scholarly journals The complex of phosphatidylinositol 4,5-bisphosphate and calcium ions is not responsible for Ca(2+)-induced loss of phospholipid asymmetry in the human erythrocyte: a study in Scott syndrome, a disorder of calcium- induced phospholipid scrambling

Blood ◽  
1995 ◽  
Vol 86 (5) ◽  
pp. 1983-1991 ◽  
Author(s):  
EM Bevers ◽  
T Wiedmer ◽  
P Comfurius ◽  
J Zhao ◽  
EF Smeets ◽  
...  
Keyword(s):  
Lipid Vesicles ◽  
Human Erythrocytes ◽  
Mechanism Analysis ◽  
Membrane Phospholipids ◽  
Mole Percent ◽  
Cellular Processes ◽  
Phospholipid Asymmetry ◽  
Dependent Cell ◽  
Normal Human ◽  
State Concentration

Elevation of cytoplasmic Ca2+ levels in human erythrocytes induces a progressive loss of membrane phospholipid asymmetry, a process that is impaired in erythrocytes from a patient with Scott syndrome. We show here that porcine erythrocytes are similarly incapable of Ca(2+)- induced redistribution of membrane phospholipids. Because a complex of phosphatidylinositol 4,5-bisphosphate (PIP2) and Ca2+ has been proposed as the mediator of enhanced transbilayer movement of lipids (J Biol Chem 269:6347,1994), these cell systems offer a unique opportunity for testing this mechanism. Analysis of both total PIP2 content and the metabolic-resistant pool of PIP2 that remains after incubation with Ca2+ ionophore showed no appreciable differences between normal and Scott erythrocytes. Moreover, porcine erythrocytes were found to have slightly higher levels of both total and metabolic-resistant PIP2 in comparison with normal human erythrocytes. Although loading of normal erythrocytes with exogenously added PIP2 gave rise to a Ca(2+)-induced increase in prothrombinase activity and apparent transbilayer movement of nitrobenzoxadiazolyl (NBD)-phospholipids, these PIP2-loaded cells were also found to undergo progressive Ca(2+)-dependent cell lysis, which seriously hampers interpretation of these data. Moreover, loading Scott cells with PIP2 did not abolish their impaired lipid scrambling, even in the presence of a Ca(2+)-ionophore. Finally, artificial lipid vesicles containing no PIP2 or 1 mole percent of PIP2 were indistinguishable with respect to transbilayer movement of NBD- phosphatidylcholine in the presence of Ca2+. Our findings suggest that Ca(2+)-induced redistribution of membrane phospholipids cannot simply be attributed to the steady-state concentration of PIP2, and imply that such lipid movement is regulated by other cellular processes.

scholarly journals CBIO-05. LIPID METABOLIC REPROGRAMMING SENSITIZES A MOLECULARLY-DEFINED SUBSET OF GLIOBLASTOMAS TO FERROPTOSIS

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii16-ii16
Author(s):  
Danielle Morrow ◽  
David Nathanson ◽  
Timothy Cloughesy ◽  
Robert Prins ◽  
Nicholas Bayley ◽  
...  
Keyword(s):  
Lipid Metabolism ◽  
Lipid Droplets ◽  
Metabolic Reprogramming ◽  
Membrane Phospholipids ◽  
Molecular Alterations ◽  
Lipidomic Analysis ◽  
Novel Association ◽  
Therapeutic Opportunity ◽  
Dependent Cell ◽  
Main Components

