATPasy MeSH
• protozoální proteiny MeSH
• suramin MeSH

The possibility to use leptin therapeutically for lowering glucose levels in patients with type 1 diabetes has attracted interest. However, earlier animal models of type 1 diabetes are severely catabolic with very low endogenous leptin levels, unlike most patients with diabetes. Here, we aim to test glucose-lowering effects of leptin in novel, more human-like murine models. We examined the glucose-lowering potential of leptin in diabetic models of two types: streptozotocin-treated mice and mice treated with the insulin receptor antagonist S961. To prevent hypoleptinemia, we used combinations of thermoneutral temperature and high-fat feeding. Leptin fully normalized hyperglycemia in standard chow-fed streptozotocin-treated diabetic mice. However, more humanized physiological conditions (high-fat diets or thermoneutral temperatures) that increased adiposity - and thus also leptin levels - in the diabetic mice abrogated the effects of leptin, i.e., the mice developed leptin resistance also in this respect. The glucose-lowering effect of leptin was not dependent on the presence of the uncoupling protein-1 and was not associated with alterations in plasma insulin, insulin-like growth factor 1, food intake or corticosterone but fully correlated with decreased plasma glucagon levels and gluconeogenesis. An important implication of these observations is that the therapeutic potential of leptin as an additional treatment in patients with type 1 diabetes is probably limited. This is because such patients are treated with insulin and do not display low leptin levels. Thus, the potential for a glucose-lowering effect of leptin would already have been attained with standard insulin therapy, and further effects on blood glucose level through additional leptin cannot be anticipated.

• Klíčová slova
• glucagon, insulin receptor antagonist, leptin, thermoneutrality, type 1 diabetes, uncoupling protein 1,
• MeSH
• bílá tuková tkáň metabolismus   MeSH
• diabetes mellitus 1. typu metabolismus   MeSH
• experimentální diabetes mellitus metabolismus   MeSH
• glukagon metabolismus   MeSH
• glukoneogeneze MeSH
• hnědá tuková tkáň metabolismus   MeSH
• insulinu podobný růstový faktor I metabolismus   MeSH
• inzulin metabolismus   MeSH
• kortikosteron metabolismus   MeSH
• krevní glukóza účinky léků   metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• leptin metabolismus   farmakologie   MeSH
• lidé MeSH
• modely nemocí na zvířatech MeSH
• myši knockoutované MeSH
• myši MeSH
• peptidy farmakologie   MeSH
• přijímání potravy MeSH
• receptor inzulinu antagonisté a inhibitory   MeSH
• spotřeba kyslíku MeSH
• transkriptom MeSH
• uncoupling protein 1 genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glukagon MeSH
• insulin-like growth factor-1, mouse MeSH Prohlížeč
• insulinu podobný růstový faktor I MeSH
• inzulin MeSH
• kortikosteron MeSH
• krevní glukóza MeSH
• kyselina pyrohroznová MeSH
• LEP protein, human MeSH Prohlížeč
• leptin MeSH
• peptidy MeSH
• receptor inzulinu MeSH
• S961 peptide MeSH Prohlížeč
• Ucp1 protein, mouse MeSH Prohlížeč
• uncoupling protein 1 MeSH

The aim of the study was to screen Yarrowia lipolytica strains for keto acid production and determine optimal conditions for pyruvic acid biosynthesis from glycerol by the best producer. The analyzed parameters were thiamine concentration, medium pH, stirring speed, and substrate concentration. The screening was performed in flask cultures, whereas pyruvic acid production was carried out in 5-L stirred-tank reactor with 2 L of working volume. In total, 24 Y. lipolytica strains were compared for their abilities to produce pyruvic and α-ketoglutaric acids. The total concentration of both acids ranged from 0.1 to 15.03 g/L. Ten strains were selected for keto acid biosynthesis in bioreactor. The Y. lipolytica SKO 6 strain was identified as the best producer of pyruvic acid. In the selected conditions (thiamine concentration 1.5 μg/L, pH 4.0, stirring speed 800 rpm, 150 g/L of glycerol), the strain Y. lipolytica SKO 6 produced 99.3 g/L of pyruvic acid, with process yield of 0.63 g/g and volumetric production rate of 1.18 g/L/h. Higher titer of pyruvic acid was obtained during fed-batch culture with 200 g/L of glycerol, reaching 125.8 g/L from pure glycerol (yield 0.68 g/g) and 124.4 g/L from crude glycerol (yield 0.62 g/g). Results obtained for the strain Y. lipolytica SKO 6 proved the suitability of microbial production of pyruvic acid at industrial scale.

