The differential regulation of placenta trophoblast bisphosphoglycerate mutase in fetal growth restriction: preclinical study in mice and observational histological study of human placenta.

Background: Fetal growth restriction (FGR) is a pregnancy complication in which a newborn fails to achieve its growth potential, increasing the risk of perinatal morbidity and mortality. Chronic maternal gestational hypoxia, as well as placental insufficiency are associated with increased FGR incidence; however, the molecular mechanisms underlying FGR remain unknown. Methods: Pregnant mice were subjected to acute or chronic hypoxia (12.5% O2) resulting in reduced fetal weight. Placenta oxygen transport was assessed by blood oxygenation level dependent (BOLD) contrast magnetic resonance imaging (MRI). The placentae were analyzed via immunohistochemistry and in situ hybridization. Human placentae were selected from FGR and matched controls and analyzed by immunohistochemistry (IHC). Maternal and cord sera were analyzed by mass spectrometry. Results: We show that murine acute and chronic gestational hypoxia recapitulates FGR phenotype and affects placental structure and morphology. Gestational hypoxia decreased labyrinth area, increased the incidence of red blood cells (RBCs) in the labyrinth while expanding the placental spiral arteries (SpA) diameter. Hypoxic placentae exhibited higher hemoglobin-oxygen affinity compared to the control. Placental abundance of Bisphosphoglycerate mutase (BPGM) was upregulated in the syncytiotrophoblast and spiral artery trophoblast cells (SpA TGCs) in the murine gestational hypoxia groups compared to the control. Hif1α levels were higher in the acute hypoxia group compared to the control. In contrast, human FGR placentae exhibited reduced BPGM levels in the syncytiotrophoblast layer compared to placentae from healthy uncomplicated pregnancies. Levels of 2,3 BPG, the product of BPGM, were lower in cord serum of human FGR placentae compared to control. Polar expression of BPGM was found in both human and mouse placentae syncytiotrophoblast, with higher expression facing the maternal circulation. Moreover, in the murine SpA TGCs expression of BPGM was concentrated exclusively in the apical cell side, in direct proximity to the maternal circulation. Conclusions: This study suggests a possible involvement of placental BPGM in maternal-fetal oxygen transfer, and in the pathophysiology of FGR. Funding: This work was supported by the Weizmann Krenter Foundation and the Weizmann - Ichilov (Tel Aviv Sourasky Medical Center) Collaborative Grant in Biomedical Research, by the Minerva Foundation, by the ISF KillCorona grant 3777/19.

Bisphosphoglycerate mutase predicts myocardial dysfunction and adverse outcome in sepsis: an observational cohort study.

BACKGROUND: Sepsis not only causes inflammation, but also damages the heart and increases the risk of death. The glycolytic pathway plays a crucial role in the pathogenesis of sepsis-induced cardiac injury. This study aims to investigate the value of bisphosphoglycerate mutase (BPGM), an intermediate in the glycolytic pathway, in evaluating cardiac injury in septic patients and predicting poor prognosis in sepsis. METHODS: This prospective study included 85 patients with sepsis. Serum BPGM was measured at the time of enrollment, and the patients were divided into a BPGM-positive group (n = 35) and a BPGM-negative group (n = 50) according to their serum BPGM levels. Baseline clinical and echocardiographic parameters, and clinical outcomes were analyzed and compared between the two groups. Kaplan-Meier analysis was used to compare the 28-day survival rate between BPGM-negative and BPGM-positive patients. Multivariate logistic regression analysis was conducted to explore the independent risk factors for 28-day mortality in septic patients. The predictive value of serum BPGM for sepsis-induced myocardial injury and poor prognosis in sepsis was evaluated using receiver operating characteristic (ROC)curve analysis. RESULT: The serum level of BPGM was significantly higher in patients who died within 28 days compared to survivors (p < 0.001). Kaplan-Meier analysis showed that serum BPGM-positive sepsis patients had a significantly shorter 28-day survival time (p < 0.001). Multivariate logistic regression analysis showed that serum BPGM (OR = 9.853, 95%CI 1.844-52.655, p = 0.007) and left ventricular ejection fraction-simpson(LVEF-S) (OR = 0.032, 95% CI 0.002-0.43, p = 0.009) were independent risk factors for 28-day mortality in sepsis patients. Furthermore, BPGM levels was negatively correlated with LVEF-S (p = 0.005) and positively correlated with the myocardial performance (Tei) index (p < 0.001) in sepsis patients. ROC curve analysis showed that serum BPGM was a good predictor of septic myocardial injury and 28-day mortality in sepsis patients. CONCLUSION: The level of BPGM in the serum of sepsis patients can serve as a monitoring indicator for myocardial injury, with its high level indicating the occurrence of secondary myocardial injury events and adverse outcomes in sepsis patients.

