Platelet-leukocyte interactions on activated endothelial cells play an important role during microvascular occlusion under oxidative stress conditions. However, it remains poorly understood how ...neutrophil-platelet interactions are regulated during vascular inflammation. By using intravital microscopy with mice lacking nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) and their bone marrow chimera, we demonstrated that NOX2 from both hematopoietic and endothelial cells is crucial for neutrophil-platelet interactions during tumor necrosis factor alpha-induced venular inflammation. Platelet NOX2-produced reactive oxygen species (ROS) regulated P-selectin exposure upon agonist stimulation and the ligand-binding function of glycoprotein Ibα. Furthermore, neutrophil NOX2-generated ROS enhanced the activation and ligand-binding activity of αMβ2 integrin following N-formyl-methionyl-leucyl phenylalanine stimulation. Studies with isolated cells and a mouse model of hepatic ischemia/reperfusion injury revealed that NOX2 from both platelets and neutrophils is required for cell-cell interactions, which contribute to the pathology of hepatic ischemia/reperfusion injury. Platelet NOX2 modulated intracellular Ca2+ release but not store-operated Ca2+ entry (SOCE), whereas neutrophil NOX2 was crucial for SOCE but not intracellular Ca2+ release. Different regulation of Ca2+ signaling by platelet and neutrophil NOX2 correlated with differences in the phosphorylation of AKT, ERK, and p38MAPK. Our results indicate that platelet and neutrophil NOX2-produced ROS are critical for the function of surface receptors essential for neutrophil-platelet interactions during vascular inflammation.
•NOX2-generated ROS regulate the function of surface receptors required for platelet-neutrophil interactions during vascular inflammation.
Mutations in the isocitrate dehydrogenase-1 gene (IDH1) are common drivers of acute myeloid leukemia (AML) but their mechanism is not fully understood. It is thought that IDH1 mutants act by ...inhibiting TET2 to alter DNA methylation, but there are significant unexplained clinical differences between IDH1- and TET2-mutant diseases. We have discovered that mice expressing endogenous mutant IDH1 have reduced numbers of hematopoietic stem cells (HSCs), in contrast to Tet2 knockout (TET2-KO) mice. Mutant IDH1 downregulates the DNA damage (DD) sensor ATM by altering histone methylation, leading to impaired DNA repair, increased sensitivity to DD, and reduced HSC self-renewal, independent of TET2. ATM expression is also decreased in human IDH1-mutated AML. These findings may have implications for treatment of IDH-mutant leukemia.
•Mutant IDH1 decreases hematopoietic stem cell (HSC) number and impairs self-renewal•Mutant IDH1 causes TET2-independent downregulation of ATM via methylation of H3K9•Mutant IDH1 causes accumulation of DNA damage and impairs DNA repair in HSCs•Mutant IDH1 increases HSC sensitivity to radiation and daunorubicin
Although strong evidence supports that IDH1 mutants act by inhibiting TET2 in hematological malignancies, there are clear clinical differences between mutations of these genes. Inoue et al. show that mutant IDH1 decreases ATM independent of TET2, leading to impaired DNA repair and reduced hematopoietic stem cells.
Deregulated Skp2 function promotes cell transformation, and this is consistent with observations of Skp2 overexpression in many human cancers. However, the mechanisms underlying elevated Skp2 ...expression are still unknown. Here we show that the serine/threonine protein kinase Akt1, but not Akt2, directly controls Skp2 stability by a mechanism that involves degradation by the APC-Cdh1 ubiquitin ligase complex. We show further that Akt1 phosphorylates Skp2 at Ser 72, which is required to disrupt the interaction between Cdh1 and Skp2. In addition, we show that Ser 72 is localized within a putative nuclear localization sequence and that phosphorylation of Ser 72 by Akt leads to cytoplasmic translocation of Skp2. This finding expands our knowledge of how specific signalling kinase cascades influence proteolysis governed by APC-Cdh1 complexes, and provides evidence that elevated Akt activity and cytoplasmic Skp2 expression may be causative for cancer progression.
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Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Recently, it has been shown that multiple mammalian cell types express daily rhythms in vitro. Although the suprachiasmatic nucleus (SCN) of the hypothalamus is known to regulate a wide range of ...circadian behaviors, the role for intrinsic rhythmicity in other tissues is unknown. We tested whether the main olfactory bulb (OB) of mice mediates daily changes in olfaction. We found circadian rhythms in cedar oil-induced c-Fos, a protein marker of cellular excitation, in the mitral and granular layers of the OB and in the piriform cortex (PC). These oscillations persisted in constant darkness with a fourfold change in amplitude and a peak approximately 4 h after the onset of daily locomotor activity. Electrolytic lesions of the SCN abolished circadian locomotor rhythms, but not odor-induced c-Fos rhythms in the OB or PC. Furthermore, removal of the OB abolished spontaneous circadian cycling of c-Fos in the PC, shortened the free-running period of locomotor rhythms, and accelerated re-entrainment after a 6 h advance and slowed re-entrainment after a 6 h delay in the light schedule. OB ablation or odorant altered the amplitude of c-Fos rhythms in the SCN and ablation of one OB abolished c-Fos rhythms in the ipsilateral PC, but not in the contralateral OB and PC. We conclude that the OB comprises a master circadian pacemaker, which enhances olfactory responsivity each night, drives rhythms in the PC, and interacts with the SCN to coordinate other daily behaviors.
