Hypoxic Animals Research Articles

Neuroprotective effects of hypothermia on synaptic actin cytoskeletal changes induced by perinatal asphyxia

Cerebral hypoxia–ischemia damages synaptic proteins, resulting in cytoskeletal alterations, protein aggregation and neuronal death. In the previous works, we have shown neuronal and synaptic changes in rat neostriatum subjected to hypoxia that leads to ubi-protein accumulation. Recently, we also showed that, changes in F-actin organization could be related to early alterations induced by hypoxia in the Central Nervous System. However, little is known about effective treatment to diminish the damage. The main aim of this work is to study the effects of birth hypothermia on the actin cytoskeleton of neostriatal post-synaptic densities (PSD) in 60 days olds rats by immunohistochemistry, photooxidation and western blot. We used 2 different protocols of hypothermia: (a) intrahypoxic hypothermia at 15°C and (b) post-hypoxia hypothermia at 32°C. Consistent with previous data at 30 days, staining with phalloidin–Alexa488 followed by confocal microscopy analysis showed an increase of F-actin fluorescent staining in the neostriatum of hypoxic animals. Correlative photooxidation electron microscopy confirmed these observations showing an increment in the number of mushroom-shaped F-actin staining spines in neostriatal excitatory synapses in rats subjected to hypoxia. In addition, western blot revealed β-actin increase in PSDs in hypoxic animals. The optic relative density measurement showed a significant difference between controls and hypoxic animals. When hypoxia was induced under hypothermic conditions, the changes observed in actin cytoskeleton were blocked. Post-hypoxic hypothermia showed similar answer but actin cytoskeleton modifications were not totally reverted as we observed at 15°C. These data suggest that the decrease of the body temperature decreases the actin modifications in dendritic spines preventing the neuronal death.

Prolonged Hypoxia Increases Survival Even in Zebrafish (Danio rerio) Showing Cardiac Arrhythmia

Tolerance towards hypoxia is highly pronounced in zebrafish. In this study even beneficial effects of hypoxia, specifically enhanced survival of zebrafish larvae, could be demonstrated. This effect was actually more pronounced in breakdance mutants, which phenotypically show cardiac arrhythmia. Breakdance mutants (bre) are characterized by chronically reduced cardiac output. Despite an about 50% heart rate reduction, they become adults, but survival rate significantly drops to 40%. Normoxic bre animals demonstrate increased hypoxia inducible factor 1 a (Hif-1α) expression, which indicates an activated hypoxic signaling pathway. Consequently, cardiovascular acclimation, like cardiac hypertrophy and increased erythrocyte concentration, occurs. Thus, it was hypothesized, that under hypoxic conditions survival might be even more reduced. When bre mutants were exposed to hypoxic conditions, they surprisingly showed higher survival rates than under normoxic conditions and even reached wildtype values. In hypoxic wildtype zebrafish, survival yet exceeded normoxic control values. To specify physiological acclimation, cardiovascular and metabolic parameters were measured before hypoxia started (3 dpf), when the first differences in survival rate occurred (7 dpf) and when survival rate plateaued (15 dpf). Hypoxic animals expectedly demonstrated Hif-1α accumulation and consequently enhanced convective oxygen carrying capacity. Moreover, bre animals showed a significantly enhanced heart rate under hypoxic conditions, which reached normoxic wildtype values. This improvement in convective oxygen transport ensured a sufficient oxygen and nutrient supply and was also reflected in the significantly higher mitochondrial activity. The highly optimized energy metabolism observed in hypoxic zebrafish larvae might be decisive for periods of higher energy demand due to organ development, growth and increased activity. However, hypoxia increased survival only during a short period of development and starting hypoxia before or after this phase reduced survival, particularly in bre animals. Thus, the physiological plasticity, which enables zebrafish larvae to benefit from a hypoxia, occurs only within a narrow developmental window.