Enhanced BPGM/2,3-DPG pathway activity suppresses glycolysis in hypoxic astrocytes via FIH-1 and TET2.

OBJECTIVE: Bisphosphoglycerate mutase (BPGM) is expressed in human erythrocytes and responsible for the production of 2,3-bisphosphoglycerate (2,3-DPG). However, the expression and role of BPGM in other cells have not been reported. In this work, we found that BPGM was significantly upregulated in astrocytes upon acute hypoxia, and the role of this phenomenon will be clarified in the following report. METHODS: The mRNA and protein expression levels of BPGM and the content of 2,3-DPG with hypoxia treatment were determined in vitro and in vivo. Furthermore, glycolysis was evaluated upon in hypoxic astrocytes with BPGM knockdown and in normoxic astrocytes with BPGM overexpression or 2,3-DPG treatment. To investigate the mechanism by which BPGM/2,3-DPG regulated glycolysis in hypoxic astrocytes, we detected the expression of HIF-1α, FIH-1 and TET2 with silencing or overexpression of BPGM and 2,3-DPG treatment. RESULTS: The expression of glycolytic genes and the capacity of lactate markedly increased with 6 h, 12 h, 24 h, 36 h and 48 h 1 % O2 hypoxic treatment in astrocytes. The expression of BPGM was upregulated, and the production of 2,3-DPG was accelerated upon hypoxia. Moreover, when BPGM expression was knocked down, glycolysis was promoted in HEB cells. However, overexpression of BPGM and addition of 2,3-DPG to the cellular medium in normoxic cells could downregulate glycolytic genes. Furthermore, HIF-1α and TET2 exhibited higher expression levels and FIH-1 showed a lower expression level upon BPGM silencing, while these changes were reversed under BPGM overexpression and 2,3-DPG treatment. CONCLUSIONS: Our study revealed that the BPGM/2,3-DPG pathway presented a suppressive effect on glycolysis in hypoxic astrocytes by negatively regulating HIF-1α and TET2.

Heterozygosity for bisphosphoglycerate mutase deficiency expressing clinically as congenital erythrocytosis: A case series and literature review.

Erythrocytosis is associated with increased red blood cell mass and can be either congenital or acquired. Congenital secondary causes are rare and include germline variants increasing haemoglobin (Hb)-oxygen affinity (e.g., Hb or bisphosphoglycerate mutase (BPGM) variants) or affecting oxygen-sensing pathway proteins. Here, we describe five adults from three kindreds with erythrocytosis associated with heterozygosity for BPGM variants, including one novel. Functional analyses showed partial BPGM deficiency, reduced 2,3-bisphosphoglycerate levels and/or increased Hb-oxygen affinity. We also review currently known BPGM variants. This study contributes to raising awareness of BPGM variants, and in particular that heterozygosity for BPGM deficiency may already manifest clinically.

Molecular insight into 2-phosphoglycolate activation of the phosphatase activity of bisphosphoglycerate mutase.

Bisphosphoglycerate mutase (BPGM) is an erythrocyte-specific multifunctional enzyme that is responsible for the regulation of 2,3-bisphosphoglycerate (2,3-BPG) in red blood cells through its synthase and phosphatase activities; the latter enzymatic function is stimulated by the endogenous activator 2-phosphoglycolate (2-PG). 2,3-BPG is a natural allosteric effector of hemoglobin (Hb) that is responsible for decreasing the affinity of Hb for oxygen to facilitate tissue oxygenation. Here, crystal structures of BPGM with 2-PG in the presence and absence of 3-phosphoglycerate are reported at 2.25 and 2.48 Šresolution, respectively. Structure analysis revealed a new binding site for 2-PG at the dimer interface for the first time, in addition to the expected active-site binding. Also, conformational non-equivalence of the two active sites was observed as one of the sites was found in an open conformation, with the residues at the active-site entrance, including Arg100, Arg116 and Arg117, and the C-terminus disordered. The kinetic result is consistent with the binding of 2-PG to an allosteric or noncatalytic site as well as the active site. This study paves the way for the rational targeting of BPGM for therapeutic purposes, especially for the treatment of sickle cell disease.

