Levels Of Hypoxia Research Articles

Effect of exercise during pregnancy on offspring development through ameliorating high glucose and hypoxia in gestational diabetes mellitus

BACKGROUND Gestational diabetes mellitus (GDM) women require prenatal care to minimize short- and long-term complications. The mechanism by which exercise during pregnancy affects organ development and whether glucose transporter (GLUT) 1 plays a role in GDM offspring organ development remains unknown. AIM To determine the effect of exercise during pregnancy on the cardiac, hepatic and renal development of GDM mother’s offspring. METHODS Placenta samples were collected from humans and mice. GDM mouse models were created using streptozotocin along with a GDM with exercise group. The hearts, livers and kidneys of 3- and 8-week-old offspring were collected for body composition analysis and staining. The effects of high glucose levels and hypoxia were investigated using HTR8/SVneo. Transwell and wound-healing assays were performed to assess cell migration. Immunofluorescence accompanied with TUNEL and Ki67 staining was used to explore apoptosis and proliferation. RESULTS Exercise during pregnancy downregulated the GLUT1 and hypoxia inducible factor-1α expression in placenta from individuals with GDM. Cobalt chloride-induced hypoxia and high glucose levels also significantly decreased migration and apoptosis of HTR8/SVneo cells. In addition, exercise reduced inflammatory cell infiltration in the liver and decreased the tubular vacuolar area in the kidneys of offspring. CONCLUSION GDM affects the growth and development of organs in offspring. Exercise during pregnancy can reverse adverse effects of GDM on the development of the heart, liver, and kidney in offspring.

Integrated analysis of single-cell and bulk transcriptome reveals hypoxia-induced immunosuppressive microenvironment to predict immunotherapy response in high-grade serous ovarian cancer

BackgroundHypoxia is significantly associated with cancer progression and treatment outcomes. Nevertheless, the precise molecular mechanisms underlying the hypoxia-induced immunosuppressive microenvironment in high-grade serous ovarian cancer (HGSOC) are still not fully understood.MethodsBy analyzing five independent transcriptomic datasets, we investigated the effect of hypoxia on prognosis and tumor microenvironment (TME) in HGSOC. The hypoxia levels and the intercellular communication signaling pathways were studied by using single-cell analysis. Furthermore, the Hypoxia-TME classifier was developed and then validated in the multiple HGSOC datasets. In addition, we also investigated the prognostic significance, genetic variations, signaling pathways, and the potential for immunotherapy benefits in different Hypoxia-TME subgroups.ResultsHypoxia was identified as a crucial risk factor in HGSOC, and strongly correlated with an immunosuppressive microenvironment characterized by alterations in the composition and distribution of immune cells. Single-cell analysis elucidated the heterogeneity inherent within the TME in HGSOC, and demonstrated an association between the hypoxic TME and fibroblasts as well as macrophages. CellChat analysis identified SPP1-CD44 and CXCL12-CXCR4 as the principal signaling axes through which macrophages and fibroblasts interact with T cells, respectively. Moreover, a personalized Hypoxia-TME classifier was constructed and validated through the integration of the hypoxia (18 genes) and TME (7 immune cells) scores. It was observed that patients in the Hypoxialow/TMEhigh subgroup displayed a significantly better prognosis than other subgroups. Different subgroups exhibited unique genomic alterations and variations in signaling pathway differences, including TGF-β and Wnt/β-catenin pathways, which are closely associated with various biological functions. Finally, our results indicated that patients in the Hypoxialow/TMEhigh subgroup exhibit a better response to immunotherapy, suggesting the potential utility of the Hypoxia-TME classifier as a new biomarker in HGSOC.ConclusionOur study revealed hypoxia-induced immunosuppressive microenvironment, and developed Hypoxia-TME classifier to distinguish the prognosis, immune characteristics, and potential benefits of immunotherapy in HGSOC.

K2FeO4-Enhanced Photodynamic Therapy of Breast Cancer via In Situ Synthesis of Fe2O3 and O2.

