losartan-potassium and oxalylglycine

Prolyl hydroxylase (PHD) inhibitors are being developed as alternatives to recombinant human erythropoietin (rHuEPO) for the treatment of anemia in patients with chronic kidney disease (CKD). However, the effects of PHD inhibitors and rHuEPO on blood pressure and CKD in animal models susceptible to hypertension and nephropathy have not been studied. The present study compared the effects of dimethyloxaloylglycine (DMOG), a PHD inhibitor, and rHuEPO on the development of hypertension and renal injury in Dahl salt-sensitive rats fed an 8% salt diet for 3 weeks. DMOG and rHuEPO were equally effective at raising hemoglobin levels. Systolic blood pressure rose to a greater extent in rHuEPO-treated rats (267 ± 10 vs. 226 ± 4 mm Hg) than in rats given DMOG (189 ± 8 mm Hg). Urinary protein excretion increased to 568 ± 54 versus 353 ± 25 mg/day in rats treated with rHuEPO and vehicle; however, it only rose to 207 ± 21 mg/day in rats receiving DMOG. DMOG significantly attenuated the degree of glomerulosclerosis and renal interstitial fibrosis as compared with that in vehicle and rHuEPO-treated rats. This was associated with lower renal levels of monocyte chemoattractant protein-1 and interleukin-1

Topics: Amino Acids, Dicarboxylic; Anemia; Animals; Blood Pressure; Dose-Response Relationship, Drug; Erythropoietin; Fibrosis; Hemoglobins; Hypertension; Kidney; Male; Oxidative Stress; Prolyl Hydroxylases; Prolyl-Hydroxylase Inhibitors; Rats; Rats, Inbred Dahl; Recombinant Proteins; Renal Insufficiency, Chronic; Sodium Chloride, Dietary; Vascular Endothelial Growth Factor A

Traumatic brain injury (TBI) is one of the major cause of morbidity and mortality and it affects more than 1.7 million people in the USA. A couple of regenerative pathways including activation of hypoxia-inducible transcription factor 1 alpha (HIF-1α) are initiated to reduce cellular damage following TBI; however endogenous activation of these pathways is not enough to provide neuroprotection after TBI. Thus we aimed to see whether sustained activation of HIF-1α can provide neuroprotection and neurorepair following TBI. We found that chronic treatment with dimethyloxaloylglycine (DMOG) markedly increases the expression level of HIF-1α and mRNA levels of its downstream proteins such as Vascular endothelial growth factor (VEGF), Phosphoinositide-dependent kinase-1 and 4 (PDK1, PDK4) and Erythropoietin (EPO). Treatment of DMOG activates a major cell survival protein kinase Akt and reduces both cell death and lesion volume following TBI. Moreover, administration of DMOG augments cluster of differentiation 31 (CD31) staining in pericontusional cortex after TBI, which suggests that DMOG stimulates angiogenesis after TBI. Treatment with DMOG also improves both memory and motor functions after TBI. Taken together our results suggest that sustained activation of HIF-1α provides significant neuroprotection following TBI.

Topics: 3-Phosphoinositide-Dependent Protein Kinases; Amino Acids, Dicarboxylic; Angiogenesis Inducing Agents; Animals; Brain Injuries, Traumatic; Cell Death; Disease Models, Animal; Erythropoietin; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Maze Learning; Memory Disorders; Mice, Inbred C57BL; Motor Activity; Neuroprotective Agents; Nootropic Agents; Protein Serine-Threonine Kinases; Pyruvate Dehydrogenase Acetyl-Transferring Kinase; RNA, Messenger; Vascular Endothelial Growth Factor A

