Elimination of Young Erythrocytes from Blood Circulation and Altered Erythropoietic Patterns during Paraquat Induced Anemic Phase in Mice
Double biotinylation technique was used to label erythrocytes in vivo as described in methods. Mice were administered three daily i.v. doses of 1 mg biotin-X-NHS Ester (BXN) (first biotinylation step). After 5 days rest, a single i.v. dose of 0.6 mg BXN was administered (second biotinylation step). Repeated doses of paraquat (10 mg/kg) were administrated i.p. on alternate days starting the day after the second biotinylation step. Blood samples were collected on day 0 just before the first paraquat injection, and at 7, 14, 21 and 28 day time points during the paraquat administration. Erythrocytes in the blood samples were stained with streptavidin-APC and analyzed on flow cytometer. Box-Y in panel A and B contain erythrocytes that entered the circulation between the first and the second biotinylation step in control and paraquat treated mice respectively. Erythrocytes younger than those in box-Y are in box-Z (biotinnegative population) whereas the erythrocyte population older than the population in box-Y is represented as events in box-X (biotinhigh population). Panel C shows the survival kinetics of older erythrocytes (box-X) in control and paraquat treated mice. Panel D shows the kinetics of survival of the tracked cohort of box-Y erythrocytes in control and paraquat treated mice. Panel E shows the kinetics of accumulation of younger box-Z erythrocytes in control and paraquat treated mice. Data is represented as mean ± SEM; n = 6 in control and 4–5 in treated groups. *p<0.05, **p<0.005 and ***p<0.0005.