Photodynamic Therapy (PDT) offers a promising minimally invasive treatment for breast cancer, but its efficacy is limited by the hostile tumor microenvironment (TME), including hypoxia and high glutathione (GSH) levels. Although various strategies to improve oxygen concentration or reduce reactive oxygen species (ROS) resistance for enhanced PDT have been explored, they typically require intricate design and complex synthesis of multifunctional nanocarriers. Thus, this study introduces a facile K2FeO4-induced strategy to enhance PDT efficiency in breast cancer through the tumor in situ synthesis of Fe2O3 and O2. Inspired by the successful application of K2FeO4 in ecological remediation and hemostasis, K2FeO4 reacts with GSH, biological system, H2O2, and water, to generate Fe2O3 and O2. Intratumoral injection of K2FeO4 improves the TME, followed by Ce6 administration to enhance PDT through synergistic ferroptosis. This approach boosts PDT efficacy significantly by increasing ROS generation, lipid peroxidation, and inhibiting GSH and GPX4. Proteomic analysis revealed alterations in key pathways, including endocytosis and energy metabolism. This K2FeO4-PDT strategy creates a positive feedback loop by enhancing oxidative stress, providing an interesting and promising approach to PDT.

In vitro/In vivo oxygen electrochemical nanosensor for bioanalysis

Direct in vivo monitoring of O2 is critical in the study of numerous biological processes, and the development of novel highly sensitive techniques for O2in vivo detection is of great potential importance. Electrochemical amperometric sensors are some of the most promising, easy to operate, inexpensive, sensitive and have a high temporal resolution. As part of this work, we have demonstrated a direct rapid method for molecular oxygen detection inside multicellular spheroids in vitro and rat brain in vivo. External and internal oxygen profiles in human adenocarcinoma MCF-7 spheroids were studied using developed nanoelectrode. The size of spheroids was shown to affect the level of hypoxia inside them, and also affected the oxygen consumption near spheroids in solution. To demonstrate the in vivo relevance of the novel sensor we used it to measure the oxygen concentrations in the superficial layers of the rat brain. This study demonstrates a minimally invasive electrochemical method for real-time oxygen profiling in vivo.

Endometriosis development in relation to hypoxia: a murine model study.

Endometriosis, due to its ambiguous symptoms, still remains one of the most difficult female diseases to treat, with an average diagnosis time of 7-9years. The changing level of hypoxia plays an important role in a healthy endometrium during menstruation and an elevated expression of the hypoxia-inducible factor 1-alpha (HIF-1α) has been demonstrated in ectopic endometria. HIF-1α mediates the induction of proangiogenic factors and the development of angiogenesis is a critical step in the establishment and pathogenesis of endometriosis. Although the inhibition of angiogenesis has been proposed as one of the actionable therapeutic modalities, vascular normalization and re-oxygenation may become a possible new approach for therapeutic intervention. Our goal was to investigate whether a selected murine model of endometriosis would be suitable for future studies on new methods for treating endometriosis. Non-invasive, high-resolution ultrasound-monitored observation was selected as the preclinical approach to obtain imaging of the presence and volume of the endometriotic-like lesions. The EF5 (2-(2-Nitro-1H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide) compound that selectively binds to reduced proteins in hypoxic cells was used for hypoxia detection. The expression of Pten and other crucial genes linking endometriosis and hypoxia were also assessed. Using EF5, a pentafluorinated derivative of the 2-nitroimidazole that is metabolically reduced by oxygen-inhibitable nitroreductase, we confirmed that hypoxia did develop in the selected model and was detected in uterine and ectopic endometriotic lesions. Moreover, the changes in oxygen tension also influenced the expression level of significant genes related to endometriosis, like Pten, Trp53, Hif1a, Epas1, and Vegfa. Their strong modulation evidenced here is indicative of model reliability. Using high-resolution ultrasound-based imaging, we present a non-invasive method of visualization that enables the detection and observation of lesion evolution throughout the duration of the experiment, which is fundamental for further preclinical studies and treatment evaluation. The selected model and method of visualization appear to be suitable for the study of new treatment strategies based on hypoxia alleviation and blood flow restoration.

