Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.14.11.2 (prolyl hydroxylase)
1,814 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The purpose of this study was to test for the presence of liver hypoxia and recovery after reperfusion when blood alcohol levels (BAL) are high. Male rats were fed ethanol intragastrically at a constant rate for 1 month. The pO(2) levels were then measured on the liver surface of these rats, in vivo during laparatomy under isoflurane anesthesia. To measure the response to acute hypoxia, the hepatic blood flow was clamped off at the porta hepatis. When the clamp was released, recovery from hypoxia was measured. A number of hypoxic-inducible genes in the liver were analyzed by means of quantitative RT-PCR as a measure of increased activation of hypoxia initiated transcription. The mRNA levels of genes for adrenomedullin, adrenergic receptor alpha, 1a and 1d, CDK inhibitor 1a, and erythropoietin were all significantly higher at the peaks than troughs. Expression of these same genes in the livers of control rats fed dextrose was lower than at the troughs. Although the mRNA level of the hypoxia-inducible factor (HIF-1alpha) was higher at the trough than at the peak, its protein concentration in the nuclear fraction was not increased at the troughs compared with the peaks. In fact, the nuclear protein level of HIF-1alpha at the peak was significantly higher than in control samples, which is consistent with the presence of hypoxia at the peaks. Further analysis of the HIF-alpha degradation regulation revealed that prolyl 4-hydroxylase (P4ha1) and von Hippel-Lindau syndrome homolog (Vhl) were both up-regulated at the troughs compared with the peaks. The liver surface oxygen levels at the peaks were reduced compared with the control samples. The pO(2) levels fell abruptly when the vessels at the porta hepatis were clamped. When the clamp was removed, allowing reperfusion of the liver, pO(2) returned to baseline levels in the control, and at the troughs but not at the peaks. These results support the hypothesis that hypoxia occurs at the peaks of the BAL cycle and recovery from ischemia is impaired at the peaks.
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PMID:Liver hypoxia and lack of recovery after reperfusion at high blood alcohol levels in the intragastric feeding model of alcohol liver disease. 1550 34

The CXC chemokine IL-8, which promotes adhesion, activation, and transmigration of polymorphonuclear neutrophils (PMN), has been associated with production of tissue injury in reperfused myocardium. Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric peptide that is a key regulator of genes such as heme oxygenase (HO)-1 expressed under hypoxic conditions. We hypothesized that HO-1 plays an important role in regulating proinflammatory mediator production under conditions of ischemia-reperfusion. HIF-1 was activated in the human microvascular endothelial cell line (HMEC-1) with the prolyl hydroxylase inhibitor dimethyloxalylglycine (DMOG). DMOG significantly attenuated cytokine-induced IL-8 promoter activity and protein secretion and cytokine-induced PMN migration across human microvascular endothelial cell line HMEC-1 monolayers. In vivo studies in a rabbit model of myocardial ischemia-reperfusion showed that rabbits pretreated with a 20 mg/kg DMOG infusion (n = 6) 24 h before study exhibited a 21.58 +/- 1.76% infarct size compared with 35.25 +/- 2.06% in saline-treated ischemia-reperfusion animals (n = 6, change in reduction = 39%; P < 0.001). In DMOG-pretreated (20 mg/kg) animals, plasma IL-8 levels at 3 h after onset of reperfusion were 405 +/- 40 pg/ml vs. 790 +/- 40 pg/ml in saline-treated ischemia-reperfusion animals (P < 0.001). DMOG pretreatment reduced myocardial myeloperoxidase activity, expressed as number of PMN per gram of myocardium, to 1.43 +/- 0.59 vs. 4.86 +/- 1.1 (P = 0.012) in saline-treated ischemia-reperfused hearts. Both in vitro and in vivo DMOG-attenuated IL-8 production was associated with robust HO-1 expression. Thus our data show that HIF-1 activation induces substantial HO-1 expression that is associated with attenuated proinflammatory chemokine production by microvascular endothelium in vitro and in vivo.
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PMID:HIF-1 activation attenuates postischemic myocardial injury: role for heme oxygenase-1 in modulating microvascular chemokine generation. 1601 14

