Showing papers on "Pyruvate dehydrogenase kinase published in 2014"
TL;DR: Because most major diseases involve aberrant metabolism, understanding and exploiting pyruvate carbon flux may yield novel treatments that enhance human health.
Abstract: Pyruvate is a keystone molecule critical for numerous aspects of eukaryotic and human metabolism. Pyruvate is the end-product of glycolysis, is derived from additional sources in the cellular cytoplasm, and is ultimately destined for transport into mitochondria as a master fuel input undergirding citric acid cycle carbon flux. In mitochondria, pyruvate drives ATP production by oxidative phosphorylation and multiple biosynthetic pathways intersecting the citric acid cycle. Mitochondrial pyruvate metabolism is regulated by many enzymes, including the recently discovered mitochondria pyruvate carrier, pyruvate dehydrogenase, and pyruvate carboxylase, to modulate overall pyruvate carbon flux. Mutations in any of the genes encoding for proteins regulating pyruvate metabolism may lead to disease. Numerous cases have been described. Aberrant pyruvate metabolism plays an especially prominent role in cancer, heart failure, and neurodegeneration. Because most major diseases involve aberrant metabolism, understanding and exploiting pyruvate carbon flux may yield novel treatments that enhance human health.
524 citations
TL;DR: Available evidence suggests an association between defective mitochondrial oxidative capacity in the pancreatic β-cells and diminished insulin secretion that may trigger the development of diabetes in patients already affected with insulin resistance, although the pathogenesis remains unsettled.
417 citations
TL;DR: Bacterial and human PDCs are compared, as they represent the two major classes of the superfamily of 2-oxo acid dehydrogenase complexes with different assembly of, and interactions among components.
392 citations
TL;DR: Testing the regulation and role of glucose uptake and metabolism in the proliferation and Ab production of control, anergic, and autoimmune-prone B cells shows that activated B cells require Glut1-dependent metabolic reprogramming to support proliferation andAb production that is distinct from T cells and that this glycolytic reprograming is regulated in tolerance.
Abstract: B cell activation leads to proliferation and Ab production that can protect from pathogens or promote autoimmunity. Regulation of cell metabolism is essential to support the demands of lymphocyte growth and effector function and may regulate tolerance. In this study, we tested the regulation and role of glucose uptake and metabolism in the proliferation and Ab production of control, anergic, and autoimmune-prone B cells. Control B cells had a balanced increase in lactate production and oxygen consumption following activation, with proportionally increased glucose transporter Glut1 expression and mitochondrial mass upon either LPS or BCR stimulation. This contrasted with metabolic reprogramming of T cells, which had lower glycolytic flux when resting but disproportionately increased this pathway upon activation. Importantly, tolerance greatly affected B cell metabolic reprogramming. Anergic B cells remained metabolically quiescent, with only a modest increase in glycolysis and oxygen consumption with LPS stimulation. B cells chronically stimulated with elevated BAFF, however, rapidly increased glycolysis and Ab production upon stimulation. Induction of glycolysis was critical for Ab production, as glycolytic inhibition with the pyruvate dehydrogenase kinase inhibitor dichloroacetate sharply suppressed B cell proliferation and Ab secretion in vitro and in vivo. Furthermore, B cell-specific deletion of Glut1 led to reduced B cell numbers and impaired Ab production in vivo. Together, these data show that activated B cells require Glut1-dependent metabolic reprogramming to support proliferation and Ab production that is distinct from T cells and that this glycolytic reprogramming is regulated in tolerance.
385 citations
TL;DR: An important mechanism by which Wnt‐driven Warburg metabolism directs the use of glucose for cancer cell proliferation and links it to vessel delivery of oxygen and nutrients is identified.