Mesenchymal stem cells (MSCs), as well as osteoblastic cells derived from these MSCs, have been shown to be key components of the hematopoietic stem cell (HSC) niche. In this study, we wished to ...examine whether other cell types that are known to differentiate from MSCs similarly regulate the stem cell niche, namely cells of the adipocyte lineage. Recent studies have examined the role that adipocytes play in the biology of the HSCs in different bone locations and in transplantation settings; however, none have examined their role under homeostatic conditions. We compared the ability of adipocytic and nonadipocytic cell lines to support primitive hematopoietic cells in vitro. Preadipocytic cell lines demonstrated enhanced support of hematopoietic cells. Similarly, primary bone marrow (BM) cells treated with troglitazone, a drug that enhances adipogenesis, also demonstrated augmented support over control-treated stromal cells. We further examined the effects of increased adipocyte number in vivo under homeostatic conditions using troglitazone treatment and found that these alterations had no effect on HSC frequency. Taken together, we demonstrate that cells of the adipocyte lineage promote the ability of stromal cells to support primitive hematopoietic cells in vitro, yet alterations of adipocyte number and volume in vivo have no effect. These data suggest that adipocytes are not a component of the adult BM HSC niche under homeostatic conditions.
Adherent neutrophils on vascular endothelium positively contribute to cell-cell aggregation and vaso-occlusion in sickle cell disease. In the present study, we demonstrated that pyridoxamine, a ...derivative of vitamin B6, might be a therapeutic agent to alleviate intravascular cell-cell aggregation in sickle cell disease. Using real-time intravital microscopy, we found that one oral administration of pyridoxamine dose-dependently increased the rolling influx of neutrophils and reduced neutrophil adhesion to endothelial cells in cremaster microvessels of sickle cell disease mice challenged with hypoxia-reoxygenation. Short-term treatment also mitigated neutrophil-endothelial cell and neutrophil-platelet interactions in the microvessels and improved the survival of sickle cell disease mice challenged with tumor necrosis factor-α. The inhibitory effects of pyridoxamine on intravascular cell-cell interactions were potentiated by co-treatment with hydroxyurea. We observed that long-term (5.5 months) oral treatment with pyridoxamine significantly diminished the adhesive function of neutrophils and platelets and down-regulated the expression of E-selectin and intercellular adhesion molecule-1 on the vascular endothelium in tumor necrosis factor-α-challenged sickle cell disease mice. Ex vivo studies revealed that the surface amount of αMβ2 integrin was significantly decreased in stimulated neutrophils isolated from sickle cell disease mice treated with pyridoxamine-containing water. Studies using platelets and neutrophils from sickle cell disease mice and patients suggested that treatment with pyridoxamine reduced the activation state of platelets and neutrophils. These results suggest that pyridoxamine may be a novel therapeutic and a supplement to hydroxyurea to prevent and treat vaco-occlusion events in sickle cell disease.
Previous studies identified the Ser/Thr protein kinase, AKT, as a therapeutic target in thrombo-inflammatory diseases. Here we report that specific inhibition of AKT with ARQ 092, an orally-available ...AKT inhibitor currently in phase Ib clinical trials as an anti-cancer drug, attenuates the adhesive function of neutrophils and platelets from sickle cell disease patients in vitro and cell-cell interactions in a mouse model of sickle cell disease. Studies using neutrophils and platelets isolated from sickle cell disease patients revealed that treatment with 50-500 nM ARQ 092 significantly blocks αMβ2 integrin function in neutrophils and reduces P-selectin exposure and glycoprotein Ib/IX/V-mediated agglutination in platelets. Treatment of isolated platelets and neutrophils with ARQ 092 inhibited heterotypic cell-cell aggregation under shear conditions. Intravital microscopic studies demonstrated that short-term oral administration of ARQ 092 or hydroxyurea, a major therapy for sickle cell disease, diminishes heterotypic cell-cell interactions in venules of sickle cell disease mice challenged with tumor necrosis factor-α. Co-administration of hydroxyurea and ARQ 092 further reduced the adhesive function of neutrophils in venules and neutrophil transmigration into alveoli, inhibited expression of E-selectin and intercellular adhesion molecule-1 in cremaster vessels, and improved survival in these mice. Ex vivo studies in sickle cell disease mice suggested that co-administration of hydroxyurea and ARQ 092 efficiently blocks neutrophil and platelet activation and that the beneficial effect of hydroxyurea results from nitric oxide production. Our results provide important evidence that ARQ 092 could be a novel drug for the prevention and treatment of acute vaso-occlusive complications in patients with sickle cell disease.