A Case for Co-oximetry: Methemoglobinemia secondary to a Rare Hemoglobin Variant

Abstract Introduction/Objective The presence of dyshemoglobins and some inherited hemoglobin (Hb) variants can cause over or underestimation of oxygen saturation (O2sat) when using a pulse oximeter (ox). These noninvasive medical devices rely on two wavelengths, a pulsing arterial bed, and light absorption properties of Hb to measure the difference between deoxy- and oxy-Hb. In contrast, a co-oximeter (co-ox) requires arterial blood. By using at least four wavelengths, co-ox measures the functional and fractional O2sat to detect all forms of Hb, including those that do not bind oxygen (O2), such as methemoglobin (metHb). Methemoglobinemia may be congenital or acquired, resulting from exposure to certain toxins, metabolic disorders, enzymatic deficiencies such as cytochrome b5 reductase deficiency, or hemoglobinopathies. Methods/Case Report A healthy, adolescent male undergoing a sports physical at his pediatrician’s office was hypoxic by pulse ox and sent to the emergency department (ED) for evaluation. On exam, he had cold, blue tinged fingertips and lips but was otherwise asymptomatic. His O2sat was 88-90% on room air with minimal improvement with 6L of supplemental O2. Co-ox revealed a metHb level > 20%. The father also mentioned that the mother had frequent ED visits for hypoxia. Capillary Hb electrophoresis showed peaks in the HbA, HbF, and HbA2 zones of 78.3%, 17.9%, and 3.6%, respectively. All CBC parameters were within the reference intervals. The large peak in the HbF zone would suggest hereditary persistence of fetal hemoglobin (HPFH); however, the clinical picture was not consistent. Further reference laboratory investigation determined that the patient was heterozygous for Hb Tübingen (HbTüb), a beta globin chain variant resulting from a point mutation at position 106. By mass spectrometry, the variant quantified as 38% of the total hemoglobin, indicating underestimation by electrophoretic techniques. Results (if a Case Study enter NA) NA Conclusion This case presents an essentially asymptomatic patient with hypoxia and elevated methemoglobin levels. Electrophoretic hemoglobinopathy and CBC workup were clinically inconsistent with HPFH and the presence of an aberrant hemoglobin migrating in the HbF must be considered in the differential diagnosis. The manifestation of HbTüb as methemoglobinemia in this context adds complexity to the diagnostic puzzle. Further laboratory exploration into the underlying mechanisms driving this unique presentation is crucial for a comprehensive understanding and tailored management approach.

Suppressing the expression of steroidogenic acute regulatory protein (StAR) in the myocardium by spironolactone contributes to the improvement of right ventricular remodeling in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a progressive condition that frequently leads to right ventricular (RV) remodeling. Aldosterone promotes vascular and RV remodeling. The upregulation of steroidogenic acute regulatory protein (StAR) stimulates aldosterone synthesis. However, the expression of StAR in the myocardium under PAH conditions remains unknown. To investigate the expression of StAR in the myocardium and its association with RV remodeling in PAH, utilizing spironolactone as a treatment. A PAH model was created using male Sprague-Dawley rats, which received a subcutaneous injection of Sugen5416 (20 mg/kg) and were exposed to hypoxia (10% O2) for 2 weeks, followed by 2 weeks of normoxia. The animals were then divided into two groups, with one group receiving spironolactone (25 mg/kg/day) for an additional 4 weeks, while the other group did not. H9c2 cells were cultured under hypoxic conditions (37 °C, 1% O2, 5% CO2) with or without spironolactone treatment. In the model rats, RV systolic pressure and the Fulton index, both of which increased upon exposure to Sugen5416 and hypoxia, significantly decreased with spironolactone treatment. In H9c2 cells, hypoxic exposure elevated aldosterone levels, while spironolactone treatment significantly suppressed aldosterone production. Suppression of StAR expression in the myocardium via spironolactone contributes to the improvement of RV remodeling in PAH. Spironolactone may offer a valuable therapeutic strategy for RV remodeling in patients with PAH.

Local adaptation, plasticity, and evolved resistance to hypoxic cold stress in high-altitude deer mice

A fundamental question in evolutionary biology concerns the relative contributions of phenotypic plasticity vs. local adaptation (genotypic specialization) in enabling wide-ranging species to inhabit diverse environmental conditions. Here, we conduct a long-term hypoxia acclimation experiment to assess the relative roles of local adaptation and plasticity in enabling highland and lowland deer mice (Peromyscus maniculatus) to sustain aerobic thermogenesis at progressively increasing elevations. We assessed the relative physiological performance capacities of highland and lowland natives as they were exposed to progressive, stepwise increases in hypoxia, simulating the gradual ascent from sea level to an elevation of 6,000 m. The final elevation of 6,000 m far exceeds the highest attainable elevations within the species' range, and therefore tests the animals' ability to tolerate levels of hypoxia that surpass the prevailing conditions within their current distributional limits. Our results demonstrate that highland natives exhibit superior thermogenic capacities at the most severe levels of hypoxia, suggesting that the species' broad fundamental niche and its ability to inhabit such a broad range of elevational zones is attributable to genetically based local adaptation, including evolved changes in plasticity. Transcriptomic and physiological measurements identify evolved changes in the acclimation response to hypoxia that contribute to the enhanced thermogenic capacity of highland natives.

