Oncometabolite d-2-hydroxyglutarate impairs α-ketoglutarate dehydrogenase and contractile function in rodent heart

Hematologic malignancies are frequently associated with cardiac pathologies. Mutations of isocitrate dehydrogenase 1 and 2 (IDH1/2) occur in a subset of acute myeloid leukemia patients, causing metabolic and epigenetic derangements. We have now discovered that altered metabolism in leukemic cells has a profound effect on cardiac metabolism. Combining mathematical modeling and in vivo as well as ex vivo studies, we found that increased amounts of the oncometabolite d-2-hydroxyglutarate (D2-HG), produced by IDH2 mutant leukemic cells, cause contractile dysfunction in the heart. This contractile dysfunction is associated with impaired oxidative decarboxylation of α-ketoglutarate, a redirection of Krebs cycle intermediates, and increased ATP citrate lyase (ACL) activity. Increased availability of D2-HG also leads to altered histone methylation and acetylation in the heart. We propose that D2-HG promotes cardiac dysfunction by impairing α-ketoglutarate dehydrogenase and induces histone modifications in an ACL-dependent manner. Collectively, our results highlight the impact of cancer cell metabolism on function and metabolism of the heart.

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Abstract 385: D-2-hydroxyglutaric acid Acutely Impairs Oxidative Decarboxylation in the Heart

Circulation Research ◽

10.1161/res.117.suppl_1.385 ◽

2015 ◽

Vol 117 (suppl_1) ◽

Author(s):

Anja Karlstaedt ◽

Xiaotian Zhang ◽

Hernan Vasquez ◽

Margaret A Goodell ◽

Heinrich Taegtmeyer

Keyword(s):

Contractile Function ◽

Energy Charge ◽

Cardiac Metabolism ◽

Oxidative Decarboxylation ◽

Low Grade ◽

Dependent Manner ◽

Isocitrate Dehydrogenase 1 ◽

Concentration Dependent Manner ◽

Cardiac Energy ◽

Hydroxyglutaric Acid

Mutations in isocitrate dehydrogenase 1 and 2 (IDH1, IDH2) have been described in low-grade glioma and in acute myeloid leukemia. Accumulation of the oncometabolite D-2-hydroxyglutaric acid (2HG) and its release into the blood is associated with dilated cardiomyopathy. The mechanisms leading to changes in cardiac metabolism and contractile function are unknown. We studied in the isolated working rat heart preparation metabolic consequences of increased 2HG supply and its impact on cardiac energy provision. In combination with physiological levels of glucose (5mM) and lactate (0.5mM), hearts were perfused at different concentrations with 2HG (0.5mM, 1.0mM). We confirmed the uptake and enrichment of 2HG in the tissue through liquid chromatography followed by mass spectrometry (LC/MS). 2HG markedly decreased cardiac power and cardiac efficiency in a concentration dependent manner. At the same time glucose oxidation increased significantly (1.4±0.1 μmol/min/g dry wt, p<0.05) at higher workloads. The alpha-Ketoglutarate Dehydrogenase activity was reduced by two-fold (2.84±0.56 μmol/min/g dry wt, p<0.01) and production of reactive oxygen species (e.g. H2O2) increased by three-fold (1.06±0.07 μmol/min/g dry wt, p<0.01) in presence of 2HG. Consistent with this reduction in oxidative decarboxylation, the cytosolic NAD+/NADH redox state increased (3.1±0.1, p<0.001), while the cellular energy charge declined (AMP:ATP ratio, 0.15±0.1, p<0.001). Together, our results demonstrate a direct impairment of cardiac energy substrate metabolism with a resulting decline in contractile function. The data suggests that 2HG directly suppresses cardiac function by specific metabolic alterations in a range of pathologies.

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Prenatal THC Does Not Affect Female Mesolimbic Dopaminergic System in Preadolescent Rats

International Journal of Molecular Sciences ◽

10.3390/ijms22041666 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1666

Author(s):

Francesco Traccis ◽

Valeria Serra ◽

Claudia Sagheddu ◽

Mauro Congiu ◽

Pierluigi Saba ◽

...

