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Scandinavian Journal of Clinical and... Apr 2024The Immature Platelet Fraction (IPF) is an indicator of thrombopoiesis which is a useful parameter in thrombocytopenia. It demonstrates compensatory mechanisms in...
BACKGROUND
The Immature Platelet Fraction (IPF) is an indicator of thrombopoiesis which is a useful parameter in thrombocytopenia. It demonstrates compensatory mechanisms in production of platelets, but currently not implemented in routine clinical practice. The aim of this study was to establish the reproducibility and stability of IPF, for both percentage (%-IPF) and absolute (A-IPF) measurements. A total of 71 samples, of which 45 for reproducibility and 26 for stability analysis, were assayed for full blood count using the Sysmex XN-10 analyser at room temperature (RT:19-25 °C). For reproducibility analysis, IPF measurements were analysed 11 times by different appraisers using the same sample, while for stability analysis, IPF was measured over fourteen hourly-intervals up to 24 h ( = 21) and then separately extended beyond the point of stability to 72 h ( = 5).
RESULTS
Reproducibility analysis of %-IPF and A-IPF ( = 45) showed very reliable results, with the range of mean CV% values between 1.25-8.90% and 1.70-9.96%, respectively. On the other hand, overall, stability analysis of %-IPF and A-IPF ( = 21) at RT over 24 h showed reliable results, with pooled mean CV% values of 1.32% and 1.43%, respectively, with no significant difference between %-IPF and A-IPF ( = 0.767 and = 0.821). All %-IPF and A-IPF values had exceeded the set acceptance criterion of stability (CV% ≥ 10.0%) before 72 h.
CONCLUSIONS
Overall, %-IPF and A-IPF reproducibility and storage at RT for 24 h predominantly demonstrates the suitability of their usage for testing on the Sysmex XN-series analysers.
Topics: Humans; Reproducibility of Results; Blood Platelets; Platelet Count; Thrombocytopenia; Thrombopoiesis
PubMed: 38420711
DOI: 10.1080/00365513.2024.2321590 -
Recent Patents on Biotechnology 2024Immune thrombocytopenic purpura (ITP) is an autoimmune disorder determined by immune-mediated platelet demolition and reduction of platelet production. Romiplostim is a... (Review)
Review
Immune thrombocytopenic purpura (ITP) is an autoimmune disorder determined by immune-mediated platelet demolition and reduction of platelet production. Romiplostim is a new thrombopoiesis motivating peptibody that binds and stimulates the human thrombopoietin receptor the patent of which was registered in 2008. It is used to treat thrombocytopenia in patients with chronic immune thrombocytopenic purpura. Romiplostim is a 60 kDa peptibody designed to inhibit cross-reacting immune responses. It consists of four high-affinity TPO-receptor binding domains for the Mpl receptor and one human IgG1 Fc domain. is a good host for the fabrication of recombinant proteins such as romiplostim. The expression of a gene intended in is dependent on many factors such as a protein's inherent ability to fold, mRNA's secondary structure, its solubility, its toxicity preferential codon use, and its need for post-translational modification (PTM). This review focuses on the structure, function, mechanism of action, and expressive approach to romiplostim in .
Topics: Humans; Purpura, Thrombocytopenic, Idiopathic; Escherichia coli; Patents as Topic; Blood Platelets; Thrombopoietin; Receptors, Fc; Recombinant Fusion Proteins
PubMed: 38282441
DOI: 10.2174/1872208317666230503094451 -
Regenerative Therapy Mar 2024The production of platelets from induced pluripotent cells (iPSCs) may offer a safer and sustainable alternative for transfusions and drug delivery systems (DDS).... (Review)
Review
The production of platelets from induced pluripotent cells (iPSCs) may offer a safer and sustainable alternative for transfusions and drug delivery systems (DDS). However, the mass production of the clinically required number of iPSC-derived platelets (iPSC-PLTs) is challenging. Here, we introduce recent technologies for mass production and the first-in-human clinical trial using iPSC-PLTs. To this end, we established immortalized megakaryocyte progenitor cell lines (imMKCLs) as an expandable master cell bank (MCB) through the overexpression of c-MYC, BMI1 and BCL-XL, which modulated megakaryopoiesis and thrombopoiesis. We also optimized a culture cocktail for maturation of the imMKCLs by mixing an aryl hydrocarbon receptor (AhR) antagonist, SR1/GNF-316; a Rho-associated protein kinase (ROCK) inhibitor, Y-27632/Y-39983; and a small-molecule compound replacing recombinant thrombopoietin (TPO), TA-316. Finally, we discovered the importance of turbulence on the manufacturing of intact iPSC-PLTs, allowing us to develop a turbulence-based bioreactor, VerMES. Combination of the MCB and VerMES enabled us to produce more than 100 billion iPSC-PLTs, leading to the first-in-human clinical trial. Despite these advancements, many challenges remain before expanding the clinical implementation of this strategy.
