Red blood cells

Red blood cells are derived from haemopoietic stem cells in bone marrow. Following a series of maturation steps, directed largely by the hormone erythropoietin (Epo), red cells enucleate and enter the circulatory system. In circulation these small, flexible biconcave cells containing haemoglobin transport O₂ from the lungs to the periphery, and CO₂ back from the periphery to the lungs. The most common disorders associated with red blood cells are anaemias. While there are numerous causes of anaemia, the reduced capacity for gaseous exchange is the underlying theme. Over the past 15 years, recombinant Epo has been used extremely successfully in the treatment of several forms of anaemia. The single gene disorders collectively known as haemoglobinopathies represent one of the best opportunities for gene therapy.     

Red blood cells under mechanical stress

The effect of mechanical stress on erythrocytes suspended in various media was studied. The ability of the cells to increase their glucose consumption was found to be the major criterion allowing to divide the media into two groups. In plasma, serum or in Ringer's solution supplemented with albumin and glucose the energy consumption by mechanically stressed erythrocytes increased 20 to 50%; no morphological changes of the cells were observed either in suspension or on Giemsa smears. The cells behaved in the same way in Mg2(+)-free medium. The other group included protein-free medium (Ringer's solution supplemented with glucose) and Ca2(+)-free Ringer's solution supplemented with albumin and glucose; under these conditions erythrocytes were unable to raise their energy consumption in response to mechanical stress, and after some period structural impairment of the membrane could be observed on Giemsa smears. No differences in metabolism-associated nucleotide concentrations (ATP, ADP, NAD, NADP) were observed between the samples. Resealed red cell ghosts with high concentrations of intracellular components were prepared as a model of cells with damaged membrane. In these ghosts (with low ATP concentration) mechanical stress produced increased proportions of echinocytes, even in the "native" suspension. These results have confirmed the vital role of the energy-consuming contractile apparatus in the erythrocyte membrane, and supplied a clue to the role of Ca2+ in its activation and to the influence of extracellular proteins on the maintenance of in red cell shape.     

Red blood cells imaging and antigen-antibody interaction measurement

In the present study the atomic force microscope (AFM) was used to image the surface morphology of red blood cells (RBC) for the first time. The AFM yielded very reproducible images without appreciable modifications of the sample surfaces. In addition to this topographical imaging, we have developed an experimental approach to measure the binding strength between antibody (anti-A), and the RBC antigen A, when reversible bonds between specific molecules such as antigen and antibody mediate the adhesion. The experimental results suggest that the procedure established here may be used for specific antibody detection. This study has also enhanced our understanding under physiological conditions of molecular interaction in particular antigen-antibody.     

Red blood cells that do and red blood cells that don't: how to resist a persistent parasite

Sixty years ago, Haldane proposed that certain abnormalities in red blood cells could be selected as malaria-resistance genes. Population studies have confirmed that many human polymorphisms confer resistance to severe malaria, although the mechanisms of protection remain unknown. A recent article proposes a new mechanism for explaining the protective effects of hemoglobin C (HbC). HbC-containing red blood cells have modified displays of malaria surface proteins that reduce parasite adhesiveness and could reduce the risk of severe disease.     

Red blood cells as an antigen-delivery system.

The use of adjuvants is usually required to induce strong immunological responses to protein antigens. However, in many cases these adjuvants cannot be extensively applied in human and veterinary vaccinations because of associated inflammatory reactions or granuloma formation. We show here that protein antigens (bovine serum albumin, hog liver uricase, and yeast hexokinase), coupled to autologous red blood cells by way of a biotin-avidin-biotin bridge, elicit an immunological response in mice similar to or higher than that obtained by the use of Freund's adjuvant. Quantities as low as 0.5 micrograms/mouse are high enough to generate these immunological responses. Furthermore, splenocytes of mice immunized by red blood cell-coupled antigens can be used to generate hybridomas secreting monoclonal antibodies. Thus, the delivery of antigens by autologous red blood cells is an effective way to avoid the use of adjuvants for producing anti-peptide antibodies and possibly to generate peptide vaccines.     