Keyword(s):

Ex Vivo ◽

Female Offspring ◽

Dopamine Neurons ◽

Male Rat ◽

Dependent Manner ◽

Mesolimbic Dopamine ◽

Dopamine System ◽

Mesolimbic Dopamine System ◽

The Impact

Cannabis use among pregnant women is increasing worldwide along with permissive sociocultural attitudes toward it. Prenatal cannabis exposure (PCE), however, is associated with adverse outcome among offspring, ranging from reduced birth weight to child psychopathology. We have previously shown that male rat offspring prenatally exposed to Δ9-tetrahydrocannabinol (THC), a rat model of PCE, exhibit extensive molecular, cellular, and synaptic changes in dopamine neurons of the ventral tegmental area (VTA), resulting in a susceptible mesolimbic dopamine system associated with a psychotic-like endophenotype. This phenotype only reveals itself upon a single exposure to THC in males but not females. Here, we characterized the impact of PCE on female behaviors and mesolimbic dopamine system function by combining in vivo single-unit extracellular recordings in anesthetized animals and ex vivo patch clamp recordings, along with neurochemical and behavioral analyses. We find that PCE female offspring do not show any spontaneous or THC-induced behavioral disease-relevant phenotypes. The THC-induced increase in dopamine levels in nucleus accumbens was reduced in PCE female offspring, even when VTA dopamine activity in vivo and ex vivo did not differ compared to control. These findings indicate that PCE impacts mesolimbic dopamine function and its related behavioral domains in a sex-dependent manner and warrant further investigations to decipher the mechanisms determining this sex-related protective effect from intrauterine THC exposure.

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Establishment of a Robust and Simple Corneal Organ Culture Model to Monitor Wound Healing

Journal of Clinical Medicine ◽

10.3390/jcm10163486 ◽

2021 ◽

Vol 10 (16) ◽

pp. 3486

Author(s):

Sandra Schumann ◽

Eva Dietrich ◽

Charli Kruse ◽

Salvatore Grisanti ◽

Mahdy Ranjbar

Keyword(s):

Wound Healing ◽

Organ Culture ◽

Mitomycin C ◽

Ex Vivo ◽

Corneal Tissue ◽

Dependent Manner ◽

Toxicological Studies ◽

Culture Model ◽

The Impact

The use of in vitro systems to investigate the process of corneal wound healing offers the opportunity to reduce animal pain inflicted during in vivo experimentation. This study aimed to establish an easy-to-handle ex vivo organ culture model with porcine corneas for the evaluation and modulation of epithelial wound healing. Cultured free-floating cornea disks with a punch defect were observed by stereomicroscopic photo documentation. We analysed the effects of different cell culture media and investigated the impact of different wound sizes as well as the role of the limbus. Modulation of the wound healing process was carried out with the cytostatic agent Mitomycin C. The wound area calculation revealed that after three days over 90% of the lesion was healed. As analysed with TUNEL and lactate dehydrogenase assay, the culture conditions were cell protecting and preserved the viability of the corneal tissue. Wound healing rates differ dependent on the culture medium used. Mitomycin C hampered wound healing in a concentration-dependent manner. The porcine cornea ex vivo culture ideally mimics the in vivo situation and allows investigations of cellular behaviour in the course of wound healing. The effect of substances can be studied, as we have documented for a mitosis inhibitor. This model might aid in toxicological studies as well as in the evaluation of drug efficacy and could offer a platform for therapeutic approaches based on regenerative medicine.

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Ethanol Extract of Achillea fragrantissima Enhances Angiogenesis through Stimulation of VEGF Production

Endocrine Metabolic & Immune Disorders - Drug Targets ◽

10.2174/1871530321666201230113018 ◽

2020 ◽

Vol 21 ◽

Author(s):

Hana M. Hammad ◽

Amer Imraish ◽

Maysa Al-Hussaini ◽

Malek Zihlif ◽

Amani A. Harb ◽

...

