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Biomedical subjects

D T Chiu

Publications and source records attributed to D T Chiu.

At least 91 records · Page 5Linked to original sources
PubMed1989

Enrichment of two glycosyl-phosphatidylinositol-anchored proteins, acetylcholinesterase and decay accelerating factor, in vesicles released from human red blood cells.

Several proteins are attached to the cell membrane by a glycosyl-phosphatidylinositol (GPI) anchor. In this report, we show that during vesiculation of human RBCs in vitro, two of these proteins, acetylcholinesterase and decay accelerating factor, redistribute on the cell surface and become enriched in the released vesicles. As a result, the remnant cells are depleted of these proteins. We suggest that alterations in the architecture of the RBC membrane that precede vesiculation lead to selective polarization of GPI-anchored proteins within the domain of the membrane destined to become a vesicle. Since vesiculation occurs in many cell types, and if the loss of GPI-anchored proteins accompanies this process, it may have important biologic significance.

Acetylcholinesterase↗
PubMed1989

Red cell deformability and lipid composition in two forms of acanthocytosis: enrichment of acanthocytic populations by density gradient centrifugation.

Whole cell deformability and lipid determinations were performed on red cells from two patients who had acanthocytes in the peripheral blood (10% and 20% to 30%) and normal serum lipoprotein levels. One patient had typical chorea-acanthocytosis and the other had no clinical abnormalities associated with acanthocytosis. Red cells from the patient with chorea-acanthocytosis showed reduced deformability, as measured by a visco-diffractometric method (ektacytometry), which could be explained by the presence of increased numbers of dehydrated cells containing high concentrations of hemoglobin. The total cell population showed a modest increase in potassium efflux, which may be responsible for reduced cation content and dehydration in a subpopulation of cells. When the patients' red cells were separated into different density populations by centrifugation on density gradients, the cells of classic acanthocyte morphology were concentrated in the high-density layers. This was true for both patients, although the subject with acanthocytes and no clinical disorder had a normal red cell density distribution. Lipid analysis of both types of acanthocytes showed an increase in the relative proportion of sphingomyelin with respect to the glycerophospholipids. Total cholesterol and phospholipid levels were reduced in the chorea-acanthocytosis red cells, but the other acanthocytes did not differ significantly in total lipid content from normal control samples. Thus, the one common abnormal feature in these two forms of acanthocytosis is the increase in the proportion of red cell sphingomyelin. Because this is a very stable, immobile component of the membrane, we suggest that its relative enrichment could result from a defect in the transport and maintenance of glycerophospholipids. Further study of the dynamics of glycerophospholipid organization in acanthocytes may be useful in increasing our understanding of the genesis of abnormal, acanthocytic morphology.

Acanthocytes↗
PubMed1989

Qinghaosu-mediated oxidation in normal and abnormal erythrocytes.

Qinghaosu, a potent antimalarial agent, has recently been shown to act via oxidative mechanisms. Hence, we have investigated what effect qinghaosu has on cellular oxidation in normal and oxidant-sensitive red blood cells (RBCs). At 500 mumol/L, qinghaosu was found to directly alter red cell deformability (DI) in both normal (hemoglobin AA) and abnormal (hemoglobins SS, AE, and EE) RBCs, with the maximum DI being 70% to 80% of that of untreated RBCs. Although concentrations of less than or equal to 200 mumol/L qinghaosu had a minimal effect on the maximum DI, qinghaosu was found to act as an efficient prooxidant at these concentrations. Cellular deformability was lost more rapidly in response to exogenous oxidants in the qinghaosu-treated RBCs than in control cells. Hemoglobin SS and EE RBCs pretreated with qinghaosu demonstrated a much more rapid decrease in cellular deformability than did the control RBCs in response to exogenous oxidants. Additionally, qinghaosu resulted in a dose-dependent increase in red cell lysis and methemoglobin generation while decreasing reduced glutathione concentration. As a consequence of qinghaosu challenge, a decrease in unsaturated fatty acids was noted. Deoxyqinghaosu, which lacks the endoperoxide bridge and is pharmacologically inactive, did not affect cellular deformability and did not function as a prooxidant. Additionally, deoxyqinghaosu had no effect on fatty acid composition, red cell lysis, methemoglobin generation, or reduced glutathione concentration. In conclusion, although the oxidative effects of qinghaosu on uninfected erythrocytes were only seen at concentrations much greater than that necessary for antimalarial activity, these results confirm the proposition that qinghaosu may act via oxidative mechanisms. Furthermore, qinghaosu-mediated oxidation was significantly increased in erythrocytes characterized by enhanced oxidant sensitivity caused by unstable hemoglobins.

