Morphologic studies of sickle erythrocytes by image analysis.

An automated image analysis system was used to characterize the morphology of sickle cells under hypoxic conditions. Images observed by light microscopy were transferred to the image analysis system and were processed to binary (black and white) images, and were then analyzed by area, perimeter, and shape. A circular shape factor (CSF = 4 pi X[area]/[perimeter]2) was found to be useful for elucidating the degree of deformation, but it could not differentiate elongated sickle cells from maple leaf-shaped cells. By combining an elliptical shape factor (ESF = [short axis]/[long axis]) with CSF we could separate deoxygenated homozygous sickle red cells (SS cells) into several morphologically distinct groups, including non-sickled cells, maple leaf-shaped cells, and elongated sickle cells. Using this automated image analysis system, we studied morphologic changes of SS cells exposed to deoxygenation-oxygenation (d-o) cycles between PO2 of 0 and 100 mm Hg (one cycle = 12 minutes) at pHs of 6.9 and 7.4. We found that at both pHs the morphology of sickled cells after the first deoxygenation was predominantly maple leaf-shaped. The number of elongated sickled cells increased as the number of d-o cycles increased, indicating that SS cells changed from maple leaf morphology to classic elongated sickle shape during d-o cycles. Desickling occurred less during the oxygenation phase at pH 6.9 than at pH 7.4 and as the number of d-o cycles increased. These results suggest that during d-o cycles deoxyhemoglobin S fibers may align to form large bundles that do not depolymerize completely even at the arterial oxygen pressure.