Screening of activators of 2,3-diphosphoglycerate mutase from traditional Chinese herb medicines.

OBJECTIVE: To screen activators of 2,3-diphosphoglycerate (BPG) mutase (BPGM) from Chinese herb medicines, so as to improve the hypoxia tolerance of erythrocytes. METHODS: BPGM was used as the receptor and Chinese medicine ingredients database was used as the ligand in the study. After Lipinski rule of five screening, LibDock and CDOCKER docking were used for virtual screening. The effect of the screened compounds on the affinity of BPGM in erythrocytes was verified. Finally, the erythrocytes were incubated in vitro to establish the erythrocyte hypoxia model, and the effect of the compound on the activity of BPGM in the erythrocyte hypoxia model was verified. RESULTS: Ten compounds with highest binding affinity to BPGM were selected by LibDock and CDOCKER, and the cytoplasm protein was incubated with the ten compounds. Compared with blank control group, methyl rosmarinate group, dihydrocurcumin high dose group, octahydrocurcumin medium dose group and coniferyl ferulate high dose group were able to further activate the BPGM, significantly increase the levels of 2, 3-BPG in normal erythrocytes (all P<0.05); while the low dose of tetrahydrocurcumin, high dose and low dose of aurantiamide, hexahydrocurcumin and medium dose of N- (p-coumaroyl) serotonin had a tendency to increase the contents of 2,3-BPG in normal erythrocytes (all P>0.05). In the hypoxic red blood cells, the medium dose methyl rosmarinate, medium dose octahydrocurcumin, high dose hexahydrocurcumin and medium dose N-(p-coumaroyl) serotonin could significantly increase the contents of 2,3-BPG (all P<0.05). CONCLUSION: The methyl rosmarinate, octahydrocurcumin, hexahydrocurcumin and N-(p-coumaroyl) serotonin could activate BPGM and increase the contents of 2,3-BPG in hypoxic erythrocytes.

Erythrocyte adenosine A2B receptor prevents cognitive and auditory dysfunction by promoting hypoxic and metabolic reprogramming.

Hypoxia drives aging and promotes age-related cognition and hearing functional decline. Despite the role of erythrocytes in oxygen (O2) transport, their role in the onset of aging and age-related cognitive decline and hearing loss (HL) remains undetermined. Recent studies revealed that signaling through the erythrocyte adenosine A2B receptor (ADORA2B) promotes O2 release to counteract hypoxia at high altitude. However, nothing is known about a role for erythrocyte ADORA2B in age-related functional decline. Here, we report that loss of murine erythrocyte-specific ADORA2B (eAdora2b-/-) accelerates early onset of age-related impairments in spatial learning, memory, and hearing ability. eAdora2b-/- mice display the early aging-like cellular and molecular features including the proliferation and activation of microglia and macrophages, elevation of pro-inflammatory cytokines, and attenuation of hypoxia-induced glycolytic gene expression to counteract hypoxia in the hippocampus (HIP), cortex, or cochlea. Hypoxia sufficiently accelerates early onset of cognitive and cochlear functional decline and inflammatory response in eAdora2b-/- mice. Mechanistically, erythrocyte ADORA2B-mediated activation of AMP-activated protein kinase (AMPK) and bisphosphoglycerate mutase (BPGM) promotes hypoxic and metabolic reprogramming to enhance production of 2,3-bisphosphoglycerate (2,3-BPG), an erythrocyte-specific metabolite triggering O2 delivery. Significantly, this finding led us to further discover that murine erythroblast ADORA2B and BPGM mRNA levels and erythrocyte BPGM activity are reduced during normal aging. Overall, we determined that erythrocyte ADORA2B-BPGM axis is a key component for anti-aging and anti-age-related functional decline.

Dysregulation of bisphosphoglycerate mutase during in vitro maturation of oocytes.

