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Red blood cells of a transgenic mouse expressing high levels of human hemoglobin S exhibit deoxy-stimulated cation flux.
J Membr Biol 1997; 159(3):187-96JM

Abstract

Deoxy-stimulated cation fluxes have been implicated in the generation of the dense and irreversibly sickled red blood cells (RBCs) in patients homozygous for hemoglobin S (SS). We now report on the effect of short term deoxygenation on K+ and Na+ transport in RBCs from control mice (C57Bl/6J) and a transgenic (alphaHbetaS[betaMDD]) mouse line that expresses high levels of human alphaH and betaS-chains and has a small percent dense cells but does not exhibit anemia. In transgenic mouse RBCs (n = 5) under oxygenated conditions, K+ efflux was 0.22 +/- 0.01 mmol/L cell x min and Na+ influx was 0.17 +/- 0.02 mmol/L cell x min. Both fluxes were stimulated by 10 min deoxygenation in transgenic but not in control mice. The deoxy-stimulated K+ efflux from transgenic mouse RBCs was about 55% inhibited by 5 nm charybdotoxin (CTX), a blocker of the calcium activated K+-channel. To compare the fluxes between human and mouse RBCs, we measured the area of mouse RBCs and normalized values to area per liter of cells. The deoxy-simulated CTX-sensitive K+ efflux was larger than the CTX-sensitive K+ efflux observed in RBCs from SS patients. These results suggest that in transgenic mice, deoxygenation increases cytosolic Ca2+ to levels which open Ca2+-activated K+ channels. The presence of these channels was confirmed in both control and transgenic mice by clamping intracellular Ca2+ at 10 microM with the ionophore A23187 and measuring Ca2+-activated K+ efflux. Both types of mouse had similar maximal rates of CTX-sensitive, Ca2+-activated K+ efflux that were similar to those in human SS cells. The capacity of the mouse red cell membrane to regulate cytosolic Ca2+ levels was examined by measurements of the maximal rate of calmodulin activated Ca2+-ATPase activity. This activity was 3-fold greater than that observed in human RBCs thus indicating that mouse RBC membranes have more capacity to regulate cytosolic Ca2+ levels. In summary, transgenic mouse RBCs exhibit larger values of deoxy-stimulated K+ efflux and Na+ influx when compared to human SS cells. They have a similar Ca2+-activated K+ channel activity to human SS cells while expressing a very high Ca2+ pump activity. These properties may contribute to the smaller percent of very dense cells and to the lack of adult anemia in this animal model.

Authors+Show Affiliations

Endocrine-Hypertension Division, Brigham and Women's Hospital, Department of Medicine, Harvard Medical School, 221 Longwood Avenue, Boston, MA 02115, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

9312208

Citation

Romero, J R., et al. "Red Blood Cells of a Transgenic Mouse Expressing High Levels of Human Hemoglobin S Exhibit Deoxy-stimulated Cation Flux." The Journal of Membrane Biology, vol. 159, no. 3, 1997, pp. 187-96.
Romero JR, Fabry ME, Suzuka S, et al. Red blood cells of a transgenic mouse expressing high levels of human hemoglobin S exhibit deoxy-stimulated cation flux. J Membr Biol. 1997;159(3):187-96.
Romero, J. R., Fabry, M. E., Suzuka, S., Nagel, R. L., & Canessa, M. (1997). Red blood cells of a transgenic mouse expressing high levels of human hemoglobin S exhibit deoxy-stimulated cation flux. The Journal of Membrane Biology, 159(3), pp. 187-96.
Romero JR, et al. Red Blood Cells of a Transgenic Mouse Expressing High Levels of Human Hemoglobin S Exhibit Deoxy-stimulated Cation Flux. J Membr Biol. 1997 Oct 1;159(3):187-96. PubMed PMID: 9312208.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Red blood cells of a transgenic mouse expressing high levels of human hemoglobin S exhibit deoxy-stimulated cation flux. AU - Romero,J R, AU - Fabry,M E, AU - Suzuka,S, AU - Nagel,R L, AU - Canessa,M, PY - 1997/10/6/pubmed PY - 1997/10/6/medline PY - 1997/10/6/entrez SP - 187 EP - 96 JF - The Journal of membrane biology JO - J. Membr. Biol. VL - 159 IS - 3 N2 - Deoxy-stimulated cation fluxes have been implicated in the generation of the dense and irreversibly sickled red blood cells (RBCs) in patients homozygous for hemoglobin S (SS). We now report on the effect of short term deoxygenation on K+ and Na+ transport in RBCs from control mice (C57Bl/6J) and a transgenic (alphaHbetaS[betaMDD]) mouse line that expresses high levels of human alphaH and betaS-chains and has a small percent dense cells but does not exhibit anemia. In transgenic mouse RBCs (n = 5) under oxygenated conditions, K+ efflux was 0.22 +/- 0.01 mmol/L cell x min and Na+ influx was 0.17 +/- 0.02 mmol/L cell x min. Both fluxes were stimulated by 10 min deoxygenation in transgenic but not in control mice. The deoxy-stimulated K+ efflux from transgenic mouse RBCs was about 55% inhibited by 5 nm charybdotoxin (CTX), a blocker of the calcium activated K+-channel. To compare the fluxes between human and mouse RBCs, we measured the area of mouse RBCs and normalized values to area per liter of cells. The deoxy-simulated CTX-sensitive K+ efflux was larger than the CTX-sensitive K+ efflux observed in RBCs from SS patients. These results suggest that in transgenic mice, deoxygenation increases cytosolic Ca2+ to levels which open Ca2+-activated K+ channels. The presence of these channels was confirmed in both control and transgenic mice by clamping intracellular Ca2+ at 10 microM with the ionophore A23187 and measuring Ca2+-activated K+ efflux. Both types of mouse had similar maximal rates of CTX-sensitive, Ca2+-activated K+ efflux that were similar to those in human SS cells. The capacity of the mouse red cell membrane to regulate cytosolic Ca2+ levels was examined by measurements of the maximal rate of calmodulin activated Ca2+-ATPase activity. This activity was 3-fold greater than that observed in human RBCs thus indicating that mouse RBC membranes have more capacity to regulate cytosolic Ca2+ levels. In summary, transgenic mouse RBCs exhibit larger values of deoxy-stimulated K+ efflux and Na+ influx when compared to human SS cells. They have a similar Ca2+-activated K+ channel activity to human SS cells while expressing a very high Ca2+ pump activity. These properties may contribute to the smaller percent of very dense cells and to the lack of adult anemia in this animal model. SN - 0022-2631 UR - https://www.unboundmedicine.com/medline/citation/9312208/Red_blood_cells_of_a_transgenic_mouse_expressing_high_levels_of_human_hemoglobin_S_exhibit_deoxy_stimulated_cation_flux_ L2 - https://dx.doi.org/10.1007/s002329900282 DB - PRIME DP - Unbound Medicine ER -