Decrease In Oxygen Consumption Research Articles

Hepatocyte-specific Selenoi deficiency predisposes mice to hepatic steatosis and obesity.

Selenoprotein I (Selenoi) is highly expressed in liver and plays a key role in lipid metabolism as a phosphatidylethanolamine (PE) synthase. However, the precise function of Selenoi in the liver remains elusive. In the study, we generated hepatocyte-specific Selenoi conditional knockout (cKO) mice on a high-fat diet to identify the physiological function of Selenoi. The cKO group exhibited a significant increase in body weight, with a 15.6% and 13.7% increase in fat accumulation in white adipose tissue (WAT) and the liver, respectively. Downregulation of the lipolysis-related protein (p-Hsl) and upregulation of the adipogenesis-related protein (Fasn) were observed in the liver of cKO mice. The cKO group also showed decreased oxygen consumption (VO2), carbon dioxide production (VCO2), and energy expenditure (p < .05). Moreover, various metabolites of the steroid hormone synthesis pathway were affected in the liver of cKO mice. A potential cascade of Selenoi-phosphatidylethanolamine-steroid hormone synthesis might serve as a core mechanism that links hepatocyte-specific Selenoi cKO to biochemical and molecular reactions. In conclusion, we revealed that Selenoi inhibits body fat accumulation and hepatic steatosis and elevates energy consumption; this protein could also be considered a therapeutic target for such related diseases.

Lactate-induced IGF1R protein lactylation promotes proliferation and metabolic reprogramming of lung cancer cells.

Lung cancer (LC) is regarded as a fatal cancer, and insulin-like growth factor 1 (IGF1) and its receptor (IGF1R) have been found to play a key role in regulating tumor glycolytic metabolism. The aim of this study is to investigate LC proliferation regulated by metabolite-mediated IGF1R lactylation. IGF1R was highly expressed in LC tissues and cells, and the effects of IGF1R on protein stability were inhibited by Lactate dehydrogenase A (LDHA) inhibition. Moreover, the tightness of IGF1R binding to IGF1 was also enhanced by exogenous lactic acid but suppressed by LDHA silencing, while cell viability and proliferation were promoted by over-expression of IGF1R. Exogenous lactic acid further exacerbated the effects of the IGF1R gene, while LDHA knocking down reduced the IGF1R-induced malignant behaviors. The IGF1R and exogenous lactic acid were also found to increase extracellular acidification rate (ECAR) and decrease oxygen consumption rate to regulate glycolysis, which was inhibited by LDHA deficiency in LC cells. The study concluded that IGF1R-mediated aggressive behaviors of LC cells were associated with higher levels of IGF1R lactylation. Moreover, lactic acid can improve the protein stability of the IGF1R oncogene, thus promoting glycolysis and generating lactic acid, forming a closed loop. Therefore, targeting IGF1R is envisaged to provide a novel strategy for developing therapeutic agents against LC.

Heat stress upregulates arachidonic acid to trigger autophagy in sertoli cells via dysfunctional mitochondrial respiratory chain function

As a key factor in determining testis size and sperm number, sertoli cells (SCs) play a crucial role in male infertility. Heat stress (HS) reduces SCs counts, negatively impacting nutrient transport and supply to germ cells, and leading to spermatogenesis failure in humans and animals. However, how HS affects the number of SCs remains unclear. We hypothesized that changes in SC metabolism contribute to the adverse effects of HS. In this study, we first observed an upregulation of arachidonic acid (AA), an unsaturated fatty acid after HS exposure by LC-MS/MS metabolome detection. By increasing ROS levels, expression of KEAP1 and NRF2 proteins as well as LC3 and LAMP2, 100 µM AA induced autophagy in SCs by activating oxidative stress (OS). We observed adverse effects of AA on mitochondria under HS with a decrease of mitochondrial number and an increase of mitochondrial membrane potential (MMP). We also found that AA alternated the oxygen transport and absorption function of mitochondria by increasing glycolysis flux and decreasing oxygen consumption rate as well as the expression of mitochondrial electron transport chain (ETC) proteins Complex I, II, V. However, pretreatment with 5 mM NAC (ROS inhibitor) and 2 µM Rotenone (mitochondrial ETC inhibitor) reversed the autophagy induced by AA. In summary, AA modulates autophagy in SCs during HS by disrupting mitochondrial ETC function, inferring that the release of AA is a switch-like response, and providing insight into the underlying mechanism of high temperatures causing male infertility.