Keyword(s):

Wound Healing ◽

Rural Communities ◽

Ex Vivo ◽

Ethanol Extract ◽

Aortic Ring ◽

Treated Group ◽

Control Group ◽

Dependent Manner ◽

Achillea Fragrantissima

Objective: Achillea fragrantissima L. (Asteraceae) is a traditionally used medicinal herb in the rural communities of Jordan. Methods: The present study evaluated the efficacy of the ethanol extract of this species on angiogenesis in both, ex vivo using rat aortic ring assay and in vivo using rat excision wound model. Results: In concentrations of 50 and 100 µg/ml, the ethanol extract showed angiogenic stimulatory effect and significantly increased length of capillary protrusions around aorta rings of about 60% in comparison to those of untreated aorta rings. In MCF-7 cells, the ethanol extract of A. fragrantissima stimulates the production of VEGF in a dose-dependent manner. 1% and 5% of ethanol extract of A. fragrantissima containing vaseline based ointment was applied on rat excision wounds for six days and was found to be effective in wound healing and maturation of the scar. Both preparations resulted in better wound healing when compared to the untreated control group and vaseline-treated group. This effect was comparable to that induced by MEBO, the positive control. Conclusion: The results indicate that A. fragrantissima has a pro-angiogenic effect, which may act through the VEGF signaling pathway.

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Histological, Biomechanical, and Biological Properties of Genipin-Crosslinked Decellularized Peripheral Nerves

International Journal of Molecular Sciences ◽

10.3390/ijms22020674 ◽

2021 ◽

Vol 22 (2) ◽

pp. 674

Author(s):

Óscar Darío García-García ◽

Marwa El Soury ◽

David González-Quevedo ◽

David Sánchez-Porras ◽

Jesús Chato-Astrain ◽

...

Keyword(s):

Ex Vivo ◽

Nerve Repair ◽

Wistar Rat ◽

Biomechanical Properties ◽

Biological Properties ◽

Suitable Alternative ◽

Promising Alternative ◽

Histological Pattern ◽

The Impact

Acellular nerve allografts (ANGs) represent a promising alternative in nerve repair. Our aim is to improve the structural and biomechanical properties of biocompatible Sondell (SD) and Roosens (RS) based ANGs using genipin (GP) as a crosslinker agent ex vivo. The impact of two concentrations of GP (0.10% and 0.25%) on Wistar rat sciatic nerve-derived ANGs was assessed at the histological, biomechanical, and biocompatibility levels. Histology confirmed the differences between SD and RS procedures, but not remarkable changes were induced by GP, which helped to preserve the nerve histological pattern. Tensile test revealed that GP enhanced the biomechanical properties of SD and RS ANGs, being the crosslinked RS ANGs more comparable to the native nerves used as control. The evaluation of the ANGs biocompatibility conducted with adipose-derived mesenchymal stem cells cultured within the ANGs confirmed a high degree of biocompatibility in all ANGs, especially in RS and RS-GP 0.10% ANGs. Finally, this study demonstrates that the use of GP could be an efficient alternative to improve the biomechanical properties of ANGs with a slight impact on the biocompatibility and histological pattern. For these reasons, we hypothesize that our novel crosslinked ANGs could be a suitable alternative for future in vivo preclinical studies.

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The impact of indole-3-lactic acid on immature intestinal innate immunity and development: a transcriptomic analysis

Scientific Reports ◽

10.1038/s41598-021-87353-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Wuyang Huang ◽

Ky Young Cho ◽

Di Meng ◽

W. Allan Walker

Keyword(s):

Lactic Acid ◽

Mechanistic Model ◽

Transcriptomic Analysis ◽

Dependent Manner ◽

Protective Effects ◽

Very Preterm Infants ◽

Anti Inflammatory ◽

The Impact

AbstractAn excessive intestinal inflammatory response may have a role in the pathogenesis of necrotizing enterocolitis (NEC) in very preterm infants. Indole-3-lactic acid (ILA) of breastmilk tryptophan was identified as the anti-inflammatory metabolite involved in probiotic conditioned media from Bifidobacteria longum subsp infantis. This study aimed to explore the molecular endocytic pathways involved in the protective ILA effect against inflammation. H4 cells, Caco-2 cells, C57BL/6 pup and adult mice were used to compare the anti-inflammatory mechanisms between immature and mature enterocytes in vitro and in vivo. The results show that ILA has pleiotropic protective effects on immature enterocytes including anti-inflammatory, anti-viral, and developmental regulatory potentials in a region-dependent and an age-dependent manner. Quantitative transcriptomic analysis revealed a new mechanistic model in which STAT1 pathways play an important role in IL-1β-induced inflammation and ILA has a regulatory effect on STAT1 pathways. These studies were validated by real-time RT-qPCR and STAT1 inhibitor experiments. Different protective reactions of ILA between immature and mature enterocytes indicated that ILA’s effects are developmentally regulated. These findings may be helpful in preventing NEC for premature infants.