PURPOSE: Oxygen is vital for oocyte maturation; however, oxygen regulation within ovarian follicles is not fully understood. Hemoglobin is abundant within the in vivo matured oocyte, indicating potential function as an oxygen regulator. However, hemoglobin is significantly reduced following in vitro maturation (IVM). The molecule 2,3-bisphosphoglycerate (2,3-BPG) is essential in red blood cells, facilitating release of oxygen from hemoglobin. Towards understanding the role of 2,3-BPG in the oocyte, we characterized gene expression and protein abundance of bisphosphoglycerate mutase (Bpgm), which synthesizes 2,3-BPG, and whether this is altered under low oxygen or hemoglobin addition during IVM. METHODS: Hemoglobin and Bpgm expression within in vivo matured human cumulus cells and mouse cumulus-oocyte complexes (COCs) were evaluated to determine physiological levels of Bpgm. During IVM, Bpgm gene expression and protein abundance were analyzed in the presence or absence of low oxygen (2% and 5% oxygen) or exogenous hemoglobin. RESULTS: The expression of Bpgm was significantly lower than hemoglobin when mouse COCs were matured in vivo. Following IVM at 20% oxygen, Bpgm gene expression and protein abundance were significantly higher compared to in vivo. At 2% oxygen, Bpgm was significantly higher compared to 20% oxygen, while exogenous hemoglobin resulted in significantly lower Bpgm in the COC. CONCLUSION: Hemoglobin and 2,3-BPG may play a role within the maturing COC. This study shows that IVM increases Bpgm within COCs compared to in vivo. Decreasing oxygen concentration and the addition of hemoglobin altered Bpgm, albeit not to levels observed in vivo.

Congenital erythrocytosis.

Erythrocytosis, or increased red cell mass, may be labeled as primary or secondary, depending on whether the molecular defect is intrinsic to the red blood cells/their precursors or extrinsic to them, the latter being typically associated with elevated erythropoietin (EPO) levels. Inherited/congenital erythrocytosis (CE) of both primary and secondary types is increasingly recognized as the cause in many patients in whom acquired, especially neoplastic causes have been excluded. During the past two decades, the underlying molecular mechanisms of CE are increasingly getting unraveled. Gain-in-function mutations in the erythropoietin receptor gene were among the first to be characterized in a disorder termed primary familial and congenital polycythemia. Another set of mutations affect the components of the oxygen-sensing pathway. Under normoxic conditions, the hypoxia-inducible factor (HIF), upon hydroxylation by the prolyl-4-hydroxylase domain protein 2 (PHD2) enzyme, is degraded by the von Hippel-Lindau protein. In hypoxic conditions, failure of prolyl hydroxylation leads to stabilization of HIF and activation of the EPO gene. CE has been found to be caused by loss-of-function mutations in VHL and PHD2/EGLN1 as well as gain-of-function mutations in HIF-2α (EPAS1), all resulting in constitutive activation of EPO signaling. Apart from these, globin gene mutations leading to formation of high oxygen affinity hemoglobins also cause CE. Rarely, bisphosphoglycerate mutate mutations, affecting the 2,3-bisphosphoglycerate levels, can increase the oxygen affinity of hemoglobin and cause CE. This narrative review examines the current mutational spectrum of CE and the distinctive pathogenetic mechanisms that give rise to this increasingly recognized condition in various parts of the world.

Hydrogen Sulfide Is a Regulator of Hemoglobin Oxygen-Carrying Capacity via Controlling 2,3-BPG Production in Erythrocytes.

Hydrogen sulfide (H2S) is naturally synthesized in a wide range of mammalian tissues. Whether H2S is involved in the regulation of erythrocyte functions remains unknown. Using mice with a genetic deficiency in a H2S natural synthesis enzyme cystathionine-γ-lyase (CSE) and high-throughput metabolomic profiling, we found that levels of erythrocyte 2,3-bisphosphoglycerate (2,3-BPG), an erythroid-specific metabolite negatively regulating hemoglobin- (Hb-) oxygen (O2) binding affinity, were increased in CSE knockout (Cse -/-) mice under normoxia. Consistently, the 50% oxygen saturation (P50) value was increased in erythrocytes of Cse -/- mice. These effects were reversed by treatment with H2S donor GYY4137. In the models of cultured mouse and human erythrocytes, we found that H2S directly acts on erythrocytes to decrease 2,3-BPG production, thereby enhancing Hb-O2 binding affinity. Mouse genetic studies showed that H2S produced by peripheral tissues has a tonic inhibitory effect on 2,3-BPG production and consequently maintains Hb-O2 binding affinity in erythrocytes. We further revealed that H2S promotes Hb release from the membrane to the cytosol and consequently enhances bisphosphoglycerate mutase (BPGM) anchoring to the membrane. These processes might be associated with S-sulfhydration of Hb. Moreover, hypoxia decreased the circulatory H2S level and increased the erythrocyte 2,3-BPG content in mice, which could be reversed by GYY4137 treatment. Altogether, our study revealed a novel signaling pathway that regulates oxygen-carrying capacity in erythrocytes and highlights a previously unrecognized role of H2S in erythrocyte 2,3-BPG production.