High proliferative potential endothelial colony-forming cells contribute to hypoxia-induced pulmonary artery vasa vasorum neovascularization

Angiogenic expansion of the vasa vasorum (VV) is an important contributor to pulmonary vascular remodeling in the pathogenesis of pulmonary hypertension (PH). High proliferative potential endothelial progenitor-like cells have been described in vascular remodeling and angiogenesis in both systemic and pulmonary circulations. However, their role in hypoxia-induced pulmonary artery (PA) VV expansion in PH is not known. We hypothesized that profound PA VV neovascularization observed in a neonatal calf model of hypoxia-induced PH is due to increased numbers of subsets of high proliferative cells within the PA adventitial VV endothelial cells (VVEC). Using a single cell clonogenic assay, we found that high proliferative potential colony-forming cells (HPP-CFC) comprise a markedly higher percentage in VVEC populations isolated from the PA of hypoxic (VVEC-Hx) compared with control (VVEC-Co) calves. VVEC-Hx populations that comprised higher numbers of HPP-CFC also demonstrated markedly higher expression levels of CD31, CD105, and c-kit than VVEC-Co. In addition, significantly higher expression of CD31, CD105, and c-kit was observed in HPP-CFC vs. the VVEC of the control but not of hypoxic animals. HPP-CFC exhibited migratory and tube formation capabilities, two important attributes of angiogenic phenotype. Furthermore, HPP-CFC-Co and some HPP-CFC-Hx exhibited elevated telomerase activity, consistent with their high replicative potential, whereas a number of HPP-CFC-Hx exhibited impaired telomerase activity, suggestive of their senescence state. In conclusion, our data suggest that hypoxia-induced VV expansion involves an emergence of HPP-CFC populations of a distinct phenotype with increased angiogenic capabilities. These cells may serve as a potential target for regulating VVEC neovascularization.

Adaptogenic effects of dihydroquercetin-chitosan composition during modeling of acute hypoxia.

The course of preventive oral treatment with a dihydroquercetin-chitosan composition produced a strong antihypoxic effect under conditions of experimental hypobaric hypoxia (arbitrary altitude 12,000m). The lactate/pyruvate ratio in composition-receiving rats was much lower than in hypoxic animals (by 83%), but higher than in intact specimens (by 29%). The composition of chitosan and dihydroquercetin also possessed a strong antioxidant activity.

Effect of ghrelin on renal erythropoietin production in chronic hypoxic rats

Ghrelin, a 28 amino acid peptide, has diverse effects in body organs. Erythropoietin is a key mediator in increasing the red blood cells during hypoxia. Previously, we have shown that ghrelin has a polycythemic effect. In the present study, we evaluated the effect of ghrelin on erythropoietin gene expression with the aim to find out the mechanism of its effect. Thirty two adult male Wistar rats were divided randomly into four groups. The hypoxic condition was induced by placing the rats into the hypoxic chamber with 11% oxygen for two weeks. Saline- and ghrelin-treated control rats remained in room with a regular air conditions. Erythropoietin gene expression was measured by real-time reverse transcription-polymerase chain reaction (RT-PCR). Plasma erythropoietin was measured by enzyme linked immunosorbent assay (ELISA). After 2-weeks of hypoxia, erythropoietin transcripts and erythropoietin plasma levels were significantly increased in hypoxic animals compared with control animals. Ghrelin treatment decreased both plasma erythropoietin and erythropoietin gene expression only in the hypoxic rats. Our data indicate that ghrelin might induce polycythemia through an erythropoietin-independent manner. However, to confirm this hypothesis and to find out the precise mechanism of this phenomenon further investigations are needed.

