Sulfhydryl oxidation and activation of red cell K(+)-Cl- cotransport in the transgenic SAD mouse
L De Franceschi1, Y Beuzard, C Brugnara
1Department of Internal Medicine, University of Verona, Italy.
Insights
Mouse erythrocytes with human SAD hemoglobin show dehydration and increased K(+)-Cl- cotransport. This suggests sulfhydryl oxidation activates this transport in SAD hemoglobin red blood cells.
Area of Science:
- Hematology
- Molecular Biology
- Physiology
Background:
- The SAD mouse model expresses human sickle cell disease (SCD) hemoglobin (Hb SAD), which has increased polymerization tendency due to Antilles and D Punjab mutations.
- Understanding red blood cell (RBC) cation transport is crucial for SCD pathophysiology.
Purpose of the Study:
- To investigate monovalent cation transport in erythrocytes of SAD-1 and beta-thal/SAD-1 mice.
- To elucidate the role of K(+)-Cl- cotransport in SAD hemoglobin-expressing erythrocytes.
Main Methods:
- Erythrocytes from SAD-1 and beta-thal/SAD-1 mice were analyzed for cation transport.
- K(+)-Cl- cotransport was characterized by Cl(-)-dependent, okadaic acid-sensitive K+ efflux.
- Effects of staurosporine and dithiothreitol (DTT) on K(+)-Cl- cotransport were assessed.
Main Results:
- SAD erythrocytes exhibited dehydration and increased Na(+)-K+ pump activity.
- K(+)-Cl- cotransport was significantly increased in SAD erythrocytes.
- Volume regulatory decrease via K(+)-Cl- cotransport was enhanced in swollen SAD erythrocytes.
- DTT treatment normalized elevated K(+)-Cl- cotransport in SAD erythrocytes, indicating reversible sulfhydryl oxidation.
Conclusions:
- Erythrocytes expressing human Hb SAD display altered cation transport, including enhanced K(+)-Cl- cotransport.
- Reversible sulfhydryl oxidation contributes to the activation of K(+)-Cl- cotransport in SAD hemoglobin red blood cells.
- These findings provide insights into the cellular mechanisms underlying SAD hemoglobin pathophysiology.
Abstract:
The SAD mouse is characterized by the expression of human SAD hemoglobin (Hb), a super S Hb with a higher tendency to polymerize than HbS due to the presence of two additional mutations, Antilles beta 23Ile and D Punjab beta 121Glu. Monovalent cation transport was studied in erythrocytes from SAD-1 (Hb SAD = 19%) and beta-thal/SAD-1 (Hb SAD = 26%) mice. Erythrocytes containing Hb SAD exhibited dehydration, increased maximal rate of Na(+)-K+ pump, unchanged Rb+ flux via the Gardos channel, and increased K(+)-Cl- cotransport. K(+)-Cl- cotransport was defined as Cl(-)-dependent (substitution with sulfamate or methanesulfonate) okadaic acid-sensitive K+ efflux. Volume regulatory decrease via K(+)-Cl- cotransport was also increased in swollen SAD erythrocytes compared with controls. K(+)-Cl- cotransport was stimulated by staurosporine in all mouse strains, but the extent of stimulation was reduced in beta-thal/SAD-1 mice. Treatment with dithiothreitol reduced K(+)-Cl- cotransport activity in SAD-1 and beta-thal/SAD-1 mice to levels similar to that of control strains, indicating that reversible sulfhydryl oxidation contributes to the activated state of K(+)-Cl- cotransport in mouse erythrocytes that express transgenic human Hb SAD.
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