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Journal of Translational Medicine Aug 2023Cyclooxygenase (COX)-2 is a rate-limiting enzyme in the biosynthesis of prostanoids, which is mostly inducible by inflammatory cytokines. The participation of COX-2 in...
BACKGROUND
Cyclooxygenase (COX)-2 is a rate-limiting enzyme in the biosynthesis of prostanoids, which is mostly inducible by inflammatory cytokines. The participation of COX-2 in the maturation of megakaryocytes has been reported but barely studied in primary immune thrombocytopenia (ITP).
METHODS
The expressions of COX-2 and Caspase-1, Caspase-3 and Caspase-3 p17 subunit in platelets from ITP patients and healthy controls (HC), and the expressions of COX-2 and CD41 in bone marrow (BM) of ITP patients were measured and analyzed for correlations. The effects of COX-2 inhibitor on megakaryopoiesis and thrombopoiesis were assessed by in vitro culture of Meg01 cells and murine BM-derived megakaryocytes and in vivo experiments of passive ITP mice.
RESULTS
The expression of COX-2 was decreased and Caspase-1 and Caspase-3 p17 were increased in platelets from ITP patients compared to HC. In platelets from ITP patients, the COX-2 expression was positively correlated with platelet count and negatively correlated to the expression of Caspase-1. In ITP patients BM, the expression of CD41 was positively correlated with the expression of COX-2. COX-2 inhibitor inhibited the count of megakaryocytes and impaired the maturation and platelet production in Meg01 cells and bone marrow-derived megakaryocytes. COX-2 inhibitor aggravated thrombocytopenia and damaged megakaryopoiesis in ITP murine model.
CONCLUSION
COX-2 plays a vital role in the physiologic and pathologic conditions of ITP by intervening the survival of platelets and impairing the megakaryopoiesis and thrombopoiesis of megakaryocytes.
Topics: Animals; Mice; Blood Platelets; Caspase 3; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Megakaryocytes; Purpura, Thrombocytopenic, Idiopathic; Thrombopoiesis
PubMed: 37573325
DOI: 10.1186/s12967-023-04389-9 -
Radiation Research Sep 2021Thrombocytopenia is a major complication in hematopoietic-acute radiation syndrome (H-ARS) that increases the risk of mortality from uncontrolled hemorrhage. There is a... (Comparative Study)
Comparative Study
Thrombocytopenia is a major complication in hematopoietic-acute radiation syndrome (H-ARS) that increases the risk of mortality from uncontrolled hemorrhage. There is a great demand for new therapies to improve survival and mitigate bleeding in H-ARS. Thrombopoiesis requires interactions between megakaryocytes (MKs) and endothelial cells. 16, 16-dimethyl prostaglandin E2 (dmPGE2), a longer-acting analogue of PGE2, promotes hematopoietic recovery after total-body irradiation (TBI), and various angiotensin-converting enzyme (ACE) inhibitors mitigate endothelial injury after radiation exposure. Here, we tested a combination therapy of dmPGE2 and lisinopril to mitigate thrombocytopenia in murine models of H-ARS following TBI. After 7.75 Gy TBI, dmPGE2 and lisinopril each increased survival relative to vehicle controls. Importantly, combined dmPGE2 and lisinopril therapy enhanced survival greater than either individual agent. Studies performed after 4 Gy TBI revealed reduced numbers of marrow MKs and circulating platelets. In addition, sublethal TBI induced abnormalities both in MK maturation and in in vitro and in vivo platelet function. dmPGE2, alone and in combination with lisinopril, improved recovery of marrow MKs and peripheral platelets. Finally, sublethal TBI transiently reduced the number of marrow Lin-CD45-CD31+Sca-1- sinusoidal endothelial cells, while combined dmPGE2 and lisinopril treatment, but not single-agent treatment, accelerated their recovery. Taken together, these data support the concept that combined dmPGE2 and lisinopril therapy improves thrombocytopenia and survival by promoting recovery of the MK lineage, as well as the MK niche, in the setting of H-ARS.
