A Double In Vivo Biotinylation Technique to Assess Erythrocyte Turnover in Blood Circulation

Abstract

We have developed a new double in vivo biotinylation (DIB) technique that may be used for assessing turnover patterns of erythrocytes in circulation. This technique involves two successive in vivo biotinylation steps, interspersed by a period of 5-30 days, which would enable us to tag with biotin a population of erythrocytes entering blood circulation over a defined period of time, between the two biotinylation steps. As such we can track the age-related changes in a lifetime of the circulating erythrocytes, or we can simultaneously study two defined age cohorts of aged as well as young erythrocytes in circulation. We have extensively used this technique to look at erythrocyte loss in mouse models of anemia induced by (a) heavy metal cadmium (Cd), (b) herbicide Paraquat (PQ), (c) carbon nanotubes (CNTs), and (d) autoantibody in autoimmune hemolytic anemia (AIHA). We have found that the pattern of erythrocyte removal is distinctly different in different models of murine anemia. In certain types of anemia (CNT and AIHA), younger erythrocytes in blood circulation are preferentially removed, whereas in other cases (Cd and PQ), old erythrocytes are specifically eliminated.

Murine models of autoimmune hemolytic anemia

“…Saxena et al has developed a technique called double in-vivo biotinylation (DIB) to delineate erythrocyte age, which has been instrumental in elucidating age-related changes and clearance. Incorporation of DIB in multiple models of anemia has demonstrated that clearance patterns of RBC precursors, reticulocytes, and mature RBCs are distinct, depending on how anemia is induced (e.g., heavy metal, autoantibodies) [ 39 , 40 ▪ ]. With regard to the Playfair and Marshall–Clarke model, reticulocytes have more autoantibodies bound to their surface, produce more ROS, and are preferentially cleared from circulation [ 41 ▪ ].…”