Artemisinins↗
PubMed1989

Erythrocyte phospholipid organization and vesiculation in hereditary high red cell membrane phosphatidylcholine hemolytic anemia.

We have studied the erythrocyte membrane phospholipid organization in hereditary high red cell membrane phosphatidylcholine hemolytic anemia (HPCHA) and the response of these red cells during incubation with sonicated suspensions of dimyristoylphosphatidylcholine (DMPC). Although both the absolute and relative amounts of phosphatidylcholine were elevated in these red cells, the relative distribution of phosphatidylcholine on both sides of the membrane bilayer and the transbilayer mobility of phosphatidylcholine were normal. HPCHA erythrocytes showed elevated absolute amounts per cell of both protein and lipid and an increased cellular ratio of protein to phospholipid. Incubation of normal red cells with DMPC led to the formation of echinocytes, followed by the release of acetylcholinesterase-containing vesicles. Both echinocyte formation and vesiculation were markedly reduced in red cells from patients with HPCHA. Studies with red cells from patients with liver disease, cells that also have elevated relative amounts of membrane phosphatidylcholine and increased amounts of lipid per cell, revealed normal echinocyte formation and normal DMPC-induced vesiculation. We conclude that the altered lipid composition of HPCHA erythrocytes per se is not responsible for the observed reduction in DMPC-induced vesiculation, but that it is more likely the result of a modification in the protein moiety of these cells. This putative protein abnormality could enhance binding of phosphatidylcholine to red cell membranes and could explain the elevated phosphatidylcholine content of HPCHA erythrocytes and their inability to vesiculate.

Anemia, Hemolytic, Congenital↗
PubMed1988

Studies on sickled erythrocytes provide evidence that the asymmetric distribution of phosphatidylserine in the red cell membrane is maintained by both ATP-dependent translocation and interaction with membrane skeletal proteins.

In order to study factors which are involved in maintenance of phosphatidylserine (PS) asymmetry within the human red cell membrane, we measured the effect of ATP-depletion and of membrane skeleton/lipid bilayer uncoupling induced by sickling on the distribution of PS within the membrane bilayer of sickle cells. Trace amounts of radiolabeled PS were introduced into the outer membrane leaflet of both fresh and ATP-depleted reversibly sickled cells (RSCs), using a non-specific lipid transfer protein purified from bovine liver. The equilibration of the newly introduced PS over the two halves of the bilayer was monitored by treatment of the cells with phospholipase A2 which selectively hydrolyzes only those molecules present in the outer membrane leaflet. Within 1 h after insertion into fresh RSCs, only 10% of the labeled PS was accessible to the action of phospholipase A2. This fraction was markedly increased when the cells were subsequently deoxygenated. Prolonged deoxygenation of RSCs, deprived of their ATP after incorporation of radiolabeled PS, caused enhanced phospholipase A2-induced hydrolysis of radiolabeled PS. Similarly, phospholipase A2-induced hydrolysis of endogenous PS in intact RSCs was markedly enhanced when ATP-depleted, but not when fresh cells, were incubated under nitrogen for 3.5 h. Deoxygenated ATP-depleted RSCs markedly enhanced the rate of thrombin formation in the presence of purified coagulation factors Xa, Va, prothrombin and Ca2+. This enhancement appeared to be dependent on the duration of incubation under nitrogen. This phenomenon, indicating the presence of increasing amounts of endogenous PS in the outer membrane leaflet, was not observed when either fresh RSCs or ATP-depleted normal erythrocytes were incubated under nitrogen. Our present observations provide evidence that, in addition to the interaction of PS with the skeletal proteins, an ATP-dependent translocation of PS is required to maintain its absolute asymmetric distribution in the human erythrocyte membrane.

Adenosine Triphosphate↗
PubMed1988

Comparative electrophysiologic evaluation of nerve grafts and autogenous vein grafts as nerve conduits: an experimental study.