Sodium hydrosulfide prevents hypoxia-induced behavioral impairment in neonatal mice

Hypoxic encephalopathy is a common cause of neonatal seizures and long-term neurological abnormalities. Endogenous hydrogen sulfide (H2S) may have multiple functions in brain. The aim of this study is to investigate whether sodium hydrosulfide (NaHS), a H2S donor, provides protection against neonatal hypoxia-induced neurobehavioral deficits. Neonatal mice were subjected to hypoxia (5% oxygen for 120min) at postnatal day 1 and received NaHS (5.6mg/kg) once daily for 3d. Neurobehavioral toxicity was examined at 3–30d after hypoxia. Treatment with NaHS significantly attenuated the delayed development of sensory and motor reflexes induced by hypoxia up to two weeks after the insult. Moreover, NaHS improved the learning and memory performance of hypoxic animals as indicated in Morris water maze test at 30d after hypoxia. In mice exposed to hypoxia, treatment with NaHS enhanced expression of brain derived neurotrophic factor (BDNF) in the hippocampus. Furthermore, the protective effects of NaHS were associated with its ability to repress the hypoxia-induced nitric oxide synthase (NOS) activity and nitric oxide production in the hippocampus of mice brain. Taken together, these results suggest that the long-lasting beneficial effects of NaHS on hypoxia-induced neurobehavioral deficits are mediated, at least in part, by inducing BDNF expression and suppressing NOS activity in the brain of mice.

Melatonin improves short and long-term neurobehavioral deficits and attenuates hippocampal impairments after hypoxia in neonatal mice

Hypoxic encephalopathy is a common cause of neonatal seizures and long-term neurobehavioral abnormalities. The purpose of this study was to determine whether administration of melatonin, starting at 1h before hypoxia and then every 24h for 3 days, influences short and long-term neurobehavioral development and hippocampal impairments in postnatal day 1 mice subjected to hypoxia (5% oxygen and 95% nitrogen for 120min). Melatonin significantly attenuated hypoxia-induced neurobehavioral deficits, including sensorimotor performance, locomotor functions, and hyperactivity up to two weeks after hypoxia insult. The above-mentioned functional benefits of melatonin were associated with attenuation of cell death in the hippocampus. Importantly, melatonin improved learning and memory performance in the Morris water test, as associated with significantly increased proliferating cells (BrdU-positive cells) and differentiating neuroblasts (doublecortin-positive neuroblasts) in the hippocampus of hypoxic animals at 30 days after hypoxia. In addition, melatonin significantly decreased microglial activation and overproduction of pro-inflammatory mediators (tumor necrosis factor-α, interleukin-1β and nitric oxide) from 3 to 30 days after hypoxia, possibly by inhibiting NF-κB activation in the hippocampus. The present results show that melatonin has short- and long-term protective effects against hypoxia-induced neurobehavioral deficits in the neonatal mouse. These beneficial effects are associated with increasing neurogenesis and attenuation of cell death and inflammatory responses in the hippocampus.

Serotonin receptors take the TRiPV4 highway in chronic hypoxic pulmonary hypertension. Focus on “TRPV4 channel contributes to serotonin-induced pulmonary vasoconstriction and the enhanced vascular reactivity in chronic hypoxic pulmonary hypertension”

pulmonary arterial hypertension (PAH) is a rare human disease displaying a very poor prognosis of survival. The chronic increase in pulmonary vascular resistance associated with PAH eventually leads to right ventricular hypertrophy and failure, and ultimately death. It is well accepted that the

Identifying Hypoxia in a Newborn Piglet Model Using Urinary NMR Metabolomic Profiling

Establishing the severity of hypoxic insult during the delivery of a neonate is key step in the determining the type of therapy administered. While successful therapy is present, current methods for assessing hypoxic injuries in the neonate are limited. Urine Nuclear Magnetic Resonance (NMR) metabolomics allows for the rapid non-invasive assessment of a multitude breakdown products of physiological processes. In a newborn piglet model of hypoxia, we used NMR spectroscopy to determine the levels of metabolites in urine samples, which were correlated with physiological measurements. Using PLS-DA analysis, we identified 13 urinary metabolites that differentiated hypoxic versus nonhypoxic animals (1-methylnicotinamide, 2-oxoglutarate, alanine, asparagine, betaine, citrate, creatine, fumarate, hippurate, lactate, N-acetylglycine, N-carbamoyl-β-alanine, and valine). Using this metabolomic profile, we then were able to blindly identify hypoxic animals correctly 84% of the time compared to nonhypoxic controls. This was better than using physiologic measures alone. Metabolomic profiling of urine has potential for identifying neonates that have undergone episodes of hypoxia.