Mediating role of obstructive sleep apnea in altering slow-wave activity and elevating Alzheimer’s disease risk: Pilot study from a northern Taiwan cohort

ObjectivesObstructive sleep apnea is associated with alterations in slow-wave activity during sleep, potentially increasing the risk of Alzheimer’s disease. This study investigated the associations between obstructive sleep apnea manifestations such as respiratory events, hypoxia, arousal, slow-wave patterns, and neurochemical biomarker levels. MethodsIndividuals with suspected obstructive sleep apnea underwent polysomnography. Sleep disorder indices, oxygen metrics, and slow-wave activity data were obtained from the polysomnography, and blood samples were taken the following morning to determine the plasma levels of total tau (T-Tau) and amyloid beta-peptide 42 (Aβ42) by using an ultrasensitive immunomagnetic reduction assay. Subsequently, the participants were categorized into groups with low and high Alzheimer’s disease risk on the basis of their computed product Aβ42 × T-Tau. Intergroup differences and the associations and mediation effects between sleep-related parameters and neurochemical biomarkers were analyzed. ResultsForty-two participants were enrolled, with 21 assigned to each of the low- and high-risk groups. High-risk individuals had a higher apnea–hypopnea index, oxygen desaturation index (≥3%, ODI-3%), fraction of total sleep time with oxygen desaturation (SpO2-90% TST), and arousal index and greater peak-to-peak amplitude and slope in slow-wave activity, with a correspondingly shorter duration, than did low-risk individuals. Furthermore, indices such as the apnea–hypopnea index, ODI-3% and SpO2-90% TST were found to indirectly affect slow-wave activity, thereby raising the Aβ42 × T-Tau level. ConclusionsObstructive sleep apnea manifestations, such as respiratory events and hypoxia, may influence slow-wave sleep activity (functioning as intermediaries) and may be linked to elevated neurochemical biomarker levels. However, a longitudinal study is necessary to determine causal relationships among these factors. Statement of significanceThis research aims to bridge gaps in understanding how obstructive sleep apnea is associated with an elevated risk of Alzheimer’s disease, providing valuable knowledge for sleep and cognitive health.

Hypoxia-associated gene signatures are not prognostic in high-risk localised prostate cancers undergoing androgen deprivation therapy with radiotherapy

PurposeMen with high-risk prostate cancer (PCa) are treated with androgen deprivation therapy (ADT) and radiotherapy, but the disease reoccurs in 30% of patients. Biochemical recurrence of PCa after treatment is influenced by tumour hypoxia. Tumours with high levels of hypoxia are aggressive, resistant to treatment, and have increased metastatic capacity. Gene expression signatures derived from diagnostic biopsies can predict tumour hypoxia and radiosensitivity, but none are in routine clinical use, due to concerns about the applicability of these biomarkers to new patient cohorts. There has been no or limited testing in cohorts of high-risk PCa. Methods and materialsWe generated transcriptomic data for cohorts of high-risk PCa patients. Patients were treated with ADT followed by external beam radiotherapy with or without a brachytherapy boost. Biomarkers curated from the literature were calculated from pre-treatment biopsy gene expression data. The primary endpoint for survival analyses was biochemical recurrence-free survival (bRFS) and the secondary endpoints were distant metastasis-free survival (DMFS) and overall survival. ResultsThe performance of the selected biomarkers was poor, with none achieving prognostic significance for bRFS or DMFS in any cohort. The brachytherapy boost cohort received shorter durations of ADT than the conventionally fractionated or hypofractionated cohorts (Wilcoxon rank sum test, p=2.1 × 10-18 and 2.3 × 10-10 respectively) and had increased risk of distant metastasis (log-rank test, p=8 × 10-4). There were no consistent relationships between biomarker score and outcome for any of the endpoints. ConclusionsHypoxia and radiosensitivity biomarkers were not prognostic in high-risk PCa patients treated with ADT plus radiotherapy. We speculate that the lack of prognostic capability could be caused by the variable hypoxia-modifying effects of the ADT that these high-risk patients received before and during definitive treatment with radiotherapy. A deeper understanding of biomarker construction, performance and inter-cohort transferability in relation to patient characteristics, sample handling and treatment modalities is required before hypoxia biomarkers can be recommended for routine clinical use in the pre-treatment setting.