Abstract: Much of the mechanism by which Wnt signaling drives proliferation during oncogenesis is attributed to its regulation of the cell cycle. Here, we show how Wnt/β-catenin signaling directs another hallmark of tumorigenesis, namely Warburg metabolism. Using biochemical assays and fluorescence lifetime imaging microscopy (FLIM) to probe metabolism in vitro and in living tumors, we observe that interference with Wnt signaling in colon cancer cells reduces glycolytic metabolism and results in small, poorly perfused tumors. We identify pyruvate dehydrogenase kinase 1 (PDK1) as an important direct target within a larger gene program for metabolism. PDK1 inhibits pyruvate flux to mitochondrial respiration and a rescue of its expression in Wnt-inhibited cancer cells rescues glycolysis as well as vessel growth in the tumor microenvironment. Thus, we identify an important mechanism by which Wnt-driven Warburg metabolism directs the use of glucose for cancer cell proliferation and links it to vessel delivery of oxygen and nutrients.
335 citations
TL;DR: The impaired glucose metabolism induced by muscle-specific Bmal1 knockout suggests that a major physiological role of the muscle clock is to prepare for the transition from the rest/fasting phase to the active/feeding phase, when glucose becomes the predominant fuel for skeletal muscle.
Abstract: Circadian rhythms control metabolism and energy homeostasis, but the role of the skeletal muscle clock has never been explored. We generated conditional and inducible mouse lines with muscle-specific ablation of the core clock gene Bmal1. Skeletal muscles from these mice showed impaired insulin-stimulated glucose uptake with reduced protein levels of GLUT4, the insulin-dependent glucose transporter, and TBC1D1, a Rab-GTPase involved in GLUT4 translocation. Pyruvate dehydrogenase (PDH) activity was also reduced due to altered expression of circadian genes Pdk4 and Pdp1, coding for PDH kinase and phosphatase, respectively. PDH inhibition leads to reduced glucose oxidation and diversion of glycolytic intermediates to alternative metabolic pathways, as revealed by metabolome analysis. The impaired glucose metabolism induced by muscle-specific Bmal1 knockout suggests that a major physiological role of the muscle clock is to prepare for the transition from the rest/fasting phase to the active/feeding phase, when glucose becomes the predominant fuel for skeletal muscle.
315 citations
TL;DR: In this article, the authors applied (13)C metabolic flux analysis (MFA) to cells after transcriptional or pharmacological inhibition of the mitochondrial pyruvate carrier (MPC).
241 citations
TL;DR: It is reported that lysine acetylation of PDHA1 and PDP1 is common in epidermal growth factor-stimulated cells and diverse human cancer cells and contributes to the Warburg effect.
227 citations
TL;DR: The findings suggest that the early induction of HIF1α targets may be instrumental in iPSC derivation via the activation of a glycolytic program, and implicate the Hif1α pathway as an enabling regulator of cellular reprogramming.
Abstract: Reprogramming somatic cells to a pluripotent state drastically reconfigures the cellular anabolic requirements, thus potentially inducing cancer-like metabolic transformation. Accordingly, we and others previously showed that somatic mitochondria and bioenergetics are extensively remodeled upon derivation of induced pluripotent stem cells (iPSCs), as the cells transit from oxidative to glycolytic metabolism. In the attempt to identify possible regulatory mechanisms underlying this metabolic restructuring, we investigated the contributing role of hypoxia-inducible factor one alpha (HIF1α), a master regulator of energy metabolism, in the induction and maintenance of pluripotency. We discovered that the ablation of HIF1α function in dermal fibroblasts dramatically hampers reprogramming efficiency, while small molecule-based activation of HIF1α significantly improves cell fate conversion. Transcriptional and bioenergetic analysis during reprogramming initiation indicated that the transduction of the four factors is sufficient to upregulate the HIF1α target pyruvate dehydrogenase kinase (PDK) one and set in motion the glycolytic shift. However, additional HIF1α activation appears critical in the early upregulation of other HIF1α-associated metabolic regulators, including PDK3 and pyruvate kinase (PK) isoform M2 (PKM2), resulting in increased glycolysis and enhanced reprogramming. Accordingly, elevated levels of PDK1, PDK3, and PKM2 and reduced PK activity could be observed in iPSCs and human embryonic stem cells in the undifferentiated state. Overall, the findings suggest that the early induction of HIF1α targets may be instrumental in iPSC derivation via the activation of a glycolytic program. These findings implicate the HIF1α pathway as an enabling regulator of cellular reprogramming.