Intermittent Fasting Alters O2 Consumption and Body Composition in Young Rats: Comparative Analysis between Females and Males

Objectives: Intermittent Fasting (IF), time-restrictive food intake, has been shown to improve cognitive functions and reduce age-related behavioral impairments in rodent models. Our prior studies have found that IF intervention has significantly reduced body weights and food intake in male Fsher-344 rats; however, females did not display IF-induced declines in body weights and food intake. The exact mechanism for the sex differences in body weight and food intake during IF is not fully understood. Factors such as shifts in body composition and metabolic demand may contribute to variations in body weight shifts and food consumption. We hypothesized that changes in body composition and metabolic shifts may contribute to sex differences in body weight and food consumption in response to IF. Methods: Thirty-six, two-month-old male and female Fisher-344 rats were randomly assigned to four groups: ad libitum (AL) male, IF males, AL females, IF females. The AL group had 24/7 access to food and water while the IF group had 24/7 access to water and full access to food on an every-other-day basis for four weeks. After four weeks of IF intervention, rats were introduced to a metabolic rate (respiratory gas analyzer) and a grip strength test. In addition, body scans were performed using a Lunar Piximus rodent densitometer, (GE Lunar corporation). Group differences were analyzed with two-way ANOVA per Prism Graph Pad software. Following body scans, animals were euthanized, and tissues were sectioned, weighted, and stored for future analysis (brain, skeletal muscles, and adipose tissues). Results: IF intervention resulted in reduced body mass in males (vs. AL males, p&lt;0.0001) but not in females. Average daily food intake tended to be lowered in IF groups compared to their AL counterparts. Densitometry results revealed that the IF males had a decreased lean mass (vs. AL males, p&lt;0.0001) and fat mass (vs. AL males, p&lt;0.0001). Neither lean mass nor fat mass were changed in females as a result of IF. After two weeks of IF, O2 consumption was lowered in IF females compared to AL (P=0.0215 fed-state, P=0.0030 fast-state); however, no significant effect was seen in males. Grip strength was not significant among groups. Conclusions: In the present study, IF males significantly reduced body weights, as well as both fat and lean mass while females did not exhibit IF-induced changes. Interestingly, IF females displayed declines in oxygen consumption while males did not change. The decrease in oxygen consumption (slower metabolism) in IF females can be seen as a compensatory mechanism by the body to conserve energy and maintain body weights and composition shifts. The sexual dimorphic effect of body composition shifts is intriguing and the metabolic adaptations in skeletal muscle/fat tissues will be the focus of follow-up investigations. Funding source: P20GM113109 Cognitive and Neurobiological Approaches to Plasticity (CNAP) center. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Renal potassium regulation in a pig model of therapeutic hypothermia

Therapeutic hypothermia is increasingly utilized clinically to reduce tissue metabolism and protect against neurological damage in cases of cardiac arrest, traumatic brain injury or perinatal hypoxic ischemia. With even mild exposure to hypothermia, hypokalemia develops purportedly due to transcellular potassium shifts resulting in decreased circulating potassium levels. Renal dysfunction with body temperature cooling has also been postulated to potentially contribute to development of sustained hypokalemia. In this study, we used a pig model of controlled acute hypothermia without confounding hypotension and hypoxemia, to test the hypothesis that renal handling of potassium remains intact in hypothermia and is not the cause of decreased circulating plasma potassium seen with body cooling. Anesthetized, mechanically ventilated Duroc cross young pigs (n=25, body weight = 10.0 ± 0.4 kg, male or female) were catheterized for hemodynamic monitoring, regional blood flow assessment via colored microspheres, and urine collection. After baseline values were obtained, pigs were divided into a control normothermia group (n=12) with body temperature maintained at 38.4 ± 0.1 oC or a hypothermia group (n=13) with core body temperature cooled to 34.5 ± 0.1 oC and hypothermia sustained for 2 hours. Hypothermia reduced tissue metabolism as indicated by a decrease in oxygen consumption (VO2= 5.2 ± 0.2 ml/kg in normothermia vs 3.7 ± 0.3 ml/kg in hypothermia. Mean arterial pressure (68± 3 mm Hg) and cardiac output (199 ± 21 ml/min/kg) were not different between groups. Even with the short duration of cold exposure, plasma potassium was lower (p&lt;0.05) within 2 hours of hypothermia (4.1 ± 0.1 mEq/L) vs control (4.8 ± 0.1 mEq/L). Cold exposure caused redistribution of blood flow to the small intestines from other internal organs. Renal blood flow tended to decrease with cold, but glomerular filtration rate was suffcient to maintain urine flow, sodium excretion and free water clearance. Potassium clearance, however, was lower (p&lt;0.05) in hypothermia (0.70 ± 0.07 ml/min/kg) than in normothermia (1.04 ± 0.07 ml/min/kg), with lower potassium excretion (hypothermia = 2997 ± 331 mEq/min vs control = 4784 ± 393 mEq/min, p&lt;0.05) and fractional excretion of potassium. Reduced renal potassium excretion is consistent with renal compensation for lower circulating potassium induced by hypothermia. Interestingly, with relatively higher blood flow to the small intestines compared to decreased blood flow of other internal organs, potassium secretion in the early colon may contribute to further potassium wasting in hypothermia. The rapid initiation of decreased potassium circulating levels with hypothermia supports the thesis of temperature sensitive cell membrane potassium channels affecting extracellular to intracellular shifts of potassium. Results of this study indicate that renal potassium regulation is maintained in hypothermia and functions to decrease urinary potassium loss in the face of hypokalemia. The views expressed are those of the authors and do not necessarily reflect the offcial policy or position of the Defense Health Agency, Department of the Army, Department of Defense, or the U.S. Government. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Changes in running economy and attainable maximal oxygen consumption in response to prolonged running: The impact of training status.

