Marine deoxygenated areas are expanding, and more hypoxic zones emerge globally. Climate change induced warming and stratification can extrapolate the biological oxygen demand, more predominantly at coastal areas and reefs with dense vegetation and high metabolic activity. The diurnal oxygen fluctuation can reach a critically low oxygen level at nighttime, exposing aquatic organisms to severe hypoxia that could interfere with viable ecosystem functions. Little is known about the impact of hypoxia on marine primary producers macroalgae, mainly the physiological adaptation of the Red Sea macroalgae under climate change scenarios is understudied. Here we investigate hypoxia thresholds at night time for conspicuous Red Sea macroalgae species calcareous Halimeda opuntia and Padina boryana and noncalcifying brown algae Sargassum latifolium. We utilized a computer-based gas system to expose the samples to different oxygen treatments (normoxia, hypoxia, and severe hypoxia) that mimic in situ water chemistry at 32 °C. We monitored algal physiological response during 12 hours of exposure to different oxygen levels in the dark by measuring photochemical efficiency, respiration rates, and cellular viability. For the duration of our experiments (12h), we did not detect lethal thresholds. In all tested species, severe hypoxia significantly decreased photochemical efficiency, and hypoxia had a limited impact on photochemical efficiency. However, both low oxygen treatments significantly decreased respiration rates and induced changes in cellular activity. We concluded a sublethal O$_2$ thresholds SLC$_{(50)}$ of 1.2 ±0.1, 1.5 ±0.1, and 1.7 ±0.1 mg O$_2$ L$^{−1}$ ±SD for H. opuntia, P. boryana, and S. latifolium responses, respectively. We also found that during 12 hours of treatments, the median time to observe a 50% reduction in photochemical efficiency under severe hypoxia treatment relative to control was 6.3 ±1.4, 3.5 ±1.0, and 0.8 ±1.3 hours ±SD for H. opuntia, P. boryana, and S. latifolium responses, respectively. This study is the first assessment of Red Sea macroalgae response to hypoxia and the first dark nighttime algal adaptation of its kind for our proposed species. Further investigation is needed to assess daytime recovery, recurring dark hypoxia, and synergic or sequential effects of other environmental stressors on hypoxia thresholds.

The role of lipids and ROS in cardioprotective mechanism of chronic hypoxia Cardiovascular diseases, mainly ischemic heart disease is one of the most frequently cause of morbidity and mortality in developed countries. Therefore effective protection of the heart against ischemia and reperfusion injury is the crucial aim of experimental and clinical cardiology. One of the main streams of cardiovascular research is looking for possibilities of natural heart resistance augmentation. Adaptation to chronic hypoxia is one possibility how to protect the heart against ischemia-reperfusion injury. Chronic hypoxia increases resistance of the myocardium to acute deficiency of oxygen leading to vetricular arrythmias, postischemic contractile dysfunction and necrotic changes in the tissue. Recently, it has been shown that reactive oxygen species (ROS) play an important role in the cardioprotective mechanism of chronic hypoxia. It is known that oxidative stress has a harmful effect in acute ischemia-reperfusion however ROS generated during the adaptation to hypobaric intermittent chronic hypoxia play a role in the induction of cardioprotection. In this study, we demonstrated that adaptation of adult rats to chronic hypoxia increased the activity and protein abundance of manganese superoxide dismutase (MnSOD) in the...

Hypoxic pulmonary vasoconstriction (HPV) is a local physiological mechanism in lungs that optimalises blood oxygenation during alveolar hypoxia. Arterioles in the affected region increase flow resistance which redirects blood to better ventilated parts of the lung. During global hypoxia - e.g. in high altitude or in chronic pulmonary illnessess - this mechanism doesn't work, as the blood cannot be redirected elsewhere. The pressure in pulmonary artery rises which leads to right heart hypertrophy and ultimately to cor pulmonale. This mechanism has been studied for decades, but specific signalling pathways still lack full description and therapeutical solutions are not available. This thesis offers description of selected properties of pulmonary circulation and patophysiological context of pulmonary hypertension, introduces the reader to HPV localization and signalization, and discusses its most important steps from decreased oxygen availability to vessel constriction. The practical part of this work explores Succinate dehydrogenase (SDH) - complex coupling Kreb's cycle to electron transport chain - as a primary detection site of hypoxia in pulmonary artery smooth muscle cells. We decided to test this hypothesis in isolated rat lungs by measuring if malonate (SDH inhibitor) causes vasoconstriction as...