Bisphosphoglycerate Mutase Deficiency Protects against Cerebral Malaria and Severe Malaria-Induced Anemia.

The replication cycle and pathogenesis of the Plasmodium malarial parasite involves rapid expansion in red blood cells (RBCs), and variants of certain RBC-specific proteins protect against malaria in humans. In RBCs, bisphosphoglycerate mutase (BPGM) acts as a key allosteric regulator of hemoglobin/oxyhemoglobin. We demonstrate here that a loss-of-function mutation in the murine Bpgm (BpgmL166P) gene confers protection against both Plasmodium-induced cerebral malaria and blood-stage malaria. The malaria protection seen in BpgmL166P mutant mice is associated with reduced blood parasitemia levels, milder clinical symptoms, and increased survival. The protective effect of BpgmL166P involves a dual mechanism that enhances the host's stress erythroid response to Plasmodium-driven RBC loss and simultaneously alters the intracellular milieu of the RBCs, including increased oxyhemoglobin and reduced energy metabolism, reducing Plasmodium maturation, and replication. Overall, our study highlights the importance of BPGM as a regulator of hemoglobin/oxyhemoglobin in malaria pathogenesis and suggests a new potential malaria therapeutic target.

Analysis of plasma metabolic profile, characteristics and enzymes in the progression from chronic hepatitis B to hepatocellular carcinoma.

Hepatitis B virus (HBV) infection is an important factor causing hepatocellular carcinoma (HCC). The aim of this study was to investigate the metabolic characteristics and related metabolic enzyme changes during the progression from chronic hepatitis B (CHB) to liver cirrhosis (LC) and, ultimately, to HCC. An untargeted metabolomics assay was performed in plasma from 50 healthy volunteers, 43 CHB patients, 67 LC patients, and 39 HCC patients. A total of 24 differential metabolites (DMs) were identified. Joint pathway analysis suggested striking changes in amino acid metabolism and lipid metabolism from CHB to HCC. The panel of L-serine, creatine and glycine distinguished LC from CHB, and L-serine, cystathionine, creatine and linoleic acid distinguished HCC from LC. Bioinformatic analysis of publicly available data showed that differential metabolite profile-associated enzyme genes, including alanine-glyoxylate aminotransferase-2 (AGXT2), D-amino-acid oxidase (DAO), and cystathionine gamma-lyase (CTH), were downregulated, while bisphosphoglycerate mutase (BPGM), cystathionine-ß-synthase (CBS), phosphoserine phosphatase (PSPH) and acyl-CoA thioesterase 7 (ACOT7) were upregulated, in HCC, all of which correlated with a poor prognosis for HCC patients. Our results indicated that serum metabolites and related enzymes are of considerable significance for the diagnosis and prognosis of HCC and can provide a theoretical basis and therapeutic index for future diagnosis and treatment.

Sickling-suppressive effects of chrysin may be associated with sequestration of deoxy-haemoglobin, 2,3-bisphosphoglycerate mutase, alteration of redox homeostasis and functional chemistry of sickle erythrocytes.