During prolonged running at moderate-to-high intensity, running economy (RE) deteriorates and attainable maximal oxygen consumption (VO2max) decreases. Whether these changes appear similarly in trained and untrained runners exercising at the same relative intensity is not clear. We recruited 10 trained runners (TR) and 10 active adults (AA), and compared RE and attainable VO2max before and after 1 h of running at 70% of VO2max. Submaximal VO2 increased more (p = 0.019) in AA (0.20 ± 0.13 L min-1) than in TR (0.07 ± 0.05 L min-1). Attainable VO2max decreased in AA (-0.21 ± 0.15 L min-1, p = 0.002), but remained unchanged in TR (-0.05 ± 0.10 L min-1, p = 0.18). Relative intensity (i.e., VO2/attainable VO2max), increased more (p = 0.001) in AA (8.3 ± 4.4%) than in TR (2.6 ± 1.9%). These results demonstrate that the ability to resist changes in RE and VO2max following prolonged running is superior in trained versus untrained runners, when exercising at the same relative intensity.

Effects of Butorphanol on Respiration in White Rhinoceros (Ceratotherium simum) Immobilized with Etorphine-Azaperone.

This article reports on respiratory function in white rhinoceros (Ceratotherium simum) immobilized with etorphine-azaperone and the changes induced by butorphanol administration as part of a multifaceted crossover study that also investigated the effects of etorphine or etorphine-butorphanol treatments. Six male white rhinoceros underwent two immobilizations by using 1) etorphine-azaperone and 2) etorphine-azaperone-butorphanol. Starting 10 min after recumbency, arterial blood gases, limb muscle tremors, expired minute ventilation, and respiratory rate were evaluated at 5-min intervals for 25 min. Alveolar to arterial oxygen gradient, expected respiratory minute volume, oxygen consumption, and carbon dioxide production were calculated. Etorphine-azaperone administration resulted in hypoxemia and hypercapnia, with increases in alveolar to arterial oxygen gradient, oxygen consumption, and carbon dioxide production, and a decrease in expired minute ventilation. Muscle tremors were also observed. Intravenous butorphanol administration in etorphine-azaperone-immobilized white rhinoceros resulted in less hypoxemia and hypercapnia; a decrease in oxygen consumption, carbon dioxide production, and expired minute ventilation; and no change in the alveolar to arterial oxygen gradient and rate of breathing. We show that the immobilization of white rhinoceros with etorphine-azaperone results in hypoxemia and hypercapnia and that the subsequent intravenous administration of butorphanol improves both arterial blood oxygen and carbon dioxide partial pressures.

Folate induces stemness and increases oxygen consumption under glucose deprivation by notch-1 pathway activation in colorectal cancer cell.

