Assessment of erythrocyte shape by flow cytometry techniques.

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.

[A patient under hemodialysis with acute distal ischaemia: cholesterol crystal embolism]. / Ischémie aiguë distale chez un patient dialysé: la maladie des emboles de cholésterol.

A 61-year old man was admitted in our hospital for distal ischaemic lesions of the right foot with necrosis of the fifth toe. The lesions appeared a few days before admission. He was known for severe atheromatous disease and multiple aortic and femoro-popliteous aneurysms. He presented also numerous cardiovascular risk factors and a terminal chronic kidney disease was found 18 months earlier. The etiology of the kidney disease was not clear. The patient received hemodialysis since one month. We suspected a severe peripheral arterial disease but the investigations showed preserved distal arterial blood flow what made us think about a thromboembolic disease. We ruled out a potential embolic cardiopathy and, after finding eosinophilia and a previous consultation report of livedo, we suggested the diagnosis of cholesterol crystal embolisation. It was confirmed by the macroscopic examination of the necrotic toe. It's a rare and under-diagnosed pathology with a variable presentation. Though, it's important to detect this affection early as possible to prevent the frequent recurrences. The acute mortality is about 15%. There is no specific treatment, but an an aggressive control of the atheromatous disease is important. We must also avoid the known potential triggering factors: vascular surgery, angiography and anticoagulant or thrombolytic medications.

Resistance to erythropoietin in iron-overloaded haemodialysis patients can be overcome by ascorbic acid administration.

Haemodialysis patients with iron overload sometimes develop resistance to erythropoietin therapy due to 'functional iron deficiency'. It is known that this resistance may be overcome by iron supplementation; however, the latter could worsen haemosiderosis. Therefore, we treated four iron-overloaded haemodialysis patients who had developed relative resistance to erythropoietin (among whom three had features of 'functional iron deficiency') with ascorbic acid (500 mg intravenously after haemodialysis, 1-3 times a week). The erythropoietin doses were voluntarily kept unchanged during the study. After a latency of 2-4 weeks, haematocrit and haemoglobin had increased respectively from 26.5 +/- 0.7 to 32.7 +/- 0.4 vol% and from 8.8 +/- 0.3 to 10.8 +/- 0.2 g/dl (means +/- SEM, P < 0.001). While serum ferritin remained unchanged, transferrin saturation increased from 27 +/- 7 to 54 +/- 12% (P < 0.05), suggesting that ascorbic acid supplementation had allowed mobilization of iron from tissue burdens. In one patient, haematocrit declined after withdrawal of vitamin C and increased again after rechallenge. Also, ascorbate supplementation was continued after the study in two patients and allowed the erythropoietin doses to be decreased, 8 and 11 weeks, respectively, after the start of the trial. When a control group of seven patients with normal iron status and without resistance to erythropoietin were challenged in the same manner with ascorbate, no elevation of haematocrit or transferrin saturation was noted. We conclude that ascorbate supplementation may circumvent resistance to erythropoietin that sometimes occurs in iron-overloaded patients, in particular, in the setting of 'functional iron deficiency'.

High haematocrit in haemodialysis patients can be controlled by theophylline administration.

Three patients on chronic maintenance haemodialysis have progressively increased their haematocrit to reach values between 40 and 45%, a situation associated with an increased risk of thrombosis of their arteriovenous fistulae. Two of them had been submitted to repeated phlebotomies, which remained unsuccessful despite the induction of a profound iron deficiency in one of them. Hence, a trial with oral theophylline was performed in the three patients, resulting in a sustained decrease of the haematocrit (from 43.6 to 33%) and endogenous erythropoietin (from 46 to 15 mU/ml) levels. In two patients, theophylline therapy was stopped transiently due to gastrointestinal side-effects, which resulted in a rapid return to previous haematocrit levels; rechallenge with a better tolerated preparation, however, was efficient again. We conclude that oral theophylline appears to be an efficient treatment to control too high haematocrit levels in dialysis patients.

Factors related to early mortality in cirrhotic patients bleeding from varices and treated by urgent sclerotherapy.

Variceal haemorrhage in cirrhotic patients carries a high early mortality even when balloon tamponade or emergency sclerotherapy are applied. The aim of this study to identify patients dying within six weeks of their first variceal haemorrhage. One hundred and twenty one patients with parenchymal cirrhosis presenting with the first variceal bleeding episode between June 1983 and December 1988 were studied. Nineteen patients were excluded for various reasons. Emergency sclerotherapy was carried out in cases of active bleeding or where there were endoscopic signs of recent bleeding, and then regularly repeated afterwards. Of the 24 variables studied and included in a multivariate analysis using a logistic regression model, three had an independent prognostic value: encephalopathy, prothrombin time, and the number of blood units transfused within the 72 hours of time zero. The subsequent regression equation was able to predict 89% of the patients who will die and 97% of the patients who will still be alive six weeks after their first variceal haemorrhage treated by sclerotherapy. Pugh score was less discriminatory than these last three variables in terms of accuracy of adjustment, goodness of fit to the model, receiver operating characteristic curves, and percentage correct prediction. To measure the accuracy of the prediction rule, our model was applied to another series of 28 cirrhotic patients admitted with their first variceal bleeding during the next period (January 1989 to May 1990). Death and survival were correctly predicted in respectively 82% and 94% of the cases. The use of this score is recommended for the selection of patients with high early mortality after variceal bleeding despite sclerotherapy, and for the design of new therapeutic trials.