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Uptake and Distribution of Administered Bone Marrow Mesenchymal Stem Cell Extracellular Vesicles in Retina

Cells ◽

10.3390/cells10040730 ◽

2021 ◽

Vol 10 (4) ◽

pp. 730

Author(s):

Biji Mathew ◽

Leianne A. Torres ◽

Lorea Gamboa Gamboa Acha ◽

Sophie Tran ◽

Alice Liu ◽

...

Keyword(s):

Stem Cell ◽

Mesenchymal Stem Cell ◽

Extracellular Vesicles ◽

Time Course ◽

Ganglion Cells ◽

Ex Vivo ◽

Dependent Manner ◽

Paracrine Effects

Cell replacement therapy using mesenchymal (MSC) and other stem cells has been evaluated for diabetic retinopathy and glaucoma. This approach has significant limitations, including few cells integrated, aberrant growth, and surgical complications. Mesenchymal Stem Cell Exosomes/Extracellular Vesicles (MSC EVs), which include exosomes and microvesicles, are an emerging alternative, promoting immunomodulation, repair, and regeneration by mediating MSC’s paracrine effects. For the clinical translation of EV therapy, it is important to determine the cellular destination and time course of EV uptake in the retina following administration. Here, we tested the cellular fate of EVs using in vivo rat retinas, ex vivo retinal explant, and primary retinal cells. Intravitreally administered fluorescent EVs were rapidly cleared from the vitreous. Retinal ganglion cells (RGCs) had maximal EV fluorescence at 14 days post administration, and microglia at 7 days. Both in vivo and in the explant model, most EVs were no deeper than the inner nuclear layer. Retinal astrocytes, microglia, and mixed neurons in vitro endocytosed EVs in a dose-dependent manner. Thus, our results indicate that intravitreal EVs are suited for the treatment of retinal diseases affecting the inner retina. Modification of the EV surface should be considered for maintaining EVs in the vitreous for prolonged delivery.

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Treatment of type 2 diabetic db/db mice with a novel PPARγ agonist improves cardiac metabolism but not contractile function

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00329.2003 ◽

2004 ◽

Vol 286 (3) ◽

pp. E449-E455 ◽

Cited By ~ 70

Author(s):

Andrew N. Carley ◽

Lisa M. Semeniuk ◽

Yakhin Shimoni ◽

Ellen Aasum ◽

Terje S. Larsen ◽

...

Keyword(s):

Contractile Function ◽

Ex Vivo ◽

Metabolic Changes ◽

Pparγ Agonist ◽

Perfused Hearts ◽

Type 2 Diabetic ◽

Contractile Performance ◽

Exogenous Substrates

Hearts from insulin-resistant type 2 diabetic db/db mice exhibit features of a diabetic cardiomyopathy with altered metabolism of exogenous substrates and reduced contractile performance. Therefore, the effect of chronic oral administration of 2-(2-(4-phenoxy-2-propylphenoxy)ethyl)indole-5-acetic acid (COOH), a novel ligand for peroxisome proliferator-activated receptor-γ that produces insulin sensitization, to db/db mice (30 mg/kg for 6 wk) on cardiac function was assessed. COOH treatment reduced blood glucose from 27 mM in untreated db/db mice to a normal level of 10 mM. Insulin-stimulated glucose uptake was enhanced in cardiomyocytes from COOH-treated db/db hearts. Working perfused hearts from COOH-treated db/db mice demonstrated metabolic changes with enhanced glucose oxidation and decreased palmitate oxidation. However, COOH treatment did not improve contractile performance assessed with ex vivo perfused hearts and in vivo by echocardiography. The reduced outward K+ currents in diabetic cardiomyocytes were still attenuated after COOH. Metabolic changes in COOH-treated db/db hearts are most likely indirect, secondary to changes in supply of exogenous substrates in vivo and insulin sensitization.