Abstract Cancers, including the universally lethal glioblastoma (GBM), have reprogrammed lipid metabolism to fuel tumor growth. However, the molecular alterations responsible for aberrant lipid metabolism, and the potential for identifying new therapeutic opportunities are not fully understood. To systematically investigate the GBM lipidome, we performed integrated transcriptomic, genomic and shotgun lipidomic analysis of an extensive library of molecularly diverse patient-derived GBM samples. Using this comprehensive approach, we discovered two GBM sub-groups defined by their combined molecular and lipidomic profile. Triacylglycerides (TAGs) enriched in polyunsaturated fatty acids (PUFAs) were among the most significantly altered lipids between the two groups of GBM tumors. TAGs are the main components of lipid droplets, which sequester PUFA-TAGs away from membrane phospholipids where their peroxidation can lead to ferroptosis – a regulated from of PUFA-peroxidation dependent cell death. Accordingly, the GBM subgroup with a depletion of PUFA TAGs showed heightened sensitivity to ferroptosis. Our findings suggest a novel association between specific molecular signatures of GBM, lipid metabolism and ferroptosis. This relationship may present a new therapeutic opportunity to target reprogrammed lipid metabolism in a molecularly-defined subset of GBMs.

Anaerobic Glycolysis in Normal Human Erythrocytes Incubated in Vitro with Sodium Salicylate

1975 ◽  
Vol 49 (5) ◽  
pp. 375-384
Author(s):  
N. Worathumrong ◽  
A. J. Grimes
Keyword(s):  
Sodium Salicylate ◽  
Lactate Production ◽  
Glucose Consumption ◽  
Human Erythrocytes ◽  
Anaerobic Glycolysis ◽  
Initial Stimulus ◽  
Sodium Efflux ◽  
Normal Human ◽  
Concentration Changes

1. Some effects of sodium salicylate upon anaerobic glycolysis have been studied in normal human erythrocytes incubated for up to 6 h at 37°C in autologous sera. 2. Both glucose consumption and lactate production were stimulated by concentrations of salicylate up to 60 mmol/l but at the highest concentration used (90 mmol/l) an initial stimulus was followed by inhibition of glycolysis. 3. Losses occurred of adenosine 5′-triphosphate (ATP), adenosine 5′-diphosphate (ADP) and adenosine 5′-phosphate (AMP) at higher concentrations of salicylate and there was a concomitant increase of inorganic phosphate. 4. Other phosphate esters underwent concentration changes at higher concentrations of salicylate that reflected inadequate concentrations of ATP for glycolysis. 5. The rates of sodium efflux from, and potassium influx into, erythrocytes were unaffected by the presence of salicylate at concentrations sufficient to stimulate glycolysis.

Purification and characterization of the yeast-expressed erythropoietin mutant Epo (R103A), a specific inhibitor of human primary hematopoietic cell erythropoiesis

Blood ◽  
2002 ◽  
Vol 99 (12) ◽  
pp. 4400-4405 ◽  
Author(s):  
Suzanne Burns ◽  
Murat O. Arcasoy ◽  
Li Li ◽  
Elizabeth Kurian ◽  
Katri Selander ◽  
...  
Keyword(s):  
Animal Models ◽  
Specific Inhibitor ◽  
Competitive Inhibitor ◽  
Erythropoietin Receptor ◽  
Single Amino Acid ◽  
Human Cd34 ◽  
Unit Formation ◽  
Dependent Cell ◽  
Normal Human

A drug that specifically inhibits erythropoiesis would be clinically useful. The erythropoietin (Epo) mutant Epo (R103A) could potentially be used for this purpose. Epo (R103A) has a single amino acid substitution of alanine for arginine at position 103. Because of this mutation, Epo (R103A) is only able to bind to one of the 2 subunits of the erythropoietin receptor (EpoR) homodimer and is thus a competitive inhibitor of Epo activity. To produce large quantities of Epo (R103A) to test in animal models of thalassemia and sickle cell disease, we expressed and purified recombinant Epo (R103A) from the yeast Pichia pastoris. Using this method milligram quantities of highly purified Epo (R103A) are obtained. The yeast-expressed Epo (R103A) is properly processed and glycosylated and specifically inhibits Epo-dependent cell growth and125I-Epo binding. Epo (R103A) does not, however, directly induce apoptosis in 32D cells expressing EpoR. Epo (R103A) inhibits erythropoiesis of human CD34+ hematopoietic cells and completely blocks erythroid burst-forming unit formation in normal human bone marrow colony assays. Yeast-expressed Epo (R103A) is a specific inhibitor of primary erythropoiesis suitable for testing in animal models.

