The circulatory physiopathology of human red blood cells investigated with a multiplatform model of cellular homeostasis. II. Capillary transits and the role of PIEZO1

AbstractIn this and the next paper of this series we apply the red cell model introduced in the previous paper to investigate the changes in RBC homeostasis during capillary transits and over the full circulatory lifespan of the cells. These are topics inaccessible to direct experimentation but rendered mature for a modelling approach by recent findings and by a large body of apparently unrelated early results which robustly constrain the parameter space offering the opportunity for an in depth study of the mechanisms involved. Capillary transit times vary between 0.5 and 1.5s during which the red blood cells squeeze and deform in the capillary stream transiently opening stress-gated PIEZO1 channels, creating minuscule quantal changes in RBC ion contents and volume. Widely accepted early views originally based on results from experimentally shear-stressed red cells suggested that quantal changes generated during capillary transits add up over time to generate the documented changes in RBC density during their long circulatory lifespan, the quantal hypothesis. Applying the new PIEZO1 extension of the RBC model (RCM) introduced in the previous paper we investigated in detail the changes in homeostatic variables that may be expected during single capillary transits resulting from transient PIEZO1 channel activation. The predicted quantal volume changes were infinitesimal in magnitude, biphasic in nature, and essentially irreversible within inter-transit periods. A sub-second transient PIEZO1 activation triggered a sharp swelling peak followed by a much slower recovery period towards lower-than-baseline volumes. The peak response was caused by net CaCl2 and fluid gain via PIEZO1 channels driven by the steep electrochemical inward Ca2+ gradient. The ensuing dehydration followed a complex time-course with sequential, but partially overlapping contributions by KCl loss via Ca2+-activated Gardos channels, PMCA mediated calcium extrusion and chloride efflux by the Jacobs-Steward mechanism. The change in relative cell volume predicted for single capillary transits was below 10−4, an infinitesimal volume change incompatible with a functional role in capillary flow. The biphasic response predicted by the RCM appears to conform to the quantal hypothesis, but whether its cumulative effects could account for the documented changes in density during RBC senescence required an investigation of the effects of myriad transits over the full period of circulatory lifespan, the subject of the next paper of this series.

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Up-down biphasic volume response of human red blood cells to PIEZO1 activation during capillary transits

PLoS Computational Biology ◽

10.1371/journal.pcbi.1008706 ◽

2021 ◽

Vol 17 (3) ◽

pp. e1008706 ◽

Cited By ~ 2

Author(s):

Simon Rogers ◽

Virgilio L. Lew

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Time Course ◽

Capillary Flow ◽

Cell Model ◽

Recovery Period ◽

Large Body ◽

Computational Modelling ◽

Companion Paper ◽

Human Red Blood Cells

In this paper we apply a novel JAVA version of a model on the homeostasis of human red blood cells (RBCs) to investigate the changes RBCs experience during single capillary transits. In the companion paper we apply a model extension to investigate the changes in RBC homeostasis over the approximately 200000 capillary transits during the ~120 days lifespan of the cells. These are topics inaccessible to direct experimentation but rendered mature for a computational modelling approach by the large body of recent and early experimental results which robustly constrain the range of parameter values and model outcomes, offering a unique opportunity for an in depth study of the mechanisms involved. Capillary transit times vary between 0.5 and 1.5s during which the red blood cells squeeze and deform in the capillary stream transiently opening stress-gated PIEZO1 channels allowing ion gradient dissipation and creating minuscule quantal changes in RBC ion contents and volume. Widely accepted views, based on the effects of experimental shear stress on human RBCs, suggested that quantal changes generated during capillary transits add up over time to develop the documented changes in RBC density and composition during their long circulatory lifespan, the quantal hypothesis. Applying the new red cell model (RCM) we investigated here the changes in homeostatic variables that may be expected during single capillary transits resulting from transient PIEZO1 channel activation. The predicted quantal volume changes were infinitesimal in magnitude, biphasic in nature, and essentially irreversible within inter-transit periods. A sub-second transient PIEZO1 activation triggered a sharp swelling peak followed by a much slower recovery period towards lower-than-baseline volumes. The peak response was caused by net CaCl2 and fluid gain via PIEZO1 channels driven by the steep electrochemical inward Ca2+ gradient. The ensuing dehydration followed a complex time-course with sequential, but partially overlapping contributions by KCl loss via Ca2+-activated Gardos channels, restorative Ca2+ extrusion by the plasma membrane calcium pump, and chloride efflux by the Jacobs-Steward mechanism. The change in relative cell volume predicted for single capillary transits was around 10−5, an infinitesimal volume change incompatible with a functional role in capillary flow. The biphasic response predicted by the RCM appears to conform to the quantal hypothesis, but whether its cumulative effects could account for the documented changes in density during RBC senescence required an investigation of the effects of myriad transits over the full four months circulatory lifespan of the cells, the subject of the next paper.