Around 70% of the world is covered with water but only 2.5% of it is freshwater and even less is available for the ecosystem and humanity. The limited available fresh water is facing increasing challenges from water pollutions and eutrophication is one of the major concerns worldwide. The reason of eutrophication is the presence of excessive amounts of phosphorus and nitrogen in water bodies, which may cause algal blooms and a variety of harms to aquatic ecosystem in association with algal blooms. Among these two components, phosphorus plays a major role in eutrophication control and recovery since atmospheric N2 can be fixed by biological nitrogen fixation (BNF) processes and is therefore of little meaning to control. In this study, we investigated for the first time the potential of using controlled growth of algae and, in particular, filamentous cyanobacterium Spirulina platensis, for eutrophic water restoration. This study investigated the algal cell growth, algal by-product production, and removal of phosphate by S. platensis at different phosphate levels in artificial wastewaters and eutrophic waters. Results indicate that S. platensis could remove 90.17% of phosphorus from artificial wastewaters containing 10 mg/L phosphate in a 16-day cultivation period. When tested for eutrophic water restoration, S. platensis was able to convert hypo-eutrophic, eutrophic, and meso-eutrophic waters to oligotrophic water. It was shown that by using 100- micron nylon mesh cloth we could keep biomass concentration to be lower than 0.30±0.02 g/L. In the meantime, light/dark tests indicate that the dissolved oxygen level would not go below the hypoxic level, i.e., 4 mg/L after a 12-hour dark period at biomass concentration up to 1 g/L. These results indicate that it is possible to use S. platensis for both control of point source discharge and eutrophic water restoration.

Coral reef ecosystems are under increasing threat from ocean warming and deoxygenation. Mass coral bleaching events in recent years have been linked to marine heatwaves but reporting of hypoxia-induced bleaching has also been increasing. Oxygen availability in coral reefs is driven by community metabolism and they experience a dynamic range of oxygen concentrations throughout diel cycles, hyperoxia during the day and hypoxia during the night. It has been suggested that the highest oxygen concentrations coincide with the hottest part of the day and this may protect marine taxa from high temperatures. We evaluated experimentally whether excess oxygen availability would increase the thermal threshold of the branching coral Stylophora pistillata, from the Southern Red Sea. We did this by exposing coral fragments of this species to varying dissolved oxygen concentrations (hypoxia, normoxia and hyperoxia) and a short-term temperature ramping regime (1˚C h-1). Hyperoxia did extend the thermal tolerance of S. pistillata fragments, with an LT50 of 39.1˚C as opposed to 39.0˚C for the normoxic treatment and 38.7˚C for the hypoxic treatment. Hyperoxia also increased respiration and gross photosynthesis and had a negative effect on photochemical efficiency at high temperatures. Net photosynthesis, P:R ratio and symbiont density were not significantly affected by oxygen concentration. Corals in this experiment displayed exceedingly high thermal thresholds, which were at least 2˚C higher than previously reported for the same species in the Central Red Sea. The corals used in the experiment had previously survived mass bleaching events in 2015 and hence we may have selected for individuals adapted to thermal stress. This is the first study to investigate the role of oxygen in the thermal tolerance of hermatypic corals and the first assessment of thermal thresholds from corals in the Southern Red Sea, where previously thermal thresholds have been based on a 1-2˚C increase in maximum mean monthly temperatures and visual bleaching observations. This highlights the need for increased experimental assessments of thermal thresholds in the Southern regions of the Red Sea and the important role of oxygen in moderating thermal stress.