E-cadherin is the major mediator of human melanocyte adhesion to keratinocytes in vitro

1994 ◽  
Vol 107 (4) ◽  
pp. 983-992 ◽  
Author(s):  
A. Tang ◽  
M.S. Eller ◽  
M. Hara ◽  
M. Yaar ◽  
S. Hirohashi ◽  
...  
Keyword(s):  
Cell Types ◽  
Melanoma Metastasis ◽  
Normal Keratinocytes ◽  
Human Melanocyte ◽  
Human Melanocytes ◽  
Dependent Cell ◽  
Normal Human ◽  
Pre Treatment ◽  
E Cadherin

E- and P-cadherin are calcium (Ca2+)-dependent cell adhesion molecules important in the morphogenesis and maintenance of skin structure. By use of flow cytometry and specific antibodies, we now show that cultured human melanocytes express E- and P-cadherin on their surfaces, and that these molecules have the same characteristics as reported for other cell types. Specifically, melanocyte cadherins are sensitive to trypsin digestion in the absence of Ca2+ and are protected from trypsin degradation by Ca2+, and are functional at 37 degrees C but not at 4 degrees C. We further show that melanocytes contain mRNA transcripts encoding both E- and P-cadherin. Adhesion of cultured melanocytes to keratinocyte monolayers is abolished by pre-treatment of the melanocytes with trypsin/EDTA, which degrades E- and P-cadherins, is greatly reduced by anti-E-cadherin antibodies and is slightly reduced by antibodies to P-cadherin, alpha 2, alpha 3 and beta 1 integrins. In contrast to normal melanocytes, eight of nine melanoma cell lines lacked E-cadherin (or expressed markedly reduced levels) and five were negative for P-cadherin. Melanoma cells also failed to adhere to keratinocyte monolayers. These results demonstrate that normal human melanocytes express functional E- and P-cadherin and that E-cadherin is primarily responsible for adhesion of human melanocytes to keratinocytes in vitro. In addition, transformed melanocytes express markedly reduced levels of E- and P-cadherin, and exhibit decreased affinity for normal keratinocytes in vitro, suggesting that loss of cadherins may play a role in melanoma metastasis.

Oxidative Damage Does Not Alter Membrane Phospholipid Asymmetry in Human Erythrocytes†

Biochemistry ◽  
1997 ◽  
Vol 36 (22) ◽  
pp. 6768-6776 ◽  
Author(s):  
Kitty de Jong ◽  
Danielle Geldwerth ◽  
Frans A. Kuypers
Keyword(s):  
Oxidative Damage ◽  
Human Erythrocytes ◽  
Membrane Phospholipid ◽  
Phospholipid Asymmetry

scholarly journals Combinatorial transcription factor profiles predict mature and functional human islet α and β cells

2021 ◽  
Author(s):  
Shristi Shrestha ◽  
Diane C. Saunders ◽  
John T. Walker ◽  
Joan Camunas-Soler ◽  
Xiao-Qing Dai ◽  
...  
Keyword(s):  
Islet Cells ◽  
Hormone Secretion ◽  
Developmental State ◽  
Human Islets ◽  
Glucose Sensing ◽  
Pancreatic Tissue ◽  
Β Cells ◽  
Cellular Processes ◽  
Normal Human ◽  
Multiple Donors

ABSTRACTIslet-enriched transcription factors (TFs) exert broad control over cellular processes in pancreatic α and β cells and changes in their expression are associated with developmental state and diabetes. However, the implications of heterogeneity in TF expression across islet cell populations are not well understood. To define this TF heterogeneity and its consequences for cellular function, we profiled >40,000 cells from normal human islets by scRNA-seq and stratified α and β cells based on combinatorial TF expression. Subpopulations of islet cells co-expressing ARX/MAFB (α cells) and MAFA/MAFB (β cells) exhibited greater expression of key genes related to glucose sensing and hormone secretion relative to subpopulations expressing only one or neither TF. Moreover, all subpopulations were identified in native pancreatic tissue from multiple donors. By Patch-seq, MAFA/MAFB co-expressing β cells showed enhanced electrophysiological activity. Thus, these results indicate combinatorial TF expression in islet α and β cells predicts highly functional, mature subpopulations.

