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Biomedical subjects

M R Clark

Publications and source records attributed to M R Clark.

At least 181 records · Page 10Linked to original sources
PubMed1981

Separate mechanisms of deformability loss in ATP-depleted and Ca-loaded erythrocytes.

Membrane rigidity has been widely accepted as the dominant cause of reduced deformability both of ATP-depleted erythrocytes and erythrocytes containing excess calcium (Ca). However, recent studies have shown normal membrane deformability in ATP-depleted erythrocytes. In addition, Ca accumulation causes massive ion and water loss, and it has been shown that extensive dehydration causes an increase in intracellular viscosity with attendant loss of whole cell deformability. To obtain a detailed understanding of the processes accompanying ATP depletion and/or Ca accumulation that limit cell deformability, we have used a viscodiffractometric method to identify the cellular factors contributing to reduced whole cell deformability. Analysis of the influence of the suspending medium osmolality on deformability showed the presence of two independent processes. One was a Ca-independent reduction in cell surface area/volume ratio, resulting from the spheroechinocyte formation that follows total ATP consumption. The other was a Ca-dependent increase in intracellular viscosity resulting from a Ca-induced loss of intracellular potassium and water. This deformability loss due to increased intracellular viscosity was found for cells depleted of ATP in the presence of Ca and in cells treated with Ca and A23187 without prior depletion. Ionophore-treated cells at high Ca concentration (>500 muM) formed spheroechinocytes with reduced surface area and a further loss of whole cell deformability. The rate of deformability loss associated with Ca-induced spheroechinocytosis was much more rapid than that associated with ATP-depletion-induced spheroechinocytosis, suggesting different mechanisms for the morphologic changes. No major effects of altered membrane elasticity on the reduced deformability of either ATP-depleted or Ca-loaded cells were observed.

Adenosine Triphosphate↗
PubMed1981

Factors that limit whole cell deformability in erythrocytes after calcium loading and ATP depletion.

Whole cell deformability of ATP-depleted and Ca-loaded red cells has been measured at various osmolalities to determine those cellular factors responsible for the reduce deformability of these cells. For cells depleted of ATP in Ca-free medium, a progressive loss of hypotonic deformability identified membrane loss with reduced surface area-to-volume ratio as the dominant mechanism of deformability loss. For cells treated with Ca and the ionophore A23187 without prior depletion, a rapid loss of isotonic deformability, reversible in hypotonic medium, identified dehydration with increased internal viscosity as the dominant mechanism of deformability loss. In contrast to previously held concepts, increased membrane rigidity was not found to have a major influence.

Adenosine Triphosphate↗
PubMed1980

Deformability and spectrin properties in three types of elongated red cells.

Three types of unusual, elongated erythrocytes have been studied in our effort to understand factors that regulate the shape and deformability of red blood cells. Measurements of deformability, morphologic response to membrane-shape-altering drugs, spectrin phosphorylation, spectrin extractability, and casein kinase activity were measured in llama cells, irreversibly sickled cells, and nonhemolytic human elliptocytes. The results of these studies support the concept that the physical properties of spectrin are important in maintaining the shape and deformability of the erythrocyte. Furthermore, changes in spectrin extractability and phosphorylation appear to be associated with changes in the viscoelastic properties of the membrane. However, it seems that spectrin's regulatory role is permissive rather than direct and active.

Anemia, Sickle Cell↗
PubMed1980

Deformability of oxygenated irreversibly sickled cells.

The deformability characteristics of isolated subpopulations of irreversibly sickled cells (ISC) have been studied in an ektacytometer. Analysis of laser diffraction patterns of well-oxygenated cells subjected to shear stress in solutions of varying osmolality has demonstrated a profound influence of mean corpuscular hemoglobin concentration and intracellular viscosity on the deformability of ISC. Virtually undeformable at 290 mosM, ISC became almost totally deformable at 130 mosM. In addition, when ISC membranes were loaded with normal hemoglobin at low concentration, they deformed easily in isotonic medium, as did resealed normal cell membranes. The restoration of deformability of ISC upon reduction of their hemoglobin concentration and internal viscosity to normal levels suggests that altered membrane properties are not the primary determinant of decreased deformability in these cells. Rather, cellular dehydration induced by previous sickling would appear to contribute in a major way to their abnormal rheological behavior.