Oxygen loss compromises growth and cognition of cuttlefish newborns.

Ocean deoxygenation and standing levels of hypoxia are shrinking fundamental niches, particularly in coastal areas, yet documented repercussions on species development and behaviour are limited. Here, we tackled the impacts of deoxygenation (7 mg O2 l-1), mild hypoxia (nocturnal 5 mg O2 l-1) and severe hypoxia (nocturnal 2 mg O2 l-1) on cuttlefish (Sepia officinalis) development (hatching success, development time, mantle length), cognition (ability to learn individually and socially) and behaviour (ability to camouflage and to explore its surroundings spatially). We found that hypoxia yielded lower survival rates, smaller body sizes and inhibited predatory (increased latency to attack the prey) and anti-predator (camouflage) behaviours. Acute and chronic exposure to low oxygen produced similar effects on cognition (inability to socially learn, increased open-field activity levels, no changes in thigmotaxis). It is thus expected that, although cuttlefish can withstand oxygen limitation to a certain degree, expanding hypoxic zones will diminish current habitat suitability.

Preconditioning with Ginsenoside Rg3 mitigates cardiac injury induced by high-altitude hypobaric hypoxia exposure in mice by suppressing ferroptosis through inhibition of the RhoA/ROCK signaling pathway

Ethnopharmacological relevanceGinseng has historically been utilized as a conventional herbal remedy and dietary supplement to enhance physical stamina and alleviate fatigue. The primary active component of Ginseng, Ginsenoside Rg3 (GS-Rg3), possesses diverse pharmacological properties including immune modulation and anti-inflammatory effects. Furthermore, GS-Rg3 has demonstrated efficacy in mitigating tissue and organ damage associated with metabolic disorders such as hypertension, hyperglycemia, and hyperlipidemia. Nevertheless, its potential impact on high-altitude cardiac injury (HACI) remains insufficiently explored. Aim of the studyThe aim of this study was to examine the potential cardioprotective effects of Ginsenoside Rg3, and to investigate how Ginsenoside Rg3 preconditioning can enhance high-altitude cardiac injury by inhibiting the RhoA/ROCK pathway and ferroptosis in cardiac tissue. The findings of this study may contribute to the development of novel therapeutic strategies using traditional Chinese medicine for high-altitude cardiac injury, based on experimental evidence. Materials and methodsA hypobaric hypoxia chamber was employed to simulate hypobaric hypoxia conditions equivalent to an altitude of 6000 m. Through a randomization process, groups of six male mice were assigned to receive either saline, Ginsenoside Rg3 at doses of 15 mg/kg or 30 mg/kg, or lysophosphatidic acid (LPA) at 1 mg/kg. The impact of Ginsenoside Rg3 on high altitude-induced arrhythmias was evaluated using electrocardiography. Cardiac pathology sections stained with hematoxylin and eosin were evaluated for damage, with the extent of cardiomyocyte damage observed via transmission electron microscopy. The impact of Ginsenoside Rg3 on high-altitude cardiac injury was investigated through analysis of serum biomarkers for cardiac injury (CK-MB, BNP), inflammatory cytokines (TNF, IL-6, IL-1β), reactive oxygen species (ROS) and glutathione (GSH). The expression levels of hypoxia and hypoxia-related proteins in myocardial tissues from each experimental group were assessed using Western blot analysis. Following a review of the existing literature, the traditional regulatory mechanisms of ferroptosis were examined. Immunofluorescence staining of cardiac tissues and Western blotting techniques were utilized to investigate the impact of Ginsenoside Rg3 on cardiomyocyte ferroptosis through the RhoA/ROCK signaling pathway under conditions of hypobaric hypoxia exposure. ResultsPre-treatment with Ginsenoside Rg3 improved high altitude-induced arrhythmias, reduced cardiomyocyte damage, decreased cardiac injury biomarkers and inflammatory cytokines, and lowered the expression of hypoxia-related proteins in myocardial tissues. Both Western blotting and immunofluorescence staining of cardiac tissues demonstrated that exposure to high-altitude hypobaric hypoxia results in elevated expression of ferroptosis and proteins related to the RhoA/ROCK pathway. Experimental validation corroborated that the role of the RhoA/ROCK signaling pathway in mediating ferroptosis. ConclusionsThe findings of our study suggest that preconditioning with Ginsenoside Rg3 may attenuate cardiac injury caused by high-altitude hypobaric hypoxia exposure in mice by inhibiting ferroptosis through the suppression of the RhoA/ROCK signaling pathway. These findings contribute to the current knowledge of Ginsenoside Rg3 and high-altitude cardiac injury, suggesting that Ginsenoside Rg3 shows potential as a therapeutic agent for high-altitude cardiac injury.