This study was carried out to compare electrophysiologically the efficacy of autogenous vein grafts, with autogenous nerve grafts as conduits for nerve regeneration. A 0.75-cm segment of sciatic nerve was resected in two groups of Sprague-Dawley rats of equivalent maturity. The nerve gaps were bridged with an autogenous vein graft in the first group (31 rats), and an autogenous nerve graft in the second group (24 rats). Serial in vivo nerve conduction velocity studies and terminal in vitro nerve conduction velocity and nerve action potential measurements were performed. An additional group of 21 animals who had undergone no surgical procedures, were similarly studied to establish an age-adjusted baseline for comparison. Twelve animals in the first group, 14 in the second group, and 13 in the baseline group survived the full year of study. In vivo conduction velocities between the two experimental groups compared favorably. Nerve conduction velocity determined by in vitro technique confirmed this finding and measured similarly at about 78 percent of the baseline. Nerve action potential amplitude in the vein-grafted group was 12.0 percent of the baseline, while the nerve-grafted group was 23.9 percent of the baseline. This study demonstrated that the vein graft compares well with the nerve graft in nerve conduction velocity, but only one-half as well in nerve action potential.

Action Potentials↗
PubMed1987

Spectrin oxidation correlates with membrane vesiculation in stored RBCs.

An increase in spectrin oxidation in a variety of erythrocytes displaying a tendency to vesiculate has been previously described. To explore this relationship in more detail, we have studied blood stored in citrate-phosphate-dextrose-adenine under blood bank conditions because, in this system, vesiculation occurs slowly. Vesiculation was quantitated by measuring acetylcholinesterase release, and the extent of spectrin oxidation was detected by using thiol-disulfide exchange chromatography. A strong correlation (r = .92) was found between the extent of spectrin oxidation and vesiculation when blood from five donors was analyzed at weekly intervals during storage. This strongly suggests that spectrin oxidation plays a role in the formation of spectrin-free vesicles, thereby limiting the shelf life of stored blood.

Erythrocyte Membrane↗
PubMed1986

Effect of sickling on dimyristoylphosphatidylcholine-induced vesiculation in sickle red blood cells.

To study the effect of sickling on dimyristoylphosphatidylcholine (DMPC)-induced vesiculation, sickle (SS) red blood cells were incubated with sonicated suspensions of DMPC under either room air or nitrogen. Like normal red cells, when sickle cells were incubated with DMPC under oxygenated conditions, incorporation of DMPC into the erythrocyte membrane occurred, followed by echinocytic shape transformation and subsequent release of membrane vesicles. On the other hand, when SS cells were induced to sickle by deoxygenation, DMPC-induced vesiculation of these cells was dramatically reduced. However, upon reoxygenation, release of vesicles from these sickle erythrocytes occurred immediately. When SS cells were incubated under hypertonic (500 mosM) and deoxygenated conditions (where hemoglobin polymerization occurs but red cells do not show the typical sickle morphology), a similar decrease in the extent of vesiculation was observed. Experiments with radiolabelled lipid vesicles indicated that incorporation of DMPC into erythrocyte membranes occurred in all cases and therefore was not the limiting factor in the reduction of vesiculation in deoxygenated SS cells. Taken together, these results indicate that cellular viscosity and membrane rigidity, both of which are influenced by hemoglobin polymerization, are two important factors in process of vesicle release from sickle erythrocytes.

Anemia, Sickle Cell↗
PubMed1986

Spiral interrupted suturing technique for microvascular anastomosis: a comparative study.

This is an experimental study comparing the suturing time and patency rate of a spiral interrupted suturing technique to those of conventional interrupted and continuous suturing technique in end-to-end as well as end-to-side anastomosis. The spiral interrupted suturing technique requires less time for either end-to-end or end-to-side anastomosis than conventional interrupted suturing technique, and does not result in stenosis as shown in the venous end-to-end anastomosis by continuous suturing technique.

Animals↗
PubMed1986

A successful replantation of rat ear.

This article presents a successful replantation of a completely severed rat ear. The procedure include re-union of the tubal cartilage and anastomosis of the surgically divided external carotid artery and posterior facial vein.

Animals↗
PubMed1986

Bone and joint surgery--looking ahead.

Looking ahead, one envisions an era of remarkable progress in the study of the healing of bone and cartilage. The physical, biochemical, and biomechanical factors governing bone and cartilage healing will be defined and manipulated to accelerate the healing process. Joint stiffness, the undesirable sequela of many simple fractures of the hand as a result of prolonged immobilization, will be obviated. For cases of limited joint damage, techniques for joint reconstruction will be refined. For non-salvageable bone and joint destruction, microvascular allogenic transplantation of skeletal units will become a reality. Silastic implants in the wrist will become historical oddities.

Arthroplasty↗
PubMed1986

Management of peripheral nerve injuries.