The mitochondria are organelles where energy in the form of ATP (Adenosine Triphosphate) is produced. During low oxygen supply (hypoxia) and mitochondrial uncoupling, ATP synthesis is reduced and AMP (Adenosine Monophosphate) accumulates in the cells. This increase in AMP: ATP ratio stimulates the AMP-Activated Kinase (AMPK) pathway, known to improve healthspan and lifespan by increasing mitochondrial biogenesis (making new mitochondria), decreasing oxidative stress and inflammation, and proteotoxicity (by degrading non-functional organelles and proteins). Here, the life/healthspan extending potential of chronic mild intermittent hypoxia (CMIH) and mitochondrial uncoupling using 2,4 -Dinitrophenol (DNP) was investigated in an emerging model organism, an aquatic crustacean, Daphnia magna. First, the effect of CMIH (4mgO2/L) on longevity in four different genotypes of Daphnia magna was investigated. All individuals were kept in similar conditions with controls in normoxia (8mgO2/L). Hypoxia was created by bubbling compressed nitrogen gas through the water twice daily. Survival was assessing through censuses conducted every 3 days and gene expression changes in response to CMIH were assessed by RNA sequencing using Oxford Nanopore Technology. Briefly, RNA was isolated from genotypes after hypoxic treatments and reverse transcribed to cDNA, libraries were multiplexed and sequenced using Oxford Nanopore MinION for 24-48 hours. Lastly, the effect of prolonged exposure to DNP on longevity was evaluated. Daphnia were chronically exposed to either 0 (control), or 0.1, 1, and 5μM of DNP. Genotypes displayed different tolerance to hypoxia and DNP treatments. Contrary to the expectations, CMIH and DNP reduced longevity, but only in genotypes from permanent ponds, while having no effect on the survival of genotypes from intermittent ponds, arguably better adapted to naturally occurring hypoxic conditions. We uncovered 11 candidate genes that were differentially expressed in these genotypes. In particular, genes involved in mTOR, p53, and sirtuin pathways showed patterns of expression consistent with protection against hypoxia. These pathways are known to regulate autophagy, apoptosis, inflammation, and cell cycle. Because our findings elucidate genotype-specific physiological and transcriptomic responses to respiratory perturbations (CMIH and DNP) we may be able to make a step towards the understanding of a model organism’s response to respiratory stress.

L’hypoxie est l’une des principales causes de résistance aux traitements dans les glioblastomes. Des stratégies permettant de lever l’hypoxie ou de limiter ses effets sont de ce fait nécessaires. Ces travaux de thèse s’intéressent à deux stratégies de vectorisation ciblée agissant contre l’hypoxie. La première stratégie vise à lutter contre les effets de l’hypoxie par une approche combinée de vectorisation cellulaire et moléculaire ciblant une protéine à action pro-tumorale : l’érythropoïétine. Des macrophages ont ainsi été modifiés génétiquement pour leur permettre de surexprimer une forme tronquée de récepteur à l’érythropoïétine conduisant à un ralentissement de la croissance d’un modèle de glioblastome. Les constructions moléculaires ont ensuite été modifiées pour rendre la surexpression inductible par l’hypoxie. La deuxième stratégie vise à réoxygéner spécifiquement la tumeur en se basant sur des nanozéolithes vectrices de gaz hyperoxiques. Les résultats montrent que ces nanoparticules microporeuses ne présentent pas d’effets toxiques majeurs in vitro et in vivo. L’incorporation de fer dans les zéolithes augmente significativement la capacité de transport d’O2 et le gadolinium permet leur utilisation comme agent de contraste en imagerie par résonance magnétique. D’autre part, les nanozéolithes vectrices de CO2/O2 s’accumulent spécifiquement dans le tissu tumoral et augmentent localement le volume sanguin et la quantité d’oxygène. Ainsi, les deux approches développées au cours de cette thèse démontrent le potentiel des stratégies ciblées dirigées contre l’hypoxie dans les glioblastomes. / Hypoxia is one of the main causes of resistance to treatments in glioblastoma, the worst primary brain tumor in term of survival. Two targeted vectorization strategies directed against hypoxia are presented in this thesis work. The first strategy was designed to inhibit hypoxia-induced erythropoietin through the use of macrophages. Macrophages were genetically engineered to overexpress a truncated form of the erythropoietin receptor resulting in a decrease in the tumor volume in a hypoxic model of glioblastoma in vivo. Hypoxia-inducible constructs were then developed. The second strategy aimed to use nanozeolites carrying hyperoxic gases as a tool to specifically reoxygenate the tumor. Results show that these microporous nanoparticles have no adverse effects in vitro and in vivo. The incorporation of iron in the zeolites significantly increases their oxygen transport capacity and the gadolinium allows their use as a contrast agent for magnetic resonance imaging. In addition, nanozeolites carrying CO2/O2 accumulate specifically in the tumor tissue and locally increase the blood volume/oxygenation. Thus, the two strategies developed during this thesis demonstrate the potential to fight against hypoxia specifically in glioblastoma.