Evidence for folate's protective effects on neural tube defects led the USA and Chile to start mandatory folic acid (FA) fortification programs, decreasing up to 50%. However, ∼30% of the population consuming fortified foods reach supraphysiologic serum levels. Although controversial, several epidemiological and clinical observations suggest that folate increases cancer risk, giving concern about the risks of FA supplementation. The Cancer stem cells (CSCs) model has been used to explain survival to anticancer therapies. The Notch-1 pathway plays a role in several cancers and is associated with the stemness process. Different studies show that modulation of metabolic pathways regulates stemness capacity in cancer. Supraphysiologic concentrations of FA increase the proliferation of HT-29 cells by Notch-1 activation. However, whether folate can induce a stemness-like phenotype in cancer is not known. We hypothesized that FA protects from glucose deprivation-induced cell death through Notch-1 activation. HT-29 cells were challenged with glucose deprivation at basal (20 nM) and supraphysiological (400 nM) FA and 5-MTHF concentrations. We analyzed changes in stemness-like gene expression, cell death and different energetic metabolic functions. Supraphysiological concentrations of FA increased stemness-like genes, and improved survival and oxygen consumption, inducing AMPK phosphorylation and HSP-70 protein expression. We evaluated the Notch-1 pathway using the DAPT and siRNA as inhibitors, decreasing the stemness-like gene expression and preventing the FA protection against glucose deprivation-induced cell death. Moreover, they decreased oxygen consumption and AMPK phosphorylation. These results suggest that FA protects against glucose deprivation. These effects were associated with AMPK activation, a critical metabolic mediator in nutrient consumption and availability that activates the Notch-1 pathway.

Sera from people with HIV and depression induce commensurate metabolic alterations in astrocytes: toward precision diagnoses and therapies

Abstract Objectives People with HIV (PWH) have high rates of depression and neurocognitive impairment (NCI) despite viral suppression on antiretroviral therapy (ART). Mounting evidence suggests that immunometabolic disruptions may contribute to these conditions in some PWH. We hypothesized that metabolic dysfunction in astrocytes is associated with depressive symptoms and cognitive function in PWH. Methods Human astrocytes were exposed to sera from PWH (n=40) with varying degrees of depressive symptomatology and cognitive function. MitoTrackerTM Deep Red FM (MT) was used to visualize mitochondrial activity and glial fibrillary acidic protein (GFAP) as an indicator of astrocyte reactivity using the high-throughput fluorescent microscopy and image analyses platform, CellInsight CX5 (CX5). The Seahorse platform was used to assess glycolytic and mitochondrial metabolism. Results More severe depression, as indexed by higher Beck's Depression Inventory (BDI-II) scores, was associated with lower MT signal measures. Better cognitive function, as assessed by neuropsychiatric testing t-scores, was associated with increased MT signal measures. GFAP intensity negatively correlated with several cognitive t-scores. Age positively correlated with (higher) MT signal measures and GFAP intensity. Worse depressive symptoms (higher BDI-II scores) were associated with decreased oxygen consumption rate and spare respiratory capacity, concomitant with increased extracellular acidification rate in astrocytes. Conclusions These findings show that factors in the sera of PWH alter mitochondrial activity in cultured human astrocytes, suggesting that mechanisms that alter mitochondrial and astrocyte homeostasis can be detected peripherally. Thus, in vitro cultures may provide a model to identify neuropathogenic mechanisms of depression or neurocognitive impairment in PWH and test personalized therapeutics for neurologic and psychiatric disorders.

Abstract 3082: The liver microenvironment promotes glycolysis in metastatic colorectal cancer by activating her3

Abstract Background: Liver metastasis occurs in ~80% of all metastatic colorectal cancer (mCRC) and the liver has a unique endothelial cell (EC)-rich microenvironment that promotes cancer cell survival. We previously reported that liver ECs secrete soluble factors to promote CRC growth via activating CRC-associated the HER3 signaling pathway. The present study reports that ECs promote mCRC growth by shifting cancer metabolism towards glycolysis and elucidates the involved signaling pathway. Methods: We performed mass spectrometry to profile the metabolic changes in mCRC treated with/without HER3 inhibition in a syngeneic, orthotopic mCRC model. We also performed Seahorse and lactate ELISA assays to determine the effect of liver EC-secreted factors on CRC metabolism. To further determine the role of HER3 in promoting glycolysis and CRC growth in the liver, we used siRNA silencing of HER3 and pharmacological inhibition of HER3 downstream signaling proteins to identify the key mediator(s) of HER3-induced metabolic shift in CRC. Results: We determined that HER3 inhibition leads to increased oxidative phosphorylation metabolites (OXPHOS) and decreased glycolysis metabolites in CRC liver metastases. Conditioned medium containing EC-secreted factors decreased oxygen consumption (determined by Seahorse, a readout of OXPHOS) and increased lactate secretion (determined by ELISA, a readout of glycolysis) in CRC. As HER3 inhibition reversed the metabolic shift and increased OXPHOS as an alternative survival strategy, we found that simultaneous inhibition of HER3 and OXPHOS synergistically blocked CRC cell and tumor growth. Lastly, we found that PFK2 is activated by HER3 signaling, determined by phosphorylation, and determined that the HER3-AKT-PFK2 signaling axis is the key mediator of EC-induced metabolic shifts. Conclusions: We identified that the liver microenvironment activates HER3 and the downstream AKT-PFK2 signaling to promote glycolysis and growth in CRC liver metastases by activating AKT-PFK2-dependent glycolysis. Our findings highlighted the potential of combining HER3 and OXPHOS inhibitions as a potential strategy for treating patients with mCRC. Citation Format: Moeez Ghani Rathore, Kimberly Curry, Christina Boutros, Zhenghe Wang, Jordan Winter, Rui Wang. The liver microenvironment promotes glycolysis in metastatic colorectal cancer by activating her3 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3082.