225 citations
TL;DR: The results suggest that the acetylation of PDHA1 provides another layer of enzymatic regulation, in addition to phosphorylation, involving a reversible acetyllysine, suggesting that theacetylome, as well as the kinome, links glycolysis to respiration.
148 citations
TL;DR: It is demonstrated that PDK1 is critical for Lin28A- and Lin28B-mediated cancer proliferation both in vitro and in vivo, establishing a previously unappreciated mechanism by which Lin28/let-7 axis facilitates Warburg effect to promote cancer progression.
Abstract: The RNA-binding proteins Lin28A and Lin28B are known to have key roles in a variety of pathological states including cancer, obesity and diabetes. Here the authors show that Lin28A and -B alter cancer metabolism through let-7-mediated upregulation of pyruvate dehydrogenase kinase 1.
TL;DR: It is demonstrated that HIF1α stabilization in osteoblast precursors of postnatal mice markedly increases osteobasts number and bone mass, and alterations in cellular metabolism may be a broadly applicable mechanism for regulating cell differentiation.
Abstract: The bone marrow environment is among the most hypoxic in the body, but how hypoxia affects bone formation is not known. Because low oxygen tension stabilizes hypoxia-inducible factor alpha (HIFα) proteins, we have investigated the effect of expressing a stabilized form of HIF1α in osteoblast precursors. Brief stabilization of HIF1α in SP7-positive cells in postnatal mice dramatically stimulated cancellous bone formation via marked expansion of the osteoblast population. Remarkably, concomitant deletion of vascular endothelial growth factor A (VEGFA) in the mouse did not diminish bone accrual caused by HIF1α stabilization. Thus, HIF1α-driven bone formation is independent of VEGFA up-regulation and increased angiogenesis. On the other hand, HIF1α stabilization stimulated glycolysis in bone through up-regulation of key glycolytic enzymes including pyruvate dehydrogenase kinase 1 (PDK1). Pharmacological inhibition of PDK1 completely reversed HIF1α-driven bone formation in vivo. Thus, HIF1α stimulates osteoblast formation through direct activation of glycolysis, and alterations in cellular metabolism may be a broadly applicable mechanism for regulating cell differentiation.
TL;DR: Evidence of metabolic rewiring during EMT that diverts glucose to the TCA cycle is revealed and a novel interaction between PDK4 and apoptosis-inducing factor (AIF), an inner mitochondrial protein that appears to play a role in mediating this resistance.
Abstract: Accumulating preclinical and clinical evidence implicates epithelial-mesenchymal transition (EMT) in acquired resistance to anticancer drugs; however, mechanisms by which the mesenchymal state determines drug resistance remain unknown. To explore a potential role for altered cellular metabolism in EMT and associated drug resistance, we analyzed the metabolome and transcriptome of three lung cancer cell lines that were rendered drug resistant following experimental induction of EMT. This analysis revealed evidence of metabolic rewiring during EMT that diverts glucose to the TCA cycle. Such rewiring was at least partially mediated by the reduced expression of pyruvate dehydrogenase kinase 4 (PDK4), which serves as a gatekeeper of the TCA cycle by inactivating pyruvate dehydrogenase (PDH). Overexpression of PDK4 partially blocked TGFβ-induced EMT; conversely, PDK4 inhibition via RNAi-mediated knockdown was sufficient to drive EMT and promoted erlotinib resistance in EGFR mutant lung cancer cells. We identified a novel interaction between PDK4 and apoptosis-inducing factor (AIF), an inner mitochondrial protein that appears to play a role in mediating this resistance. In addition, analysis of human tumor samples revealed PDK4-low as a predictor of poor prognosis in lung cancer and that PDK4 expression is dramatically downregulated in most tumor types. Together, these findings implicate PDK4 as a critical metabolic regulator of EMT and associated drug resistance.