The choice between repair or reconstruction in the management of nerve injury, and the determination on timing and the type of neurorrhaphy technique are clinical decisions that should be based on thorough understanding of the pathophysiology of nerve injury, the physiology of nerve healing, and the anatomy of peripheral nerves. The formulation of a treatment plan should be individualized.

Arm Injuries↗
PubMed1986

Red cell vesiculation--a common membrane physiologic event.

Loss of red blood cell membrane material in the form of microvesicles has been noted in sickle cells, in Ca++-loaded and adenosine triphosphate (ATP)-depleted normal red blood cells; and during storage of normal red blood cells. To further understand the vesiculation process, we have studied vesicles generated by a variety of perturbations of the red blood cell membrane. Vesicles were isolated by centrifugation at 30,000 X g from plasma of heparinized pathologic blood samples (sickle cell anemia, hemoglobin H disease, hereditary spherocytosis, hereditary elliptocytosis, and protein 4.1 deficiency) incubated overnight at 4 degrees C. Vesicle formation also was induced in normal erythrocytes by ATP depletion, by heating to 49 degrees C, by incubation at pH 5.4, and by incubation in 5 mmol/L diamide. Membrane protein composition was characterized on denaturing polyacrylamide gels and by immunoblot. The vesicles all contained band 3, glycophorin A, and band 4.1. Spectrin was depleted in all vesicles. Thiol disulfide exchange chromatography revealed evidence of oxidative cross-linking of spectrin in pathologic and normal red blood cells that had undergone vesiculation. This suggests that the mechanism of vesiculation may be related to cross-linking of membrane proteins. Membrane phospholipid composition of sickle cell and acid-induced vesicles was similar to that of normal red cells as determined by thin-layer chromatography. Possible pathophysiologic effects of vesiculation were assessed by using a modified Russell's viper venom assay. All vesicles examined shortened Russell's viper venom clotting time by 55% to 70% of control values. In addition, ektacytometer studies reveal that cells remaining after acid-induced vesiculation are rigid. These observations indicate that the vesicles may play a role in the hypercoagulation seen in some hemolytic disorders and that the process of vesiculation itself may contribute to increased rigidity of red cells and their subsequent removal from the circulation.

Chromatography↗
PubMed1985

Uncoupling of the membrane skeleton from the lipid bilayer. The cause of accelerated phospholipid flip-flop leading to an enhanced procoagulant activity of sickled cells.

We have previously reported that the normal membrane phospholipid organization is altered in sickled erythrocytes. More recently, we presented evidence of enhanced transbilayer movement of phosphatidylcholine (PC) in deoxygenated reversibly sickled cells (RSC) and put forward the hypothesis that these abnormalities in phospholipid organization are confined to the characteristic protrusions of these cells. To test this hypothesis, we studied the free spicules released from RSC by repeated sickling and unsickling as well as the remnant despiculated cells. The rate of transbilayer movement of PC in the membrane of deoxygenated remnant despiculated cells was determined by following the fate of 14C-labelled PC, previously introduced into the outer monolayer under fully oxygenated conditions using a PC-specific phospholipid exchange protein from beef liver. The rate of transbilayer movement of PC in the remnant despiculated cells was significantly slower than in deoxygenated native RSC and was not very much different from that in oxygenated native RSC or irreversibly sickled cells. The free spicules had the same lipid composition as the native cells, but were deficient in spectrin. These spicules markedly enhanced the rate of thrombin formation in the presence of purified prothrombinase (Factor Xa, Factor Va, and Ca2+) and prothrombin, indicating the exposure of a significant fraction of phosphatidylserine (PS) in the outer monolayer. This effect was not observed when the spicules in this assay were replaced by normal erythrocytes, deoxygenated native RSC, or a deoxygenated sample of RSC after repetitive sickling/unsickling. The results are interpreted to indicate that the destabilization of the lipid bilayer in sickled cells, expressed by the enhanced flip-flop of PC and the exposure of PS in the outer monolayer, occurs predominantly in those parts of the membrane that are in spicular form.

Anemia, Sickle Cell↗
PubMed1985

Increased adherence of sickled and phosphatidylserine-enriched human erythrocytes to cultured human peripheral blood monocytes.

