Adhesion Of Erythrocytes Research Articles

Endothelial Function in Patients with Sickle Cell Anemia During and After Sickle Cell Crises

Prior studies have demonstrated increased adherence of sickle cell erythrocytes to vascular endothelial cells. While decreased production of nitric oxide and increased production of adhesion molecules have been implicated in this pathophysiology, the relative contribution of these mechanisms during acute sickle cell crises as compared to steady state conditions have not been elucidated. We studied 10 consecutive young adult patients presenting with a sickle cell crisis. Endothelial function was evaluated by a non-invasive brachial artery shear stress method. Serum levels of adhesion molecules were obtained during the crisis. Both brachial artery responsiveness and serum levels of adhesion molecules were then repeated at steady state. Ten age and gender matched volunteers served as a control group. Impaired endothelial function and impaired endothelium-independent vasodilatation were observed in all sickle cell patients during both steady state and during crisis. Flow mediated dilation (FMD)% was 3.25+/- 2.76% during crisis, 4.57+/- 4.11 at steady state, compared with the control group FMD of 11.64+/- 7.69% (p< 0.001). Flow independent dilation was 10.35+/- 11.3% during crisis, 10.03+/- 6.52% at steady state, compared with control group FID of 24.17+/- 11.87% (p< 0.001). Levels of cell adhesion molecules and markers of inflammation were increased in sickle cell crisis patients compared with the control group: sCD40 ligand levels during the acute crisis were over twice the level of normal matched volunteers (p=0.02), and similarly significant increases were seen for E-selectin (p=0.008), ICAM-1 (p=0.037) and VCAM-1 levels (p=0.01). The levels of each of these biomarkers was not significantly increased during acute crises as compared to patients' recovery state. Sickle cell anemia patients have severe systemic endothelial dysfunction as demonstrated by both brachial artery assessment and increased serum levels of adhesion molecules. These abnormalities characterize not only the sickle cell crisis but also the steady state pathophysiology of sickle cell anemia.

Transgenic Sickle Mice Are Markedly Sensitive to Renal Ischemia-Reperfusion Injury

Ischemic injury is invoked as a mechanism contributing to end-organ damage and other complications of sickle cell disease (SCD). However, the intrinsic sensitivity of tissues in SCD to ischemic insults has never been addressed. We examined the effect of renal ischemia in a transgenic mouse expressing human sickle hemoglobin. Twenty-four hours after bilateral, total renal artery occlusion for 15 minutes, transgenic sickle mice exhibited worse renal function and more marked histological injury. With bilateral renal ischemia of greater duration (22.5 minutes), and after 6 hours, transgenic sickle mice exhibited massive vascular congestion, sickling of red blood cells, more marked histological injury in the kidney, and more prominent congestion in the capillary beds in the lungs and heart. Additionally, serum amyloid P-component, the murine homologue of C-reactive protein, was markedly increased in transgenic sickle mice as compared to wild-type mice. Twenty-four hours after bilateral renal ischemia for 22.5 minutes, transgenic sickle mice exhibited 28% mortality, with no mortality observed in any other group. With bilateral renal ischemia of short or long duration, renal expression of caspase-3 was most prominent in transgenic sickle mice subjected to ischemia. Thus, renal ischemia in this murine model induces more severe renal injury and extrarenal complications. We conclude that tissues in SCD exhibit heightened vascular congestion and sensitivity to ischemia and that clinically apparent or silent episodes of ischemia may contribute to the complications of SCD.

Identifying Plasmodium falciparum cytoadherence‐linked asexual protein 3 (CLAG 3) sequences that specifically bind to C32 cells and erythrocytes