TL;DR: In this paper, a total of 1,060 nonredundant proteins were identified in a quantitative manner using normalized spectral counts including as many as 5-fold more “extreme” proteins (low mass, high isoelectric point, hydrophobic) than previous mitochondrial proteome studies.
Abstract: Mitochondria are called the powerhouses of the cell. To better understand the role of mitochondria in maintaining and regulating metabolism in storage tissues, highly purified mitochondria were isolated from dormant potato tubers (Solanum tuberosum ‘Folva’) and their proteome investigated. Proteins were resolved by one-dimensional gel electrophoresis, and tryptic peptides were extracted from gel slices and analyzed by liquid chromatography-tandem mass spectrometry using an Orbitrap XL. Using four different search programs, a total of 1,060 nonredundant proteins were identified in a quantitative manner using normalized spectral counts including as many as 5-fold more “extreme” proteins (low mass, high isoelectric point, hydrophobic) than previous mitochondrial proteome studies. We estimate that this compendium of proteins represents a high coverage of the potato tuber mitochondrial proteome (possibly as high as 85%). The dynamic range of protein expression spanned 1,800-fold and included nearly all components of the electron transport chain, tricarboxylic acid cycle, and protein import apparatus. Additionally, we identified 71 pentatricopeptide repeat proteins, 29 membrane carriers/transporters, a number of new proteins involved in coenzyme biosynthesis and iron metabolism, the pyruvate dehydrogenase kinase, and a type 2C protein phosphatase that may catalyze the dephosphorylation of the pyruvate dehydrogenase complex. Systematic analysis of prominent posttranslational modifications revealed that more than 50% of the identified proteins harbor at least one modification. The most prominently observed class of posttranslational modifications was oxidative modifications. This study reveals approximately 500 new or previously unconfirmed plant mitochondrial proteins and outlines a facile strategy for unbiased, near-comprehensive identification of mitochondrial proteins and their modified forms.
TL;DR: Compared with wild-type controls, Mpc2(Δ16) mice exhibited normal insulin sensitivity but elevated blood glucose after bolus pyruvate or glucose injection and this was attributable to reduced glucose-stimulated insulin secretion and was corrected by sulfonylurea KATP channel inhibitor administration.
TL;DR: It is demonstrated that DCA and metformin combine to synergistically induce caspase-dependent apoptosis involving oxidative damage with simultaneous attenuation of meetformin promoted lactate production.
Abstract: The unique metabolism of breast cancer cells provides interest in exploiting this phenomenon therapeutically. Metformin, a promising breast cancer therapeutic, targets complex I of the electron transport chain leading to an accumulation of reactive oxygen species (ROS) that eventually lead to cell death. Inhibition of complex I leads to lactate production, a metabolic byproduct already highly produced by reprogrammed cancer cells and associated with a poor prognosis. While metformin remains a promising cancer therapeutic, we sought a complementary agent to increase apoptotic promoting effects of metformin while attenuating lactate production possibly leading to greatly improved efficacy. Dichloroacetate (DCA) is a well-established drug used in the treatment of lactic acidosis which functions through inhibition of pyruvate dehydrogenase kinase (PDK) promoting mitochondrial metabolism. Our purpose was to examine the synergy and mechanisms by which these two drugs kill breast cancer cells. Cell lines were subjected to the indicated treatments and analyzed for cell death and various aspects of metabolism. Cell death and ROS production were analyzed using flow cytometry, Western blot analysis, and cell counting methods. Images of cells were taken with phase contrast microscopy or confocal microscopy. Metabolism of cells was analyzed using the Seahorse XF24 analyzer, lactate assays, and pH analysis. We show that when DCA and metformin are used in combination, synergistic induction of apoptosis of breast cancer cells occurs. Metformin-induced oxidative damage is enhanced by DCA through PDK1 inhibition which also diminishes metformin promoted lactate production. We demonstrate that DCA and metformin combine to synergistically induce caspase-dependent apoptosis involving oxidative damage with simultaneous attenuation of metformin promoted lactate production. Innovative combinations such as metformin and DCA show promise in expanding breast cancer therapies.