Chronic Hypoxia Inhibits Pregnancy-Induced Upregulation of SK Ca Channel Expression and Function in Uterine Arteries

Small-conductance Ca(2+)-activated K(+) (SKCa) channels are crucial in regulating vascular tone and blood pressure. The present study tested the hypothesis that SKCa channels play an important role in uterine vascular adaptation in pregnancy, which is inhibited by chronic hypoxia during gestation. Uterine arteries were isolated from nonpregnant and near-term pregnant sheep maintained at sea level (≈300 m) or exposed to high-altitude (3801 m) hypoxia for 110 days. Immunohistochemistry revealed the presence of SKCa channels type 2 (SK2) and type 3 (SK3) in both smooth muscles and endothelium of uterine arteries. The expression of SK2 and SK3 channels was significantly increased during pregnancy, which was inhibited by chronic hypoxia. In normoxic animals, both SKCa channel opener NS309 and a large-conductance (BKCa) channel opener NS1619 relaxed norepinephrine-contracted uterine arteries in pregnant but not nonpregnant sheep. These relaxations were inhibited by selective SKCa and BKCa channel blockers, respectively. NS309-induced relaxation was largely endothelium-independent. In high-altitude hypoxic animals, neither NS1691 nor NS309 produced significant relaxation of uterine arteries in either nonpregnant or pregnant sheep. Similarly, the role of SKCa channels in regulating the myogenic reactivity of uterine arteries in pregnant animals was abrogated by chronic hypoxia. Accordingly, the enhanced SKCa channel activity in uterine arterial myocytes of pregnant animals was ablated by chronic hypoxia. The findings suggest a novel mechanism of SKCa channels in regulating myogenic adaptation of uterine arteries in pregnancy and in the maladaptation of uteroplacental circulation caused by chronic hypoxia during gestation.

254 CHRONIC HYPOXIA CAUSES NO CHANGE IN CARDIAC PYRUVATE DEHYDROGENASE FLUX IN THE CONTROL OR DIABETIC RAT: AN IN VIVO STUDY

Introduction In the healthy heart, approximately 30% of the cells’ energy comes from glucose oxidation and the remaining 70% from fatty acids. On exposure to hypoxia, the balance is altered, with increased reliance on glycolysis due to reduced oxygen availability. Pyruvate dehydrogenase (PDH) is the enzyme that stands as the gatekeeper between glycolysis and the Krebs’ cycle. It has previously been demonstrated that hypoxia causes increased expression of pyruvate dehydrogenase kinase (PDK) 1, an enzyme that inhibits PDH. Upregulation of PDK1 enhances inhibition, and it can therefore be hypothesised that PDH flux will be decreased. In the diabetic heart there is a preference for fatty acid use even in the presence of elevated circulating glucose. Exposure to hypoxia and the associated upregulation of glycolysis, may to some extent rebalance fuel selection. However, it is not known if this would be the case as it has also been suggested that the diabetic heart is compromised in terms of its ability to respond to hypoxia. The advent of hyperpolarized carbon-13 ( 13 C) magnetic resonance spectroscopy (MRS) has enabled the visualisation of metabolism in real-time and in vivo . 13 C-labelled pyruvate can be injected into an anaesthetised animal in an MRS system, and spectra acquired (one per second over one minute) to show the metabolism of pyruvate and therefore the flux through PDH. Using hyperpolarized 13 C MRS, this study has investigated in vivo alterations in glucose metabolism in hypoxia in comparison with normoxia, and how these alterations are affected by diabetes. Methods A rat model of late-stage type 2 diabetes was induced by high fat feeding and injection of streptozotocin (25mg/kg). Diabetic and control (chow-fed) animals (n=8) were housed for 3 weeks either in hypoxia (one week gradual acclimatisation and two further weeks at 11% oxygen) or normoxia. Animals were then anaesthetised and scanned in a 7T MRS system, with an injection of hyperpolarized [1- 13 C]-labelled pyruvate and cardiac spectra acquired over ∼1 min. Animals were terminated after scanning, blood samples taken and tissue samples freeze-clamped for later analysis. Data were analysed using jMRUI and the AMARES package, and the conversion of pyruvate into bicarbonate taken as a measure of PDH flux. Results There were no alterations in cardiac PDH flux (Kpyr-bic) observed in hypoxic control animals when compared to normoxic controls. There were also no changes in 13 C label transfer to lactate (Kpyr-lac). Diabetic animals in both normoxia and hypoxia demonstrated decreased PDH flux when compared to controls, however there was no interaction in terms of PDH flux between the hypoxic and diabetic conditions (Figure 1). Conclusions We have observed no alteration in cardiac PDH flux caused by three weeks of chronic hypoxia. In addition, whilst there was a significant reduction in PDH flux in diabetes, exposure to chronic hypoxia had no further metabolic effect on PDH flux.