Adhesion of mature asexual stage Plasmodium falciparum parasite-infected erythrocytes (iRBC) to the vascular endothelium is a critical event in the pathology of Plasmodium falciparum malaria. It has been suggested that the clag gene family is essential in cytoadherence to endothelial receptors. Primers used in PCR and RT-PCR assays allowed us to determine that the gene encoding CLAG 3 (GenBank accession no. NP_473155) is transcribed in the Plasmodium falciparum FCB2 strain. Western blot showed that antisera produced against polymerized synthetic peptides from this protein recognized a 142-kDa band in P. falciparum schizont lysate. Seventy-one 20-amino-acid-long nonoverlapping peptides, spanning the CLAG 3 (cytoadherence-linked asexual protein on chromosome 3) sequence were tested in C32 cell and erythrocyte binding assays. Twelve CLAG peptides specifically bound to C32 cells (which mainly express CD36) with high affinity, hereafter referred to as high-affinity binding peptides (HABPs). Five of them also bound to erythrocytes. HABP binding to C32 cells and erythrocytes was independent of peptide charge or peptide structure. Affinity constants were between 100 nM and 800 nM. Cross-linking and SDS-PAGE analysis allowed two erythrocyte binding proteins of around 26 kDa and 59 kDa to be identified, while proteins of around 53 kDa were identified as possible receptor sites for C-32 cells. The HABPs' role in Plasmodium falciparum invasion inhibition was determined. Such an approach analyzing various CLAG 3 regions may elucidate their functions and may help in the search for new antigens important for developing antimalarial vaccines.

Erythrocyte Adhesion Receptors: Blood Group Antigens and Related Molecules

During the second half of the 20th century, blood bankers quickly expanded our knowledge of human erythrocyte blood group antigens. By the dawn of the 21st century, several hundred blood group antigen polymorphisms had been identified. Hot on the heels of the serologists, membrane biochemists and molecular geneticists defined both the biochemical and genetic bases of most of these antigens. Perhaps to their surprise, this work has led to the discovery of functionally diverse and important membrane proteins expressed on the surface of red cells, including numerous adhesion molecules. Red cells express an unexpected number of such adhesion receptors, some of which contribute to human disease, as well as to normal red cell development. And perhaps most interestingly, study of these molecules has elucidated ways in which even mature red cells respond to external stimuli, such as adrenergic hormones.

Epinephrine acts through erythroid signaling pathways to activate sickle cell adhesion to endothelium via LW-αvβ3 interactions

The possible role of physiologic stress hormones in enhancing adhesion of sickle erythrocytes (SS RBCs) to endothelial cells (ECs) in sickle cell disease (SCD) has not been previously explored. We have now found that up-regulation of intracellular cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) by epinephrine significantly increased sickle but not normal erythrocyte adhesion to both primary and immortalized ECs. Inhibition of serine/threonine phosphatases also enhanced sickle erythrocyte adhesion at least partially through a PKA-dependent mechanism. Adhesion was mediated through LW (intercellular adhesion molecule-4 [ICAM-4], CD242) blood group glycoprotein, and immunoprecipitation studies showed that LW on sickle but not on normal erythrocytes undergoes increased PKA-dependent serine phosphorylation as a result of activation. The major counter receptor for LW was identified as the alphavbeta3 integrin on ECs. These data suggest that adrenergic hormones such as epinephrine may initiate or exacerbate vaso-occlusion and thus contribute to the association of vaso-occlusive events with physiologic stress.

Two Distinct Domains Mediate Self-Assembly of Knob Protein in Plasmodium Falciparum-Infected Human Erythrocytes.