01 Jan 2014
TL;DR: This study reveals approximately 500 new or previously unconfirmed plant mitochondrial proteins and outlines a facile strategy for unbiased, near-comprehensive identification of mitochondria and their modified forms.
Abstract: Mitochondria are called the powerhouses of the cell. To better understand the role of mitochondria in maintaining and regulating metabolism in storage tissues, highly purified mitochondria were isolated from dormant potato tubers (Solanum tuberosum ‘Folva’) and their proteome investigated. Proteins were resolved by one-dimensional gel electrophoresis, and tryptic peptides were extracted from gel slices and analyzed by liquid chromatography-tandem mass spectrometry using an Orbitrap XL. Using four different search programs, a total of 1,060 nonredundant proteins were identified in a quantitative manner using normalized spectral counts including as many as 5-fold more “extreme” proteins (low mass, high isoelectric point, hydrophobic) than previous mitochondrial proteome studies. We estimate that this compendium of proteins represents a high coverage of the potato tuber mitochondrial proteome (possibly as high as 85%). The dynamic range of protein expression spanned 1,800-fold and included nearly all components of the electron transport chain, tricarboxylic acid cycle, and protein import apparatus. Additionally, we identified 71 pentatricopeptide repeat proteins, 29 membrane carriers/transporters, a number of new proteins involved in coenzyme biosynthesis and iron metabolism, the pyruvate dehydrogenase kinase, and a type 2C protein phosphatase that may catalyze the dephosphorylation of the pyruvate dehydrogenase complex. Systematic analysis of prominent posttranslational modifications revealed that more than 50% of the identified proteins harbor at least one modification. The most prominently observed class of posttranslational modifications was oxidative modifications. This study reveals approximately 500 new or previously unconfirmed plant mitochondrial proteins and outlines a facile strategy for unbiased, near-comprehensive identification of mitochondrial proteins and their modified forms.
TL;DR: The pharmacological approach of targeting PDK to control both glucose and fat levels in obesity and type 2 diabetes is supported, and obesity phenotypes are effectively treated by chemical intervention with PDK inhibitors.
TL;DR: This work first obtained a successful engineered diatom regulating a key gene involved in lipid metabolism, and provides powerful indications in enhancing microalgal lipid production by metabolic engineering for biofuel industry.
Abstract: Microalgae have been an emerging biofuel resource; however, the germplasm improvement has been slow due to the lack of molecular tools. Pyruvate dehydrogenase kinase (PDK) deactivates the pyruvate dehydrogenase complex (PDC) which catalyzes the oxidative decarboxylation of pyruvate. Acetyl-CoA production via PDC is important in plant tissues that are active in fatty acid synthesis. A 1261-bp cDNA of a putative PDK gene (PtPDK) was cloned from a diatom Phaeodactylum tricornutum, and PtPDK antisense knockdown transgenic diatoms were generated. Both PtPDK transcript abundance and enzyme activity were reduced significantly due to antisense knockdown of PtPDK. Neutral lipid content of transgenic diatom cells increased up to 82% as determined by Nile red staining, and fatty acid composition was not altered. Transgenic cells showed slightly lower growth rate but similar cell size with the wild type, hence retaining similar biomass productivity. This work first obtained a successful engineered diatom regulating a key gene involved in lipid metabolism. Our findings also provide powerful indications in enhancing microalgal lipid production by metabolic engineering for biofuel industry.
TL;DR: It is suggested that phosphorylation at distinct serine and tyrosine residues inhibits PDHA1 through distinct mechanisms to impact active site accessibility, which act in concert to regulate PDC activity and promote the Warburg effect.