Anemia, Sickle Cell↗
PubMed1980

Analysis of factors regulating erythrocyte deformability.

Using a laser diffraction technique, we have studied factors that influence the deformability of erythrocytes. Variations in suspending medium osmolality and applied shear stress were employed to isolate the individual contributions to whole cell deformability of internal viscosity, surface area-to-volume ratio, and viscoelastic properties of the membrane. An experimental system was devised in which normal cells were modified in vitro to induce specific alterations in each factor. Measurements of deformability as a function of medium osmolality showed characteristic behavior of the modified cells. Reduced surface area-to-volume ratio was detected by an exaggeration of the normal decrease in deformability as medium osmolality was decreased. In contrast, increased internal viscosity was detected by an increase in deformability as osmolality was decreased. Finally, decreased membrane flexibility was detected by reduced deformation at low shear stress. These methods of analysis were applied to cells from patients with hereditary spherocytosis, hereditary pyropoikilocytosis, and hemoglobin CC disease to define the basis of reduced deformability. Hereditary spherocytes showed the combined effects of reduced surface area and increased internal viscosity. Hereditary pyropoikilocytes revealed the effects of severely reduced surface area-to-volume ratio. Hemoglobin CC cells showed only the effects of high internal viscosity. An increase in the membrane shear modulus (decreased membrane deformability) was not evident in these disorders.

Anemia, Hemolytic, Congenital↗
PubMed1980

Stimulation of prostaglandin accumulation by luteinizing hormone-releasing hormone (LHRH) and LHRH analogs in rat granulosa cells in vitro.

LHRH and LHRH agonists have been reported to exhibit somewhat paradoxical antireproductive effects. The current studies were undertaken to examine the actions of these substances on one parameter of ovarian function which is required for ovulation, i.e. prostaglandin production. Granulosa cells were obtained from immature rats 48 h after injection of 20 IU PMSG and incubated for up to 5 h in vitro. LHRH, [D-Ala6, des-Gly-NH2(10)]LHRH-ethylamide (Analog I) and [D-Leu6, des-Gly-NH2(10)]LHRH-ethylamide (Analog II) were observed to stimulate prostaglandin accumulation by granulosa cells. Analog I, for example, at 100 ng/ml increased PGE from 0.19 +/- 0.06 in the control to 7.7 +/- 2.01 ng/2 x 10(6) cells (n = 7; P less than 0.01) after 5 h. TRH, on the other hand, had no effect on prostaglandin accumulation. When added with LH or FSH, the stimulation by maximal concentrations of Analog I and the gonadotropins appeared additive. Although the stimulation of prostaglandin accumulation by LH appears to be mediated by cAMP, no effect of Analog I on the amount of cAMP could be detected. cAMP was determined in cells plus medium at times of 1.5 min to 5 h and at concentrations of 10-2000 ng/ml Analog I, but did not change from the control. LHRH and LHRH agonists, therefore, stimulate the accumulation of ovarian granulosa cell prostaglandins and do so in a manner apparently distinct from that of LH or FSH.

Animals↗
PubMed1980

Inhibition by estradiol of human chorionic gonadotropin-induced progesterone accumulation in isolated human luteal cells: lack of mediation by prostaglandin F.

Estradiol has been previously reported to be a potent inhibitor of hCG-stimulated progesterone accumulation in isolated cells from human corpora lutea of the menstrual cycle. We assessed the role of prostaglandin (PG) in this in vitro estradiol-induced inhibition by measuring PGF and adding a blocker of PG synthesis (indomethacin) to the incubation medium. The inhibition by estradiol of hCG-stimulated progesterone accumulation occurred without any increase in PGF accumulation in the incubations. Furthermore, PGF accumulation was markedly reduced (P less than 0.05) at all concentrations of indomethacin testes (0.1, 1, and 10 microgram/ml), but the inhibitory effect of estradiol on hCG-stimulated progesterone accumulation was not prevented. These data suggest that the inhibitory effect of estradiol observed in vitro is not mediated by PGs.