Cell volume, K transport, and cell density in human erythrocytes

1987 ◽  
Vol 252 (3) ◽  
pp. C269-C276 ◽  
Author(s):  
C. Brugnara ◽  
D. C. Tosteson
Keyword(s):  
Cell Volume ◽  
Cell Density ◽  
Human Erythrocytes ◽  
Ph Optimum ◽  
Cation Content ◽  
Original Volume ◽  
Dense Fraction ◽  
Normal Human ◽  
K Transport ◽  
Hypotonic Swelling

We report here studies on the regulation of cell volume and K transport in human erythrocytes separated according to density. When cell volume was increased (isosmotic swelling, nystatin technique), erythrocytes of the least dense but not of the densest fraction shrunk back toward their original volume. This process was due to a ouabain (0.1 mM) and bumetanide (0.01 mM) (OB)-resistant K loss. OB-resistant K+ efflux from the least dense fraction was stimulated by hypotonic swelling and had a bell-shaped dependence on pH (pH optimum 6.75–7.0). These pH and volume effects were not evident in the densest fraction. The swelling-induced K+ efflux from the least dense fraction was inhibited when chloride was substituted by nitrate, thiocyanate, and acetate, whereas it was stimulated by bromide. Increasing cell Mg2+ content also markedly inhibited K+ efflux from isosmotically swollen cells. N-ethylmaleimide (NEM, 1 mM) greatly increased OB-resistant K+ efflux from the least dense fraction but not from the densest fraction. These data reveal the presence, in the lease dense fraction of normal human erythrocytes, of a pathway for K+ transport that is dependent on volume, pH, and chloride, is inhibited by internal Mg2+, and possibly plays a role in determining the erythrocyte water and cation content.

Adherence of phosphatidylserine-exposing erythrocytes to endothelial matrix thrombospondin

Blood ◽  
2000 ◽  
Vol 95 (4) ◽  
pp. 1293-1300 ◽  
Author(s):  
Annamaria B. Manodori ◽  
Gilda A. Barabino ◽  
Bertram H. Lubin ◽  
Frans A. Kuypers
Keyword(s):  
Endothelial Cells ◽  
Sickle Cell ◽  
Binding Sites ◽  
Umbilical Vein ◽  
Calcium Ionophore ◽  
Annexin V ◽  
Lipid Vesicles ◽  
Cell Disease ◽  
Phospholipid Asymmetry

Phospholipid asymmetry is well maintained in erythrocyte (RBC) membranes with phosphatidylserine (PS) exclusively present in the inner leaflet. The appearance of PS on the surface of the cell can have major physiologic consequences, including increased cell-cell interactions. Because increased adherence of PS-exposing RBCs to endothelial cells (ECs) may be pathologically important in hemoglobinopathies such as sickle cell disease and thalassemia, we studied the role of PS exposure in calcium ionophore-treated normal RBC adherence to human umbilical vein endothelial cell (HUVEC) monolayers. When HUVEC monolayers were incubated with these PS-exposing RBCs, the ECs retracted and the RBCs adhered primarily in the gaps opened between the ECs. A linear correlation was found between the number of PS-exposing RBCs in the population and the number of adhering RBCs to the monolayer. Pretreatment of RBCs with annexin V significantly decreased adherence by shielding PS on the RBCs. Similarly, PS-containing lipid vesicles decreased RBC binding by competing for the PS binding sites in the monolayer. PS-exposing RBCs and PS-containing lipid vesicles adhered to immobilized thrombospondin (TSP) and matrix TSP, respectively, and adherence of PS-exposing RBCs to EC monolayers was reduced by antibodies to TSP and to its EC receptor, vβ3. Together, these results indicate a role for PS and matrix TSP in the adherence of PS-exposing RBCs to EC monolayers, and suggest an important contribution of PS-exposing RBCs in pathologies with reported vascular damage, such as sickle cell anemia.

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