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Rehydration of high-density sickle erythrocytes in vitro.
Blood 2002; 100(8):3017-25Blood

Abstract

Recent studies have identified older, low-density sickle red blood cells (SSRBCs) that were resistant to dehydration by valinomycin, a K(+) ionophore. These cells, thought to derive from dense SSRBCs that have rehydrated, may represent a terminal cellular phase. To study rehydration, we subjected dense SSRBCs (rho > 1.107 g/cc) to either oxygenated incubation or rapid oxygenated/deoxygenated (oxy/deoxy) cycling (70 seconds per cycle). Light cells (rho < 1.087 g/cc) were generated during both oxy incubation (2.9% +/- 2.1%; n = 42) and oxy/deoxy cycling (5.3% +/- 2.4%; n = 42). The rehydrated cells were K(+)-depleted (K(+) = 20 +/- 14 mmol/kg hemoglobin [Hb]) and Na(+)-loaded (Na(+) = 394 +/- 106 mmol/kg Hb), and had high levels of external phosphatidylserine. In the presence of external calcium, the generation of rehydrated SSRBCs was inhibited during oxy/deoxy cycling, but the percentage with external phosphatidylserine increased. The calcium-mediated inhibition of rehydration was reversed by charybdotoxin, implying that rehydration was delayed in some cells by the Ca(++)-activated K(+) channel. Preincubation of dense SSRBCs with DIDS (4,4'-di-isothiocyanato-2,2'-disulfostilbene) inhibited the generation of light cells during fast oxy/deoxy cycling, but not during oxy incubation. These results suggest that the sickling-induced pathway, previously implicated in SSRBC dehydration, may be involved in the deoxy-dependent component of rehydration for dense, K(+)-depleted cells. Light-cell generation was inhibited by 1 mM bumetanide during both oxy incubation and oxy/deoxy cycling, providing evidence that a bumetanide-sensitive, deoxy-independent pathway, previously described in circulating light SSRBCs, also contributes to the rehydration of high-density SSRBCs.

Authors+Show Affiliations

Department of Aerospace Engineering, University of Cincinnati College of Engineering; the Cincinnati Comprehensive Sickle Cell Center, Cincinnati, OH 4526. -0508, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

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

Language

eng

PubMed ID

12351416

Citation

Holtzclaw, J David, et al. "Rehydration of High-density Sickle Erythrocytes in Vitro." Blood, vol. 100, no. 8, 2002, pp. 3017-25.
Holtzclaw JD, Jiang M, Yasin Z, et al. Rehydration of high-density sickle erythrocytes in vitro. Blood. 2002;100(8):3017-25.
Holtzclaw, J. D., Jiang, M., Yasin, Z., Joiner, C. H., & Franco, R. S. (2002). Rehydration of high-density sickle erythrocytes in vitro. Blood, 100(8), pp. 3017-25.
Holtzclaw JD, et al. Rehydration of High-density Sickle Erythrocytes in Vitro. Blood. 2002 Oct 15;100(8):3017-25. PubMed PMID: 12351416.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Rehydration of high-density sickle erythrocytes in vitro. AU - Holtzclaw,J David, AU - Jiang,Maorong, AU - Yasin,Zahida, AU - Joiner,Clinton H, AU - Franco,Robert S, PY - 2002/9/28/pubmed PY - 2002/12/6/medline PY - 2002/9/28/entrez SP - 3017 EP - 25 JF - Blood JO - Blood VL - 100 IS - 8 N2 - Recent studies have identified older, low-density sickle red blood cells (SSRBCs) that were resistant to dehydration by valinomycin, a K(+) ionophore. These cells, thought to derive from dense SSRBCs that have rehydrated, may represent a terminal cellular phase. To study rehydration, we subjected dense SSRBCs (rho > 1.107 g/cc) to either oxygenated incubation or rapid oxygenated/deoxygenated (oxy/deoxy) cycling (70 seconds per cycle). Light cells (rho < 1.087 g/cc) were generated during both oxy incubation (2.9% +/- 2.1%; n = 42) and oxy/deoxy cycling (5.3% +/- 2.4%; n = 42). The rehydrated cells were K(+)-depleted (K(+) = 20 +/- 14 mmol/kg hemoglobin [Hb]) and Na(+)-loaded (Na(+) = 394 +/- 106 mmol/kg Hb), and had high levels of external phosphatidylserine. In the presence of external calcium, the generation of rehydrated SSRBCs was inhibited during oxy/deoxy cycling, but the percentage with external phosphatidylserine increased. The calcium-mediated inhibition of rehydration was reversed by charybdotoxin, implying that rehydration was delayed in some cells by the Ca(++)-activated K(+) channel. Preincubation of dense SSRBCs with DIDS (4,4'-di-isothiocyanato-2,2'-disulfostilbene) inhibited the generation of light cells during fast oxy/deoxy cycling, but not during oxy incubation. These results suggest that the sickling-induced pathway, previously implicated in SSRBC dehydration, may be involved in the deoxy-dependent component of rehydration for dense, K(+)-depleted cells. Light-cell generation was inhibited by 1 mM bumetanide during both oxy incubation and oxy/deoxy cycling, providing evidence that a bumetanide-sensitive, deoxy-independent pathway, previously described in circulating light SSRBCs, also contributes to the rehydration of high-density SSRBCs. SN - 0006-4971 UR - https://www.unboundmedicine.com/medline/citation/12351416/Rehydration_of_high_density_sickle_erythrocytes_in_vitro_ L2 - http://www.bloodjournal.org/cgi/pmidlookup?view=long&amp;pmid=12351416 DB - PRIME DP - Unbound Medicine ER -