Perfluorocarbon Enhanced Glasgow Oxygen Level Dependent (GOLD) magnetic resonance metabolic imaging identifies the penumbra following acute ischemic stroke

The ability to identify metabolically active and potentially salvageable ischaemic penumbra is crucial for improving treatment decisions in acute stroke patients. Our solution involves two complementary novel MRI techniques (Glasgow Oxygen Level Dependant (GOLD) Metabolic Imaging), which when combined with a perfluorocarbon (PFC) based oxygen carrier and hyperoxia can identify penumbra due to dynamic changes related to continued metabolism within this tissue compartment. Our aims were (i) to investigate whether PFC offers similar enhancement of the second technique (Lactate Change) as previously demonstrated for the T2*OC technique (ii) to demonstrate both GOLD metabolic imaging techniques working concurrently to identify penumbra, following administration of Oxycyte® (O-PFC) with hyperoxia. Methods: An established rat stroke model was utilised. Part-1: Following either saline or PFC, magnetic resonance spectroscopy was applied to investigate the effect of hyperoxia on lactate change in presumed penumbra. Part-2; rats received O-PFC prior to T2*OC (technique 1) and MR spectroscopic imaging, which was used to identify regions of tissue lactate change (technique 2) in response to hyperoxia. In order to validate the techniques, imaging was followed by [14C]2-deoxyglucose autoradiography to correlate tissue metabolic status to areas identified as penumbra. Results: Part-1: PFC+hyperoxia resulted in an enhanced reduction of lactate in the penumbra when compared to saline+hyperoxia. Part-2: Regions of brain tissue identified as potential penumbra by both GOLD metabolic imaging techniques utilising O-PFC, demonstrated maintained glucose metabolism as compared to adjacent core tissue. Conclusion: For the first time in vivo, enhancement of both GOLD metabolic imaging techniques has been demonstrated following intravenous O-PFC+hyperoxia to identify ischaemic penumbra. We have also presented preliminary evidence of the potential therapeutic benefit offered by O-PFC. These unique theranostic applications would enable treatment based on metabolic status of the brain tissue, independent of time from stroke onset, leading to increased uptake and safer use of currently available treatment options

Hyperoxia Causes Mitochondrial Fragmentation in Pulmonary Endothelial Cells by Increasing Expression of Pro-Fission Proteins

Objective—We explored mechanisms that alter mitochondrial structure and function in pulmonary endothelial cells (PEC) function after hyperoxia. Approach and Results—Mitochondrial structures of PECs exposed to hyperoxia or normoxia were visualized and mitochondrial fragmentation quantified. Expression of pro-fission or fusion proteins or autophagy-related proteins were assessed by Western blot. Mitochondrial oxidative state was determined using mito-roGFP. Tetramethylrhodamine methyl ester estimated mitochondrial polarization in treatment groups. The role of mitochondrially derived reactive oxygen species in mt-fragmentation was investigated with mito-TEMPOL and mitochondrial DNA (mtDNA) damage studied by using ENDO III (mt-tat-endonuclease III), a protein that repairs mDNA damage. Drp-1 (dynamin-related protein 1) was overexpressed or silenced to test the role of this protein in cell survival or transwell resistance. Hyperoxia increased fragmentation of PEC mitochondria in a time-dependent manner through 48 hours of exposure. Hyperoxic PECs exhibited increased phosphorylation of Drp-1 (serine 616), decreases in Mfn1 (mitofusion protein 1), but increases in OPA-1 (optic atrophy 1). Pro-autophagy proteins p62 (LC3 adapter–binding protein SQSTM1/p62), PINK-1 (PTEN-induced putative kinase 1), and LC3B (microtubule-associated protein 1A/1B-light chain 3) were increased. Returning cells to normoxia for 24 hours reversed the increased mt-fragmentation and changes in expression of pro-fission proteins. Hyperoxia-induced changes in mitochondrial structure or cell survival were mitigated by antioxidants mito-TEMPOL, Drp-1 silencing, or inhibition or protection by the mitochondrial endonuclease ENDO III. Hyperoxia induced oxidation and mitochondrial depolarization and impaired transwell resistance. Decrease in resistance was mitigated by mito-TEMPOL or ENDO III and reproduced by overexpression of Drp-1. Conclusions—Because hyperoxia evoked mt-fragmentation, cell survival and transwell resistance are prevented by ENDO III and mito-TEMPOL and Drp-1 silencing, and these data link hyperoxia-induced mt-DNA damage, Drp-1 expression, mt-fragmentation, and PEC dysfunction

