Electronic Measurement of Erythrocyte Volume and Volume Heterogeneity in Horses During Erythrocyte Regeneration Associated with Experimental Anemias

Anemia was induced in three groups of horses by moderate or severe acute hemorrhage, or by acetyl phenylhydrazine-induced hemolysis (Groups I, 11, and 111, respectively). Serial hemograms were done on a multichannel automated blood cell counter with histogram capability. Changes in hematocrit, mean cell volume, erythrocyte number, red cell distribution width (RDW), and standard deviation of erythrocyte volume were examined over time. Significant increases in mean cell volume were first detectable by days 17, 20, and 14 and reached maximum by days 43, 41, and 29, in Groups I, 11, and 111, respectively (P 0.05). In some horses effective regeneration occurs without development of macrocytosis. Mean cell volume remained increased after other parameters returned to control values, suggesting that mean cell volume values may provide retrospective evidence of altered erythrocyte turnover. Anisocytosis as indicated by significant increases in the standard deviation was greatest during the early part of the regenerative response, reaching maximum values on days 30,28, and 21 in Groups I, 11, and 111, respectively, and began to decrease as homogeneous repopulation with macrocytes occurred. Red cell distribution width increased significantly only in severe hemorrhage and hemolysis groups, reaching mean maximum values of 24.3 on day 20 and of 26.4 on day 21 in Groups I1 and 111, respectively (P < 0.05). Red cell distribution width did not detect increased erythrocyte volume heterogeneity in Group I. The horse does not release reticulocytes into circu- lation which precludes use of the reticulocyte count in evaluating erythrocyte regeneration. Assessment of the equine regenerative response has been attempted using several methods. Sequential packed cell volume values rising over time indicate effective regeneration, but evidence for erythrogenesis is observed only after re- covery. Bone marrow aspirate examination has been used to estimate effective erythr~genesis.~ ~~~ Changes in erythrocyte chemical constituents have been cor- related with the rate of erythrogenesis. Erythrocyte concentrations of creatine, glucose-6-phosphate de- hydrogena~e,'~ and adenosine-5-triphosphate15 in- crease in immature erythrocytes during a regenerative response. Increases in erythrocyte lactate dehydroge- nase concentration will occur during regenerative he- molytic anemia but not blood None of the above parameters have been correlated with the degree of anemia and most have limited availability. Mean cell volume increases during accelerated eryth- rogenesis due to release of macrocytes in numerous species.'OJ4J5 This parameter may be measured either by calculation from manually determined hematocrit and red blood cell count or by direct electronic cell sizing. The reference range of values for electronically determined mean cell volume in the horse is narrower and more reproducible than that determined manual- ly." A large number of macrocytes is required to shift the calculated mean cell volume above the normal range while erythrocyte volume distribution histo- grams (erythrograms) are sensitive in detecting shifts in erythrocyte population size before the mean cell volume is abnormal, enabling earlier detection of erythrocyte size disturbances. In two horses with im- mune-mediated hemolytic anemia, erythrograms were used to demonstrate a gradual increase in macrocyte number and decrease in normocyte number over a 4 to 5 week period.I9 In one horse, increased numbers of macrocytes and a shift in the mean cell volume could be detected before an increase in hematocrit. Repop- ulation with macrocytes correlated with rising hemat- ocrit and bone marrow erythroid hyperplasia and ap- peared to be a useful indicator of regenerative response. Some newer automated, multichannel blood cell counters routinely determine the red cell volume dis- tribution width (RDW value) as an index of anisocy- tosis. The RDW value, a coefficient of variation of erythrocyte volume, is calculated by dividing the stan-