Plasmodium falciparum associated mortality is believed to originate from the adhesion of infected erythrocytes to the endothelial cells. The adhesion of mature parasite infected erythrocytes is mediated by the surface protrusions called knobs. A dominant constituent of knobs is the parasite derived knob-associated histidine-rich protein termed KAHRP or knob protein. Gene deletion experiments have shown that the knob protein is essential in malaria pathogenesis. Without the knob protein (KAHRP), adherence of malaria-infected erythrocytes is not established under physiological shear stress. The adhesive properties of infected erythrocytes are propagated by the direct interaction of KAHRP, a peripheral membrane protein, with the cytoplasmic domain of the P. falciparum erythrocyte membrane protein-1 (PfEMP1). Unlike the highly polymorphic extracellular domain, the cytoplasmic domain of PfEMP1 is conserved across species, hence the interest in delineating the mechanism of its interaction with KAHRP and components of the erythrocyte membrane. Our studies have observed recombinant KAHRP to self-associate into large aggregates in vitro. Under transmission electron microscopy, both knobs isolated from malaria-infected erythrocytes and self-assembled recombinant KAHRP in vitro have revealed a unique spiral shaped morphology. Based on these observations, we hypothesized that the unique knob spiral is formed by the self-assembly of KAHRP monomers and therefore must be encoded by distinct sequence elements within KAHRP. To investigate this speculation, we devised a solution binding assay and engineered a series of GST (glutathione S-transferase) and Trx (thioredoxin) fusion peptides to encompass the entire length of KAHRP. Binding measurements using heterologous combination of defined fusion peptides reveal a core histidine-rich amino terminal segment associating with a lysine-rich segment located within the carboxyl half of KAHRP. These findings suggest a novel head-to-tail interaction strategy employed by the intracellular parasite to assemble the spiral-shaped scaffold of KAHRP under the surface of electron dense knobs. Significantly, our results identify a novel self-interaction site within KAHRP that could be amenable to therapeutic intervention against cerebral malaria.

Effect of Hydroxyurea Treatment on In Vitro Adhesion of Sickle Erythrocytes to Sickle Leukocyte Subpopulations.

Microvascular vaso-occlusion in sickle cell disease is thought to involve adhesive interactions among erythrocytes (RBCs), leukocytes and vascular endothelial cells. Recent studies have demonstrated the presence of a significant inflammatory response in sickle cell disease, including changes in the cell surface adhesion molecules that mediate cell-cell interactions in the microvasculature. In this study, we used a parallel-plate flow chamber assay to determine the subpopulations of leukocytes that are involved in sickle leukocyte-RBC interactions. We also studied the effect of treatment with hydroxyurea (HU) on these adhesive interactions. Populations of monocytes, neutrophils (PMNs) and T cells were isolated by negative selection from the peripheral blood of untreated patients with sickle cell disease (SS), sickle patients receiving HU (SS-HU), and healthy control subjects (AA). Adhesive interactions involving these leukocyte subpopulations, human umbilical vein endothelial cells (HUVECs) pretreated with tumor necrosis factor-α (TNF-α ), and autologous RBCs were measured under a shear stress of 1 dyne/cm2. Compared to the corresponding cell populations from AA individuals, PMNs, monocytes, and T cells from SS individuals were significantly more adherent to TNF-α-treated HUVECs (774±59 vs. 502±27 cells/mm2, p=0.001; 533±66 vs. 348±36 cells/mm2, p=0.024; and 470±75 vs. 227±26 cells/mm2, p=0.009, respectively). HU therapy significantly decreased the adhesion of SS PMNs to HUVECs (774±59 cells/mm2 for SS vs. 604±36 for SS-HU, p=0.025). Compared to adherent AA leukocytes, adherent SS leukocytes exhibited greater participation in adhesive interactions with autologous RBCs (41±3% for SS vs. 27±3% for AA, p=0.002), and HU treatment decreased the fraction of leukocytes that captured autologous RBCs to the control level (29±3% for SS-HU, p=0.006 vs. SS). Compared to adherent PMNs from SS individuals, adherent PMNs from SS-HU individuals showed significantly reduced participation in the capture of RBCs (53±6% for SS vs. 35±5% for SS-HU, p=0.021). Although adherent T cells from SS individuals participated significantly more in RBC capture than adherent T cells from AA individuals (28±5% for SS vs. 10±2% for AA, p=0.007), HU therapy did not have a significant effect on this parameter (21±5% for SS-HU, p=0.373). Compared to AA leukocytes, SS leukocytes captured more RBCs per participating adherent leukocyte (2.8±0.2 vs. 1.9±0.1 RBCs/cell, p=0.001). HU therapy reduced the number of RBCs captured per PMN but not the number captured per T cell. Compared to AA T cells, SS T cells captured adherent RBCs for a significantly longer period of time (51±9 vs. 26±6 seconds, p=0.035). Our data suggest that sickle neutrophils, monocytes and T cells may all be involved in adhesive interactions with sickle RBCs. PMN-RBC and monocyte-RBC interactions appear to be more numerous than T cell-RBC interactions, although T cell-RBC interactions may be stronger. HU therapy appears to target PMN-RBC and monocyte-RBC interactions preferentially. Future studies will focus on the role of particular adhesion molecules in mediating these interactions and on the potential for therapeutic interventions targeting cell-cell adhesion.