670 nm light mitigates oxygen-induced degeneration in C57BL/6J mouse retina

BACKGROUND Irradiation with light wavelengths from the far red (FR) to the near infrared (NIR) spectrum (600 nm -1000 nm) has been shown to have beneficial effects in several disease models. In this study, we aim to examine whether 670 nm red light pretreatment can provide protection against hyperoxia-induced damage in the C57BL/6J mouse retina. Adult mice (90-110 days) were pretreated with 9 J/cm2 of 670 nm light once daily for 5 consecutive days prior to being placed in hyperoxic environment (75% oxygen). Control groups were exposed to hyperoxia, but received no 670 nm light pretreatment. Retinas were collected after 0, 3, 7, 10 or 14 days of hyperoxia exposure (n = 12/group) and prepared either for histological analysis, or RNA extraction and quantitative polymerase chain reaction (qPCR). Photoreceptor damage and loss were quantified by counting photoreceptors undergoing cell death and measuring photoreceptor layer thickness. Localization of acrolein, and cytochrome c oxidase subunit Va (Cox Va) were identified through immunohistochemistry. Expression of heme oxygenase-1 (Hmox-1), complement component 3 (C3) and fibroblast growth factor 2 (Fgf-2) genes were quantified using qPCR. RESULTS The hyperoxia-induced photoreceptor loss was accompanied by reduction of metabolic marker, Cox Va, and increased expression of oxidative stress indicator, acrolein and Hmox-1. Pretreatment with 670 nm red light reduced expression of markers of oxidative stress and C3, and slowed, but did not prevent, photoreceptor loss over the time course of hyperoxia exposure. CONCLUSION The damaging effects of hyperoxia on photoreceptors were ameliorated following pretreatment with 670 nm light in hyperoxic mouse retinas. These results suggest that pretreatment with 670 nm light may provide stability to photoreceptors in conditions of oxidative stress.This work was supported by the Australian Research Council Centre of Excellence in Vision Science

Protection by Inhaled Hydrogen Therapy in a Rat Model of Acute Lung Injury can be Tracked \u3cem\u3ein vivo\u3c/em\u3e Using Molecular Imaging

Inhaled hydrogen gas (H2) provides protection in rat models of human acute lung injury (ALI). We previously reported that biomarker imaging can detect oxidative stress and endothelial cell death in vivo in a rat model of ALI. Our objective was to evaluate the ability of 99mTc-hexamethylpropyleneamineoxime (HMPAO) and 99mTc-duramycin to track the effectiveness of H2 therapy in vivo in the hyperoxia rat model of ALI. Rats were exposed to room air (normoxia), 98% O2 + 2% N2 (hyperoxia) or 98% O2 + 2% H2 (hyperoxia+H2) for up to 60 h. In vivo scintigraphy images were acquired following injection of 99mTc-HMPAO or 99mTc-duramycin. For hyperoxiarats, 99mTc-HMPAO and 99mTc-duramycin lung uptake increased in a time-dependent manner, reaching a maximum increase of 270% and 150% at 60 h, respectively. These increases were reduced to 120% and 70%, respectively, in hyperoxia+H2 rats. Hyperoxia exposure increased glutathione content in lung homogenate (36%) more than hyperoxia+H2 (21%), consistent with increases measured in 99mTc-HMPAO lung uptake. In 60-h hyperoxia rats, pleural effusion, which was undetectable in normoxia rats, averaged 9.3 gram/rat, and lung tissue 3-nitrotyrosine expression increased by 790%. Increases were reduced by 69% and 59%, respectively, in 60-h hyperoxia+H2 rats. This study detects and tracks the anti-oxidant and anti-apoptotic properties of H2 therapy in vivo after as early as 24 h of hyperoxia exposure. The results suggest the potential utility of these SPECT biomarkers for in vivo assessment of key cellular pathways in the pathogenesis of ALI and for monitoring responses to therapies