• MeSH
• bioreaktory MeSH
• glycerol analýza   metabolismus   MeSH
• kultivační média chemie   MeSH
• kyselina pyrohroznová analýza   metabolismus   MeSH
• kyseliny ketoglutarové analýza   metabolismus   MeSH
• techniky vsádkové kultivace MeSH
• thiamin analýza   MeSH
• Yarrowia růst a vývoj   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• glycerol MeSH
• kultivační média MeSH
• kyselina pyrohroznová MeSH
• kyseliny ketoglutarové MeSH
• thiamin MeSH

Adequate concentrations of ATP are required to preserve physiological cell functions and protect tissue from hypoxic damage. Decreased oxygen concentration results in ATP synthesis relying increasingly on the presence of phosphocreatine. The lack of ATP through hypoxic insult to neurons that generate or regulate respiratory function, would lead to the cessation of breathing (apnea). It is not clear whether creatine plays a role in maintaining respiratory phrenic nerve (PN) activity during hypoxic challenge. The aim of the study was to test the effects of exogenously applied creatine or creatine pyruvate in maintaining PN induced respiratory rhythm against the deleterious effects of severe hypoxic insult using Working Heart-Brainstem (WHB) preparations of juvenile Swiss type mice. WHB's were perfused with control perfusate or perfusate containing either creatine [100μM] or creatine pyruvate [100μM] prior to hypoxic challenge and PN activity recorded throughout. Results showed that severe hypoxic challenge resulted in an initial transient increase in PN activity, followed by a reduction in that activity leading to respiratory apnea. The results demonstrated that perfusing the WHB preparation with creatine or creatine pyruvate, significantly reduced the onset of apnea compared to control conditions, with creatine pyruvate being the more effective substance. Overall, creatine and creatine pyruvate each produced time-dependent degrees of protection against severe hypoxic-induced disturbances of PN activity. The underlying protective mechanisms are unknown and need further investigations.

• Klíčová slova
• Creatine, Creatine pyruvate, Hypoxia, Working heart-brainstem preparation,
• MeSH
• dýchací soustava inervace   metabolismus   MeSH
• hypoxie metabolismus   MeSH
• kreatin metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• myši MeSH
• nervus phrenicus metabolismus   MeSH
• pohybová aktivita MeSH
• stárnutí metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kreatin MeSH
• kyselina pyrohroznová MeSH

Remodeling of the distal pulmonary artery wall is a characteristic feature of pulmonary hypertension (PH). In hypoxic PH, the most substantial pathologic changes occur in the adventitia. Here, there is marked fibroblast proliferation and profound macrophage accumulation. These PH fibroblasts (PH-Fibs) maintain a hyperproliferative, apoptotic-resistant, and proinflammatory phenotype in ex vivo culture. Considering that a similar phenotype is observed in cancer cells, where it has been associated, at least in part, with specific alterations in mitochondrial metabolism, we sought to define the state of mitochondrial metabolism in PH-Fibs. In PH-Fibs, pyruvate dehydrogenase was markedly inhibited, resulting in metabolism of pyruvate to lactate, thus consistent with a Warburg-like phenotype. In addition, mitochondrial bioenergetics were suppressed and mitochondrial fragmentation was increased in PH-Fibs. Most importantly, complex I activity was substantially decreased, which was associated with down-regulation of the accessory subunit nicotinamide adenine dinucleotide reduced dehydrogenase (ubiquinone) Fe-S protein 4 (NDUFS4). Owing to less-efficient ATP synthesis, mitochondria were hyperpolarized and mitochondrial superoxide production was increased. This pro-oxidative status was further augmented by simultaneous induction of cytosolic nicotinamide adenine dinucleotide phosphate reduced oxidase 4. Although acute and chronic exposure to hypoxia of adventitial fibroblasts from healthy control vessels induced increased glycolysis, it did not induce complex I deficiency as observed in PH-Fibs. This suggests that hypoxia alone is insufficient to induce NDUFS4 down-regulation and constitutive abnormalities in complex I. In conclusion, our study provides evidence that, in the pathogenesis of vascular remodeling in PH, alterations in fibroblast mitochondrial metabolism drive distinct changes in cellular behavior, which potentially occur independently of hypoxia.