Sensitive Cancer Hypoxia Detection via a Dual-Locking Fluorescence Response System Using Two Hypoxia Indicators.

Hypoxia is intricately associated with various diseases, including ischemia, vascular disorders, and cancer. Particularly in cancer cells, hypoxia promotes tumor growth, cell proliferation, migration, and invasion and enhances treatment resistance, making its detection crucial for cancer diagnosis and therapy. However, methods for detecting hypoxia are limited, often relying on single-detection systems. In this study, we developed a dual-lock-based fluorescent probe that selectively exhibits strong green fluorescence under hypoxic conditions due to simultaneous activity of nitroreductases (NTRs) and hydrogen sulfide (H2S), with a high signal-to-background ratio. The biocompatibility and photophysical properties of the probes were thoroughly investigated through both extracellular and intracellular experimental analyses. Among the synthesized naphthalimide-based probes, the dual-detection probe DNNC demonstrated excellent selectivity and sensitivity to the simultaneous activity of NTR/H2S compared to other single-detection probes. The performance of DNNC was applied to various organ-derived cancer cells and tumor tissue models such as HeLa cell sparoids, enabling spatiotemporal confocal fluorescence imaging and quantitative analysis of hypoxic levels in cancer. Our development of DNNC is expected to significantly advance cancer diagnosis and treatment by molecularly detecting hypoxia associated with cancer aggressiveness, therapy resistance, and unfavorable prognosis.

Activated Endothelium Changes The Activity Of Multipotent Mesenchymal Stromal Cells During Physiological Hypoxia Or Short Hypoxic Stress In Vitro

Multipotent mesenchymal stromal cells (MSCs) are used for supplemental therapy of ischemic and inflammatory diseases. After systemic administration, transmigration of MSCs to the target tissue is accompanied by interaction with activated endothelial cells (ECs) at the site of injury. In this study, we investigated the influence of TNF-α-activated ECs on the functions of MSCs under different levels of hypoxia. For this purpose, MSCs and TNF-α activated ECs were cocultured in a direct cell-to-cell setting for a short period of time. MSCs retained their stromal phenotype and multilineage differentiation potential after interaction with activated ECs. At the same time, changes in molecules involved in MSC-cell and MSC-extracellular matrix interaction were detected. The paracrine activity of MSCs and activated ECs after interaction was demonstrated by both upregulated transcription and increased levels of pleiotropic IL-6 and IL-8. Proteases/antiproteases profiles were also altered after interaction. These data suggest that short-term interaction of MSCs with activated ECs may play an important role in tissue repair and remodeling processes. In particular, it may promote the migratory phenotype of MSCs. In comparison to physiological hypoxia – 5% O2, acute hypoxic stress (0.1% O2, 24 h) attenuated the stimulatory effects of ECs on MSCs.

Microfluidic Device with an Oxygen Gradient Generator for Investigating Effects of Specific Hypoxia Conditions on Responses of Tumor Cells.