Red blood cell targeting to smooth muscle cells

Monoclonal antibody discriminating between endothelial and smooth muscle cells is suggested to be used as a vector for directed transport of drugs to injured (denuded) areas of blood vessel wall. An in vitro model system was used in the studies: vascular smooth muscle or endothelial cells grown on plastic surface were treated with specific mouse monoclonal antibody recognizing an antigen localized on the surface of smooth muscle rather than endothelial cells; then erythrocytes coated with secondary (rabbit antimouse) antibodies were added. The results were analyzed spectrophotometrically or with scanning electron microscopy. Under the experimental conditions, erythrocytes, possible 'containers' for carrying the drugs, were found to bind only to smooth muscle cells. The data show that antibody provides absolute discrimination between endothelial and smooth muscle cells and, thus, may be used as a vector for drug targeting.     

Red blood cells augment leukocyte rolling in a virtual blood vessel

Leukocyte rolling and arrest on the vascular endothelium is a central event in normal and pathological immune responses. However, rigorous estimation of the fluid and surface forces involved in leukocyte-endothelial interactions has been difficult due to the particulate, non-Newtonian nature of blood. Here we present a Lattice-Boltzmann approach to quantify forces exerted on rolling leukocytes by red blood cells in a "virtual blood vessel." We report that the normal force imparted by erythrocytes is sufficient to increase leukocyte binding and that increases in tangential force and torque can promote rolling of previously adherent leukocytes. By simulating changes in hematocrit we show that a close "envelopment" of the leukocyte by the red blood cells is necessary to produce significant changes in the forces. This novel approach can be applied to a large number of biological and industrial problems involving the complex flow of particulate suspensions.     

Positional specificity in the incorporation of isomeric cis- and trans-octadecenoic acids into glycerolipids of cultured soya cells

Heterotrophically grown cell suspension cultures of soya (Glycine max L.) were incubated with two different mixed substrates consisting of positional isomers of either cis-[1-¹⁴C]octadecenoic acids (8 to 15) or trans-[1-¹⁴C]octadecenoic acids (8 to 16), each with known composition. With both substrates, about one-fourth of the radioactivity supplied was incorporated into the diacylglycerophosphocholines, while another one-fourth of the radioactivity was almost equally distributed between diacylglycerophos-phoethanolamines and triacylglycerols. All the positional isomers of cis-and trans-octadecenoic acids supplied to the cells were readily incorporated into various classes of glycerolipids. None of the octadecenoic acids was isomerized, elongated or desaturated during incubation. From the cis-octadecenoic acids, only the naturally occurring 9-isomer (oleic acid) was preferentially incorporated into position 2 of diacylglycerophosphocholines, diacylglycerophospho-ethanolamines, and triacyglycerols; all the other isomers exhibited a strong affinity for position 1 of the glycerophospholipids and positions 1 and 3 of the triacylglycerols. From the trans-octadecenoic acids, only the 9-isomer (elaidic acid) was preferentially incorporated into position 2 of diacylglycerophospho-cholines and triacylglycerols; all the other isomers preferred position 1 and positions 1 and 3, respectively, of these lipids. In diacylglycerophospho-ethanolamines, however, each of the trans-octadecenoic acids, including the 9-isomer, exhibited a strong affinity for position 1. Apparently, the enzymes involved in the incorporation of exogenous monounsaturated fatty acids into membrane lipids of plant cells can recognize the preferred substrate in a mixture of closely related isomers.     

Red blood cells protect albumin from cigarette smoke-induced oxidation

Different studies reported the presence of oxidized (carbonylated) albumin in the extravascular pool, but not in the intravascular one of cigarette smokers. In this study we attempted to explain this apparent discrepancy exposing human serum albumin (HSA) to aqueous cigarette smoke extract (CSE). CSE induces HSA carbonylation and oxidation of the HSA Cys34 sulfhydryl group. An antioxidant action of glutathione, cysteine, and its synthetic derivative N-acetylcysteine was observed only at supra-physiological concentrations, suggesting that physiological (plasma) concentrations of glutathione and cysteine in the low micromolar range are ineffective in preventing cigarette smoke-induced oxidation of HSA. Differently, human erythrocytes resulted to be protective towards CSE-induced oxidation (carbonylation and thiol oxidation) of both HSA and total human plasma proteins.     