Tissue hypoxia/ischemia are major pathophysiological determinants. Conditions of decreased oxygen availability provoke accumulation and activation of hypoxia-inducible factor-1 (HIF-1). Recent reports demonstrate a crucial role of HIF-1 for inflammatory events. Regulation of hypoxic responses by the inflammatory mediators nitric oxide (NO) and reactive oxygen species (ROS) is believed to be of pathophysiolgical relevance. It is reported that hypoxic stabilization of HIF-1alpha can be antagonized by NO due to its ability to attenuate mitochondrial electron transport. Likely, the formation of ROS could contribute to this effect. As conflicting results emerged from several studies showing either decreased or increased ROS production during hypoxia, we used experiments mimicking hypoxic intracellular ROS changes by using the redox cycling agent 2,3-dimethoxy-1,4-naphthoquinone (DMNQ), which generates superoxide inside cells. Treatment of A549, HEK293, HepG2, and COS cells with DMNQ resulted in a concentration-dependent raise in ROS which correlated with HIF-1alpha accumulation. By using a HIF-1alpha-von Hippel-Lindau tumor suppressor protein binding assay, we show that ROS produced by DMNQ impaired prolyl hydroxylase activity. When HIF-1alpha is stabilized by NO, low concentrations of DMNQ (<1 microM) revealed no effect, intermediate concentrations of 1 to 40 microM DMNQ attenuated HIF-1alpha accumulation and higher concentrations of DMNQ promoted HIF-1alpha stability. Attenuation of NO-induced HIF-1alpha stability regulation by ROS was mediated by an active proteasomal degradation pathway. In conclusion, we propose that scavenging of NO by ROS and vice versa attenuate HIF-1alpha accumulation in a concentration-dependent manner. This is important to fully elucidate HIF-1alpha regulation under inflammatory conditions.
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PMID:Reactive oxygen species attenuate nitric-oxide-mediated hypoxia-inducible factor-1alpha stabilization. 1663 33

Hypoxia-inducible factor-1, HIF1, transcriptionally activates over 200 genes vital for cell homeostasis and angiogenesis. We developed a computational model to gain a detailed quantitative understanding of how HIF1 acts to sense oxygen and respond to hypoxia. The model consists of kinetic equations describing the intracellular variation of 17 compounds, including HIF1, iron, prolyl hydroxylase, oxygen, ascorbate, 2-oxoglutarate, von Hippel Lindau protein and associated complexes. We tested an existing hypothesis of a switch-like change in HIF1 expression in response to a gradual decrease in O2 concentration. Our model predicts that depending on the molecular environment, such as intracellular iron levels, the hypoxic response varies considerably. We show HIF1-activated cellular responses can be divided into two categories: a steep, switch-like response to O2 and a gradual one. Discovery of this dual response prompted comparison of two therapeutic strategies, ascorbate and iron supplementation, and prolyl hydroxylase targeting, to predict under what microenvironments either effectively increases HIF1alpha hydroxylation. Results provide crucial insight into the effects of iron and prolyl hydroxylase on oxygen sensing. The model advances quantitative molecular level understanding of HIF1 pathways--an endeavor that will help elucidate the diverse responses to hypoxia found in cancer, ischemia and exercise.
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PMID:A computational model of intracellular oxygen sensing by hypoxia-inducible factor HIF1 alpha. 1689 21

Ischemia/Reperfusion injury and hemolysis are characterized by erythrocyte lysis and release of free heme into the microcirculation. Following substantial erythrocyte lysis, heme overwhelms circulatory heme-binding protein networks rapidly forming hemin, the oxidized form of iron protoporphyrin IX. Hemin's role in modulating inflammatory responses in microvascular endothelium (MVEC) remains ill-defined. We studied the impact of hemin exposure on human MVEC interleukin-8 (IL-8) expression. Hemin significantly up-regulated MVEC IL-8 secretion and was associated with cellular iron loading. Hemin-induced IL-8 up-regulation was significantly attenuated by increasing environmental serum concentrations. As well, hemin-induced IL-8 secretion was significantly reduced in a concentration-dependent fashion following pyrrolidine dithiocarbamate exposure, suggesting that induction occurred via an oxidant-sensitive mechanism. Interestingly, transfection studies revealed that oxidant-driven transcription factors NF-kappaB and AP-1 played no role in hemin-induced IL-8 transcription. In studies employing actinomycin D, hemin was found to dramatically lengthen IL-8 mRNA half-life. Of major importance in the current report was the finding that hypoxia inducible factor-1 (HIF-1), a powerful transcription factor mediating tissue responses to hypoxia, potently regulated hemin-induced IL-8 secretion in human MVEC. Activation of HIF-1 via the prolyl hydroxylase inhibitor dimethyloxalylglycine attenuated hemin-induced IL-8 secretion. These studies were confirmed via DNA-directed siRNA silencing of HIF-1alpha. In conclusion, hemin induces a serum protein-sensitive pro-inflammatory phenotype in MVEC via an oxidant-sensitive mechanism that is powerfully regulated by HIF-1.
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PMID:Hypoxia inducible factor-1 modulates hemin-induced IL-8 secretion in microvascular endothelium. 1733 40