Constitutive Innate Immunity and Systemic Responses to Infection of the American Alligator (Alligator mississippiensis).

Uninfected alligators (Alligator mississippiensis) exhibited high constitutive levels of hepatic gene expression related to immune function, whereas the highest-expressed hepatic genes of uninfected mice were related to metabolism. Intraperitoneal challenge of mice with bacterial lipopolysaccharide results in dramatic inflammatory effects including peritoneal ascites, febrile response, dramatic alterations in electrophoretic serum profile, and mortality. In contrast, coelomic injection of alligators with 200× the murine LD50 of intraperitoneal bacterial lipopolysaccharide resulted in no changes in serum protein profiles, behavioral effects, mortality, and no coelomic ascites. However, injection of juvenile alligators with live bacteria resulted in a titer-dependent decrease in metabolic rate, as measured by oxygen consumption. These results are the opposite of those observed for mammalian and avian species. The decreased oxygen consumption was not accompanied by changes in heart or respiration rate, indicating that this phenomenon was not due to bradycardia or bradypnea. Interestingly, challenge of alligators with bacteria resulted in the complete expulsion of digestive tract contents within four hours. We interpret these activities as temporary minimization of other biological systemic activities to redirect and devote energy to immune function. The reallocation of resources within an organism to fight infection without increases in metabolic rate has not been described in other animals.

Mutations of GEMIN5 are associated with coenzyme Q10 deficiency: long-term follow-up after treatment.

GEMIN5 exerts key biological functions regulating pre-mRNAs intron removal to generate mature mRNAs. A series of patients were reported harboring mutations in GEMIN5. No treatments are currently available for this disease. We treated two of these patients with oral Coenzyme Q10 (CoQ10), which resulted in neurological improvements, although MRI abnormalities remained. Whole Exome Sequencing demonstrated compound heterozygosity at the GEMIN5 gene in both cases: Case one: p.Lys742* and p.Arg1016Cys; Case two: p.Arg1016Cys and p.Ser411Hisfs*6. Functional studies in fibroblasts revealed a decrease in CoQ10 biosynthesis compared to controls. Supplementation with exogenous CoQ10 restored it to control intracellular CoQ10 levels. Mitochondrial function was compromised, as indicated by the decrease in oxygen consumption, restored by CoQ10 supplementation. Transcriptomic analysis of GEMIN5 patients compared with controls showed general repression of genes involved in CoQ10 biosynthesis. In the rigor mortis defective flies, CoQ10 levels were decreased, and CoQ10 supplementation led to an improvement in the adult climbing assay performance, a reduction in the number of motionless flies, and partial restoration of survival. Overall, we report the association between GEMIN5 dysfunction and CoQ10 deficiency for the first time. This association opens the possibility of oral CoQ10 therapy, which is safe and has no observed side effects after long-term therapy.

Capsaicin Enhanced the Efficacy of Photodynamic Therapy Against Osteosarcoma via a Pro-Death Strategy by Inducing Ferroptosis and Alleviating Hypoxia.