Enzyme catalyzed hydrolysis of the diesters of cis- and trans-cyclohexanedicarboxylic acids

Summary Pig liver esterase (EC 3.1.1.1) catalyzed hydrolysis of the dimetrhy ester of meso-cis-1,2-cyclohexanedicarboxylic acid yielded the optically pure (1S,2R)-monoester. The corresponding diethyl ester yielded racemic monoester.The diethyl ester of racemic trans-1,2-cyclohexanedicarboxylic acid was kinetically resolved by partial hydrolysis with subtilisin (EC 3.4.21.14) or pig liver esterase. The (1R,2R)-monoester had an enantiomeric excess of 45% and was obtained in an enantiomerically pure form through recrystallisation. The remaining (1S,2S)-diester exhibited an enantiomeric excess of 83%. The nature of the ester function (methyl, ethyl, and propyl esters) had a great influence on the enantiomeric excess obtained and on the kinetic parameters.     

Red blood cells in the metabolism of nitric oxide-derived peroxynitrite

In this review we have analyzed the reactions of nitric oxide (.NO) with superoxide radical (O(2).-) at the vascular compartment which results in limitation of the bioavailability of .NO and the formation of peroxynitrite (ONOO-), a strong oxidant species. The intravascular formation of peroxynitrite can result in oxidative modifications of plasma and vessel wall proteins including the formation of protein-3-nitrotyrosine. The role of red blood cells (RBC) and oxyhemoglobin in the metabolism of intravascular peroxynitrite will be discussed. While RBC constitute an important 'sink' of both .NO and peroxynitrite, redox reactions of these species with oxyhemoglobin may in part contribute to erythrocyte aging. The intravascular formation, reactions and detoxification of peroxynitrite are revealed as important factors controlling vascular dysfunction and degeneration in a variety of pathophysiologically-relevant conditions.     

Red blood cells initiate leukocyte rolling in postcapillary expansions: a lattice Boltzmann analysis

Leukocyte rolling on the vascular endothelium requires initial contact between leukocytes circulating in the blood and the vessel wall. Although specific adhesion mechanisms are involved in leukocyte-endothelium interactions, adhesion patterns in vivo suggest other rheological mechanisms also play a role. Previous studies have proposed that the abundance of leukocyte rolling in postcapillary venules is due to interactions between red blood cells (RBCs) and leukocytes as they enter postcapillary expansions, but the details of the fluid dynamics have not been elucidated. We have analyzed the interactions of red and white blood cells as they flow from a capillary into a postcapillary venule using a lattice Boltzmann approach. This technique provides the complete solution of the flow field and quantification of the particle-particle forces in a relevant geometry. Our results show that capillary-postcapillary venule diameter ratio, RBC configuration, and RBC shape are critical determinants of the initiation of cell rolling in postcapillary venules. The model predicts that an optimal configuration of the trailing red blood cells is required to drive the white blood cell to the wall.     

Lipase from Brassica napus L. discriminates against cis-4 and cis-6 unsaturated fatty acids and secondary and tertiary alcohols

Lipase (EC 3.1.1.3) from oilseed rape (Brassica napus L., cv Ceres) hydrolyzes triacylglycerols containing a broad range of fatty acids at similar rates. In esterification reactions carried out in hexane, rape lipase also uses a wide range of fatty acids and alcohols as reaction partners. However, the rates of esterification of petroselinic, gamma-linolenic, stearidonic and docosahexaenoic acids are only between 2 and 7% that of oleic acid. The common feature of these fatty acids is that the first double bond is cis-4 or cis-6. Petroselaidic acid with a trans-6 double bond is esterified about 10-times faster than petroselinic acid. Arachidonic and eicosapentaenoic acids, both with the first double bond being cis-5, are esterified about 20-times faster than docosahexaenoic acid. By analogy, tripetroselinin and tri-gamma-linolenin are hydrolyzed at 14% and 1.5%, respectively, of the rate of triolein hydrolysis. The rape lipase esterifies primary alcohols but cannot esterify secondary and tertiary alcohols.