TL;DR: The studies clearly show that the newly identified mitochondrial pyruvate carrier sits at an important branching point in nutrient metabolism and that it is an essential regulator of insulin secretion.
TL;DR: A new molecular pathway is highlighted through which PKM2 can manage gliomagenesis via the control of glioma stemness by Oct4 through its interaction with Oct4, a major regulator of self-renewal and differentiation in stem cells.
Abstract: Glioma stem cells are highly resistant to cell death and as such are supposed to contribute to tumor recurrence by eluding anticancer treatments. Here, we show that spheroids that contain rat neural stem cells (NSCs) or rat glioma stem cells (cancer stem cells, CSCs) express isoforms 1 and 2 of pyruvate kinase (PKM1 and PKM2); however, the expression of PKM2 is considerably higher in glioma spheroids. Silencing of PKM2 enhances both apoptosis and differentiation of rat and human glioma spheroids. We establish that PKM2 was implicated in glioma spheroid differentiation through its interaction with Oct4, a major regulator of self-renewal and differentiation in stem cells. The small molecule Dichloroacetate (DCA), a pyruvate dehydrogenase kinase inhibitor, increases the amount of PKM2/Oct4 complexes and thus inhibited Oct4-dependent gene expression. Taken together, our results highlight a new molecular pathway through which PKM2 can manage gliomagenesis via the control of glioma stemness by Oct4.
TL;DR: The pyruvate dehydrogenase complex (PDC) is an emerging target for the treatment of metabolic syndrome and is a key intermediate of glucose oxidation and an important precursor for the synthesis of glucose, glycerol, fatty acids, and nonessential amino acids.
Abstract: The pyruvate dehydrogenase complex (PDC) is an emerging target for the treatment of metabolic syndrome. To maintain a steady-state concentration of adenosine triphosphate during the feed-fast cycle, cells require efficient utilization of fatty acid and glucose, which is controlled by the PDC. The PDC converts pyruvate, coenzyme A (CoA), and oxidized nicotinamide adenine dinucleotide (NAD(+)) into acetyl-CoA, reduced form of nicotinamide adenine dinucleotide (NADH), and carbon dioxide. The activity of the PDC is up- and down-regulated by pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase, respectively. In addition, pyruvate is a key intermediate of glucose oxidation and an important precursor for the synthesis of glucose, glycerol, fatty acids, and nonessential amino acids.
TL;DR: DCA treatment was able to re-sensitize gastric cancer cells with hypoxia-induced resistance to 5-FU through the alteration of glucose metabolism, as well as in PDK1 knockdown cell lines.
TL;DR: DCA sensitized metformin-cytotoxicity by reprogramming glucose metabolism in part from aerobic glycolysis to mitochondrial oxidation, evidenced by measurements of glucose consumption, lactate release, and the ratio of oxygen consumption rate/extracellular acidification rate.
TL;DR: Higher oxidative damage with suboptimal increases in DNA repair, anti‐oxidant capacity, mitochondrial biogenesis and glucose metabolism may be implicated in SLE deterioration, and this impairment might be improved by targeted biological therapy.
