1. Identification of signalling cascades involved in red blood cell shrinkage and vesiculation
- Author
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Timo K. van den Berg, Elena Kostova, Robin van Bruggen, Pasi Halonen, Thomas R. L. Klei, Boukje M. Beuger, Cor Lieftink, Roderick L. Beijersbergen, Molecular cell biology and Immunology, Graduate School, AII - Amsterdam institute for Infection and Immunity, General Internal Medicine, and Landsteiner Laboratory
- Subjects
Erythrocytes ,lcsh:Life ,lcsh:QR1-502 ,red blood cell ,Cellular Senescence/drug effects ,Biochemistry ,lcsh:Microbiology ,PC, phosphatidylcholine ,ERK, extracellular signal-regulated kinase ,Cell-Derived Microparticles ,Casein Kinase II/metabolism ,Jak, Janus kinase ,Anaplastic lymphoma kinase ,ASK1 ,Casein Kinase II ,Cellular Senescence ,CaM, calmodulin ,PDGFR, platelet-derived growth factor receptor ,AMPK, AMP-activated kinase ,Kinase ,Ionomycin ,RBC, red blood cell ,PS, phosphatidylserine ,Cell biology ,raf Kinases/metabolism ,STAT, signal transducer and activator of transcription ,Calcium Ionophores ,VEGFR, vascular endothelial growth factor receptor ,raf Kinases ,Casein kinase 2 ,RTK, receptor tyrosine kinase ,SCD, sickle cell disease ,Cell aging ,PI3K, phosphoinositide 3-kinase ,SMase, acid sphingomyelinase ,kinase inhibitor ,Calcium Ionophores/pharmacology ,SAGM, saline-adenine-glucose-mannitol ,Biophysics ,Biology ,ATA, aurintricarboxylic acid ,Cell-Derived Microparticles/metabolism ,PLC, phospholipase C ,bioactive small molecule ,PKC, protein kinase C ,MEK, mitogen-activated protein kinase kinase ,Humans ,Protein kinase A ,Molecular Biology ,Protein kinase B ,GPCR, G protein-coupled receptor ,BCR-ABL, breakpoint cluster region protein–Abelson murine leukaemia viral oncogene homologue 1 ,Original Paper ,NO, nitric oxide ,compound screen ,Ionomycin/pharmacology ,β-AR, β-adrenergic receptor ,Kinase insert domain receptor ,Cell Biology ,Erythrocytes/cytology ,vesiculation ,LOPAC, Library of Pharmacologically Active Compounds ,nRTK, non-receptor tyrosine kinase ,CK2, casein kinase 2 ,lcsh:QH501-531 ,Epo, erythropoietin ,MAPK, mitogen-activated protein kinase - Abstract
Even though red blood cell (RBC) vesiculation is a well-documented phenomenon, notably in the context of RBC aging and blood transfusion, the exact signalling pathways and kinases involved in this process remain largely unknown. We have established a screening method for RBC vesicle shedding using the Ca2+ ionophore ionomycin which is a rapid and efficient method to promote vesiculation. In order to identify novel pathways stimulating vesiculation in RBC, we screened two libraries: the Library of Pharmacologically Active Compounds (LOPAC) and the Selleckchem Kinase Inhibitor Library for their effects on RBC from healthy donors. We investigated compounds triggering vesiculation and compounds inhibiting vesiculation induced by ionomycin. We identified 12 LOPAC compounds, nine kinase inhibitors and one kinase activator which induced RBC shrinkage and vesiculation. Thus, we discovered several novel pathways involved in vesiculation including G protein-coupled receptor (GPCR) signalling, the phosphoinositide 3-kinase (PI3K)–Akt (protein kinase B) pathway, the Jak–STAT (Janus kinase–signal transducer and activator of transcription) pathway and the Raf–MEK (mitogen-activated protein kinase kinase)–ERK (extracellular signal-regulated kinase) pathway. Moreover, we demonstrated a link between casein kinase 2 (CK2) and RBC shrinkage via regulation of the Gardos channel activity. In addition, our data showed that inhibition of several kinases with unknown functions in mature RBC, including Alk (anaplastic lymphoma kinase) kinase and vascular endothelial growth factor receptor 2 (VEGFR-2), induced RBC shrinkage and vesiculation., After screening two libraries of small bioactive molecules and kinase inhibitors, we identified several signalling pathways to be involved in red blood cell (RBC) shrinkage and vesiculation. These include the Jak (Janus kinase)–STAT (signal transducer and activator of transcription) pathway, phosphoinositide 3-kinase (PI3K)–Akt pathway, the Raf–MEK (mitogen-activated protein kinase kinase)–ERK (extracellular signal-regulated kinase) pathway and GPCR (G protein-coupled receptor) signalling.
- Published
- 2015