Modulation of Erythrocyte Adhesion by Changes in Cellular Tonicity and Volume.

The sickle red blood cell (SS-RBC) membrane manifests many abnormal properties, including oxidative damage of membrane proteins and lipids, disruption of the lipid bilayer, abnormal clustering of surface proteins, dehydration, and increased adhesive properties. Conditions that alter the RBC membrane may modify or abnormally expose its native components and contribute to the adhesive properties of the RBC. We have previously reported that sickle RBCs have enhanced adhesion to the plasma and subendothelial matrix protein thrombospondin-1 (TSP) under conditions of flow in vitro. Since a significant proportion of sickle RBCs are inherently dehydrated, in part due to intracellular K+ loss via the Gardos and K-Cl co-transport channels, we test the hypothesis that dehydration-induced alterations in membrane organization or composition contribute to sickle cell adhesion in sickle cell disease (SCD). To examine the role of RBC hydration in adhesion to the subendothelial matrix protein thrombospondin-1 (TSP), normal and sickle RBCs were incubated in buffers of varying tonicity and tested for adhesion to immobilized TSP under flow conditions. Expanding on our previous preliminary results, we found that sickle RBCs exhibited a decrease in TSP binding with increasing cell hydration (p<0.005), suggesting that cellular dehydration may contribute to its adhesive phenotype. Consistent with this hypothesis, normal RBCs showed an increase in TSP adhesion with increasing dehydration (p<0.01). Furthermore, TSP adhesion of both sickle RBCs and dehydrated normal RBCs was inhibited by the anionic polysaccharides chondroitin sulfate A and high molecular weight dextran sulfate, but not by the anionic polysaccharide chondroitin sulfate C. TSP adhesion of sickle and dehydrated normal RBCs in our in vitro system was also not inhibited by competitors of CD47-, band 3-, or RBC phosphatidylserine-meditaed adhesion. Increased TSP adhesion of normal RBCs could also be induced by isotonic dehydration using nystatin-sucrose buffers; this adhesion was also inhibited by chondroitin sulfate A and high molecular weight dextran sulfate but not by chondroitin sulfate C. More importantly, we found increased adhesion of nystatin-sucrose dehydrated normal mouse RBCs, as compared to untreated normal murine RBCs, to kidney capillaries following reinfusion in vivo. In summary, these findings demonstrate that changes in hydration can significantly impact adhesion, and suggest that changes in volume are more important than changes in intracellular tonicity in the adhesion of RBCs to TSP. There are striking similarities in the adhesive properties of sickle and dehydrated normal RBCs, supporting the theory that adhesion of sickle RBCs arises as the result of exposure or modification of normal erythroid membrane components. Characterization of the membrane epitope(s) responsible for adhesion of dehydrated normal erythrocytes to TSP may thus provide a parallel avenue to identify reagents that can inhibit RBC adhesion and resultant vasoocclusion in SCD.

Sickle cell adhesion depends on hemodynamics and endothelial activation

Venular microvascular circulation in patients with sickle cell anemia exhibits reduced and episodic blood flow. Sickle erythrocyte adhesion to postcapillary venule endothelium is postulated to initiate and propagate episodes of vasoocclusive pain. Hemodynamics likely mediate the adherence of sickle cells to endothelium, controlling delivery of potentially adherent erythrocytes and removal of loosely adherent erythrocytes on the endothelium. This study found a high dependence on shear stress of sickle erythrocyte adhesion to vascular cell adhesion molecule-1 (VCAM-1) on endothelium stimulated by tumor necrosis factor-alpha (TNF-alpha). Shear stress varied from 1.0 dyne/cm 2 (microvascular venular flow), in which VCAM-1 ligand interactions induced by TNF-alpha primarily controlled adherence, to 0.1 dyne/cm 2 (low flow), in which stimulation had little effect on adherence. At shear stresses analogous to in vivo velocities from laser Doppler ultrasound studies (0.8 and 0.6 dyne/cm 2 ), TNF-alpha promoted 1.9- and 2.7-fold increases in adhesion compared with unstimulated (baseline) adherence. These findings suggest a dynamic vasoocclusive process that depends on both receptor expression and shear stress. These results indicate that, in the microvasculature, slightly reduced inflow rate, increased receptor expression, or both may result in large increases in sickle erythrocyte adhesion.