Topics: 16,16-Dimethylprostaglandin E2; Acute Radiation Syndrome; Angiotensin-Converting Enzyme Inhibitors; Animals; Blood Platelets; Bone Marrow; C-Reactive Protein; Cesium Radioisotopes; Drug Evaluation, Preclinical; Endothelial Cells; Endothelium, Vascular; Female; Gamma Rays; Hemorrhagic Disorders; Lisinopril; Megakaryocytes; Mice; Mice, Inbred C57BL; P-Selectin; Platelet Aggregation; Platelet Factor 4; Radiation Injuries, Experimental; Thrombocytopenia; Thrombopoiesis; Whole-Body Irradiation; von Willebrand Factor
PubMed: 34153091
DOI: 10.1667/RADE-20-00113.1 -
Hamostaseologie Aug 2019Management of patients with corticosteroid-refractory immune thrombocytopaenia (ITP) possesses a significant challenge to practitioners. Until recently, options included... (Comparative Study)
Comparative Study Review
Management of patients with corticosteroid-refractory immune thrombocytopaenia (ITP) possesses a significant challenge to practitioners. Until recently, options included splenectomy and immunosuppression. With improved knowledge of both thrombopoiesis and the pathophysiology of ITP, novel drug development with thrombopoietin-receptor agonists (TPO-RAs) was undertaken. Two agents, romiplostim and eltrombopag, are currently approved for use in patients with chronic ITP. Both agents have been shown to increase the platelet count, improve health-related quality of life and reduce bleeding symptoms and concomitant medication use. This review will highlight the discovery of TPO-RA agents, appraise key clinical trials and explore future directions.
Topics: Administration, Oral; Adult; Benzoates; Child; Child, Preschool; Clinical Trials as Topic; Hemorrhage; Humans; Hydrazines; Immunosuppression Therapy; Infant; Infant, Newborn; Platelet Count; Purpura, Thrombocytopenic, Idiopathic; Pyrazoles; Quality of Life; Receptors, Fc; Receptors, Thrombopoietin; Recombinant Fusion Proteins; Safety; Splenectomy; Thrombopoiesis; Thrombopoietin
PubMed: 30646404
DOI: 10.1055/s-0038-1676129 -
Blood Jan 2023Glycosylation is recognized as a key process for proper megakaryopoiesis and platelet formation. The enzyme uridine diphosphate (UDP)-galactose-4-epimerase, encoded by...
Glycosylation is recognized as a key process for proper megakaryopoiesis and platelet formation. The enzyme uridine diphosphate (UDP)-galactose-4-epimerase, encoded by GALE, is involved in galactose metabolism and protein glycosylation. Here, we studied 3 patients from 2 unrelated families who showed lifelong severe thrombocytopenia, bleeding diathesis, mental retardation, mitral valve prolapse, and jaundice. Whole-exome sequencing revealed 4 variants that affect GALE, 3 of those previously unreported (Pedigree A, p.Lys78ValfsX32 and p.Thr150Met; Pedigree B, p.Val128Met; and p.Leu223Pro). Platelet phenotype analysis showed giant and/or grey platelets, impaired platelet aggregation, and severely reduced alpha and dense granule secretion. Enzymatic activity of the UDP-galactose-4-epimerase enzyme was severely decreased in all patients. Immunoblotting of platelet lysates revealed reduced GALE protein levels, a significant decrease in N-acetyl-lactosamine (LacNAc), showing a hypoglycosylation pattern, reduced surface expression of gylcoprotein Ibα-IX-V (GPIbα-IX-V) complex and mature β1 integrin, and increased apoptosis. In vitro studies performed with patients-derived megakaryocytes showed normal ploidy and maturation but decreased proplatelet formation because of the impaired glycosylation of the GPIbα and β1 integrin, and reduced externalization to megakaryocyte and platelet membranes. Altered distribution of filamin A and actin and delocalization of the von Willebrand factor were also shown. Overall, this study expands our knowledge of GALE-related thrombocytopenia and emphasizes the critical role of GALE in the physiological glycosylation of key proteins involved in platelet production and function.