Chronic intermittent hypoxia (CIH) is an underlying component of obstructive sleep apnoea and has been shown to have deleterious and damaging effects on central neurons and to impair synaptic plasticity in the CA1 region of the rat hippocampus. CIH has previously been shown to impair synaptic plasticity and working memory. CIH is a potent inducer of hypoxia inducible factor (HIF), a key regulator in a cell's adaptation to hypoxia that plays an important role in the fate of neurons during ischemia. Levels of HIF-1α are regulated by the activity of a group of enzymes called HIF-prolyl 4-hydroxylases (PHDs) and these have become potential pharmacological targets for preconditioning against ischemia. However little is known about the effects of prolyl hydroxylase inhibition and CIH on synaptic transmission and plasticity in sub-regions of the hippocampus. Male Wistar rats were treated for 7-days with either saline, CIH or PHD inhibition (dimethyloxaloylglycine, DMOG; 50mg/kg, i.p.). At the end of treatment all three groups showed no change in synaptic excitability using paired pulse paradigms. However long-term potentiation (LTP) was impaired in the CA1 region of the hippocampus in both CIH and DMOG treated animals. LTP induced in the dentate gyrus was not significantly affected by either CIH or DMOG treatment. We also investigated the effect of 7-day CIH and DMOG treatment on the recovery of synaptic transmission following an acute 30min hypoxic insult. CIH treated animals showed an improved rate of recovery of synaptic transmission following re-oxygenation in both the CA1 and the dentate gyrus. These results suggest that LTP induction in the CA1 region is more sensitive to both CIH and DMOG treatments than the dentate gyrus.

Topics: Amino Acids, Dicarboxylic; Animals; CA1 Region, Hippocampal; CREB-Binding Protein; Dentate Gyrus; Erythropoietin; Hematocrit; Hippocampus; Hypoxia-Inducible Factor 1, alpha Subunit; Hypoxia-Inducible Factor-Proline Dioxygenases; Hypoxia, Brain; Male; Neuronal Plasticity; Rats; Rats, Wistar; Synaptic Transmission

Activation of hypoxia-inducible factor (HIF) can prevent oxygen-induced retinopathy in rodents. Here we demonstrate that dimethyloxaloylglycine (DMOG)-induced retinovascular protection is dependent on hepatic HIF-1 because mice deficient in liver-specific HIF-1α experience hyperoxia-induced damage even with DMOG treatment, whereas DMOG-treated wild-type mice have 50% less avascular retina (P < 0.0001). Hepatic HIF stabilization protects retinal function because DMOG normalizes the b-wave on electroretinography in wild-type mice. The localization of DMOG action to the liver is further supported by evidence that i) mRNA and protein erythropoietin levels within liver and serum increased in DMOG-treated wild-type animals but are reduced by 60% in liver-specific HIF-1α knockout mice treated with DMOG, ii) triple-positive (Sca1/cKit/VEGFR2), bone-marrow-derived endothelial precursor cells increased twofold in DMOG-treated wild-type mice (P < 0.001) but are unchanged in hepatic HIF-1α knockout mice in response to DMOG, and iii) hepatic luminescence in the luciferase oxygen-dependent degradation domain mouse was induced by subcutaneous and intraperitoneal DMOG. These findings uncover a novel endocrine mechanism for retinovascular protection. Activating HIF in visceral organs such as the liver may be a simple strategy to protect capillary beds in the retina and in other peripheral tissues.

Topics: Amino Acids, Dicarboxylic; Animals; Erythropoietin; Hyperoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Liver; Mice; Mice, Knockout; Oxygen; Retinal Diseases

