Biochemical characterization of size-separated human red blood cells.

Human red blood cells (RBC) are heterogeneous with respect to their size; the physiological significance of this heterogeneity has not yet been fully elucidated. To further investigate this problem, some characteristics of human RBC fractionated according to their mean corpuscular volume (MCV) by counterflow centrifugation were determined. Larger RBC are more prone to hypotonic lysis. The membrane protein content per cell increases with the MCV, but no obvious difference in the distribution of the major proteins can be demonstrated. The lipid content per cell also rises with the RBC size, while the percentages of the main lipid components do not significantly vary. However, the variations of sialic acid content per RBC according to MCV are more important than those of protein or lipid; thus, the sialic acid-to-protein ratio gradually increases with the MCV. This indicates that, in spite of the lack of major changes in the membrane composition, some qualitative differences exist between large and small cells.

Variations in serum alpha-L-fucosidase activity during childhood and pregnancy.

Variations in serum alpha-L-fucosidase activity (AFU) have been studied during childhood and pregnancy. 994 children, ages 1 day to 15 yr, were examined; no sex-linked difference was found, but significant variations according to age were demonstrated. AFU activity rose during the first 10-15 days after birth, remained high during the second month then decreased till the end of the first year, thereafter no significant changes were observed. In pregnancy, AFU activity rose and dropped quickly after delivery; neither hypertension nor fetal distress led to AFU activity changes during pregnancy. Thus, in addition to the great variability of AFU linked to the genetic polymorphism, the physiological factors such as age or pregnancy have to be taken into account to establish the significance of AFU variations in pathological situations.

Saliva flow and composition in humans exposed to acute altitude hypoxia.

The effects of acute hypoxia (2 days at 4350 m) on whole saliva flow and composition were studied on 12 sea-level natives, at rest and following a maximal exercise. Exercise, performed in normoxia and hypoxia, did not induce variations in saliva flow rate, saliva potassium or alpha-amylase concentrations. In contrast, acute hypoxia did lead to an increase in mean saliva flow rate both at rest (0.63 ml.min-1 to 0.93 ml.min-1, P less than 0.01) and after exercise (0.56 ml.min-1 to 1.06 ml.min-1, P less than 0.05) and a decrease in mean saliva potassium concentration at rest (20.8 mmol.l-1 to 14.7 mmol.l-1, P less than 0.01) as well as after exercise (21.7 mmol.l-1 to 16.5 mmol.l-1, P less than 0.05). This effect might be the consequence of a hypoxia-induced stimulation of the parasympathetic nervous system.

Does red blood cell size correlate with red blood cell age in mouse?

Mouse red blood cells (RBC) can be fractionated according to their size by counterflow centrifugation. The mean corpuscular hemoglobin content and the enzyme activities (ASAT, LDH, PK and acetylcholinesterase) increase when the mean corpuscular volume (MCV) rises. However, the in-vivo survival of size-separated RBC is similar whatever their MCV is; thus, counterflow centrifugation is not a suitable procedure to achieve an age fractionation of mouse RBC. Moreover, RBC subpopulations collected by counterflow centrifugation are different from those obtained when RBC are fractionated according to their density.

Some characteristics of human red blood cells separated according to their size: a comparison with density-fractionated red blood cells.

During their in vivo ageing, red blood cells (RBC) increase in density and become smaller. Age-defined RBC subpopulations are usually collected by centrifugation. A fractionation according to RBC volume has been proposed as an improved alternative to such age separation. Because a few data reported in the literature indicate some discrepancies between the two methods, blood samples were separated either by centrifugation or by counterflow centrifugation, and some characteristics of the RBC thus fractionated were studied. The enzyme activities decrease either when the density rises or when the volume (MCV) decreases. However, the comparison of other RBC characteristics strongly suggests that these two procedures do not lead to the collection of the same RBC subpopulations: for instance, the hemoglobin content increases when the MCV rises, whereas it remains constant whatever the RBC density is. With radiolabelled cells, it is shown 1) that the most dense RBC are recovered in all the size-separated RBC subpopulations, even though they tend to concentrate in the fractions with the largest MCV, and 2) that the smallest RBC are almost fairly distributed in all the RBC subpopulations, whatever their density, whereas the largest RBC are mainly, but not exclusively, present in the high-density fractions. Thus, fractionation according to size does not match separation according to density. Taken together with results from in vivo experiments carried out in mice and with the fact that reticulocytes are present in all the size-separated fractions, these data suggest that counterflow centrifugation may be a very questionable procedure to achieve a RBC fractionation according to age and therefore that RBC volume might not be a reliable criterion of RBC age.

Effect of diosmin upon red blood cell deformability and osmotic fragility. Relationship with lipid content.

Rats were treated by ingesting forcibly 2 ml of a suspension of four different doses (100, 200, 300 and 400 mg/kg) of diosmin in carboxymethylcellulose (CMC) and were killed immediately or after 3 or 6 hours of fasting. Animals treated by CMC only in a similar way (gavage) exhibited a fall in red blood cell (RBC) membrane cholesterol and an increase in RBC rigidity at the third hour while osmotic fragility remained stable. Diosmin treatment opposed the rise in RBC rigidity evoked by the gavage and induced a dose-dependent decrease of the RBC membrane cholesterol over phospholipid ratio.

Osmotic properties of human erythrocytes treated by phospholipase A2 from bee venom.

During treatment of human red cells with phospholipase A2 from bee venom, a linear increase of the MCV and of the osmotic fragility occurs in parallel with the cleavage of the accessible phospholipids. However, even after maximal hydrolysis, i.e. degradation of up to 65% of the phosphatidylcholines and up to 6% of the phosphatidyl-ethanolamines, almost no haemolysis is observed and the median corpuscular fragility is only 7% higher than that of control cells incubated without enzyme. Addition of albumin to the medium results in an important rise of the susceptibility to hypotonic saline solutions. Osmotic fragility curves obtained with red cells submitted to mild phospholipase action show evidence of subpopulations of cells with various sensitivities to osmotic lysis. This phenomenon can be partly explained by the heterogeneity of the cleavage intensity among the cell population. This hypothesis is supported by the studies of the lipid composition of phospholipase treated red cells fractionated according to their sensitivity to hypotonic lysis or to their size.

Properties of rabbit erythrocytes treated with phospholipase A2 from bee venom.

In isotonic saline-buffered conditions, phospholipase A2 from bee venom cleaves up to 65% of the phosphatidylcholine of the rabbit RBC membrane without causing significant hemolysis; however, the volume and the osmotic fragility of the treated RBC is modified. The osmotic behaviour of PLA-treated RBC after reincubation in autologous plasma suggests an heterogeneity in the enzymatic attack among the RBC population. In vivo RBC survival is strongly impaired by PLA treatment.