Ferroptosis, a novel form of nonapoptotic cell death, can effectively enhance photodynamic therapy (PDT) performance by disrupting intracellular redox homeostasis and promoting apoptosis. However, the extremely hypoxic tumor microenvironment (TME) together with highly expressed hypoxia-inducible factor-1α (HIF-1α) presents a considerable challenge for clinical PDT against osteosarcoma (OS). Hence, an innovative nanoplatform that enhances antitumor PDT by inducing ferroptosis and alleviating hypoxia is fabricated. Capsaicin (CAP) is widely reported to specifically activate transient receptor potential vanilloid 1 (TRPV1) channel, trigger an increase in intracellular Ca2+ concentration, which is closely linked with ferroptosis, and participate in decreased oxygen consumption by inhibiting HIF-1α in tumor cells, potentiating PDT antitumor efficiency. Thus, CAP and the photosensitizer IR780 are coencapsulated into highly biocompatible human serum albumin (HSA) to construct a nanoplatform (CI@HSA NPs) for synergistic tumor treatment under near-infrared (NIR) irradiation. Furthermore, the potential underlying signaling pathways of the combination therapy are investigated. CI@HSA NPs achieve real-time dynamic distribution monitoring and exhibit excellent antitumor efficacy with superior biosafety in vivo. Overall, this work highlights a promising NIR imaging-guided "pro-death" strategy to overcome the limitations of PDT for OS by promoting ferroptosis and alleviating hypoxia, providing inspiration and support for future innovative tumor therapy approaches.

Antitumour effect of the mitochondrial complex III inhibitor Atovaquone in combination with anti-PD-L1 therapy in mouse cancer models

Immune checkpoint blockade (ICB) provides effective and durable responses for several tumour types by unleashing an immune response directed against cancer cells. However, a substantial number of patients treated with ICB develop relapse or do not respond, which has been partly attributed to the immune-suppressive effect of tumour hypoxia. We have previously demonstrated that the mitochondrial complex III inhibitor atovaquone alleviates tumour hypoxia both in human xenografts and in cancer patients by decreasing oxygen consumption and consequently increasing oxygen availability in the tumour. Here, we show that atovaquone alleviates hypoxia and synergises with the ICB antibody anti-PD-L1, significantly improving the rates of tumour eradication in the syngeneic CT26 model of colorectal cancer. The synergistic effect between atovaquone and anti-PD-L1 relied on CD8+ T cells, resulted in the establishment of a tumour-specific memory immune response, and was not associated with any toxicity. We also tested atovaquone in combination with anti-PD-L1 in the LLC (lung) and MC38 (colorectal) cancer syngeneic models but, despite causing a considerable reduction in tumour hypoxia, atovaquone did not add any therapeutic benefit to ICB in these models. These results suggest that atovaquone has the potential to improve the outcomes of patients treated with ICB, but predictive biomarkers are required to identify individuals likely to benefit from this intervention.

Metabolic perturbation studies using a Nash Equilibrium model of liver machine perfusion: modeling oxidative stress and effect of glutathione supplementation

The current clinical standard of Static Cold Storage (SCS) which involves preservation on ice (about +4°C) in a hypoxic state limits storage to a few hours for metabolically active tissues such as the liver and the heart. This period of hypoxia during can generate superoxide and other free radicals from purine metabolism, a well-established component of ischemia/reperfusion injury (IRI). Machine perfusion is at the cutting edge of organ preservation, which provides a functional, oxygenated preservation modality that can avoid/attenuate IRI. In clinical application, perfusion usually follows a period of SCS. This presentation of oxygen following hypoxia can lead to superoxide and hydrogen peroxide generation, but machine perfusion also allows manipulation of the temperature profiles and supply of antioxidant treatments, which could be used to minimize such issues. However, metabolomic data is difficult to gather, and there are currently no mathematical models present to allow rational design of experiments or guide clinical practice. In this article, the effects of a gradual warming temperature policy and glutathione supplementation to minimize oxidative stress are studied. An optimal gradual warming temperature policy for mid-thermic machine perfusion of a liver metabolic model is determined using a combination of Nash Equilibrium and Monte Carlo optimization. Using this optimal gradual warming temperature policy, minimum GSH requirements to maintain hydrogen peroxide concentrations in the normal region are calculated using a different Monte Carlo optimization methodology. In addition, the dynamic behavior of key metabolites and cofactors are determined. Results show that the minimum GSH requirement increases and that the ratio of GSH/GSSG decreases with increasing hydrogen peroxide concentration. In addition, at high concentrations of hydrogen peroxide it is shown that cytochrome C undergoes dysfunction leading to a decrease in useful oxygen consumption and ATP synthesis from the electron transport chain and an overall reduction in the energy charge for the liver cells.

