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Mechanisms of Human Erythrocytes Volume Stabilization

Functional completeness of erythrocytes depends on high deformability of these cells, that allows them to pass through narrow tissue capillaries. The erythrocytes high deformability is provided due to maintenance of discoid shape with an optimal cell surface area to volume ratio. In its turn this ratio is maintained due to cell volume stabilization at a given cell surface area. In this work, using mathematical simulation, we studied role of Na/K-ATPase, calcium activated potassium channels and adenylate metabolism in human erythrocyte volume stabilization at increase in cell membrane permeability to cations. The simulation took into account a contribution of glycolytic metabolites and adenylates to cytoplasm osmotic pressure. It was shown that the presence in the cell of Na/K-ATPase and two opposite transmembrane gradients of Na+ and K+ ions provide a significantly improved cell volume stabilization at the increase in cell membrane permeability, compared with hypothetical cells, in which the osmotic balance between cell and extracellular compartment is provided due to a gradient of only one ion (Na+). In this case the erythrocyte volume deviates from the optimal value by less than 10% at change in cell membrane permeability from 50 to 200% of its normal value. In this case, however, the intracellular ion concentrations may change significantly (by several times). The adenylate metabolism system can provide an additional regulation of transport ATPases due to regulation of intracellular ATP levels. Under these conditions stabilization of steady-state values of intracellular ion concentrations (ion homeostasis) and of cell volume in the range of cell membrane permeability changes from 50 to 1500% of the normal value. In this case, however, the cell volume and intracellular ion concentrations may significantly deviate from the stabilized values during transitional processes. Simultaneous function of both, ion transport systems and adenylate metabolism allows to provide ion homeostasis and efficient erythrocyte volume stabilization (within 5% deviation from the optimal value) both in steady-state conditions and during transitional processes at increase in cell membrane permeability up to 10-15 times compared with the normal value.

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#red blood cells#metabolism#ion channel

Systems Biology and Physiology Reports in 2021: a yearly report

Dear All,

I am happy to greet all readers, editors, reviewers, staff members and publishers of SBPR on the eve of year 2022. I thank all of you for your efforts in creating this journal and making it work despite all the challenges faced by a new independent journal in the age dominated by huge publishers, research societies and citation systems. Please accept my best wishes for the year 2022 in your professional and personal undertakings!

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First year of open-access scientific journal publishing: our mishaps, our expectations, our plans for the bright future

In silico methods have become a versatile and effective tool for the studies of the natural phenomenon at multiple levels. However, unlike such fields as economics and physics, biology and physiology have always been slightly reluctant in acceptance of the novel systemic approaches. Since scientific articles are among the key metrics for the modern scientific society, the number of scientific journals, devoted to some topic, can be a good marker to evaluate the impact of this topic for the society. Indeed, there are esteemed journals, such as PLOS Computational Biology or Journal of Theoretical Biology, but they focus mainly on a common biological phenomenon, while questions of physiology and pathophysiology are often overlooked. On the other hand, the amount of work on computational biology is steadily growing with ~12000 works published in 2010 and ~28000 in 2020 with the keyword “computational model”, according to PubMed. With all this being said and our great experience in the different aspects of the field of computational physiology, our team has decided that it would be a mistake to lose an opportunity to cover this niche. Likewise, we, as a group of fellow scientists, at the end of 2019 decided to create our own venture and to launch our own scientific journal – Systems Biology and Physiology Reports (SBPReports) [1]. This publisher's would be an attempt to underline our first hectic year of the open access scientific publishing.

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On calcium fluorophore’s impact in platelet signaling studies

Observation of calcium signaling in platelets - blood cells designed to be involved in stopping bleeding and forming blood clots - is an important part of fundamental research in hemostasis. However, such a study is possible only with the use of calcium fluorophores - small molecules that penetrate the platelet membrane due to their hydrophobic -AM part, which is then hydrolyzed by cytosol esterases. In this work, we consider the phenomenon of inhomogeneous loading of calcium fluorophores into platelets.