P1: CD47 drives pulmonary hypertension through coordinate regulation of c-Myc and endothelin-1

Background Pulmonary hypertension (PH), regardless of etiology, remains progressive and incurable despite intensive research for over half a century. Existing therapies have only modestly enhanced survival with the only available “cure” being lung transplantation. The reasons for lack of progress in this area remain unknown but suggest a deficit in understanding the overarching mechanism (s) that drive PH. Recently we reported that the thrombospondin-1 (TSP1)-CD47 ligand receptor pathway is induced in multiple pre-clinical models of PH and human disease. Mice lacking secreted TSP1 are protected from PH. Methods Wild type and CD47 null mice were challenged with hypoxia and cardiopulmonary assessment performed. Likewise, in cell culture experiments pulmonary arterial vascular smooth muscle cells were exposed to hypoxia and molecular signal transduction assessed. Results We know report that mice mutated to lack cell receptor CD47 are highly resistant to hypoxia-mediated PH with minimal evidence of pulmonary arterial overgrowth and no right ventricular hypertrophy compared to controls. Hypoxic CD47 null mice demonstrated increased stroke volume and cardiac output compared to a significant deterioration of these functional parameters in wild type controls. Hypoxic wild type animals also displayed concurrent loss of cMyc and upregulation of endothelin-1. In contrast, hypoxic CD47 null animals demonstrated cMyc-mediated suppression of endothelin-1 signaling. Impressively, under basal conditions endothelin-1 and the cognate receptors ETA and ETB where nearly undetectable in CD47 null lungs. Disrupting TSP1-mediated activation of CD47 in monocrotaline treated rats, and in hypoxic pulmonary arterial smooth muscle cell cultures, upregulated cMyc, inhibited endothelin-1 signaling and corrected established PH. Finally, antibody blocking CD47 in lungs from patients with end-stage PH normalized the vasodilation profile of distal pulmonary arteries to acetylcholine and sodium nitroprusside. Conclusions The above preclinical results suggest CD47 is an important and proximate promoter of PH through cMyc-mediated inhibition of endothelin-1, while results in diseased human PH lung arterioles suggest that CD47 is a clinical target to restore end-stage pulmonary arteriole function. Disclosure J.S.I. is Chair of the Scientific Advisory Boards and has equity interest in Vasculox, Inc. (St. Louis, MO) and Radiation Control Technologies, Inc. (Rockville, MD).