To study the effect of systemic hypoxia-inducible factor prolyl hydroxylase inhibition (HIF PHDi) in the rat 50/10 oxygen-induced retinopathy (OIR) model.. Oxygen-induced retinopathy was created with the rat 50/10 OIR model. OIR animals received intraperitoneal injections of dimethyloxalylglycine (DMOG, 200 μg/g), an antagonist of α-ketoglutarate cofactor and inhibitor for HIF PHD, on postnatal day (P)3, P5, and P7. Control animals received intraperitoneal injections of PBS. On P14 and P21, animals were humanely killed and the effect on vascular obliteration, tortuosity, and neovascularization quantified. To analyze HIF and erythropoietin, rats at P5 were injected with DMOG (200 μg/g). Western blot or ELISA measured the levels of HIF-1 and Epo protein. Epo mRNA was measured by quantitative PCR.. Alternating hyperoxia and hypoxia in untreated rats led to peripheral vascular obliteration on day P14 and P21. Rats that were treated with systemic DMOG by intraperitoneal injections had 3 times less ischemia and greater peripheral vascularity (P = 0.001) than control animals treated with PBS injections. Neovascularization similarly decreased by a factor of 3 (P = 0.0002). Intraperitoneal DMOG administration increased the levels of HIF and Epo in the liver and brain. Serum Epo also increased 6-fold (P = 0.0016). Systemic DMOG had no adverse effect on growth of rats treated with oxygen.. One of the many controversies in the study of retinopathy of prematurity is whether hyperoxia or alternating hyperoxia and hypoxia creates the disease phenotype in humans. We have previously demonstrated that PHDi prevents OIR in mice exposed to 5 days of sustained 75% oxygen followed by 5 days of 21% oxygen. The 50/10 rat experiments demonstrate that PHDi is also effective in a 24-hour alternating hyperoxia-hypoxia model. The rat OIR model further validates the therapeutic value of HIF PHDi to prevent retinopathy of prematurity because it reduces oxygen-induced vascular obliteration and retinovascular growth attenuation in prolonged and/or alternating hyperoxia.

Topics: Amino Acids, Dicarboxylic; Animals; Animals, Newborn; Blotting, Western; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Erythropoietin; Hyperoxia; Hypoxia-Inducible Factor 1; Injections, Intraperitoneal; Neovascularization, Pathologic; Oxygen; Procollagen-Proline Dioxygenase; Rats; Retinal Diseases; Retinal Vessels

Understanding the endogenous survival pathways induced by ischemic tolerance may yield targets for neuroprotection from stroke. One well-studied pathway reported to be evoked by preconditioning stimuli is the transcription factor HIF (hypoxia-inducible factor). However, whether HIF induction by ischemic insults is neuroprotective or toxic is still unclear. We examined the ability of three prolyl-hydroxylase inhibitors, which induce HIF, to protect hippocampal cultures from oxygen-glucose deprivation. Hippocampal cultures were exposed to ischemic preconditioning or various concentrations of cobalt chloride, deferoxamine (DFO) or dimethyloxylalyglycine (DMOG), prior to lethal oxygen-glucose deprivation (OGD). Cell survival of neurons and astrocytes was determined with dual-label immunocytochemistry. The induction of HIF targets was assessed in mixed as well as astrocyte-enriched cultures. Ischemic preconditioning, as well as low concentrations of cobalt and DFO, enhanced the survival of neurons following OGD. However, DMOG exacerbates OGD-induced neuronal death. At low concentrations, all three prolyl-hydroxylase (PHD) inhibitors increased the survival of astrocytes. Neuroprotective concentrations of cobalt induced the transcription of the cytokine erythropoietin (EPO) in astrocyte cultures. In addition, pretreatment with recombinant human erythropoietin (rH-EPO) also protected neurons from OGD. Our data suggest that HIF-induced EPO, released from astrocytes, protects neurons from OGD. However, the three PHD inhibitors each exhibited different neuroprotective profiles at low concentrations, suggesting that not all PHD inhibitors are created equal. The protective effects at low doses is reminiscent of HIF involvement in ischemic tolerance, in which sub-lethal insults induce HIF pathways resulting in neuroprotection, whereas the high-dose toxicity suggests that over-activation of HIF is not always protective. Therefore, the choice of inhibitor and dose may determine the clinical utility of these compounds. Deferoxamine exhibited little toxicity even at higher doses, and therefore appears a promising candidate for clinical use.