CELL ADHESION TO POLYMER SUBSTRATES CHARACTERIZED BY THE MICROPIPETTE ASPIRATION TECHNIQUE

A micropipette suction method was adapted for the characterization of cell adhesion to polymer surfaces. The instrument was applied in experiments probing the adhesion of human erythrocytes to polymer films which had been precoated with monoclonal antibodies against two different transmembrane proteins of the cells. Cells were impinged on the polymer substrates and subsequently removed by stepwise micropipette aspiration. Variations of shape and contact area of the cells during micropipette aspiration-driven detachment were evaluated to determine the separation energy. A strong increase of the separation energy with decreased contact area was observed and explained by the smoothing of the cell membrane at elevated membrane tensions. The results indicate that the overall strength of attachment was determined by the amount and availability of the adsorbed antibodies, while the separation of the cells from the polymer substrates occurred, in general, due to dislocation of the transmembrane proteins from the membrane.

Chondroitin sulfate proteoglycans of bovine cornea: structural characterization and assessment for the adherence of Plasmodium falciparum-infected erythrocytes

The structures of the bovine corneal chondroitin sulfate (CS) chains and the nature of core proteins to which these chains are attached have not been studied in detail. In this study, we show that structurally diverse CS chains are present in bovine cornea and that they are mainly linked to decorin core protein. DEAE-Sephacel chromatography fractionated the corneal chondroitin sulfate proteoglycans (CSPGs) into three distinct fractions, CSPG-I, CSPG-II, and CSPG-III. These CSPGs markedly differ in their CS and dermatan sulfate (DS) contents, and in particular the CS structure—the overall sulfate content and 4- to 6-sulfate ratio. In general, the CS chains of the corneal CSPGs have low to moderate levels (15–64%) of sulfated disaccharides and 0–30% DS content. Structural analysis indicated that the DS disaccharide units in the CS chains are segregated as large blocks. We have also assessed the suitability of the corneal CSPGs as an alternative to placental CSPG or the widely used bovine tracheal chondroitin sulfate A (CSA) for studying the structural interactions involved in the adherence of Plasmodium falciparum-infected red blood cells (IRBCs) to chondroitin 4-sulfate. The data demonstrate that the corneal CSPGs efficiently bind IRBCs, and that the binding strength is either comparable or significantly higher than the placental CSPG. In contrast, the IRBC binding strength of bovine tracheal CSA is markedly lower than the human placental and bovine corneal CSPGs. Thus, our data demonstrate that the bovine corneal CSPG but not tracheal CSA is suitable for studying structural interactions involved in IRBC-C4S binding.

Chemical modifications of band 3 protein affect the adhesion of Plasmodium falciparum-infected erythrocytes to CD36.

The role of the erythrocyte anion exchanger, band 3 protein (AE1), in the adhesion of Plasmodium falciparum-infected erythrocytes to CD36 and thrombospondin (TSP) was studied. Two specific anion exchange inhibitors that bind covalently to different regions of the band 3 molecule affected cytoadherence in dissimilar ways. Modification of lysine 539 by diisothiocyanostilbene sulfonic acid (DIDS) resulted in a significant reduction in the adhesive properties of parasitized erythrocytes for CD36, but not TSP, whereas treatment with fluorescein-5-maleimide, which modifies lysine 430, was without effect on both TSP and CD36 binding. The adhesive properties of the DIDS binding region (DBR) was demonstrated by competition experiments using synthetic peptides and by direct interaction of such peptides with CD36 transfected CHO cells. The results suggest that host membrane proteins such as AE1 contribute to the adhesion of malaria-infected erythrocytes to CD36.