PubMed: 38260088
DOI: 10.1016/j.reth.2023.12.009 -
Pharmaceuticals (Basel, Switzerland) Jan 2024Interleukins, a diverse family of cytokines produced by various cells, play crucial roles in immune responses, immunoregulation, and a wide range of physiological and... (Review)
Review
Interleukins, a diverse family of cytokines produced by various cells, play crucial roles in immune responses, immunoregulation, and a wide range of physiological and pathological processes. In the context of megakaryopoiesis, thrombopoiesis, and platelet function, interleukins have emerged as key regulators, exerting significant influence on the development, maturation, and activity of megakaryocytes (MKs) and platelets. While the therapeutic potential of interleukins in platelet-related diseases has been recognized for decades, their clinical application has been hindered by limitations in basic research and challenges in drug development. Recent advancements in understanding the molecular mechanisms of interleukins and their interactions with MKs and platelets, coupled with breakthroughs in cytokine engineering, have revitalized the field of interleukin-based therapeutics. These breakthroughs have paved the way for the development of more effective and specific interleukin-based therapies for the treatment of platelet disorders. This review provides a comprehensive overview of the effects of interleukins on megakaryopoiesis, thrombopoiesis, and platelet function. It highlights the potential clinical applications of interleukins in regulating megakaryopoiesis and platelet function and discusses the latest bioengineering technologies that could improve the pharmacokinetic properties of interleukins. By synthesizing the current knowledge in this field, this review aims to provide valuable insights for future research into the clinical application of interleukins in platelet-related diseases.
PubMed: 38256942
DOI: 10.3390/ph17010109 -
Blood Jan 2024
Topics: Bone Marrow; Megakaryocytes; Thrombopoiesis; Erythrocyte Membrane; Lung
PubMed: 38236613
DOI: 10.1182/blood.2023022897 -
American Journal of Hematology Mar 2024Mechanisms through which mature megakaryocytes (Mks) and their progenitors sense the bone marrow extracellular matrix to promote lineage differentiation in health and...
Mechanisms through which mature megakaryocytes (Mks) and their progenitors sense the bone marrow extracellular matrix to promote lineage differentiation in health and disease are still partially understood. We found PIEZO1, a mechanosensitive cation channel, to be expressed in mouse and human Mks. Human mutations in PIEZO1 have been described to be associated with blood cell disorders. Yet, a role for PIEZO1 in megakaryopoiesis and proplatelet formation has never been investigated. Here, we show that activation of PIEZO1 increases the number of immature Mks in mice, while the number of mature Mks and Mk ploidy level are reduced. Piezo1/2 knockout mice show an increase in Mk size and platelet count, both at basal state and upon marrow regeneration. Similarly, in human samples, PIEZO1 is expressed during megakaryopoiesis. Its activation reduces Mk size, ploidy, maturation, and proplatelet extension. Resulting effects of PIEZO1 activation on Mks resemble the profile in Primary Myelofibrosis (PMF). Intriguingly, Mks derived from Jak2 PMF mice show significantly elevated PIEZO1 expression, compared to wild-type controls. Accordingly, Mks isolated from bone marrow aspirates of JAK2 PMF patients show increased PIEZO1 expression compared to Essential Thrombocythemia. Most importantly, PIEZO1 expression in bone marrow Mks is inversely correlated with patient platelet count. The ploidy, maturation, and proplatelet formation of Mks from JAK2 PMF patients are rescued upon PIEZO1 inhibition. Together, our data suggest that PIEZO1 places a brake on Mk maturation and platelet formation in physiology, and its upregulation in PMF Mks might contribute to aggravating some hallmarks of the disease.
Topics: Humans; Animals; Mice; Megakaryocytes; Primary Myelofibrosis; Bone Marrow; Thrombopoiesis; Thrombocythemia, Essential; Blood Platelets; Ion Channels
PubMed: 38165047
DOI: 10.1002/ajh.27184 -
Scientific Reports Dec 2023The use of megakaryoblastic leukemia MEG-01 cells can help reveal the mechanisms of thrombopoiesis. However, conventional in vitro activation of platelet release from...
The use of megakaryoblastic leukemia MEG-01 cells can help reveal the mechanisms of thrombopoiesis. However, conventional in vitro activation of platelet release from MEG-01 cells requires thrombopoietin, which is costly. Here, we aim to develop a more straightforward and affordable method. Synchronization of the MEG-01 cells was initially performed using serum-free culture, followed by spontaneous cell differentiation in the presence of serum. Different stages of megakaryoblast differentiation were classified based on cell morphology, DNA content, and cell cycle. The MEG-01 cells released platelet-like particles at a level comparable to that of the thrombopoietin-activated MEG-01 cells. The platelet-like particles were distinguishable from PLP-derived extracellular vesicles and could express P-selectin following ADP activation. Importantly, the platelet-like particles induced fibrin clotting in vitro using platelet-poor plasma. Therefore, this thrombopoietin-independent cell synchronization method is an effective and straightforward method for studying megakaryopoiesis and thrombopoiesis.