Important Constituents of Heavy Water-containing Solution for Cold Storage and Subsequent Reperfusion on an Isolated Perfused Rat Liver

The University of Wisconsin (UW) solution is the most effective preservation solution currently used; however, to safely use expanded-criteria donor grafts, a new cold storage solution that alleviates graft injury more effectively is required. We prepared a heavy water (D2O)-containing buffer, Dsol, and observed strong protective effects during extended cold storage of rat hearts and livers. In the current study, we modified Dsol (mDsol) and tested its efficacy. The aim of the present study was to determine whether mDsol could protect the rat liver more effectively than the UW solution and to clarify the roles of D2O and deferoxamine (DFX). Rat livers were subjected to cold storage for 48 hours in test solutions: UW, mDsol, mDsol without D2O or DFX (mDsol-D2O[-], mDsol-DFX[-]), and subsequently reperfused on an isolated perfused rat liver for 90 minutes at 37°C. In the UW group, the liver was dehydrated during cold storage and rapidly expanded during reperfusion. Accordingly, the cumulative weight change was the highest in the UW group, together with augmented portal veinous resistance and ALT leakage and decreased oxygen consumption rate and bile production. These changes were significantly suppressed in the mDsol-treated group. In the mDsol-D2O(-) and mDsol-DFX(-) groups offered partial protection. In conclusion, mDsol appeared to be superior to the UW solution for simple cold storage of the rat liver, presumably due to improved microcirculation in the early phase of reperfusion. Both heavy water and deferoxamine are essential for alleviating seamless organ swelling that occurs during cold storage and subsequent reperfusion.

The Effects of Xanthine Oxidase Inhibitors on the Management of Cardiovascular Diseases

The Effects of Xanthine Oxidase Inhibitors on the Management of Cardiovascular Diseases

The sensitivity of the equatorial pacific ODZ to particulate organic matter remineralization in a climate model under pre-industrial conditions

Marine oxygen plays a fundamental role in regulating the transfer of organic carbon and nutrients to their dissolved inorganic forms, serving as the terminal electron acceptor for heterotrophic respiration. Oxygen can become limiting to these processes in coastal and open-ocean oxygen deficient zones (ODZs). The maintenance of ODZs depends on the balance between physical processes such as ventilation and biogeochemical processes such as remineralization. These two processes act in opposing directions on ODZs, with ventilation responsible for transporting oxygen from the surface, where it is near saturation with the partial pressure of O2 in the atmosphere, to deeper oxygen-depleted layers, and remineralization acting to consume oxygen through biogeochemical processes such as microbial degradation throughout the water column. Remineralization is represented in all CMIP6 models, but the actual parameterizations, as well as their magnitude, are widely varying.In this study, we examine the sensitivity to remineralization of the equatorial Pacific ODZ (O2<60μmol/kg) in a model; the NASA GISS Ocean Biogeochemical Model (NOBM-G) embedded in the NASA-GISS coupled atmosphere-ocean model. We find that increasing the remineralization rate shoals the ODZ onset depth (i.e. the regionally averaged upper bound of the ODZ), and decreases ODZ volume and regional-averaged thickness. Changes in biological consumption and vertical convergence of oxygen are identified as the primary contributors to changes in ODZ onset depth. ODZ thickness is mainly influenced by the shoaling of the bottom boundary of the ODZ, which decreases with maximum remineralization rate due to decreasing oxygen consumption and increasing horizontal oxygen convergence in deep waters. On the other hand, vertically-averaged ODZ area has a more complex, non-monotonic relationship with maximum remineralization rate. While these findings suggest an important role for remineralization in determining the shape of the ODZ in our model, the relative importance of remineralization vs. other physical parameterizations remains to be established. While our results reflect the structure of our ocean biogeochemical model, the relationship of remineralization to ODZ characteristics in other models should be examined to better inform model inter-comparisons.

Lower cardiotoxicity of CPX-351 relative to daunorubicin plus cytarabine free-drug combination in hiPSC-derived cardiomyocytes in vitro

Liposomal formulations are hypothesized to alleviate anthracycline cardiotoxicity, although this has only been documented clinically for doxorubicin. We developed an in vitro multiparametric model using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) to assess the relative toxicity of anthracyclines across formulations. Proof of concept was established by treating hiPSC-CM with equivalent concentrations of free and liposomal doxorubicin. The study was then repeated with free daunorubicin plus cytarabine and CPX-351, a dual-drug liposomal encapsulation of daunorubicin/cytarabine. hiPSC-CM were treated with free-drug or liposomal formulations for 24 h on Days 1, 3, and 5 at equivalent concentrations ranging from 0 to 1000 ng/mL and assessed on subsequent days. Free-drug treatment resulted in concentration-dependent cumulative cytotoxicity (microscopy), more profound decrease in ATP levels, and significant time- and concentration-dependent decreases in oxygen consumption versus liposomal formulations (p < 0.01). Repeated free-drug exposure also resulted in greater release of biomarkers (cardiac troponin I, FABP3) and lactate dehydrogenase, as well as in a biphasic rhythmicity response (initial increase followed by slowing/quiescence of beating) indicating significant injury, which was not observed after repeated exposure to liposomal formulations. Overall, liposomal formulations were considerably less toxic to hiPSC-CM than their free-drug counterparts. Clinical data will be needed to confirm findings for CPX-351.