We used platelets from healthy adult donors loaded with various fluorescent probes (CalBryte590, DiOC6 (3), Fura Red, Fluo-4 and CellTracker Violet BMQC) and immobilized on antibodies to CD31 in parallel plane flow chambers. Total internal reflection fluorescence (TIRF) microscopy was used for observations.

We demonstrated that all studied probes are loaded heterogeneously, with 30% platelets being loaded with a probe 2-6 times higher than the population median value. Using the CalBryte590 probe as an example, we have shown that a decrease in the incubation temperature, the addition of Pluronic 127 to the incubation medium, or membrane cholesterol depletion significantly reduces the heterogeneity of the probe distribution in the population. By looking at platelet activation from the surface, we have shown that the probability of experiencing strong activation, as measured by the intensity of calcium oscillations, correlates with the amount of probe in the platelet.

Thus, we conclude that the type of fluorophore used and the conditions of its loading into platelets can significantly affect the results of experiments on the observation of calcium signaling in platelets.

Overall scheme of the assay. First, whole pre-processed blood is perfused through the flow chamber for 2.5 minutes. Then Tyrode's buffer is perfused through the chamber for 5 minutes to wash away unattached cells. Then, depending on the protocol, the microscopy video is recorded for 5-10 minutes.
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#platelets#calcium fluorophores#membrane#fluorescent probes#intracellular signaling

Immune thrombocytopenia: what can the systems biology and systems physiology offer?

Immune thrombocytopenia (ITP) is an acquired bleeding disorder of autoimmune pathophysiology. The causes of ITP could be related to other pathology (viral, bacterial, or systemic), or ITP could develop without any apparent reason. While the immune system dysregulation mechanisms in ITP were described, its etiology remains unclear. Moreover, all existing treatment approaches are not specific for ITP, and its action is highly patient- specific. Here we describe recent findings in the origins and development of ITP and discuss novel experimental and theoretical approaches to diagnosing ITP and predicting therapy effects.

Schematic of immune thrombocytopenia mechanisms. The thrombocytopenia in patients could be caused by both T-cells cytotoxicity (CTLs are more active because T-regulatory cells and T-helpers provide dysregulated stimuli) and B-cells antibody production (plasma cells are producing antiplatelet antibodies of IgG and IgM classes; B-regulatory cells provide more activating signals to B-cells to produce antibodies). CTLs specifically attack platelets and induce their apoptosis. Antibodies from plasma cells opsonize platelets. Consecutively, opsonized platelets are eliminated through the spleen. CTLs – cytotoxic T-lymphocytes; Treg – T-regulatory cell; Th – T-helper; Breg – B-regulatory cell; BAFF – B-cell activating factor.
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#autoimmunity#ITP#platelets#acute/chronic ITP#computational modeling#murine models

Hubs and Webs in Platelet Intracellular Signalling

In this issue of Systems Biology and Physiology Reports A.A. Martyanov and M.A. Panteleev suggested a review on platelet intracellular signalling network, which is a second part in the discussion on the molecular relationships between platelet activation and responses [1]. The review contains seven thousands words and two hundred references and yet it is not complete, as there are still unclear parts in platelet signalling, especially in its inhibition [2-4]. In an effort to comprehend the platelet activation pattern, I drew a signalling scheme based on the review and data from other authors [2, 3].

