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Assessment of erythrocyte shape by flow cytometry techniques

¹ Department of Intensive Care, Erasme University Hospital, Free University of Brussels, Brussels, Belgium
² Experimental Medicine Laboratory, ULB 222 Unit, CHU Charleroi-Vésale, Free University of Brussels, Montigny-le-Tilleul, Belgium
³ Department of Nephrology, CHU Charleroi-Vésale, Free University of Brussels, Montigny-le Tilleul, Belgium

Correspondence to:
Professor J-L Vincent
Department of Intensive Care, Erasme University Hospital, 808, Route de Lennik, 1070 Brussels, Belgium; jlvincen{at}ulb.ac.be Background: Red blood cell (RBC) rheology is altered in different diseases, including acute conditions such as patients in intensive care units (ICU) with sepsis or with an inflammatory reaction due to postoperative states or intracerebral haemorrhage, or chronic conditions such as diabetes mellitus or terminal renal failure. Several techniques are available to assess alterations in RBC rheology, especially deformability, but they are too cumbersome to be used on a large number of cells.

Objective: To develop a new, rapid flow cytometry technique for easy assessment of RBC shape in patients.

Methods: In flow cytometry, healthy human RBC shape shows a bimodal distribution related to the biconcave form. On this histogram, the second Pearson coefficient of dissymmetry (PCD) representing the asymmetry of this histogram and the spherical index (M2:M1) were calculated, both representing the spherical shape. This technique was used in healthy volunteers (n = 17) and in diseases characterised by abnormalities in RBC rheology, including terminal renal failure requiring haemodialysis (n = 28), diabetes mellitus (n = 18), sepsis (n = 19) and acute inflammatory states (postoperative, intracerebral haemorrhage, chronic obstructive pulmonary disease, epilepsy or severe drug intoxication; n = 21). Multivariate analysis was performed to determine the factors influencing RBC shape.

Results: Measurement of RBC shape was highly reproducible. A good correlation was observed between the PCD and the spherical index, except in the critically ill patients without sepsis. RBCs were more spherical in patients with terminal renal failure (PCD –0.56 (0.14), p<0.05), diabetes mellitus (PCD –0.59 (0.23), p<0.05), sepsis (PCD –0.58 (0.22), p<0.05) or an acute inflammatory state (PCD –0.65 (0.29), p<0.05) than in healthy volunteers (PCD –0.89 (0.12)). The spherical index was also increased in all populations compared with healthy volunteers (terminal renal failure 2.30 (0.20); diabetes mellitus 2.27 (0.38); sepsis 2.28 (0.37); acute inflammatory state 2.35 (0.42) vs healthy volunteers 2.72 (0.47); all p<0.05). Multivariate analysis demonstrated that the underlying pathology (sepsis, acute inflammatory state, diabetes mellitus, terminal renal failure) was the principal cause of these RBC shape abnormalities.

Conclusion: RBCs are characterised by an increased spherical shape in many disease states. The measure of the second PCD in flow cytometry is a new, easy method to investigate RBC shape in various diseases. This technique could facilitate the investigation of abnormalities of RBC rheology.