MR imaging–derived oxygen-hemoglobin dissociation curves and fetal-placental oxygen-hemoglobin affinities

PURPOSE: To generate magnetic resonance (MR) imaging–derived, oxygen-hemoglobin dissociation curves and to map fetal-placental oxygen-hemoglobin affinity in pregnant mice noninvasively by combining blood oxygen level–dependent (BOLD) T2* and oxygen-weighted T1 contrast mechanisms under different respiration challenges. MATERIALS AND METHODS: All procedures were approved by the Weizmann Institutional Animal Care and Use Committee. Pregnant mice were analyzed with MR imaging at 9.4 T on embryonic days 14.5 (eight dams and 58 fetuses; imprinting control region ICR strain) and 17.5 (21 dams and 158 fetuses) under respiration challenges ranging from hyperoxia to hypoxia (10 levels of oxygenation, 100%–10%; total imaging time, 100 minutes). A shorter protocol with normoxia to hyperoxia was also performed (five levels of oxygenation, 20%–100%; total imaging time, 60 minutes). Fast spin-echo anatomic images were obtained, followed by sequential acquisition of three-dimensional gradient-echo T2*- and T1-weighted images. Automated registration was applied to align regions of interest of the entire placenta, fetal liver, and maternal liver. Results were compared by using a two-tailed unpaired Student t test. R1 and R2* values were derived for each tissue. MR imaging–based oxygen-hemoglobin dissociation curves were constructed by nonlinear least square fitting of 1 minus the change in R2*divided by R2*at baseline as a function of R1 to a sigmoid-shaped curve. The apparent P50 (oxygen tension at which hemoglobin is 50% saturated) value was derived from the curves, calculated as the R1 scaled value (x) at which the change in R2* divided by R2*at baseline scaled (y) equals 0.5. RESULTS: The apparent P50 values were significantly lower in fetal liver than in maternal liver for both gestation stages (day 14.5: 21% ± 5 [P = .04] and day 17.5: 41% ± 7 [P < .0001]). The placenta showed a reduction of 18% ± 4 in mean apparent P50 values from day 14.5 to day 17.5 (P = .003). Reproduction of the MR imaging–based oxygen-hemoglobin dissociation curves with a shorter protocol that excluded the hypoxic periods was demonstrated. CONCLUSION: MR imaging–based oxygen-hemoglobin dissociation curves and oxygen-hemoglobin affinity information were derived for pregnant mice by using 9.4-T MR imaging, which suggests a potential to overcome the need for direct sampling of fetal or maternal blood. Online supplemental material is available for this article

The effect of experimental hyperoxia on erythrocytes’ oxygen-transport function

The aim of this study was to investigate the effect of hyperoxia, calcium ions and pH value on the composition of major phospholipids in human erythrocyte membranes and erythrocytes’ oxygen-transport function. To create a model of hyperoxia, we saturated the incubated mixture with oxygen by constant passing of oxygen–air mixture through the incubation medium. To assess the effect of elevated calcium ion concentrations, CaCl2 was added to the incubation medium. An incubation medium with different pH was used to study the effect of various pH values. Lipids were extracted from erythrocytes and chromatographic separation was carried out in a thin layer of silica gel deposited on a glass plate. The thiobarbituric acid (TBA)-active products and the content of diene conjugates (DC) in erythrocytes were determined. The oxygen-binding capacity of haemoglobin was evaluated using Raman spectroscopy. The obtained results indicated that hyperoxia causes deep changes both in the composition and character of bilayer lipids of erythrocyte membranes, which affects the functional characteristics of erythrocytes, primarily the oxygen-transport properties of erythrocyte haemoglobin. It should be noted that a combination of Ca2+ ions and change in the pH value intensify the processes associated with disruption of phospholipids’ composition. The findings indicate that the lipid phase is one of the key elements in the functioning of erythrocytes in norm as well as during development of various pathological processes

Hooray for Hypoxia?