Targeting Sumoylation Sensitizes Acute Myeloid Leukemia to Venetoclax Treatment

Acute myeloid leukemia (AML) is a hematopoietic cancer characterized by aberrant immature myeloid progenitor blasts in bone marrow and peripheral blood. Venetoclax (VEN), a BCL-2 inhibitor, is FDA-approved for AML treatment with hypomethylating agents (HMA). Despite the improvement of VEN and HMA, not all patients respond, and most patients develop resistance. This presents an urgent need for new therapies to overcome VEN resistance and enhance VEN efficacy. We propose SUMOylation inhibition as a novel therapy to address this need. SUMOylation regulates protein function by covalently attaching Small Ubiquitin-like MOdifier (SUMO) proteins to target proteins. SUMOylation plays an important role in tumor progression and immune response. Our study aims to evaluate the effects of SUMOylation inhibition on anti-AML activity of VEN and the mechanism. We firstly generated VEN-resistant cell lines Molm13-VR and MV411-VR by culturing Molm13 and MV4-11 cells in increasing doses of VEN. SUMO E1 (SAE2) and global SUMOylation levels were remarkably upregulated in VEN-resistant cell lines, suggesting SUMOylation plays a role in VEN resistance. We evaluated the antileukemic effects of TAK-981 (subasumstat), a novel and specific SUMO E1 inhibitor, alone and in combination with VEN, using AML cell lines and primary blasts isolated from refractory/relapsed (R/R) AML patients. Different AML cell lines and blast samples showed various sensitivities to VEN (IC 50 10nM ~ 5,000nM), but significant synergism was observed with TAK-981 and VEN in cell viability assay. TAK-981 synergized with VEN at inducing AML cell apoptosis determined by annexin-V staining with flow cytometry as well as increased levels of cleaved-PARP and cleaved caspase-3 by western blotting. The mechanism of action of VEN is dependent on BCL-2 mediated mitochondrial function and metabolism. We assessed mitochondrial membrane potential using TMRM probe. TAK-981 reduced mitochondrial membrane potential in both VEN-sensitive and resistant AML lines, further in combination with VEN. Because mitochondrial membrane potential is maintained by normal mitochondrial structure, we performed electron microscopy to determine mitochondrial structure. VEN had limited effect on mitochondria in resistant cells. However, TAK-981 induced abnormal ultrastructure mitochondria with fewer cristae, worsened by TAK-981+VEN combination. Since TAK-981 treatment caused mitochondrial structure defects, we determine the mitochondrial function using Seahorse XF assay. VEN decreased oxygen consumption rate (OCR), indicating inhibition of oxidative phosphorylation (OXPHOS), further suppressed by TAK-981. Metabolomic profiling by mass spectrometry indicated TAK-981+VEN treated AML cells displayed striking decreases in TCA cycle intermediates and reduced electron transport chain (ETC) activity. Pathway analysis of the metabolomics data revealed significant enrichment in amino acids metabolism. Notably, RNA-sequencing and GSEA analysis showed consistent findings that TAK-981+VEN showed suppressed signaling pathways in mitochondrial function, glycolysis and amino-acid metabolisms, indicating TAK-981 targeted mitochondrial metabolism to enhance VEN activity. We then determined whether TAK-981 enhanced antileukemic effect of VEN ex vivo and in vivo. TAK-981 synergized with VEN at reducing colony formation capacity in colony formation assay using leukemia stem cells (LSCs) isolated from relapse AML blasts. More importantly, TAK-981 showed remarkable therapeutic effect in patient-derived xenograft (PDX) mouse model engrafted with blasts isolated from R/R AML patients, and significantly synergized with VEN at extending mice survival in vivo. In conclusion, TAK-981 enhances the antileukemic activity of VEN by targeting mitochondria function and metabolism, offering a potent therapy for refractory/relapsed AML.