Sickle cell disease (SCD) is a medical condition caused by mutation in a single nucleotide in the ß-globin gene. It is a health problem for people in sub-Saharan Africa, the Middle East and India. Orthodox drugs developed so far for SCD focus largely on symptomatic respite of pain and crisis mitigation. We investigated the antisickling effects of chrysin via modulation of deoxy-haemoglobin, 2,3-bisphosphoglycerate mutase, redox homeostasis and alteration of functional chemistry in human sickle erythrocytes. In silico and in vitro methods were adopted for the studies. Chrysin was docked against deoxy-haemoglobin and 2,3-bisphosphoglycerate mutase, with binding energies (-24.064 and -18.171 kcal/mol) and inhibition constant (K i) of 0.990 µM and 0.993 µM at their active sites through strong hydrophobic and hydrogen bond interactions. Sickling was induced with 2% metabisulphite at 3 h. Chrysin was able to prevent sickling maximally at 2.5 µg/mL and reversed the same at 12.5 µg/mL, by 66.5% and 69.6%, respectively. Treatment with chrysin significantly (p < 0.05) re-established the integrity of erythrocytes membrane as evident from the observed percentage of haemolysis relative to induced erythrocytes. Chrysin also significantly (p < 0.05) prevented and reversed lipid peroxidation. Similarly, glutathione and catalase levels were observed to significantly (p < 0.05) increase with concomitant significant (p < 0.05) decrease in superoxide dismutase activity relative to untreated. From Fourier-transform infrared results, treatment with chrysin was able to favourably alter the functional chemistry, judging from the shifts and functional groups observed. Sickling-suppressive effects of chrysin may therefore be associated with sequestration of deoxy-haemoglobin, 2,3-bisphosphoglycerate mutase, alteration of redox homeostasis and functional chemistry of sickle erythrocytes.

Beneficial Role of Erythrocyte Adenosine A2B Receptor-Mediated AMP-Activated Protein Kinase Activation in High-Altitude Hypoxia.

BACKGROUND: High altitude is a challenging condition caused by insufficient oxygen supply. Inability to adjust to hypoxia may lead to pulmonary edema, stroke, cardiovascular dysfunction, and even death. Thus, understanding the molecular basis of adaptation to high altitude may reveal novel therapeutics to counteract the detrimental consequences of hypoxia. METHODS: Using high-throughput, unbiased metabolomic profiling, we report that the metabolic pathway responsible for production of erythrocyte 2,3-bisphosphoglycerate (2,3-BPG), a negative allosteric regulator of hemoglobin-O2 binding affinity, was significantly induced in 21 healthy humans within 2 hours of arrival at 5260 m and further increased after 16 days at 5260 m. RESULTS: This finding led us to discover that plasma adenosine concentrations and soluble CD73 activity rapidly increased at high altitude and were associated with elevated erythrocyte 2,3-BPG levels and O2 releasing capacity. Mouse genetic studies demonstrated that elevated CD73 contributed to hypoxia-induced adenosine accumulation and that elevated adenosine-mediated erythrocyte A2B adenosine receptor activation was beneficial by inducing 2,3-BPG production and triggering O2 release to prevent multiple tissue hypoxia, inflammation, and pulmonary vascular leakage. Mechanistically, we demonstrated that erythrocyte AMP-activated protein kinase was activated in humans at high altitude and that AMP-activated protein kinase is a key protein functioning downstream of the A2B adenosine receptor, phosphorylating and activating BPG mutase and thus inducing 2,3-BPG production and O2 release from erythrocytes. Significantly, preclinical studies demonstrated that activation of AMP-activated protein kinase enhanced BPG mutase activation, 2,3-BPG production, and O2 release capacity in CD73-deficient mice, in erythrocyte-specific A2B adenosine receptor knockouts, and in wild-type mice and in turn reduced tissue hypoxia and inflammation. CONCLUSIONS: Together, human and mouse studies reveal novel mechanisms of hypoxia adaptation and potential therapeutic approaches for counteracting hypoxia-induced tissue damage.

Microarray and Co-expression Network Analysis of Genes Associated with Acute Doxorubicin Cardiomyopathy in Mice.

Clinical use of doxorubicin (DOX) in cancer therapy is limited by its dose-dependent cardiotoxicity. But molecular mechanisms underlying this phenomenon have not been well defined. This study was to investigate the effect of DOX on the changes of global genomics in hearts. Acute cardiotoxicity was induced by giving C57BL/6J mice a single intraperitoneal injection of DOX (15 mg/kg). Cardiac function and apoptosis were monitored using echocardiography and TUNEL assay at days 1, 3 and 5. Myocardial glucose and ATP levels were measured. Microarray assays were used to screen gene expression profiles in the hearts at day 5, and the results were confirmed with qPCR analysis. DOX administration caused decreased cardiac function, increased cardiomyocyte apoptosis and decreased glucose and ATP levels. Microarrays showed 747 up-regulated genes and 438 down-regulated genes involved in seven main functional categories. Among them, metabolic pathway was the most affected by DOX. Several key genes, including 2,3-bisphosphoglycerate mutase (Bpgm), hexokinase 2, pyruvate dehydrogenase kinase, isoenzyme 4 and fructose-2,6-bisphosphate 2-phosphatase, are closely related to glucose metabolism. Gene co-expression networks suggested the core role of Bpgm in DOX cardiomyopathy. These results obtained in mice were further confirmed in cultured cardiomyocytes. In conclusion, genes involved in glucose metabolism, especially Bpgm, may play a central role in the pathogenesis of DOX-induced cardiotoxicity.