The oxygen level in the tumor microenvironment (TME) plays a critical role in regulating cell fates such as proliferation, migration, apoptosis, and so forth. To better elucidate how hypoxia affects tumor cell behaviors, a series of microfluidic strategies have been utilized to generate an oxygen gradient covering both hypoxia and normoxia conditions. However, in most studies, some chemicals are introduced into microfluidic chips, causing the potential of their poor biocompatibility. The common oxygen gradient with linear variation does not allow the effects of specific oxygen concentrations on tumor cells to be analyzed accurately. In this paper, based on the physical method of gas diffusion, a microfluidic device integrated with an oxygen gradient generator is proposed for investigating effects of different hypoxia levels on responses of tumor cells. This device consists of three layers, i.e., upper layer, thin film layer, and bottom layer. The upper layer is used for introducing the initial gas and generating an oxygen gradient in the form of gas. The bottom layer is used for introducing cells and culture medium. The thin film layer separates the former two layers, allowing the gas to diffuse from the top to the bottom through it. The oxygen gradient in the bottom layer is finally generated in the form of dissolved oxygen. The device is fabricated using microfabrication technology. The effects of structural and working parameters of the device on the oxygen gradient are evaluated by finite element simulation. The oxygen gradient in cell culture channels is characterized by using oxygen-sensitive fluorescence materials. The proliferation and morphology of HeLa cells under specific oxygen levels are compared after culturing for 48 h. The oxygen gradient with a ladder-like distribution demonstrates that this microfluidic device can provide a prospective experimental platform for in vitro cell studies and revelation of the mechanism of tumor metastasis associated with a specific hypoxic microenvironment.

Cognitive performance during exposure to moderate normobaric hypoxia after sleep restriction: Relationship to physiological and stress biomarkers

IntroductionExposure to moderate levels of simulated hypoxia has subtle cognitive effects relative to ground level, in healthy individuals. However, there are few data on the cognitive consequences of the combination of hypoxia and partial sleep deprivation, which is a classic military or civilian operational context. In this study, we tested the hypothesis that exposure to moderate hypoxia while sleep-restricted impairs several domains of cognition, and we also assessed physiological parameters and salivary concentrations of cortisol and alpha-amylase. MethodSeventeen healthy males completed two sessions of cognitive tests (sustained attention using the PVT psychomotor vigilance task and executive functions using the Go-NoGo inhibition task and N-Back working memory task) after 30 min (T + 30′) and 4 h (T + 240′) of exposure in a normobaric hypoxic tent (FIO2 = 13.6 %, ≃ 3,500 m) (HY). This was completed after one night of sleep restriction (3 a.m. to 6 a.m. bedtime, SRHY) and one night of habitual sleep (10 p.m. to 6 a.m. bedtime, HSHY) (with cross-over randomization). The two nights sleep architecture and physiological parameters (oxygen saturation (SpO2) and heart rate (HR) during T + 30′ and T + 240′sessions were analyzed. Salivary cortisol and alpha-amylase (sAA) concentrations were analyzed before hypoxia, after the T + 30′ and T + 240′ cognitive sessions, and after leaving the hypoxic tent. ResultsSustained attention (RT and number of lapses in the PVT) and executive functions (Go-NoGo and 1-Back and 2-Back parameters, as inhibition and working memory signatures) were impaired in the SRHY condition compared to HSHY. SpO2 and HR were higher after 4 h compared with 30 min of hypoxia in the HSHY condition, while only HR was statistically higher in the SRHY condition. In SRHY, salivary AA concentration was lower and cortisol was higher than in HSHY. A significant increase in sAA concentration is observed after the cognitive session at 4 h of hypoxia exposure compared to that at 30 min, only in the SRHY condition. There are significant positive correlations between reaction time and the corresponding heart rate (a non-invasive marker of physiological stress) for the executive tasks in the two sleep conditions. This was not observed for salivary levels of sAA and cortisol, respective reliable indicators of the sympathoadrenomedullary system and the hypothalamic-pituitary adrenocortical system. ConclusionExposure to moderate normobaric hypoxia (≃ 3500 m / ≃ 11,500 ft simulated) after a single night of 3-hour sleep impairs cognitive performance after 30 min and 4 h of exposure. The key determinants and/or mechanism(s) responsible for cognitive impairment when exposed to moderate hypoxia with sleep restriction, particularly on the executive function, have yet to be elucidated.