Scheme of platelet intracellular signalling with focus on cytosolic calcium as a “Hub”. Cytosolic calcium concentration is given in the shades of red. Almost all types of platelet receptor agonists lead to activation of phospholipase C (PLC), followed by calcium release from intracellular stores (DTS). Calcium concentration is rapidly reduced by calcium-dependent ATPases (SERCA and PMCA). Also, it could be reduced by binding with some buffering proteins, including its effector-proteins (indicated with red dots). Platelet mitochondria also can function as a calcium buffer and, simultaneously, be regulated by calcium concentration. Direct activation is shown by solid green arrows, direct inhibition - by solid red arrows. Indirect interactions are shown by dashed lines. Abbreviations. AC – adenylate cyclase, α2AAR - α2A-adrenergic receptor, CDGEF - CalDAGGEFI, COX – cyclooxygenase, DAG - diacylglycerol, DTS - dense tubular system, Fbg – fibrinogen, IP3R - receptor for inositol-1,4,5-trisphosphate (IP3), Mit - a mitochondrion, mPTP - mitochondrial permeability transition pore, NCLX - mitochondrial sodium/calcium exchanger, OCS - open canalicular system, P2Y - purinergic receptor, PAR - protease-activated receptor, PIP2 - phosphoinositol-4,5-bisphosphate, PIP3 - phosphoinositol-3,4,5-trisphosphate, PI(P)n – phosphoinositides, PKA – protein kinase A, PKG – protein kinase G, PKC – protein kinase C, PL – phospholipid, PLA2 – phospholipase A2, PMCA - plasma membrane calcium ATPase, PR - PGI2 receptor, P-Tyr – phosphorylated tyrosine residue, SERCA - sarcoplasmic/endoplasmic reticulum calcium ATPase, sGC – soluble guanylate cyclase, TR - thromboxane A2 (TxA2) receptor, TRPC - transient receptor potential channel, UNI - mitochondrial uniporter, vWF – von Willebrand Factor, Y-Pase – tyrosine phosphatase.
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Platelet functional responses and signalling: the molecular relationship. Part 2: receptors.

Small, non-nuclear cells, platelets, are primarily designed to form aggregates when blood vessels are damaged, stopping bleeding. To perform this function, platelets can implement several functional responses induced by various agonists and coordinated by a complex network of intracellular signaling triggered by a dozen of different receptors. This review, the second in a series, describes the known intracellular signaling pathways induced by platelet receptors in response to canonical and rare agonists. Particular focus will be on interaction points and “synergy” of platelet activation pathways and intermediate or “secondary” activation mediators that transmit a signal to functional manifestations.


Different degrees of the platelet activation in hemostasis. Upon weak stimulation, platelets pass into a weakly activated state, in which there is no clustering of platelet integrins and no significant change in the shape of platelets. This weak activation is reversible, and it corresponds to the state of platelets in the outer layers ("coat") of the thrombus. Upon stronger activation, platelet shape significantly changes. Platelets become irreversibly activated and aggregate. The secretion of platelet granules also occurs. At the maximum degree of activation, platelet mitochondria collapse, and platelets pass into a procoagulant state, exposing phosphatidylserine, which significantly accelerates blood plasma coagulation.
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#platelets#intracellular signaling#physiology

Overview of the neutralizing antibody and memory B cell response kinetics in SARS-CoV-2 convalescent and/or mRNA vaccinated individuals.

COVID-19 pandemics triggered by the SARS-CoV-2 virus have caused millions of deaths worldwide and have led to expedited developments of various effective vaccines that, if administered, could prevent and/or circumvent the infection and reduce the death toll. Since the start of the pandemics multiple research groups around the world have been involved in the analysis of immune responses of various human cohorts to the SARS-CoV-2 infection and vaccines. Now, over 1.5 years later, the scientific community has accumulated extensive data about both the development of an immune response to SARS-CoV-2 following infection, as well as its rate of fading off. Kinetic analysis of the immune response generated by vaccines is also emerging, enabling the possibility of making comparisons and predictions. In this review we will focus on the comparing B cell and antibody immune responses to the SARS-CoV-2 infection as opposed to mRNA vaccines for the SARS-CoV-2 S-protein, which have been utilized to immunize hundreds of millions of people and analyzed in multiple studies.

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#B-cells#COVID-19#vaccines#immune response

A minimal mathematical model of neutrophil pseudopodium formation during chemotaxis

The directed movement of neutrophils is provided by the rapid polymerization of actin with the formation of a protrusion growing forward. In our previous work we observed impaired neutrophil movement for patients with Wiskott-Aldrich syndrome (WAS) compared to healthy donors.