Hereditary erythrocytosis, thrombocytosis and neutrophilia.

Hereditary erythrocytosis, thrombocytosis, and neutrophilia are rare inherited syndromes which exhibit Mendelian inheritance. Some patients with primary hereditary erythrocytosis exhibit a mutation in the erythropoietin receptor (EPOR) which is associated with low serum erythropoietin (EPO) levels. Secondary congenital erythrocytosis may be characterized by normal or high serum EPO levels, and is related to high oxygen affinity haemoglobin variants, mutation of the enzyme biphosphoglycerate mutase (BPGM), or defects in components of the oxygen-sensing pathway. Hereditary thrombocytosis was first linked to mutations in genes encoding thrombopoietin (THPO) or the thrombopoietin receptor, MPL. More recently, germline mutations in JAK2, distinct from JAK2 V617F, and mutation of the gelsolin gene, were uncovered in several pedigrees of hereditary thrombocytosis. Hereditary neutrophilia has been described in one family with an activating germline mutation in CSF3R. The mutational basis for most hereditary myeloproliferative disorders has yet to be identified.

Insights into the phosphatase and the synthase activities of human bisphosphoglycerate mutase: a quantum mechanics/molecular mechanics simulation.

Bisphosphoglycerate mutase (BPGM) is a multi-activity enzyme. Its main function is to synthesize the 2,3-bisphosphoglycerate, the allosteric effector of hemoglobin. This enzyme can also catalyze the 2,3-bisphosphoglycerate to the 3-phosphoglycerate. In this study, the reaction mechanisms of both the phosphatase and the synthase activities of human bisphosphoglycerate mutase were theoretically calculated by using the quantum mechanics/molecular mechanics method based on the metadynamics and umbrella sampling simulations. The simulation results not only show the free energy curve of the phosphatase and the synthase reactions, but also reveal the important role of some residues in the active site. Additionally, the energy barriers of the two reactions indicate that the activity of the synthase in human bisphosphoglycerate mutase is much higher than that of the phosphatase. The estimated reaction barriers are consistent with the experimental data. Therefore, our work can give important information to understand the catalytic mechanism of the bisphosphoglycerate mutase family.

Usefulness of material recovered from distal embolic protection devices after carotid angioplasty for proteomic studies.

PURPOSE: To demonstrate the usefulness of biologic material obtained from distal embolic protection devices (DEPDs) used in carotid angioplasty for the study of atherosclerosis protein markers and to establish the effect of systemic inflammation on the protein expression of carotid atheromatous plaques. MATERIALS AND METHODS: Two-dimensional gel electrophoresis and mass spectrometry were used to study proteins obtained from debris captured in DEPDs from patients who underwent carotid angioplasty. In addition, protein expression obtained from angioplasty samples in patients with different types of systemic inflammation (measured by serum levels of high-sensitivity C-reactive protein [CRP] with a cutoff value of 3 mg/L) was compared. Finally, immunohistochemistry of atherosclerotic plaques obtained by endarterectomy was used to validate the results obtained using DEPDs. RESULTS: Proteomic studies were successfully performed using debris from DEPDs. Protein expression differences were found in debris from patients with high systemic inflammation compared with debris from patients with low systemic inflammation. Annexin A5 (ANXA5), haptoglobin precursor, purine nucleoside phosphorylase, transgelin-2 (TAGLN2), and bisphosphoglycerate mutase were upregulated in debris from patients with high systemic inflammation, and proteasome subunit 8 beta type and glutathione-S-transferase kappa 1 (GSTK1) levels were higher in debris from patients with low levels of systemic inflammation. CONCLUSIONS: Atherosclerotic plaque debris captured in DEPDs is a suitable and valid source of material for proteomic studies of atherosclerosis. Protein expression in DEPD debris is affected by systemic inflammation.