• Klíčová slova
• adventitial fibroblasts, complex I, mitochondria, oxidative metabolism, pulmonary hypertension,
• MeSH
• buněčné dýchání MeSH
• chronická nemoc MeSH
• citrátový cyklus MeSH
• down regulace MeSH
• energetický metabolismus MeSH
• fenotyp MeSH
• fibroblasty metabolismus   MeSH
• glykolýza MeSH
• hypoxie komplikace   patologie   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• lidé MeSH
• makrofágy metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• oxidace-redukce MeSH
• oxidativní fosforylace MeSH
• parakrinní signalizace MeSH
• plíce patologie   MeSH
• plicní hypertenze komplikace   metabolismus   patologie   MeSH
• přeprogramování buněk * MeSH
• pyruvátdehydrogenasový komplex metabolismus   MeSH
• respirační komplex I metabolismus   MeSH
• skot MeSH
• superoxidy metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• skot MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kyselina pyrohroznová MeSH
• pyruvátdehydrogenasový komplex MeSH
• respirační komplex I MeSH
• superoxidy MeSH

Pyruvate is a key product of glycolysis that regulates the energy metabolism of cells. In Trypanosoma brucei, the causative agent of sleeping sickness, the fate of pyruvate varies dramatically during the parasite life cycle. In bloodstream forms, pyruvate is mainly excreted, whereas in tsetse fly forms, pyruvate is metabolized in mitochondria yielding additional ATP molecules. The character of the molecular machinery that mediates pyruvate transport across mitochondrial membrane was elusive until the recent discovery of mitochondrial pyruvate carrier (MPC) in yeast and mammals. Here, we characterized pyruvate import into mitochondrion of T. brucei. We identified mpc1 and mpc2 homologs in the T. brucei genome with attributes of MPC protein family and we demonstrated that both proteins are present in the mitochondrial membrane of the parasite. Investigations of mpc1 or mpc2 gene knock-out cells proved that T. brucei MPC1/2 proteins facilitate mitochondrial pyruvate transport. Interestingly, MPC is expressed not only in procyclic trypanosomes with fully activated mitochondria but also in bloodstream trypanosomes in which most of pyruvate is excreted. Moreover, MPC appears to be essential for bloodstream forms, supporting the recently emerging picture that the functions of mitochondria in bloodstream forms are more diverse than it was originally thought.

• MeSH
• biologický transport fyziologie   MeSH
• buněčné linie MeSH
• energetický metabolismus fyziologie   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• membránové transportní proteiny metabolismus   MeSH
• mitochondriální membrány metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• přenašeče monokarboxylových kyselin MeSH
• proteiny přenášející anionty metabolismus   MeSH
• sekvence aminokyselin MeSH
• Trypanosoma brucei brucei genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kyselina pyrohroznová MeSH
• membránové transportní proteiny MeSH
• přenašeče monokarboxylových kyselin MeSH
• proteiny přenášející anionty MeSH

BACKGROUND: Intensive care of severe trauma patients focuses on the treatment of haemorrhagic shock. Tissues should be perfused sufficiently with blood and with sufficient oxygen content to ensure adequate tissue oxygen delivery. Tissue metabolism can be monitored by microdialysis, and the lactate/pyruvate ratio (LPR) may be used as a tissue ischemia marker. The aim of this study was to determine the adequate cardiac output and haemoglobin levels that avoid tissue ischemia. METHODS: Adult patients with serious traumatic haemorrhagic shock were enrolled in this prospective observational study. The primary observed parameters included haemoglobin, cardiac output, central venous saturation, arterial lactate and the tissue lactate/pyruvate ratio. RESULTS: Forty-eight patients were analysed. The average age of the patients was 39.8 ± 16.7, and the average ISS was 43.4 ± 12.2. Hb < 70 g/l was associated with pathologic arterial lactate, ScvO2 and LPR. Tissue ischemia (i.e., LPR over 25) developed when CI ≤ 3.2 l/min/m(2) and Hb between 70 and 90 g/l were observed. Severe tissue ischemia events were recorded when the Hb dropped below 70 g/l and CI was 3.2-4.8 l/min/m(2). CI ≥ 4.8 l/min/m(2) was not found to be connected with tissue ischemia, even when Hb ≤ 70 g/l. CONCLUSION: LPR could be a useful marker to manage traumatic haemorrhagic shock therapies. In initial traumatic haemorrhagic shock treatments, it may be better to maintain CI ≥ 3.2 l/min/m(2) and Hb ≥ 70 g/l to avoid tissue ischemia. LPR could also be a useful transfusion trigger when it may demonstrate ischemia onset due to low local DO2 and early reveal low/no tissue perfusion.

• Klíčová slova
• Cardiac output, Haemoglobin, Lactate, Microdialysis, Pyruvate, Shock, Transfusion, Trauma,
• MeSH
• dospělí MeSH
• hemoglobiny metabolismus   MeSH
• hemoragický šok patofyziologie   terapie   MeSH
• kyselina mléčná metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• kyslík metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikrodialýza metody   MeSH
• minutový srdeční výdej fyziologie   MeSH
• mladý dospělý MeSH
• prospektivní studie MeSH
• resuscitace metody   MeSH
• stupeň závažnosti nemoci MeSH
• traumatický šok patofyziologie   terapie   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• pozorovací studie MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hemoglobiny MeSH
• kyselina mléčná MeSH
• kyselina pyrohroznová MeSH
• kyslík MeSH