Adult↗
PubMed1980

Ektacytometric analysis of factors regulating red cell deformability.

Photometric analysis of laser diffraction patterns has been used to obtain quantitative measurements of deformability of specifically modified normal red cells. Variation of deformability with suspending medium osmolality and with applied shear stress was used to distinguish between changes in internal viscosity, surface area-to-volume ratio, and viscoelastic properties of the membrane in their influence on whole cell deformability.

Dextrans↗
PubMed1980

Influence of red cell water content on the morphology of sickling.

The response of sickle cells with varying water content to alterations in oxygen tension has been studied. Cells that were severely dehydrated while sickled retained the characteristic sickled morphology even after prolonged reoxygenation. When the cell water content was increased by reduction of the suspending medium osmolality, the cells unsickled. Cells that were dehydrated before deoxygenation were unable to assume the spiculated morphology typical of sicked cells. This was true both for high mean cell hemoglobin concentration (MCHC) discoid sickle cells and for irreversibly sickled cells. When such cells were resuspended in hypotonic medium before deoxygenation, they sickled with the characteristic morphology of sickle cells with normal MCHC. The morphological behavior of Ca-loaded sickled cells as well as irreversibly sickled cells showed a major influence of increased hemoglobin concentration and extremely high internal viscosity. Constraint on cell morphology by putative membrane rigidity was not observed.

Anemia, Sickle Cell↗
PubMed1980

Erythropoietic stress, macrocytosis, and hemoglobin switching in HbAA sheep.

Hemoglobin switching and macrocytosis were studied in homozygous hemoglobinAA sheep. An abrupt initiation of erythropoietic stress, accompanied by a pulsed elevation of circulating erythropoietin levels, was induced by phlebotomy. Sequential blood samples were separated according to density on Stractan gradients to isolate cells newly entering the circulation from the marrow each day. Analysis of hemoglobin phenotype and cell volume distribution in these young reticulocytes revealed a distinct temporal separation in the appearance of hemoglobin C and increased cell volume. The appearance of macrocytes within 24 hr of erythropoietin elevation suggests that macrocytosis could be the result of the action of erythropoietin during the late stages of erythroid maturation. The 72-hr delay in the appearance of hemoglobin C indicates that commitment to a particular hemoglobin phenotype occurs at an early stage of differentiation and involves immature erythroid stem cells. The results of this study show that these consequences of erythropoietic stress are initiated at two different developmental stages, resulting in the production of macrocytosis and hemoglobin switching.

Animals↗
PubMed1980

Inaccuracies associated with the automated measurement of mean cell hemoglobin concentration in dehydrated cells.

Because of discrepancies between electronically and manually measured values of mean cell hemoglobin concentration (MCHC) encountered in studies of pathologic red cells, we studied the effect of cell water content on MCHC measurements by both methods. A series of red cell samples with varying water contents (54%-164% normal) were prepared from normal cells using the antibiotic nystatin. MCHC was then measured, using the microhematocrit centrifuge and three different electronic cell counters in common laboratory use. For MCHC values above 36 g/dl as measured by the spun hematocrit method, all three electronic counters under estimmated the MCHC, with increasing error as the true MCHC increased. For MCHC values below 30 g/dl, the values from two conductivity based instruments agreed with those from the spun hematocrit method, whereas one instrument based on light scattering overestimated the MCHC. These results indicate that inaccuracies in the measured mean cell volume (MCV) of dehydrated or otherwise undeformable cells may lead to spurious values for MCHC when electronic cell counters are used.

Erythrocyte Indices↗
PubMed1978

Mechanism of luteinizing hormone regulation of prostaglandin synthesis in rat granulosa cells.