Chemical modifications of band 3 protein affect the adhesion of Plasmodium falciparum-infected erythrocytes to CD36

The role of the erythrocyte anion exchanger, band 3 protein (AE1), in the adhesion of Plasmodium falciparum -infected erythrocytes to CD36 and thrombospondin (TSP) was studied. Two specific anion exchange inhibitors that bind covalently to different regions of the band 3 molecule affected cytoadherence in dissimilar ways. Modification of lysine 539 by diisothiocyanostilbene sulfonic acid (DIDS) resulted in a significant reduction in the adhesive properties of parasitized erythrocytes for CD36, but not TSP, whereas treatment with fluorescein-5-maleimide, which modifies lysine 430, was without effect on both TSP and CD36 binding. The adhesive properties of the DIDS binding region (DBR) was demonstrated by competition experiments using synthetic peptides and by direct interaction of such peptides with CD36 transfected CHO cells. The results suggest that host membrane proteins such as AE1 contribute to the adhesion of malaria-infected erythrocytes to CD36.

Effect of Different Exercise Modes on Immune Adherence of Erythrocytes

Effect of Different Exercise Modes on Immune Adherence of Erythrocytes

Effect of Different Exercise Modes on Immune Adherence of Erythrocytes

Effect of Different Exercise Modes on Immune Adherence of Erythrocytes

An antimalarial extract from neem leaves is antiretroviral

An acetone–water neem leaf extract with antimalarial activity was evaluated in vitro at 5 μg/ml for inhibition of adhesion of malaria parasite-infected erythrocytes and cancer cells to endothelial cells, and at 10 μg/ml for protection of lymphocytes against invasion by HIV. The extract was also evaluated in 10 patients with HIV/AIDS at 1000 mg daily for 30 d. The mean binding of infected erythrocytes and cancer cells per endothelial cell was 15 and 11 respectively in the absence of the extract, and 0 and 2 respectively in with the extract. In the absence and presence of the extract, 0% and 75%, respectively, of lymphocytes were protected. In the treated patients, haemoglobin concentration, mean CD4 + cell count and erythrocyte sedimentation rate, which were initially 9.8 g/dl, 126 cells/μl and 90 mm/h respectively, improved to 12.1 g/dl, 241 cells/μl and 49 mm/h. Mean bodyweight and platelet count, initially 57 kg and 328×10 3/mm 3 respectively, increased to 60 kg and 359×10 3/mm 3. No adverse effects were observed during the study. The extract showed antiretroviral activity with a mechanism of action that may involve inhibition of cytoadhesion. The results may help in the development of novel antiretroviral and antimalarial drugs.

Placenta cryosections for study of the adhesion of Plasmodium falciparum-infected erythrocytes to chondroitin sulfate A in flow conditions

The adhesion of Plasmodium falciparum-infected erythrocytes (IEs) to chondroitin-4-sulfate (CSA) via the PfEMP1-CSA parasite ligand domain is correlated with placental malaria in primigravidae. The recent identification of parasite genes encoding CSA adhesion molecules and the development of pan-reactive monoclonal antibodies against the Pf CSA ligand have opened up new avenues for the development of anti-IE sequestration therapies for the prevention of placental malaria. A model closely mimicking placental sequestration of IEs during pregnancy is needed for the preclinical and clinical evaluation of candidate molecules for the induction of antibodies that could protect pregnant women from placental malaria. We found that normal placenta cryosections were a specific and highly consistent support for the binding of IEs to CSA in flow conditions under physiological conditions. This model makes possible the quantitative and qualitative analysis of IE adhesion. We identified distinct CSA-binding phenotypes within the FCR3 CSA-selected parasites in flow analyses, but not in static analyses. We also analyzed inhibitors of placental parasite binding such as soluble CSA and antibodies directed against the Pf CSA ligand. Our data demonstrate that placenta cryosections could be used to standardize assays between laboratories, potentially advancing the development of therapies against placental malaria.