The mortality of critically ill patients rises steadily as the partial pressure of arterial oxygen falls below about 11 kPa (80 mm Hg). A new animal study in the May 2005 issue of PLoS Biology showing a potential benefit for hypoxia is thus a challenge to current thinking

More oxygen during development enhanced flight performance but not thermal tolerance of Drosophila melanogaster

High temperatures can stress animals by raising the oxygen demand above the oxygen supply. Consequently, animals under hypoxia could be more sensitive to heating than those exposed to normoxia. Although support for this model has been limited to aquatic animals, oxygen supply might limit the heat tolerance of terrestrial animals during energetically demanding activities. We evaluated this model by studying the flight performance and heat tolerance of flies (Drosophila melanogaster) acclimated and tested at different concentrations of oxygen (12%, 21%, and 31%). We expected that flies raised at hypoxia would develop into adults that were more likely to fly under hypoxia than would flies raised at normoxia or hyperoxia. We also expected flies to benefit from greater oxygen supply during testing. These effects should have been most pronounced at high temperatures, which impair locomotor performance. Contrary to our expectations, we found little evidence that flies raised at hypoxia flew better when tested at hypoxia or tolerated extreme heat better than did flies raised at normoxia or hyperoxia. Instead, flies raised at higher oxygen levels performed better at all body temperatures and oxygen concentrations. Moreover, oxygen supply during testing had the greatest effect on flight performance at low temperature, rather than high temperature. Our results poorly support the hypothesis that oxygen supply limits performance at high temperatures, but do support the idea that hyperoxia during development improves performance of flies later in life

Influence of 100% and 40% oxygen on penumbral blood flow, oxygen level, and T2*-weighted MRI in a rat stroke model

Accurate imaging of the ischemic penumbra is a prerequisite for acute clinical stroke research. T2* magnetic resonance imaging (MRI) combined with an oxygen challenge (OC) is being developed to detect penumbra based on changes in blood deoxyhemoglobin. However, inducing OC with 100% O2 induces sinus artefacts on human scans and influences cerebral blood flow (CBF), which can affect T2* signal. Therefore, we investigated replacing 100% O2 OC with 40% O2 OC (5 minutes 40% O2 versus 100% O2) and determined the effects on blood pressure (BP), CBF, tissue pO2, and T2* signal change in presumed penumbra in a rat stroke model. Probes implanted into penumbra and contralateral cortex simultaneously recorded pO2 and CBF during 40% O2 (n=6) or 100% O2 (n=8) OC. In a separate MRI study, T2* signal change to 40% O2 (n=6) and 100% O2 (n=5) OC was compared. Oxygen challenge (40% and 100% O2) increased BP by 8.2% and 18.1%, penumbra CBF by 5% and 15%, and penumbra pO2 levels by 80% and 144%, respectively. T2* signal significantly increased by 4.56%±1.61% and 8.65%±3.66% in penumbra compared with 2.98%±1.56% and 2.79%±0.66% in contralateral cortex and 1.09%±0.82% and −0.32%±0.67% in ischemic core, respectively. For diagnostic imaging, 40% O2 OC could provide sufficient T2* signal change to detect penumbra with limited influence in BP and CBF

Neural regulation of cardiovascular response to exercise: role of central command and peripheral afferents

During dynamic exercise, mechanisms controlling the cardiovascular apparatus operate to provide adequate oxygen to fulfill metabolic demand of exercising muscles and to guarantee metabolic end-products washout. Moreover, arterial blood pressure is regulated to maintain adequate perfusion of the vital organs without excessive pressure variations. The autonomic nervous system adjustments are characterized by a parasympathetic withdrawal and a sympathetic activation. In this review, we briefly summarize neural reflexes operating during dynamic exercise. The main focus of the present review will be on the central command, the arterial baroreflex and chemoreflex, and the exercise pressure reflex. The regulation and integration of these reflexes operating during dynamic exercise and their possible role in the pathophysiology of some cardiovascular diseases are also discusse