Abstract: Summary We measured plasma levels of the oxidative DNA damage marker 8-hydroxy- 2'-deoxyguanosine (8-OHdG) and leucocyte mRNA expression levels of the genes encoding the 8-OHdG repair enzyme human 8-oxoguanine DNA glycosylase 1 (hOGG1), the anti-oxidant enzymes copper/zinc superoxide dismutase (Cu/ZnSOD), manganese superoxide dismutase (MnSOD), catalase, glutathione peroxidase-1 (GPx-1), GPx-4, glutathione reductase (GR) and glutathione synthetase (GS), the mitochondrial biogenesis-related proteins mtDNA-encoded ND 1 polypeptide (ND1), ND6, ATPase 6, mitochondrial transcription factor A (Tfam), nuclear respiratory factor 1(NRF-1), pyruvate dehydrogenase E1 component alpha subunit (PDHA1), pyruvate dehydrogenase kinase isoenzyme 1 (PDK-1) and hypoxia inducible factor-1α (HIF-1α) and the glycolytic enzymes hexokinase-II (HK-II), glucose 6-phosphate isomerase (GPI), phosphofructokinase (PFK), glyce- raldehyde 3-phosphate dehydrogenase (GAPDH) and lactate dehydrogenase A (LDHa). We analysed their relevance to oxidative damage in 85 systemic lupus erythematosus (SLE) patients, four complicated SLE patients undergo- ing rituximab treatment and 45 healthy individuals. SLE patients had higher plasma 8-OHdG levels (P < 0·01) but lower leucocyte expression of the genes encoding hOGG1(P < 0·01), anti-oxidant enzymes (P < 0·05), mitochondrial biogenesis-related proteins (P < 0·05) and glycolytic enzymes (P < 0·05) than healthy individuals. The increase in plasma 8-OHdG was correlated posi- tively with the elevation of leucocyte expression of the genes encoding hOGG1 (P < 0·05), anti-oxidant enzymes (P < 0·05), several mitochondrial biogenesis-related proteins (P < 0·05) and glycolytic enzymes (P < 0·05) in lupus patients. The patients, whose leucocyte mtDNA harboured D310 heteroplasmy, exhibited a positive correlation between the mtDNA copy number and expression of ND1, ND6 and ATPase 6 (P < 0·05) and a negative correlation between mtDNA copy number and systemic lupus erythe- matosus disease activity index (SLEDAI) (P < 0·05), as well as plasma 8-OHdG (P < 0·05). In particular, four complicated SLE patients with increased expression of the genes encoding the anti-oxidant enzymes, GAPDH, Tfam and PDHA1, experienced better therapeutic outcomes after rituximab therapy. In conclusion, higher oxidative damage with suboptimal increases in DNA repair, anti-oxidant capacity, mitochondrial biogenesis and glucose metabolism may be implicated in SLE deterioration, and this impair- ment might be improved by targeted biological therapy.
TL;DR: HIF-1α knockdown can inhibit the prolife ratio and promote apoptosis of pancreatic cancerous BxPC-3 cells in vitro.
Abstract: Objectives: The aim of this study is to explore the possible role of HIF-1α in glucose metabolism, proliferation and apoptosis of pancreatic cancerous cells. Method: The pancreatic cancerous BxPC-3 cells were cultured in normoxia or hypoxia (3% O2), respectively. Cell proliferation was determined by MTT assay, apoptosis was determined by Annexin V/PI staining. Expression of Pyruvate dehydrogenase kinase (PDK1), Lactate dehydrogenase (LDHA), pyruvate kinase M2 (PKM2) and citrate synthase (CS) was determined by Western-blot and Real-time PCR. Results: Under hypoxia, the expression of HIF-1α and the lactate production were increased. The expression of glucose metabolic enzymes PDK1, LDHA, PKM2 was also increased compared with that under aerobic condition. Hypoxia treatment had little effect on expression of CS. Under hypoxia, knockdown of HIF-1α inhibited the production of lactate and the expression of PDK1, LDHA and PKM2. Knockdown of HIF-1α repressed the growth of pancreatic cancer BxPC-3 cells and induced apoptosis of the cells under hypoxia. Conclusion: Under hypoxia, the expression of HIF-1α is induced, leading to the increase of glycolysis in BxPC-3 cells possibly through upregulation of the enzymes related to glycolysis. HIF-1α knockdown can inhibit the prolife ratio and promote apoptosis of pancreatic cancerous BxPC-3 cells in vitro.
TL;DR: It is found that modulation of oxidative substrate selection to increase muscle glucose utilization surprisingly results in muscle insulin resistance, offering genetic evidence against the glucose-fatty acid cycle hypothesis of Muscle insulin resistance.