Intermittent hypoxia increases melanoma metastasis to the lung in a mouse model of sleep apnea

Obstructive sleep apnea (OSA) has recently been associated with an increased risk of cancer incidence and mortality in humans. Experimental data in mice have also shown that intermittent hypoxia similar to that observed in OSA patients enhances tumor growth. The aim of this study was to test the hypothesis that intermittent hypoxia mimicking OSA enhances lung metastasis. A total of 75 C57BL/6J male mice (10-week-old) were subjected to either spontaneous or induced melanoma lung metastasis. Normoxic animals breathed room air and intermittent hypoxic animals were subjected to cycles of 20s of 5% O2 followed by 40s of room air for 6h/day. Spontaneous and induced lung metastases were studied after subcutaneous and intravenous injection of B16F10 melanoma cells, respectively. Compared with normoxia, intermittent hypoxia induced a significant increase in melanoma lung metastasis. These animal model results suggest that intermittent hypoxia could contribute to cancer metastasis in patients with OSA.

Deep hypothermia protects against acute hypoxia in vivo in rats: a mechanism related to the attenuation of oxidative stress

There is growing interest in using hypothermia to prevent hypoxic damage in clinical and experimental models, although the mechanisms regulated by hypothermia are still unclear. As reactive oxygen and nitrogen species are the main factors causing cellular damage, our objective was to study the scope of hypothermia in preventing hypoxia-induced oxidative damage. We analysed systemic and hepatic indicators of oxidative stress after an acute hypoxic insult (10% oxygen in breathing air) in normothermic (37°C body temperature) and hypothermic conditions (22°C) in rats. Exposure to hypoxia resulted in tissue damage (aspartate aminotransferase increased from 54.6 ± 6.9 U l(-1) in control animals to 116 ± 1.9 U l(-1) in hypoxia, and alanine aminotransferase increased from 19 ± 0.8 to 34 ± 2.9 U l(-1)), oxidative stress (nitric oxide metabolites increased from 10.8 ± 0.4 μM in control rats to 23 ± 2.7 μM in hypoxia, and thiobarbituric reactive substances increased from 3.3 ± 0.2 to 5.9 ± 0.4 nm) and antioxidant consumption (reduced/oxidized glutathione ratio changed from 9.8 ± 0.3 to 6.8 ± 0.3). In contrast, when hypothermia was applied prior to hypoxia, the situation was reversed, with a reduction in aspartate aminotransferase (from 116 ± 1.9 in hypoxic animals to 63 ± 7.8 U l(-1) in animals exposed to hypothermia followed by hypoxia), alanine aminotransferase (from 34 ± 2.9 to 19 ± 0.9 U l(-1)), oxidative stress (nitric oxide metabolites decreased from 23 ± 2.7 to 17.8 ± 1.9 μM and thiobarbituric acid-reactive substances decreased from 5.9 ± 0.4 to 4.3 ± 0.2 nm) and antioxidant preservation (reduced/oxidized glutathione ratio changed from 6.8 ± 0.3 to 11.1 ± 0.1). Hypoxia induced a decrease in liver enzymatic antioxidant activities even during hypothermia. Both treatments, hypoxia and hypothermia, produced a similar increase in hepatic caspase-3 activity. In conclusion, hypothermia prevented the tissue damage and oxidative stress elicited by hypoxia. Our results provide new evidence concerning the protective mechanism of hypothermia in vivo.

Change of Genes in Calcium Transport Channels Caused by Hypoxic Stress in the Placenta, Duodenum, and Kidney of Pregnant Rats1