2847: Regulatory role of miRNAs at different tumor hypoxia levels in prostate cancer patients

2847: Regulatory role of miRNAs at different tumor hypoxia levels in prostate cancer patients

Facile One Pot Synthesis of Hybrid Core-Shell Silica-Based Sensors for Live Imaging of Dissolved Oxygen and Hypoxia Mapping in 3D Cell Models.

Fluorescence imaging allows for noninvasively visualizing and measuring key physiological parameters like pH and dissolved oxygen. In our work, we created two ratiometric fluorescent microsensors designed for accurately tracking dissolved oxygen levels in 3D cell cultures. We developed a simple and cost-effective method to produce hybrid core-shell silica microparticles that are biocompatible and versatile. These sensors incorporate oxygen-sensitive probes (Ru(dpp) or PtOEP) and reference dyes (RBITC or A647 NHS-Ester). SEM analysis confirmed the efficient loading and distribution of the sensing dye on the outer shell. Fluorimetric and CLSM tests demonstrated the sensors' reversibility and high sensitivity to oxygen, even when integrated into 3D scaffolds. Aging and bleaching experiments validated the stability of our hybrid core-shell silica microsensors for 3D monitoring. The Ru(dpp)-RBITC microparticles showed the most promising performance, especially in a pancreatic cancer model using alginate microgels. By employing computational segmentation, we generated 3D oxygen maps during live cell imaging, revealing oxygen gradients in the extracellular matrix and indicating a significant decrease in oxygen level characteristics of solid tumors. Notably, after 12 h, the oxygen concentration dropped to a hypoxic level of PO2 2.7 ± 0.1%.

Prolonged inhibition of intratumoral mast cells enhances efficacy of low-dose antiangiogenic therapy.

Low-dose antiangiogenic therapies have demonstrated the ability to enhance normalization of tumor vessels, consequently improving hypoxia levels, drug delivery, and promoting anticancer immune responses. Mast cells have been identified as contributors to resistance against antiangiogenic therapy and facilitators of abnormal neoangiogenesis. In this study, we demonstrate that by simultaneously targeting intratumoral mast cells with Imatinib and administering low-dose anti-VEGFR2 therapy, antitumor efficacy can be enhanced in preclinical models. Thus, combinatory treatment overcomes therapy resistance, while concurrently promoting tumor vessel normalization. Notably, histomorphometric analysis of tumor sections revealed that vessel perfusion could be improved through mast cell inhibition and, despite a significantly reduced microvessel density, the combination treatment did not result in elevated tumor hypoxia levels compared to anti-VEGFR2 therapy alone. Short-term Imatinib application effectively increased antitumor efficacy, and by prolonging the application of Imatinib tumor vessel normalization was additionally improved. The combination of mast cell depletion and antiangiogenic treatments has not been investigated in detail and promises to help overcoming therapy resistance. Further studies will be required to explore their impact on other treatment approaches, and subsequently to validate these findings in a clinical setting.

Xylazine Exacerbates Fentanyl-Induced Respiratory Depression and Bradycardia.

Fatal opioid overdoses in the United States have nearly tripled during the past decade, with greater than 92% involving a synthetic opioid like fentanyl. Fentanyl potently activates the μ-opioid receptor to induce both analgesia and respiratory depression. The danger of illicit fentanyl has recently been exacerbated by adulteration with xylazine, an α2-adrenergic receptor agonist typically used as a veterinary anesthetic. In 2023, over a 1,000% increase in xylazine-positive overdoses was reported in some regions of the U.S. Xylazine has been shown to potentiate the lethality of fentanyl in mice, yet a mechanistic underpinning for this effect has not been defined. Herein, we evaluate fentanyl, xylazine, and their combination in whole-body plethysmography (to measure respiration) and pulse oximetry (to measure blood oxygen saturation and heart rate) in male and female CD-1 mice. We show that xylazine decreases breathing rate more than fentanyl by increasing the expiration time. In contrast, fentanyl primarily reduces breathing by inhibiting inspiration, and xylazine exacerbates these effects. Fentanyl but not xylazine decreased blood oxygen saturation, and when combined, xylazine did not change the maximum level of fentanyl-induced hypoxia. Xylazine also reduced heart rate more than fentanyl. Finally, loss in blood oxygen saturation correlated with the frequency of fentanyl-induced apneas, but not breathing rate. Together, these findings provide insight into how the addition of xylazine to illicit fentanyl may increase the risk of overdose.