Emerging research suggests that oxidant-driven transcription of key cytokine/chemokine networks within the myocardium plays a crucial role in producing ischemia-reperfusion (I/R) injury. We recently showed that activation of hypoxia-inducible factor-1 (HIF-1) attenuated cardiac I/R injury. Diminished injury in these prior studies was associated with significant reductions in circulating interleukin-8 levels, suggesting that HIF-1 may play an important role in modulating postischemic cardiac inflammation. In the current study, we examined the role of HIF-1 activation in modulating proinflammatory chemokine [macrophage inflammatory protein (MIP)-2, cytokine-induced neutrophil chemoattractant factor (KC), and lipopolysaccharide-induced CXC chemokine (LIX)] and adhesion molecule [intercellular adhesion molecule (ICAM)-1] expression in murine cardiomyocytes in vitro (HL-1 cell line) and in intact murine hearts following in vivo I/R injury. Our results show that HIF-1 activation induced both pharmacologically by the prolyl hydroxylase inhibitor dimethyloxallyl glycine and via small-interfering RNA (siRNA)-mediated prolyl-4 hydroxylase-2 (P4HA2) gene silencing significantly attenuated tumor necrosis factor-alpha-induced chemokine (KC and LIX) and ICAM-1 expression in cardiomyocytes. In vivo, postischemic hearts obtained from animals receiving the P4HA2 siRNA (HIF-1 activation) exhibited significantly reduced CXC chemokine (MIP-2, KC, and LIX), CC chemokine (monocyte chemoattractant protein-1), and ICAM-1 expression when compared with postischemic hearts from either saline I/R controls or postischemic hearts from animals receiving a nontargeting control siRNA (no HIF-1 activation). Diminished chemokine and adhesion molecule expression in HIF-1-activated postischemic hearts was associated with significantly reduced polymorphonuclear leukocyte infiltration and myocardial infarct size (>60% reduction P4HA2 siRNA I/R vs. saline I/R, P < 0.001, n = 6). In conclusion, these results demonstrate for the first time that HIF-1 activation following infusion of siRNA to P4HA2 plays a key role in modulating I/R-associated cardiac inflammatory responses.
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PMID:Activation of hypoxia-inducible factor-1 via prolyl-4 hydoxylase-2 gene silencing attenuates acute inflammatory responses in postischemic myocardium. 1754 79

T(4) activation into T(3) is catalyzed by type 2 deiodinase (D2) in the brain. The rapid induction of D2 in astrocytes by transient brain ischemia has prompted us to explore the effects of hypoxia on D2 in cultures of astrocytes. Hypoxia (2.5% O(2)) of cultured astrocytes increased D2 activity, alone or in association with agents stimulating the cAMP pathway. Hypoxia had no effect on D2 mRNA accumulation. Cycloheximide did not block the effect of hypoxia on D2 activity and D2 half-life was enhanced under hypoxia demonstrating a posttranslational action of hypoxia. Furthermore, the D2 activity increase by hypoxia was not additive with the increase promoted by the proteasome inhibitor carbobenzoxy-L-leucyl-L-leucyl-L-leucinal (MG132). This strongly suggests that hypoxia leads to stabilization of D2 by slowing its degradation by the proteasome pathway. Hypoxia, in contrast to MG132, did not block the T(4)-induced D2 inactivation. A contribution of prolyl hydroxylase to the hypoxia effects on D2 was also suggested on the basis of increased D2 activity after addition of different prolyl hydroxylase inhibitors (cobalt chloride, desferrioxamine, dimethyloxalylglycine, dimethylsuccinate). Specific inhibitors of ERK, p38 MAPK, or phosphatidylinositol 3-kinase pathways were without any effect on hypoxia-increased D2 activity, eliminating their role in the effects of hypoxia. Interestingly, diphenyleneiodonium, an inhibitor of nicotinamide adenine dinucleotide phosphate oxidase inhibited the hypoxia-increased D2 indicating a role for some reactive oxygen species in the mechanism of D2 increase. Further studies are required to clarify the precise molecular mechanisms involved in the D2 stabilization by hypoxia.
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PMID:Hypoxia stabilizes type 2 deiodinase activity in rat astrocytes. 1761 50

Hypoxia occurs in cancer, prolonged exercise, and long-term ischemia with durations of several hours or more, and the hypoxia-inducible factor 1 (HIF1) pathway response to these conditions differs from responses to transient hypoxia. We used computational modeling, validated by experiments, to gain a quantitative, temporal understanding of the mechanisms driving HIF1 response. To test the hypothesis that HIF1 alpha protein levels during chronic hypoxia are tightly regulated by a series of molecular feedbacks, we took into account protein synthesis and product inhibition, and analyzed HIF1 system changes in response to hypoxic exposures beyond 3 to 4 h. We show how three autocrine feedback loops together regulate HIF 1 alpha hydroxylation in different microenvironments. Results demonstrate that prolyl hydroxylase, succinate and HIF1 alpha feedback determine intracellular HIF1 alpha levels over the course of hours to days. The model provides quantitative insight critical for characterizing molecular mechanisms underlying a cell's response to long-term hypoxia.
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PMID:Three autocrine feedback loops determine HIF1 alpha expression in chronic hypoxia. 1772 Feb 60