Abstract: The pyruvate dehydrogenase complex (PDH) has been hypothesized to link lipid exposure to skeletal muscle insulin resistance through a glucose-fatty acid cycle in which increased fatty acid oxidation increases acetyl-CoA concentrations, thereby inactivating PDH and decreasing glucose oxidation. However, whether fatty acids induce insulin resistance by decreasing PDH flux remains unknown. To genetically examine this hypothesis we assessed relative rates of pyruvate dehydrogenase flux/mitochondrial oxidative flux and insulin-stimulated rates of muscle glucose metabolism in awake mice lacking pyruvate dehydrogenase kinase 2 and 4 [double knockout (DKO)], which results in constitutively activated PDH. Surprisingly, increased glucose oxidation in DKO muscle was accompanied by reduced insulin-stimulated muscle glucose uptake. Preferential myocellular glucose utilization in DKO mice decreased fatty acid oxidation, resulting in increased reesterification of acyl-CoAs into diacylglycerol and triacylglycerol, with subsequent activation of PKC-θ and inhibition of insulin signaling in muscle. In contrast, other putative mediators of muscle insulin resistance, including muscle acylcarnitines, ceramides, reactive oxygen species production, and oxidative stress markers, were not increased. These findings demonstrate that modulation of oxidative substrate selection to increase muscle glucose utilization surprisingly results in muscle insulin resistance, offering genetic evidence against the glucose-fatty acid cycle hypothesis of muscle insulin resistance.
TL;DR: Oral administration of DADA significantly restored PDH activity and ATP level in various organs, and ameliorated disorders of glucose and lipid metabolism in the blood, together with marked improvement of survival and suppression of cytokine storm, trypsin up-regulation and viral replication in mice with severe influenza.
Abstract: Severe influenza is characterized by cytokine storm and multiorgan failure with metabolic energy disorders and vascular hyperpermeability. In the regulation of energy homeostasis, the pyruvate dehydrogenase (PDH) complex plays an important role by catalyzing oxidative decarboxylation of pyruvate, linking glycolysis to the tricarboxylic acid cycle and fatty acid synthesis, and thus its activity is linked to energy homeostasis. The present study tested the effects of diisopropylamine dichloroacetate (DADA), a new PDH kinase 4 (PDK4) inhibitor, in mice with severe influenza. Infection of mice with influenza A PR/8/34(H1N1) virus resulted in marked down-regulation of PDH activity and ATP level, with selective up-regulation of PDK4 in the skeletal muscles, heart, liver and lungs. Oral administration of DADA at 12-h intervals for 14 days starting immediately after infection significantly restored PDH activity and ATP level in various organs, and ameliorated disorders of glucose and lipid metabolism in the blood, together with marked improvement of survival and suppression of cytokine storm, trypsin up-regulation and viral replication. These results indicate that through PDK4 inhibition, DADA effectively suppresses the host metabolic disorder-cytokine cycle, which is closely linked to the influenza virus-cytokine-trypsin cycle, resulting in prevention of multiorgan failure in severe influenza.
TL;DR: This work has shown that pyruvate carboxylase is an important determinate of the rates of gluconeogenesis in humans with type 2 diabetes and these enzymes are potential therapeutic targets for several diseases.
Abstract: Pyruvate is an obligatory intermediate in the oxidative disposal of glucose and a major precursor for the synthesis of glucose, glycerol, fatty acids, and non-essential amino acids. Stringent control of the fate of pyruvate is critically important for cellular homeostasis. The regulatory mechanisms for its metabolism are therefore of great interest. Recent advances include the findings that (a) the mitochondrial pyruvate carrier is sensitive to inhibition by thiazolidinediones; (b) pyruvate dehydrogenase kinases induce the Warburg effect in many disease states; and (c) pyruvate carboxylase is an important determinate of the rates of gluconeogenesis in humans with type 2 diabetes. These enzymes are potential therapeutic targets for several diseases.