Topics: Megakaryocytes; Thrombopoietin; Megakaryocyte Progenitor Cells; Blood Platelets; Thrombopoiesis
PubMed: 38110522
DOI: 10.1038/s41598-023-50111-6 -
Critical shifts in lipid metabolism promote megakaryocyte differentiation and proplatelet formation.Nature Cardiovascular Research Sep 2023During megakaryopoiesis, megakaryocytes (MK) undergo cellular morphological changes with strong modification of membrane composition and lipid signaling. Here we adopt a...
During megakaryopoiesis, megakaryocytes (MK) undergo cellular morphological changes with strong modification of membrane composition and lipid signaling. Here we adopt a lipid-centric multiomics approach to create a quantitative map of the MK lipidome during maturation and proplatelet formation. Data reveal that MK differentiation is driven by an increased fatty acyl import and lipid synthesis, resulting in an anionic membrane phenotype. Pharmacological perturbation of fatty acid import and phospholipid synthesis blocked membrane remodeling and directly reduced MK polyploidization and proplatelet formation resulting in thrombocytopenia. The anionic lipid shift during megakaryopoiesis was paralleled by lipid-dependent relocalization of the scaffold protein CKIP-1 and recruitment of the kinase CK2α to the plasma membrane, which seems to be essential for sufficient platelet biogenesis. Overall, this study provides a framework to understand how the MK lipidome is altered during maturation and the impact of MK membrane lipid remodeling on MK kinase signaling involved in thrombopoiesis.
PubMed: 38075556
DOI: 10.1038/s44161-023-00325-8 -
Cells Dec 2023The bone marrow (BM) hematopoietic system (HS) gives rise to blood cells originating from hematopoietic stem cells (HSCs), including megakaryocytes (MKs) and red blood...
The bone marrow (BM) hematopoietic system (HS) gives rise to blood cells originating from hematopoietic stem cells (HSCs), including megakaryocytes (MKs) and red blood cells (erythrocytes; RBCs). Many steps of the cell-fate decision remain to be elucidated, being important for cancer treatment. To explore the role of Wnt/β-catenin for MK and RBC differentiation, we activated β-catenin signaling in platelet-derived growth factor b (Pdgfb)-expressing cells of the HS using a Cre-lox approach (Ctnnb1). FACS analysis revealed that Pdgfb is mainly expressed by megakaryocytic progenitors (MKPs), MKs and platelets. Recombination resulted in a lethal phenotype in mutants (Ctnnb1, Ctnnb1) 3 weeks after tamoxifen injection, showing an increase in MKs in the BM and spleen, but no pronounced anemia despite reduced erythrocyte counts. BM transplantation (BMT) of Ctnnb1 BM into lethally irradiated wildtype recipients (BMT-Ctnnb1) confirmed the megakaryocytic, but not the lethal phenotype. CFU-MK assays in vitro with BM cells of Ctnnb1 mice supported MK skewing at the expense of erythroid colonies. Molecularly, the runt-related transcription factor 1 (RUNX1) mRNA, known to suppress erythropoiesis, was upregulated in Ctnnb1 BM cells. In conclusion, β-catenin activation plays a key role in cell-fate decision favoring MK development at the expense of erythroid production.
Topics: Animals; Mice; beta Catenin; Megakaryocyte-Erythroid Progenitor Cells; Megakaryocytes; Proto-Oncogene Proteins c-sis; Thrombopoiesis
PubMed: 38067194
DOI: 10.3390/cells12232765 -
Life Science Alliance Feb 2024Platelets display unexpected roles in immune and coagulation responses. Emerging evidence suggests that STING is implicated in hypercoagulation. STING is an adaptor...
Platelets display unexpected roles in immune and coagulation responses. Emerging evidence suggests that STING is implicated in hypercoagulation. STING is an adaptor protein downstream of the DNA sensor cyclic GMP-AMP synthase (cGAS) that is activated by cytosolic microbial and self-DNA during infections, and in the context of loss of cellular integrity, to instigate the production of type-I IFN and pro-inflammatory cytokines. To date, whether the cGAS-STING pathway is present in platelets and contributes to platelet functions is not defined. Using a combination of pharmacological and genetic approaches, we demonstrate here that megakaryocytes and platelets possess a functional cGAS-STING pathway. Our results suggest that in megakaryocytes, STING stimulation activates a type-I IFN response, and during thrombopoiesis, cGAS and STING are transferred to proplatelets. Finally, we show that both murine and human platelets contain cGAS and STING proteins, and the cGAS-STING pathway contributes to potentiation of platelet activation and aggregation. Taken together, these observations establish for the first time a novel role of the cGAS-STING DNA sensing axis in the megakaryocyte and platelet lineage.
Topics: Animals; Humans; Mice; Megakaryocytes; Signal Transduction; DNA; Cytokines; Nucleotidyltransferases; Interferon Type I
PubMed: 37993259
DOI: 10.26508/lsa.202302211