Oxygen-regulated protein 150 (ORP150) is an inducible endoplasmic reticulum (ER) chaperone molecule that is upregulated after numerous cellular insults and has a cytoprotective role in renal, neural, and cardiac models of ischemia-reperfusion injury. ORP150 also has been shown to play a role in cellular Ca(2+) homeostasis, and in turn, regulating calpain activity. In this study, we identified ORP150 in whole rat renal cortical mitochondria and matrix fractions, demonstrated the targeting of an ORP150-GFP construct to the mitochondria of NIH-3T3 cells, and showed that the NH(2)-terminal 13 amino acids of ORP150 are sufficient for this translocation. ORP150 expression was found to be regulated by the anti-C/enhancer-binding protein homologous protein (CHOP)/GADD153 transcription factor and ORP150 levels increased in the mitochondria and ER of COS-7 cells after diverse stresses, including hypoxia, serum starvation, prolyl hydroxylase inhibition with dimethyloxaloylglycine, and exposure to tunicamycin, ethidium, bromide, and 2-deoxyglucose. Induction of the mitochondrial specific stress response in COS-7 cells through expression of an ornithine transcarbamylase mutant (Delta OTC) increased mitochondrial ORP150 levels and mitochondrial calpain activity. To determine whether mitochondrial ORP150 and mitochondrial calpain 10 interact, rat cortical mitochondria exposed to Ca(2+) resulted in ORP150 cleavage in a calpain inhibitor-dependent manner, revealing that ORP150 is a substrate and may be regulated by calpain 10. These data reveal a novel cellular localization for ORP150 and that mitochondrial ORP150 is upregulated by CHOP/GADD153 in response to mitochondrial and ER stress. Our data also reveal that ORP150 is a substrate for mitochondrial calpain 10.
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PMID:Targeting of the molecular chaperone oxygen-regulated protein 150 (ORP150) to mitochondria and its induction by cellular stress. 1809 45

Hypoxia-inducible transcription factor-1alpha and -2alpha (HIF-alpha) proteins and regulated genes are increased in preeclamptic (PE) placentas. Although placental hypoxia likely stabilizes HIF-alpha proteins, we previously reported that there is also a defect in oxygen-dependent reduction of HIF-alpha proteins in PE relative to normal pregnant (NP) placentas that could contribute to their over-expression. After a 4-h exposure to 2% oxygen, placental villous explants were exposed to 21% oxygen over 90 min. As assessed by Western analysis, the defective oxygen-dependent reduction of HIF-1alpha protein in villous explants from PE placenta was unaffected by the protein synthesis inhibitor, cycloheximide. However, after incubation with the proteasomal inhibitor, clasto-lactacystin, oxygen-dependent reduction of HIF-1alpha protein was markedly and similarly impaired in the villous explants from both normal and PE placentas. Thus, impairment of protein degradation rather than increased synthesis causes inadequate oxygen-dependent reduction of HIF-1alpha protein in PE placentas. Immunoprecipitation studies revealed comparable association of HIF-1alpha with von Hippel Lindau (VHL) protein in placentas from NP and PE women. Furthermore, prolyl hydroxylase-3 protein was appropriately upregulated in the PE placentas as determined by Western analysis paralleling the increases of HIF-alpha proteins. These results suggest that molecular events leading to the formation of the HIF-1alpha:VHL:ubiquitin ligase complex are most likely not impaired in PE placentas. Finally, proteasomal trypsin, chymotrypsin, and peptidyl glutamyl-like activities were significantly reduced by approximately 1/3 in PE placentas by using specific peptide substrates coupled to a fluorescent tag. Unexpectedly, however, they were even further decreased in placentas from normotensive women delivering growth restricted babies >37 weeks gestation-placentas which do not have elevated HIF-alpha proteins. In conclusion, accumulation of HIF-alpha proteins in PE placentas may occur as a consequence of both increased formation secondary to relative ischemia/hypoxia and reduced degradation after reperfusion/oxygenation consequent to proteasomal dysfunction. In contrast, in placentas from normotensive women delivering growth restricted babies >37 weeks gestation, proteasomal activity, albeit markedly reduced, is adequate to cope with degradation of HIF-alpha proteins, which have not been increased by an hypoxic environment.
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PMID:Proteasomal activity in placentas from women with preeclampsia and intrauterine growth restriction: implications for expression of HIF-alpha proteins. 1822 38


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