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PIEZO1 and the mechanism of the long circulatory longevity of human red blood cells

PLoS Computational Biology ◽

10.1371/journal.pcbi.1008496 ◽

2021 ◽

Vol 17 (3) ◽

pp. e1008496

Author(s):

Simon Rogers ◽

Virgilio L. Lew

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Time Course ◽

Clinical Manifestations ◽

Cell Subpopulation ◽

Unexpected Finding ◽

Human Red Blood Cells ◽

Acceptable Model ◽

New Finding ◽

Extended Longevity

Human red blood cells (RBCs) have a circulatory lifespan of about four months. Under constant oxidative and mechanical stress, but devoid of organelles and deprived of biosynthetic capacity for protein renewal, RBCs undergo substantial homeostatic changes, progressive densification followed by late density reversal among others, changes assumed to have been harnessed by evolution to sustain the rheological competence of the RBCs for as long as possible. The unknown mechanisms by which this is achieved are the subject of this investigation. Each RBC traverses capillaries between 1000 and 2000 times per day, roughly one transit per minute. A dedicated Lifespan model of RBC homeostasis was developed as an extension of the RCM introduced in the previous paper to explore the cumulative patterns predicted for repetitive capillary transits over a standardized lifespan period of 120 days, using experimental data to constrain the range of acceptable model outcomes. Capillary transits were simulated by periods of elevated cell/medium volume ratios and by transient deformation-induced permeability changes attributed to PIEZO1 channel mediation as outlined in the previous paper. The first unexpected finding was that quantal density changes generated during single capillary transits cease accumulating after a few days and cannot account for the observed progressive densification of RBCs on their own, thus ruling out the quantal hypothesis. The second unexpected finding was that the documented patterns of RBC densification and late reversal could only be emulated by the implementation of a strict time-course of decay in the activities of the calcium and Na/K pumps, suggestive of a selective mechanism enabling the extended longevity of RBCs. The densification pattern over most of the circulatory lifespan was determined by calcium pump decay whereas late density reversal was shaped by the pattern of Na/K pump decay. A third finding was that both quantal changes and pump-decay regimes were necessary to account for the documented lifespan pattern, neither sufficient on their own. A fourth new finding revealed that RBCs exposed to levels of PIEZO1-medited calcium permeation above certain thresholds in the circulation could develop a pattern of early or late hyperdense collapse followed by delayed density reversal. When tested over much reduced lifespan periods the results reproduced the known circulatory fate of irreversible sickle cells, the cell subpopulation responsible for vaso-occlusion and for most of the clinical manifestations of sickle cell disease. Analysis of the results provided an insightful new understanding of the mechanisms driving the changes in RBC homeostasis during circulatory aging in health and disease.

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Cell volume and metabolic dependence of NEM-activated K+-Cl- flux in human red blood cells

AJP Cell Physiology ◽

10.1152/ajpcell.1985.249.1.c124 ◽

1985 ◽

Vol 249 (1) ◽

pp. C124-C128 ◽

Cited By ~ 45

Author(s):

P. K. Lauf ◽

C. M. Perkins ◽

N. C. Adragna

Keyword(s):