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Cord blood comprises antigen-experienced T cells specific for maternal minor histocompatibility antigen HA-1

Blood ◽

10.1182/blood-2004-07-2832 ◽

2005 ◽

Vol 105 (4) ◽

pp. 1823-1827 ◽

Cited By ~ 59

Author(s):

Bregje Mommaas ◽

Janine A. Stegehuis-Kamp ◽

Astrid G. van Halteren ◽

Michel Kester ◽

Jürgen Enczmann ◽

...

Keyword(s):

T Cells ◽

Cord Blood ◽

Ex Vivo ◽

Cytotoxic T Cells ◽

Cord Blood Transplantation ◽

Human Leukocyte ◽

Leukemic Cells ◽

Target Cells ◽

Graft Versus Leukemia

AbstractUmbilical cord blood transplantation is applied as treatment for mainly pediatric patients with hematologic malignancies. The clinical results show a relatively low incidence of graft-versus-host disease and leukemia relapse. Since maternal cells traffic into the fetus during pregnancy, we questioned whether cord blood has the potential to generate cytotoxic T cells specific for the hematopoietic minor histocompatibility (H) antigen HA-1 that would support the graft-versus-leukemia effect. Here, we demonstrate the feasibility of ex vivo generation of minor H antigen HA-1-specific T cells from cord blood cells. Moreover, we observed pre-existing HA-1-specific T cells in cord blood samples. Both the circulating and the ex vivo-generated HA-1-specific T cells show specific and hematopoietic restricted lysis of human leukocyte antigen-A2pos/HA-1pos (HLA-A2pos/HA-1pos) target cells, including leukemic cells. The cord blood-derived HA-1-specific cytotoxic T cells are from child origin. Thus, the so-called naive cord blood can comprise cytotoxic T cells directed at the maternal minor H antigen HA-1. The apparent immunization status of cord blood may well contribute to the in vivo graft-versus-leukemia activity after transplantation. Moreover, since the fetus cannot be primed against Y chromosome-encoded minor H antigens, cord blood is an attractive stem cell source for male patients. (Blood. 2005;105:1823-1827)

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Ex Vivo Mitochondrial Respiration Parallels Biochemical Response to Ibrutinib in CLL Cells

Cancers ◽

10.3390/cancers13020354 ◽

2021 ◽

Vol 13 (2) ◽

pp. 354

Author(s):

Subir Roy Chowdhury ◽

Cheryl Peltier ◽

Sen Hou ◽

Amandeep Singh ◽

James B. Johnston ◽

...

Keyword(s):

Mitochondrial Respiration ◽

Ex Vivo ◽

Standard Dose ◽

Cell Receptor ◽

Lymphocytic Leukemia ◽

Clinical Tool ◽

Potential Biomarker ◽

Apoptotic Pathways ◽

The Impact

Mitochondrial respiration is becoming more commonly used as a preclinical tool and potential biomarker for chronic lymphocytic leukemia (CLL) and activated B-cell receptor (BCR) signaling. However, respiration parameters have not been evaluated with respect to dose of ibrutinib given in clinical practice or the effect of progression on ibrutinib treatment on respiration of CLL cells. We evaluated the impact of low and standard dose ibrutinib on CLL cells from patients treated in vivo on mitochondrial respiration using Oroboros oxygraph. Cytokines CCL3 and CCL4 were evaluated using the Mesoscale. Western blot analysis was used to evaluate the BCR and apoptotic pathways. We observed no difference in the mitochondrial respiration rates or levels of plasma chemokine (C-C motif) ligands 3 and 4 (CCL3/CCL4), β-2 microglobulin (β-2 M) and lactate dehydrogenase (LDH) between low and standard doses of ibrutinib. This may confirm why clinical observations of the safety and efficacy of low dose ibrutinib are observed in practice. Of interest, we also observed that the mitochondrial respiration of CLL cells paralleled the increase in β-2 M and LDH at progression. Our study further supports mitochondrial respiration as a biomarker for response and progression on ibrutinib in CLL cells and a valuable pre-clinical tool.

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