BackgroundEarly reports have indicated a relationship between ABO and rhesus blood group types and infection with SARS-CoV-2. We aim to examine blood group type associations with COVID-19 mortality ...and disease severity. MethodsThis is a retrospective chart review of patients ages 18 years or older admitted to the hospital with COVID-19 between January 2020 and December 2021. The primary outcome was COVID-19 mortality with respect to ABO blood group type. The secondary outcomes were 1. Severity of COVID-19 with respect to ABO blood group type, and 2. Rhesus factor association with COVID-19 mortality and disease severity. Disease severity was defined by degree of supplemental oxygen requirements (ambient air, low-flow, high-flow, non-invasive mechanical ventilation, and invasive mechanical ventilation). ResultsThe blood type was collected on 596 patients with more than half (54%, N=322) being O+. The ABO blood type alone was not statistically associated with mortality (P=0.405), while the RH blood type was statistically associated with mortality (P<0.001). There was statistically significant association between combined ABO and RH blood type and mortality (P=0.014). Out of the mortality group, the O+ group had the highest mortality (52.3%), followed by A+ (22.8%). The combined ABO and RH blood type was statistically significantly associated with degree of supplemental oxygen requirements (P=0.005). The Kaplan-Meier curve demonstrated that Rh- patients had increased mortality. ConclusionABO blood type is not associated with COVID-19 severity and mortality. Rhesus factor status is associated with COVID-19 severity and mortality. Rhesus negative patients were associated with increased mortality risk.
Interactions of neutrophils with endothelial cells (ECs) and platelets contribute to tissue damage and vascular occlusion under sterile inflammatory conditions. However, the molecular mechanisms ...regulating the cell-cell interactions remain poorly understood. Previous studies suggest that reactive oxygen species, such as hydrogen peroxide (H
O
), produced from NADPH oxidase 2 play a critical role in platelet-neutrophil interactions by regulating the function of neutrophil αMβ2 integrin during sterile inflammation. In this study, we further demonstrate a crucial role for myeloperoxidase (MPO) in regulating the adhesive function of neutrophils through αMβ2 integrin. Using real-time fluorescence intravital microscopy and
assays, we showed that loss of MPO promoted neutrophil-EC interactions and neutrophil emigration but did not affect neutrophil-platelet interactions under inflammatory conditions. Using genetic and pharmacologic approaches, we found that following agonist stimulation, MPO knockout (KO) neutrophils exhibited a significant increase in extracellular H
O
and surface level of αMβ2 integrin and that these effects were dependent on MPO activity. Our
studies using an ischemia/reperfusion-induced hepatic inflammation model revealed that compared to wild-type mice, neutrophils from MPO KO mice-displayed a pro-migratory phenotype while ameliorating tissue damage. These results suggest that MPO plays a negative role in the adhesive and migratory function of neutrophils by impairing αMβ2 integrin function under sterile inflammatory conditions.
Proton-pump inhibitors (PPIs) are commonly utilized in the treatment of upper gastrointestinal bleeds (UGIBs) due to their ability to stabilize blood clot formation. PPIs have been shown to reduce ...rebleeding after endoscopic hemostasis and reduce signs of bleeding at index endoscopy. While PPIs are well-tolerated and commonly administered to patients suffering from acute UGIBs, significant adverse effects may occur. Patients have reported various mild systemic symptoms during short-term PPI use, including headache, rash, dizziness, nausea, abdominal pain, flatulence, constipation, and diarrhea. In general, serious side effects of PPIs tend to be mild during treatment periods under two weeks; however, as the treatment duration increases, side effects have been observed to increase in frequency and severity. PPI-induced thrombocytopenia is an exceedingly rarely reported adverse reaction that remains largely unstudied due to the dearth of patient cases. This adverse effect continues to be a diagnosis of exclusion, and there are no current evidence-based recommendations to approach this complication. Thrombocytopenia increases the risk of rebleeding and hemodynamic instability, which may be devastating to patients suffering from UGIBs. Here, we present a case of thrombocytopenia that began after the introduction of pantoprazole in the setting of a UGIB. The thrombocytopenia resolved promptly after cessation of the medication. We highlight this case to increase awareness of this rare finding given the lack of recommendations for short-term PPI-induced thrombocytopenia.
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