Red Blood Cells ◽

Cell Volume ◽

Blood Cells ◽

Time Course ◽

Transport Activity ◽

Human Red Blood Cells ◽

Atp Breakdown ◽

The Difference ◽

Metabolic Dependence ◽

K Transport

The effects of incubation in anisosmotic media and of metabolic depletion on ouabain-resistant (OR) Cl--dependent K+ influxes stimulated by N-ethylmaleimide (NEM) were studied in human red blood cells using Rb+ as K+ analogue. The NEM-stimulated but not the basal Rb+-Cl- influx measured in phosphate-buffered anisosmotic media was found to be cell volume dependent. When cellular ATP, [ATP]c, was lowered to less than 0.10 of its initial level by exposure to nonmetabolizable 2-deoxy-D-glucose, the NEM-stimulated but not the basal Cl--dependent Rb+ influxes were abolished. Metabolically depleted red blood cells subsequently repleted by incubation in glucose plus inosine regained the NEM-inducible Rb+ (K+) transport activity. The difference in the time course of ATP breakdown and Rb+ influx inhibition suggests that energization of the NEM-stimulated Rb+ flux by metabolism may involve factors additional to ATP.

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The circulatory physiopathology of human red blood cells investigated with a multiplatform model of cellular homeostasis. III. Senescence changes during the full circulatory lifespan

10.1101/2020.03.07.981803 ◽

2020 ◽

Cited By ~ 1

Author(s):

Simon Rogers ◽

Virgilio L. Lew

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Time Course ◽

Clinical Manifestations ◽

Cell Subpopulation ◽

Unexpected Finding ◽

Sickle Cells ◽

Human Red Blood Cells ◽

Decay Sequence ◽

Health And Disease

AbstractHuman red blood cells (RBCs) have a circulatory lifespan of about four months. Under constant oxidative and mechanical stress, but devoid of organelles and deprived of biosynthetic capacity for protein renewal, RBCs undergo substantial homeostatic changes, progressive densification followed by late density reversal among others, changes assumed to have been harnessed by evolution to sustain the rheological competence of the RBCs for as long as possible. The unknown mechanisms by which this is achieved are the subject of this investigation. Each RBC traverses capillaries between 1000 and 2000 times per day, roughly one transit per minute, a total of about 2•105 transits during their lifespan. A dedicated Lifespan model of RBC homeostasis was developed as an extension of the RCM introduced in the first paper of this series to explore the cumulative patterns predicted for repetitive capillary transits over a standardized lifespan period of 120 days, using experimental data to constrain the parameter space. Capillary transits were simulated by periods of elevated cell/medium volume ratios and by transient deformation-induced permeability changes attributed to PIEZO1 channel mediation as outlined in the second paper of this series. The first unexpected finding was that quantal changes generated during single capillary transits cease accumulating after a few days and cannot account for the observed progressive densification of RBCs on their own, thus ruling out the quantal hypothesis. The second unexpected finding was that the documented patterns of RBC densification and late reversal could only be emulated by the implementation of a strict time-course of decay in the activities of the calcium and Na/K pumps, but only in addition to the quantal changes. These results showed that both quantal changes and pump-decay regimes were necessary to account for the documented lifespan pattern, neither sufficient on their own. They also suggested a strong selective component in the pump decay sequence. A third finding was that RBCs exposed to levels of calcium permeation above certain thresholds in the circulation could develop a pattern of late or early hyperdense collapse followed by delayed density reversal. When tested over much reduced lifespan periods the results emulated the known circulatory fate of irreversible sickle cells, the cell subpopulation responsible for vaso-occlusion and for most of the clinical manifestations of sickle cell disease. Analysis of the results provided an insightful new understanding of the mechanisms driving the changes in RBC homeostasis during circulatory aging in health and disease.

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Quantitative time-course metabolomics in human red blood cells reveal the temperature dependence of human metabolic networks

Journal of Biological Chemistry ◽

10.1074/jbc.m117.804914 ◽

2017 ◽

Vol 292 (48) ◽

pp. 19556-19564 ◽

Cited By ~ 20

Author(s):

James T. Yurkovich ◽

Daniel C. Zielinski ◽

Laurence Yang ◽

Giuseppe Paglia ◽

Ottar Rolfsson ◽

...

Keyword(s):

Red Blood Cells ◽

Temperature Dependence ◽

Blood Cells ◽

Time Course ◽

Metabolic Networks ◽

Human Red Blood Cells

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Proton (or hydroxide) fluxes and the biphasic osmotic response of human red blood cells.