In this work, we set out to explain the impairment of neutrophil chemotaxis in patients by observation and computer modeling of the linear growth rates of the anterior pseudopodia. The neutrophil chemotaxis was observed by means of low-angle fluorescent microscopy in parallel-plate flow chambers. The computational model was constructed as a network-like 2D stochastic polymerization of actin guided by the proximity of cell membrane with branching governed by Arp2/3 and WASP proteins.

The observed linear velocity of neutrophil pseudopodium formation was 0.22 ± 0.04 μm/s for healthy donors and 0.23 ± 0.08 μm/s for WAS patients. The model described the velocity of the pseudopodium formation for healthy donors well. For the description of WAS patients data, a variation of branching velocity (governed by WASP) by an order of magnitude was applied, which did not significantly alter the linear protrusion growth velocity.

We conclude that the proposed mathematical model of neutrophil pseudopodium formation could describe the experimental data well, but the data on overall neutrophil movement could not be explained by the velocities of the pseudopodium growth.

Scheme of the computational model. (A) The scheme of stochastic events and species included in the model. A single F-actin filament is assumed to be straight and to be divided into segments. Each segment can be considered to be an actin monomer. New G-actin molecules can attach to and detach from the filament “barbed” end. It is assumed that the child filament begins to grow from the middle between two segments of F-actin at the angle of 70o.  If there is a branch growing from the F-actin segments, they are considered occupied and no branching can occur there. (B) The spatial restrictions on the filament growth and branching. The filaments can branch if the distance from the cell membrane is lesser than D. Filaments can grow if the distance from the cell membrane is lesser than H, where H > D.
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#cytoskeleton#neutrophils#actin#chemotaxis#Wiskott-Aldrich syndrome

Presence of PI-rich vesicles is required for the PLC ζ activation according to mathematical modeling

Phospholipase C ζ (PLCζ) is an enzyme found in the cytoplasm and acrosome of mammalian spermatozoa. It catalyzes the reaction of phosphatidylinositol-4,5-phosphate hydrolysis into inositol-3-phosphate and diacylglycerol. PLCζ is present in the sperm cell acrosome and cytosol but doesn’t significantly affect its metabolism. However, after the fusion of sperm and egg membranes, its activity increases as it begins to bind membranes of the egg. It is unknown why PLCζ is inactive in spermatozoa or any type of somatic cell.

In this work, the modeling approach explains the reasons for the absence of PLCζ activity in any type of mammalian cells but eggs. A model describing the activity of PLCζ in physiological calcium concentrations was developed. It was shown that the presence of phosphoinositide-rich vesicles is required for the PLC ζ activity in mature mammalian eggs.

Scheme of the full model. (A) Reactions in the oocyte, PLCζ – calcium-free PLCζ , PLCζ_Ca – PLCζ, bound with one calcium ion, PLCζ_2Ca – PLCζ, bound with two calcium ions, PLCζ_3Ca  – with three, PLCζ_4Ca  – with four. PLCζ_m  – PLCζ, bound with cell membrane, PLCζ_(Ca m ) – PLCζ, bound with the cell membrane and one calcium ion, PLCζ_(2Ca m )– PLCζ, bound with the cell membrane and two calcium ions, PLCζ_(3Ca m ) – with the cell membrane and three calcium ions, PLCζ_(4Ca m ) – with the cell membrane and four calcium ions. PIP2 and PIP2_v  – are phosphatidylinositol-4,5 – bis-phosphates on cell and vesicle membrane accordingly. DAG and DAG_v  – diacylglycerol on cell membrane and vesicles accordingly. IP3 – inositol-3-phosphate. PLCζ_v – PLCζ, bound with vesicles, PLCζ_(Ca v ) – PLCζ, bound with vesicles and one calcium ion, PLCζ_(2Ca v ) – PLCζ, bound with vesicles and two calcium ions, PLCζ_(3Ca v ) – with vesicles and three calcium ions, PLCζ_(4Ca v ) – with vesicles and four calcium ions. (B) The sperm cell model is identical to the oocyte model except for the absence of the vesicles.
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#phospholipase Cz#calcium signaling#spermatozoa#oocyte

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