Preeclampsia is a pregnancy-specific disease characterized by concurrent development of hypertension, proteinuria, and oxidative stress in the placenta. In this study, we induced hypoxic stress in rats during pregnancy to reproduce physiological conditions associated with preeclampsia. The maternal weight of hypoxic pregnant rats was lower than that of normoxic animals. The level of calcium ions were also increased in urine collected from the hypoxic animals. In contrast, urinary concentrations of sodium, chloride, and potassium ions declined in hypoxic rats, and developed to proteinuria. The expression of genes known as two biomarkers, sFLT1 (for preeclampsia) and HIF-1alpha (for hypoxia), were highly induced in the placenta, duodenum, and kidney by hypoxic stress. The overexpression of sFLT1 and HIF-1alpha demonstrated that our experimental conditions closely mimicked ones that are associated with preeclampsia. In the present study, we measured the expression of calcium transporters (TRPV5, TRPV6, PMCA1, NCKX3, NCX1, and CaBP-9k) in the placenta, duodenum, and kidney under hypoxic conditions on Gestational Day 19.5 in rats. Placental TRPV5, TRPV6, and PMCA1 expression was up-regulated in the hypoxic rats, whereas the levels of NCX1 and CaBP-9k were unchanged. In addition, NCKX3 expression was increased in the placenta of hypoxic rats. Duodenal expression of CaBP-9k, TRPV5, TRPV 6, and PMCA1 was decreased in the hypoxic rats, whereas levels of NCXs were not altered. Renal expression of NCKX3 and TRPV6 was increased, whereas NCX1 was decreased in the hypoxic rats compared to the normoxic controls. Taken together, these results indicate that physiological changes observed in the hypoxic rats were similar to ones associated with preeclampsia. Expression of calcium transport genes in the placenta, duodenum, and kidney perturbed by hypoxic stress during pregnancy may cause calcium loss in the urine, and thereby induce calcium-deficient characteristics of preeclampsia.

Fetal Hypoxia Results in Programming of Aberrant Angiotensin II Receptor Expression Patterns and Kidney Development

AIMS: The present study tested the hypothesis that fetal hypoxia adversely affects kidney development in fetal and offspring rats and alter the expression patterns of angiotensin II type 1 (AT1R) and type 2 (AT2R) receptors.METHODS: Time-dated pregnant rats were divided between normoxic and hypoxic (10.5% O2 last period of gestation) groups. Protein expression, in the offspring, was determined using western blot.RESULTS: Hypoxic treatment significantly decreased body and kidney weight in 21-day fetuses (E21) and 7-day neonates (P7). In 3-month-old offspring there were no significant differences in body and kidney weight between hypoxic and control animals. Fetal hypoxia had no effect on kidney AT1R density in E21 or P7, but significantly decreased kidney AT1R protein and mRNA abundance in both male and female adults. In contrast, kidney AT2R density was not affected by fetal hypoxia throughout the developmental stages studied. The hypoxia-mediated reduction of nephron numbers was progressively from P7 worsened into the adulthood with females affected more than males.CONCLUSION: The results suggest that fetal hypoxia causes programming of aberrant kidney development and accelerates the aging process of the kidney during the postnatal development, which may contribute to an increased risk of cardiovascular disease.

Chronic Hypoxia Differentially Up-Regulates Protein Kinase C-Mediated Ovine Uterine Arterial Contraction via Actin Polymerization Signaling in Pregnancy1

Chronic hypoxia (CH) during pregnancy is associated with increased uterine vascular tone. The present study tested the hypothesis that CH up-regulates protein kinase C (PKC)-mediated actin polymerization, resulting in enhanced uterine vascular contraction in pregnancy. Uterine arteries were isolated from nonpregnant (NPUA) and near-term (∼140 days of gestation) pregnant (PUA) sheep that had been maintained at sea level (∼300 m) or exposed to high altitude (3801 m) hypoxia for 110 days. In normoxic animals, the induced contractions by the PKC activator phorbol 12,13-dibutyrate (PDBu) were greater in NPUA than in PUA, which was abrogated by an actin polymerization inhibitor cytochalasin B (Cyto B). In hypoxic animals, PDBu-induced contractions were significantly increased in PUA but not in NPUA, which was inhibited by Cyto B. In contrast, neither pregnancy nor hypoxia affected Cyto B-mediated inhibition of norepinephrine (NE)-induced contractions. Prolonged ex vivo treatment of NPUA with 17beta-estradiol and progesterone significantly attenuated PDBu-induced actin polymerization and contractions, and the hormonal treatment did not alter the inhibitory effect of Cyto B on PDBu- or NE-induced contractions in either normoxic or hypoxic animals. 2-(2-Amino-3-methoxyphenyl)-4H-1-benzopyran-4-one potentiated PDBu-mediated actin polymerization and enhanced PDBu-induced contractions of PUA in normoxic but not hypoxic animals, which was abrogated by Cyto B. The results suggest that chronic hypoxia during pregnancy causes attenuation of steroid hormone-mediated ERK1/2 signaling and results in increased actin polymerization and uterine vascular tone, linking gestational hypoxia to aberrant uteroplacental circulation.