The Journal of General Physiology ◽

10.1085/jgp.102.1.99 ◽

1993 ◽

Vol 102 (1) ◽

pp. 99-123 ◽

Cited By ~ 9

Author(s):

J D Bisognano ◽

J A Dix ◽

P R Pratap ◽

T S Novak ◽

J C Freedman

Keyword(s):

Membrane Potential ◽

Red Blood Cells ◽

Blood Cells ◽

Time Course ◽

Rapid Development ◽

Disulfonic Acid ◽

Proton Efflux ◽

Experimental Conditions ◽

Human Red Blood Cells ◽

Hydroxide Fluxes

Upon exposure of human red blood cells to hypertonic sucrose, the fluorescence of the potentiometric indicator 3,3'-dipropylthiadicarbocyanine iodide, denoted diS-C3(5), displays a biphasic time course indicating the rapid development of an inside-positive transmembrane voltage, followed by a slow DIDS (4,4'-diisothiocyano-2,2'-disulfonic acid stilbene)-sensitive decline of the voltage. In addition to monitoring membrane potential, proton (or hydroxide) fluxes were measured by a pH stat method, cell volume was monitored by light scattering, and cell electrolytes were measured directly when red cells were shrunken either with hypertonic NaCl or sucrose. Shrinkage by sucrose induced an initial proton efflux (or OH- influx) of 5.5 mu eq/g Hb.min and a Cl shift of 21-31 mu eq/g Hb in 15 min. Upon shrinkage with hypertonic NaCl, the cells are initially close to Donnan equilibrium and exhibit no detectable shift of Cl or protons. Experiments with the carbonic anhydrase inhibitor ethoxzolamide demonstrate that for red cell suspensions exposed to air and shrunken with sucrose, proton fluxes mediated by the Jacobs-Stewart cycle contribute to dissipation of the increased outward Cl concentration gradient. With maximally inhibitory concentrations of ethoxzolamide, a residual proton efflux of 2 mu eq/g Hb.min is insensitive to manipulation of the membrane potential with valinomycin, but is completely inhibited by DIDS. The ethoxzolamide-insensitive apparent proton efflux may be driven against the electrochemical gradient, and is thus consistent with HCl cotransport (or Cl/OH exchange). The data are consistent with predictions of equations describing nonideal osmotic and ionic equilibria of human red blood cells. Thus osmotic equilibration after shrinkage of human red blood cells by hypertonic sucrose occurs in two time-resolved steps: rapid equilibration of water followed by slower equilibration of chloride and protons (or hydroxide). Under our experimental conditions, about two-thirds of the osmotically induced apparent proton efflux is mediated by the Jacobs-Stewart cycle, with the remainder being consistent with mediation via DIDS-sensitive HCl cotransport (or Cl/OH exchange).

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Morphometry and Stiffness of Red Blood Cells—Signatures of Neurodegenerative Diseases and Aging

International Journal of Molecular Sciences ◽

10.3390/ijms23010227 ◽

2021 ◽

Vol 23 (1) ◽

pp. 227

Author(s):

Velichka Strijkova-Kenderova ◽

Svetla Todinova ◽

Tonya Andreeva ◽

Desislava Bogdanova ◽

Ariana Langari ◽

...

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Cell Shape ◽

Cell Model ◽

Lower Surface ◽

Healthy Individuals ◽

Peripheral Cell ◽

Human Red Blood Cells ◽

Atomic Force ◽

Lateral Sclerosis

Human red blood cells (RBCs) are unique cells with the remarkable ability to deform, which is crucial for their oxygen transport function, and which can be significantly altered under pathophysiological conditions. Here we performed ultrastructural analysis of RBCs as a peripheral cell model, looking for specific signatures of the neurodegenerative pathologies (NDDs)—Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS) and Alzheimer’s disease (AD), utilizing atomic force (AFM) and conventional optical (OM) microscopy. We found significant differences in the morphology and stiffness of RBCs isolated from patients with the selected NDDs and those from healthy individuals. Neurodegenerative pathologies' RBCs are characterized by a reduced abundance of biconcave discoid shape, lower surface roughness and a higher Young’s modulus, compared to healthy cells. Although reduced, the biconcave is still the predominant shape in ALS and AD cells, while the morphology of PD is dominated by crenate cells. The features of RBCs underwent a marked aging-induced transformation, which followed different aging pathways for NDDs and normal healthy states. It was found that the diameter, height and volume of the different cell shape types have different values for NDDs and healthy cells. Common and specific morphological signatures of the NDDs were identified.

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