Topics: Amino Acids, Dicarboxylic; Animals; Astrocytes; Brain Ischemia; Cell Survival; Cells, Cultured; Cobalt; Deferoxamine; Erythropoietin; Hippocampus; Hypoxia-Inducible Factor 1; Immunohistochemistry; Ischemic Preconditioning; Mice; Mice, Inbred C57BL; Neurons; Neuroprotective Agents; Polymerase Chain Reaction; Prolyl-Hydroxylase Inhibitors

Hypoxia-inducible factors (HIFs) are transcriptional regulators that mediate the cellular response to low oxygen. Although HIF-1 is usually considered as the principal mediator of hypoxic adaptation, several tissues and different cell types express both HIF-1 and HIF-2 isoforms under hypoxia or when treated with hypoxia mimetic chemicals such as cobalt. However, the similarities or differences between HIF-1 and HIF-2, in terms of their tissue- and inducer-specific activation and function, are not adequately characterized. To address this issue, we investigated the effects of true hypoxia and hypoxia mimetics on HIF-1 and HIF-2 induction and specific gene transcriptional activity in two hepatic cancer cell lines, Huh7 and HepG2. Both hypoxia and cobalt caused rapid induction of both HIF-1α and HIF-2α proteins. Hypoxia induced erythropoietin (EPO) expression and secretion in a HIF-2-dependent way. Surprisingly, however, EPO expression was not induced when cells were treated with cobalt. In agreement, both HIF-1- and HIF-2-dependent promoters (of PGK and SOD2 genes, respectively) were activated by hypoxia while cobalt only activated the HIF-1-dependent PGK promoter. Unlike cobalt, other hypoxia mimetics such as DFO and DMOG activated both types of promoters. Furthermore, cobalt impaired the hypoxic stimulation of HIF-2, but not HIF-1, activity and cobalt-induced HIF-2α interacted poorly with USF-2, a HIF-2-specific co-activator. These data show that, despite similar induction of HIF-1α and HIF-2α protein expression, HIF-1 and HIF-2 specific gene activating functions respond differently to different stimuli and suggest the operation of oxygen-independent and gene- or tissue-specific regulatory mechanisms involving additional transcription factors or co-activators.

Topics: Amino Acids, Dicarboxylic; Basic Helix-Loop-Helix Transcription Factors; Cell Hypoxia; Cell Line, Tumor; Cell Nucleus; Cobalt; Deferoxamine; Erythropoietin; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Liver Neoplasms; Phosphoglycerate Kinase; Promoter Regions, Genetic; Propanolamines; Protein Binding; Pyrrolidines; RNA, Messenger; Superoxide Dismutase; Transcription, Genetic; Upstream Stimulatory Factors

Quiescent hematopoietic stem cells (HSCs) preferentially reside in poorly perfused niches that may be relatively hypoxic. Most of the cellular effects of hypoxia are mediated by O2-labile hypoxia-inducible transcription factors (HIFs). To investigate the effects of hypoxia on HSCs, we blocked O2-dependent HIF-1α degradation in vivo in mice by injecting 2 structurally unrelated prolyl hydroxylase domain (PHD) enzyme inhibitors: dimethyloxalyl glycine and FG-4497. Injection of either of these 2 PHD inhibitors stabilized HIF-1α protein expression in the BM. In vivo stabilization of HIF-1a with these PHD inhibitors increased the proportion of phenotypic HSCs and immature hematopoietic progenitor cells in phase G0 of the cell cycle and decreased their proliferation as measured by 5-bromo-2'-deoxyuridine incorporation. This effect was independent of erythropoietin, the expression of which was increased in response to PHD inhibitors. Finally, pretreatment of mice with a HIF-1α stabilizer before severe, sublethal 9.0-Gy irradiation improved blood recovery and enhanced 89-fold HSC survival in the BM of irradiated mice as measured in long-term competitive repopulation assays. The results of the present study demonstrate that the levels of HIF-1α protein can be manipulated pharmacologically in vivo to increase HSC quiescence and recovery from irradiation.. HIF-1α protein stabilization increases HSC quiescence in vivo. HIF-1α protein stabilization increases HSC resistance to irradiation and accelerates recovery.