S36 P38 MAPK: An Important Pathway in the Pathobiology of Pulmonary Hypertension and Pulmonary Vascular Remodelling

The p38 MAPK pathway is increasingly recognised as important in inflammation leading to systemic vascular disease but its role in pulmonary vascular disease is unclear. Our group has previously identified...

The role of oxygen during fracture healing

Oxygen affects the activity of multiple skeletogenic cells and is involved in many processes that are important for fracture healing. However, the role of oxygen in fracture healing has not been fully studied. Here we systematically examine the effects of oxygen tension on fracture healing and test the ability of hyperoxia to rescue healing defects in a mouse model of ischemic fracture healing. Mice with tibia fracture were housed in custom-built gas chambers and groups breathed a constant atmosphere of 13% oxygen (hypoxia), 21% oxygen (normoxia), or 50% oxygen (hyperoxia). The influx of inflammatory cells to the fracture site, stem cell differentiation, tissue vascularization, and fracture healing were analyzed. In addition, the efficacy of hyperoxia (50% oxygen) as a treatment regimen for fracture nonunion was tested. Hypoxic animals had decreased tissue vascularity, decreased bone formation, and delayed callus remodeling. Hyperoxia increased tissue vascularization, altered fracture healing in un-complicated fractures, and improved bone repair in ischemia-induced delayed fracture union. However, neither hypoxia nor hyperoxia significantly altered chondrogenesis or osteogenesis during early stages of fracture healing, and infiltration of macrophages and neutrophils was not affected by environmental oxygen after bone injury. In conclusion, our results indicate that environmental oxygen levels affect tissue vascularization and fracture healing, and that providing oxygen when fractures are accompanied by ischemia may be beneficial.

Chronic Caffeine Intake in Adult Rat Inhibits Carotid Body Sensitization Produced by Chronic Sustained Hypoxia but Maintains Intact Chemoreflex Output

Sustained hypoxia produces a carotid body (CB) sensitization, known as acclimatization, which leads to an increase in carotid sinus nerve (CSN) activity and ensuing hyperventilation greater than expected from the prevailing partial pressure of oxygen. Whether sustained hypoxia is physiological (high altitude) or pathological (lung disease), acclimatization has a homeostatic implication because it tends to minimize hypoxia. Caffeine, the most commonly ingested psychoactive drug and a nonselective adenosine receptor antagonist, alters CB function and ventilatory responses when administered acutely. Our aim was to investigate the effect of chronic caffeine intake on CB function and acclimatization using four groups of rats: normoxic, caffeine-treated normoxic, chronically hypoxic (12% O₂, 15 days), and caffeine-treated chronically hypoxic rats. Caffeine was administered in drinking water (1 mg/ml). Caffeine ameliorated ventilatory responses to acute hypoxia in normoxic animals without altering the output of the CB (CSN neural activity). Caffeine-treated chronically hypoxic rats exhibited a decrease in the CSN response to acute hypoxia tests but maintained ventilation compared with chronically hypoxic animals. The findings related to CSN neural activity combined with the ventilatory responses indicate that caffeine alters central integration of the CB input to increase the gain of the chemoreflex and that caffeine abolishes CB acclimatization. The putative mechanisms involved in sensitization and its loss were investigated: expression of adenosine receptors in CB (A(2B)) was down-regulated and that in petrosal ganglion (A(2A)) was up-regulated in caffeine-treated chronically hypoxic rats; both adenosine and dopamine release from CB chemoreceptor cells was increased in chronic hypoxia and in caffeine-treated chronic hypoxia groups.