The present study was undertaken to examine the mechanism by which luteinizing hormone (LH) stimulates prostaglandin (PG) synthesis in rat granulosa cells. Immature rats were injected with 20 IU of pregnant mare's serum gonadotropin and granulosa cells isolated 48 h later. LH (5 microgram/ml) stimulated PGE synthesis markedly over the control with no additions in 6-h incubations (1.98 +/- 0.24 and 0.24 +/- 0.05 ng/2 X 10(6) cells, respectively, n = 6). When arachidonic acid (100 microgram/ml) was included during the last hour of incubation, further increases to 15.4 +/- 2.9 and 2.48 +/- 0.48 ng of PGE/2 X 10(6) cells in LH and control incubations were observed. The cause of the increased response to a 1-h incubation with arachidonic acid in the LH-treated cells did not appear to be a stimulation of fatty acid uptake. In addition, when cellular lipids were labeled by a 2-h incubation with radioactive arachidonic acid, LH did not stimulate intra- or extracellular release of arachidonic acid. A 5-fold stimulation of prostaglandin synthetase activity, however, was observed in cells incubated with LH for 5 h. Our results, therefore, indicate that LH acts at a step in the prostaglandin pathway after hydrolysis of arachidonic acid esters and produces an increase in prostaglandin synthetase activity.

8,11,14-Eicosatrienoic Acid↗
PubMed1978

Patient care appraisal as a guide for the design of continuing medical education: 10 years' experience in the Maritime provinces.

Patient care appraisal is a process whereby physicians set down criteria and standards of patient care, prepare a summary of relevant health care activities derived from medical records and other sources of data, analyse the shortcomings discovered and design remedial programs. An attempt was made to introduce such a peer review system to 19 community hospitals in the Maritime provinces as a mechanism to improve the quality of health care. Maximum use was made of all existing quality assurance methods, complemented by innovations compatible with known educational and psychological principles. The staffs of the hospitals were willing to participate but problems were encountered in maintaining their motivation. Much of the success was related to the appropriate use of interested and competent medical record librarians in the retrieval, compilation and display of information. A positive attitude towards peer reviews and peer learning evolved in the participants. It is recommended that practising physicians, health care institutions and university divisions of continuing medical education continue and extend such cooperation and find new ways to apply the peer review mechanisms.

Canada↗
PubMed1978

Effects of abnormal cation transport on deformability of desiccytes.

We have studied the deformability of subpopulations of red cells from a patient with "desiccytosis," a disorder characterized by increased membrane permeability to potassium and associated with a probable increase in sodium-sodium exchange. Cells become increasingly dehydrated after maturation because of continued potassium loss without compensatory sodium gain, and they exhibit a progressive increase in mean cell hemoglobin concentration (MCHC). This increase in MCHC causes the cells to become underformable at shear stress values which result in extensive deformation of normal cells. Reduction of MCHC to approximately normal levels by suspending the cells in hypotonic medium restores normal deformability to all but 0.1--0.2% of the cells. These results suggest that the major factor leading to premature destruction in this disorder is whole cell rigidity conferred by increased intracellular hemoglobin concentrations, rather than any associated membrane rigidity.

Anemia, Hemolytic↗
PubMed1978

Localization and in vitro synthesis of prostaglandins in components of rabbit preovulatory graafian follicles.

Graafian follicles obtained 9 hours after the injection of human chorionic gonadotropin (hCG) into mature rabbits were dissected into a follicular fluid component, a granulosa cell-oocyte component, and a residual wall component, (the latter containing mostly theca tissue with a small and variable amount of adhering granulosa cells). The amounts of PGE and PGF were determined for each component. The follicular fluid contained approximately 4-10 times more PGE and PGF than either the granulosa cell-oocyte component or the residual wall component. The latter two components contained approximately equal amounts of these prostaglandins. The in vitro biosynthesis of PGE and PGF was also studied and it was found that the granulosa cell-oocyte component had about 4 fold the capacity of the residual wall, and that the follicular fluid synthesized no prostaglandins. There was no significant effect of LH on either PGE or PGF synthesis in any of the components.

Animals↗