The precise mechanism by which sickle erythrocytes (RBC) are removed from the circulation is controversial, although it is possible that enhanced recognition of these cells by circulating mononuclear phagocytes could contribute to this process. We investigated this possibility by interacting sickle cells with cultured human peripheral blood monocytes. Our results show that both irreversibly sickled cells (ISC) and deoxygenated reversibly sickled cells (RSC) had a higher avidity for adherence to monocytes than did oxygenated sickle and normal RBC. ISC were the most adherent cell type. Adherence of RSC to monocytes was found to be reversible; reoxygenation of deoxygenated RSC resulted in a significant decrease in RSC--monocyte adherence. Concomitant with alterations in sickle RBC adherence were alterations in the organization and bilayer distribution of membrane phospholipids in these cells. Specifically, enhanced adherence was associated with increased exposure of RBC membrane outer leaflet phosphatidylserine (PS) and phosphatidylethanolamine, whereas lack of adherence was associated with normal patterns of membrane phospholipid distribution. To investigate the possibility of whether the exposure of PS in the outer membrane leaflet of these cells might be responsible for their recognition by monocytes, the membranes of normal RBC were enriched with the fluorescent PS analogue 1-acyl-2[(N-4-nitro-benzo-2-oxa-1,3-diazole)aminocaproyl]-phosphatidy lse rine (NBD-PS) via transfer of the exogenous lipid from a population of donor phospholipid vesicles (liposomes). RBC enriched with NBD-PS exhibited enhanced adherence to monocytes, whereas adherence of RBC enriched with similar amounts of NBD-phosphatidylcholine (NBD-PC) was not increased. Furthermore, preincubation of monocytes with PS liposomes resulted in a approximately 60% inhibition of ISC adherence to monocytes, whereas no inhibition occurred when monocytes were preincubated with PC liposomes. These findings strongly suggest that erythrocyte surface PS may be a ligand recognized by receptors on human peripheral blood monocytes and that abnormal exposure of PS in the outer leaflet of the RBC membrane, as found in sickle RBC, might serve to trigger their recognition by circulating monocytes. Our results further suggest that abnormalities in the organization of erythrocyte membrane phospholipids may have significant pathophysiologic implications, possibly including shortened cell survival.

Anemia, Sickle Cell↗
PubMed1985

Plasma membrane phospholipid organization in human erythrocytes.

By weight, phospholipids make up approximately 25% of the plasma membrane of mature human erythrocytes. The four major phospholipid species present in the membrane (PC, PE, PS, and SM) are distributed asymmetrically across the bilayer leaflet resulting in an enrichment of choline-phospholipid (PC and SM) in the outer leaflet and of amino-phospholipid (PE and PS) in the cytoplasmic leaflet. This asymmetric organization is preferentially maintained through complex, and at present, poorly understood noncovalent interactions between specific membrane lipids and proteins (in particular, a stabilizing role for the skeletal protein spectrin and band 4.1 have been implicated), although other considerations such as phospholipid net charge, size, and degree of acyl chain unsaturation may also be involved. In certain red cell pathologies, or following experimental manipulation, there is a partial loss of this asymmetry (summarized in Tables XVI, XVII) often resulting in increases in the outer leaflet content of amino-phospholipids and subsequent expression of altered membrane surface properties. Some of these abnormal properties may have pathophysiologic consequence; indeed, red cell membranes displaying increased levels of surface amino-phospholipids have been shown to be potent procoagulants and demonstrate enhanced intermembrane interactions with both model (liposomes) and biologic (mononuclear phagocytes) membranes. Redistribution of membrane phospholipids may not occur homogeneously throughout an entire leaflet but may be restricted to specific membrane regions. These studies strongly suggest that the maintenance of phospholipid asymmetry in human red cell membranes is not a trivial event but probably represents a homeostatic mechanism, the failure of which may lead to alterations in normal erythrocyte functions, and ultimately, survival.

Blood Coagulation↗
PubMed1985

Membrane phospholipid organization and vesiculation of erythrocytes in sickle cell anaemia.

This review has examined the lipid composition of the RBC membrane and the methods used to determine the distribution of the phospholipids within the membrane. The importance of the membrane cytoskeletal proteins, in particular spectrin and protein 4.1, in maintaining this distribution has also been described. Membrane vesiculation and altered membrane phospholipid asymmetry in sickle cell anaemia have been reviewed. The relationships between vesiculation and hypercoagulability and an abnormal reticuloendothelial system in sickle cell disease have been examined. It is apparent that the single amino acid substitution leading to the production of sickle haemoglobin has profound effects on the entire erythrocyte, reaching to the limits of the cell, its plasma membrane.

Anemia, Sickle Cell↗