Increased erythrocyte adhesion in mice and humans with hereditary spherocytosis and hereditary elliptocytosis

AbstractMice with disruptions of the red blood cell (RBC) cytoskeleton provide severe hemolytic anemia models in which to study multiorgan thrombosis and infarction. The incidence of cerebral infarction ranges from 70% to 100% in mice with α-spectrin deficiency. To determine whether mutant RBCs abnormally bind adhesive vascular components, we measured adhesion of mouse and human RBCs to immobilized human thrombospondin (TSP) and laminin (LM) under controlled flow conditions. Mutant RBCs had at least 10-fold higher adhesion to TSP compared with normal RBCs (P < .006). Mutant relative to unaffected RBC adhesion to LM was significantly (P < .01) increased as well. Treatment of RBCs with the anionic polysaccharide dextran sulfate inhibited mutant RBC adhesion to TSP (P < .001). Treatment of RBCs with antibodies to CD47 or the CD47-binding TSP peptide 4N1K did not inhibit TSP adhesion of RBCs. Previously, we have shown that infarcts in α-spectrin–deficient sph/sph mice become histologically evident beginning at 6 weeks of age. TSP adhesion of RBCs from 3- to 4- and 6- to 8-week-old sph/sph mice was significantly higher than RBCs from adult mice (> 12 weeks old; P < .005). While the mechanism of infarction in these mice is unknown, we speculate that changes in RBC adhesive characteristics contribute to this pathology.

Plasmodium falciparum-infected erythrocytes demonstrate dual specificity for adhesion to hyaluronic acid and chondroitin sulfate A and have distinct adhesive properties.

Hyaluronic acid (HA) and chondroitin sulfate A (CSA) have been identified as receptors for adhesion of Plasmodium falciparum-infected erythrocytes (IEs) and appear to be involved in mediating parasite accumulation in the placenta. We demonstrate here that some, but not all, parasite populations can adhere to both receptors, and we identify distinguishing features of adhesion. Adhesion to HA and CSA was greatest among pigmented trophozoite-infected erythrocytes and at physiologic pH and was associated with a lack of rosette formation and little adhesion to CD36 and intercellular adhesion molecule-1. Adhesion to HA was sensitive to trypsin cleavage of the IE surface, whereas trypsin-resistant and trypsin-sensitive CSA adhesion were both observed. Soluble HA, but not CSA, could cause aggregation or clumping of IEs. Different HA types varied in adhesion-inhibitory activity, which was altered by physical treatment, suggesting that structural features of HA influence IE interactions. These findings have important implications for understanding the pathogenesis of malaria, particularly in pregnancy.

Endothelial cell P-selectin mediates a proinflammatory and prothrombogenic phenotype in cerebral venules of sickle cell transgenic mice.

Whereas the adhesion of leukocytes and erythrocytes to vascular endothelium has been implicated in the vasooclusive events associated with sickle cell disease, the role of platelet-vessel wall interactions in this process remains undefined. The objectives of this study were to: 1) determine whether the adhesion of platelets and leukocytes in cerebral venules differs between sickle cell transgenic (betaS) mice and their wild-type (WT) counterparts (C57Bl/6) under both resting and posthypoxic conditions, and 2) define the contributions of P-selectin to these adhesion processes. Animals were anesthetized, and platelet and leukocyte interactions with endothelial cells of cerebral postcapillary venules were monitored and quantified using intravital fluorescence microscopy in WT, betaS, and chimeric mice produced by transplanting bone marrow from WT or betaS mice into WT or P-selectin-deficient (P-sel(-/-)) mice. Platelet and leukocyte adhesion to endothelial cells in both unstimulated and posthypoxic betaS mice were significantly elevated over WT levels. Chimeric mice involving bone marrow transfer from betaS mice to P-sel(-/-) mice exhibited a profound attenuation of both platelet and leukocyte adhesion compared with betaS bone marrow transfer to WT mice. These findings indicate that betaS mice assume both an inflammatory and prothrombogenic phenotype, with endothelial cell P-selectin playing a major role in mediating these microvascular responses.