As selection pressures exist for not only adults, but for every life history stage, it is important to understand how environmental factors shape developing animals. Despite the significance placed on aquatic hypoxia as a driving force in the evolution of air breathing, this is the first known study to examine the effects of hypoxia on cardio-respiratory ontogeny of an air breathing fish. Blue gouramis are obligatory air breathing fish that possess a labyrinth-like structure that serves as the air breathing organ. Gouramis were reared for up to 90 d in normoxia or hypoxia, and morphological and physiological development was observed. Hypoxic larvae had increased lamellar and labyrinth organ surface areas. Bradycardia and increased gill ventilation rates were observed when larvae from either rearing group were briefly exposed to hypoxia. Hypoxic larvae also showed a reduced heart rate and gill ventilation rate in the absence of a hypoxic stimulus, possibly indicative of a more comprehensive, long-term respiratory plasticity. The similarity of routine oxygen consumption between rearing groups suggests that metabolic demand did not change for hypoxic larvae, but that they were more efficient at oxygen acquisition. This is further supported by increased resistance time of hypoxic gouramis to extreme hypoxia. The onset of air breathing was between 20 and 25 d post-fertilization, and was not affected by either rearing or exposure environment. It may be that this behavior is associated with the inability of smaller larvae to successfully overcome water surface tension, rather than with the necessity of aerial respiration at this stage. Hypoxia is commonly experienced by most air breathing fishes, and studies of hypoxia-induced developmental effects may provide critical insights into the evolution of air breathing. The studies presented here provide novel data on the plasticity of cardio-respiratory development of an air breathing fish reared in hypoxia, and can serve as a solid foundation for future studies.

Cardiovascular intervention studies are a very important issue given that the ischaemic heart disease is one of the main mortality and morbidity causes in the Western world. Cardioprotection is mediated through a variety of signalling pathways in the cell that may directly or indirectly affect energy metabolism and mitochondria. Ischaemia-reperfusion injury of the heart significantly affect mitochondrial function revealing a potential therapeutic target. The role of mitochondria in the myocardium is not only in the field of energy homeostasis, but also in mediating the cellular response to reduced oxygen supply and in apoptosis regulation. This thesis aims to elucidate the response of the hypertrophied heart of the spontaneously hypertensive rat (SHR) and the derived conplastic strain with mitochondrial genome of normotensive Brown Norway (SHR-mtBN ) to the cardioprotective regime of adaptation to chronic normobaric hypoxia (CNH, Fi 0.1). The adaptive changes were studied at the cellular, protein and gene levels using Real-time RT-PCR, Biomark Chip Analysis, Western Blot, spectrophotometric measurements of enzyme activity and quantitative immunofluorescence analyses. The present thesis was based on a different cardioprotective phenotype between SHR and SHR-mtBN strains, i.e. a significantly smaller...

Hypoxia has become a subject of interest among the many environmental stressors as its role in biology is complex and diverse. Hypoxia is a significant low oxygen condition that causes many pathologies and adaptive responses in organisms. It can lead to a moderate or dangerous loss of respiration and can be an indication of tumorigenesis as many tumors lack adequate blood supply. Organisms possess adaptive responses to hypoxia that include hypoxia-inducible factors (HIFs) that activate several downstream pathways that are responsible for altering metabolism and maintaining homeostasis. Within aquatic organisms, hypoxia is an important ecological constraint as oxygen availability within bodies of water can vary greatly over time and space. Therefore, adaptation to hypoxia is likely pervasive, especially in genotypes originating from bodies of water that are prone to hypoxia. Here we report the transcriptional response to acute hypoxia in the clonal freshwater crustacean Daphnia magna. Daphnia were subjected to 1mg/O2 for 12 hours. Then, RNA was extracted, reverse transcribed, and sequenced using Oxford Nanopore MinION. We find that severe hypoxia significantly up-regulates key enzymes in the gluconeogenesis pathway. Additionally, we report genotype-by-environment interactions showing that Daphnia clones from habitats that are hypoxia prone survive better in hypoxia.