Topics: Amino Acids, Dicarboxylic; Animals; Cell Survival; Erythropoietin; Gamma Rays; Gene Expression Regulation; Hematopoietic Stem Cells; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Mice; Protease Inhibitors; Proteolysis; Radiation Injuries, Experimental; Resting Phase, Cell Cycle

Oxygen-induced retinopathy (OIR) in the mouse, like the analogous human disease retinopathy of prematurity, is an ischemic retinopathy dependent on oxygen-induced vascular obliteration. We tested the hypothesis that chemically overriding the oxygen-induced downregulation of hypoxia-inducible factor (HIF) activity would prevent vascular obliteration and subsequent pathologic neovascularization in the OIR model. Because the degradation of HIF-1alpha is regulated by prolyl hydroxylases, we examined the effect of systemic administration of a prolyl hydroxylase inhibitor, dimethyloxalylglycine, in the OIR model. Our results determine that stabilizing HIF activity in the early phase of OIR prevents the oxygen-induced central vessel loss and subsequent vascular tortuosity and tufting that is characteristic of OIR. Overall, these findings imply that simulating hypoxia chemically by stabilizing HIF activity during the causative ischemia phase (hyperoxia) of retinopathy of prematurity may be of therapeutic value in preventing progression to the proliferative stage of the disease.

Topics: Aerobiosis; Amino Acids, Dicarboxylic; Animals; Basic Helix-Loop-Helix Transcription Factors; Disease Models, Animal; Enzyme Inhibitors; Erythropoietin; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Infant, Newborn; Kidney; Liver; Mice; Oxygen; Procollagen-Proline Dioxygenase; Retina; Retinopathy of Prematurity; Vascular Endothelial Growth Factor A

Recently a family of O(2)-dependent prolyl hydroxylase domain-containing enzymes (PHD) has been identified as a cellular oxygen-sensing mechanism. Reduced prolyl hydroxylase activity initiates a signalling cascade that includes the accumulation, as well as the activation, of hypoxia-inducible factor (HIF-1alpha). In turn the transcription factor HIF-1alpha, and other targets of the PHD, elicit a myriad of incompletely understood cellular responses. In these studies we have tested: (1) whether a small-molecule prolyl hydroxylase inhibitor (PHI) can effectively activate the oxygen-sensing pathway when administered systemically to mice, and (2) whether the activation of the PHD signalling pathway at the cellular level results in whole-animal hypoxic tolerance.. Mice received daily injections of the PHI, ethyl-3,4 dihydroxybenzoate (EDHB, 100-250 mg kg(-1)) or vehicle. Tissue levels of HIF-1alpha and the serum levels of the HIF-inducible gene, erythropoietin (EPO), were measured to evaluate PHD-pathway activation. To evaluate hypoxic tolerance, the endurance and survival ability of these animals was tested in sublethal (8% O(2)) and lethal hypoxia (5% O(2)) respectively.. Systemic treatment of mice with the PHD inhibitor, EDHB, leads to elevated levels of HIF-1alpha in liver and HIF-inducible EPO in serum, indicating activation of the cellular oxygen-sensing pathway. Animals treated with EDHB display significantly increased viability and enhanced exercise performance in hypoxia.. These results demonstrate a novel pharmacological strategy to induce hypoxic tolerance and are the first to demonstrate that the activation of the PHD oxygen-sensing pathway at the cellular level is sufficient to produce a hypoxic-tolerant phenotype at the physiological level of the whole animal.

Topics: Amino Acids, Dicarboxylic; Animals; Enzyme Inhibitors; Erythropoietin; Hematocrit; Hydroxybenzoates; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Injections, Intraperitoneal; Liver; Male; Mice; Mice, Inbred Strains; Oxygen; Physical Conditioning, Animal; Physical Endurance; Procollagen-Proline Dioxygenase; Signal Transduction