Topics: Humans; Blood Platelets; Galactose; Glycosylation; Integrin beta1; Megakaryocytes; Thrombocytopenia; Thrombopoiesis; UDPglucose 4-Epimerase; Uridine Diphosphate
PubMed: 36395340
DOI: 10.1182/blood.2022016995 -
Mini Reviews in Medicinal Chemistry 2021Pluripotent stem cells of the bone marrow are stimulated by different cytokines to proliferation and differentiation into various types of blood cells. These cytokines... (Review)
Review
Pluripotent stem cells of the bone marrow are stimulated by different cytokines to proliferation and differentiation into various types of blood cells. These cytokines are mostly glycoproteins. Erythropoietin stimulates stem cells to the formation of erythrocytes while colony-stimulating factors cause the formation of different types of white blood cells. Stem cell factors play an important role in the maintenance and survival of blood cells of all types. Thrombopoietin stimulates stem cells to proliferation and formation of blood platelets. Granulocyte colony-stimulating factor is probably the most important drug in use. It stimulates stem cells to the formation of neutrophile granulocytes. It is often used in recombinant forms such as filgrastim in the treatment of neutropenia in cancer chemotherapy or AIDS. Its pegylated conjugates such as pegfilgrastim are also available. Its activity can be supported by plerixafor, a small molecule - bicyclam derivative acting as an indirect agonist of stem cells factor. It acts as an antagonist of CXCR4 receptor activation of which brakes hematopoiesis. The treatment of conditions accompanied by thrombocytopenia such as idiopathic thrombocytopenic purpura is currently not performed by thrombopoietin but synthetic agonists of its receptor are preferred. Romiplostim is a peptibody. It consists of a protein part interacting with the thrombopoietin receptor which is, however, different from thrombopoietin, and of Fc fragment of immunoglobulin G1. In contrast, small molecule thrombopoietin receptor agonists represented by eltrombopag can be given orally unlike all of the above.
Topics: Benzoates; Blood Platelets; Cell Differentiation; Colony-Stimulating Factors; Humans; Hydrazines; Leukocytes, Mononuclear; Pyrazoles; Receptors, Thrombopoietin; Small Molecule Libraries; Stem Cell Factor; Thrombopoiesis
PubMed: 33390132
DOI: 10.2174/1389557521999201230195926 -
Blood Jan 2024
Topics: Bone Marrow; Megakaryocytes; Thrombopoiesis; Erythrocyte Membrane; Lung
PubMed: 38236613
DOI: 10.1182/blood.2023022897 -
Blood Dec 2022
Topics: Humans; Thrombopoiesis; Megakaryocytes; Blood Platelets; Thrombosis
PubMed: 36480224
DOI: 10.1182/blood.2022017936 -
Cellular and Molecular Life Sciences :... Apr 2021The discovery of iPSCs has led to the ex vivo production of differentiated cells for regenerative medicine. In the case of transfusion products, the derivation of... (Review)
Review
The discovery of iPSCs has led to the ex vivo production of differentiated cells for regenerative medicine. In the case of transfusion products, the derivation of platelets from iPSCs is expected to complement our current blood-donor supplied transfusion system through donor-independent production with complete pathogen-free assurance. This derivation can also overcome alloimmune platelet transfusion refractoriness by resulting in autologous, HLA-homologous or HLA-deficient products. Several developments were necessary to produce a massive number of platelets required for a single transfusion. First, expandable megakaryocytes were established from iPSCs through transgene expression. Second, a turbulent-type bioreactor with improved platelet yield and quality was developed. Third, novel drugs that enabled efficient feeder cell-free conditions were developed. Fourth, the platelet-containing suspension was purified and resuspended in an appropriate buffer. Finally, the platelet product needed to be assured for competency and safety including non-tumorigenicity through in vitro and in vivo preclinical tests. Based on these advancements, a clinical trial has started. The generation of human iPSC-derived platelets could evolve transfusion medicine to the next stage and assure a ubiquitous, safe supply of platelet products. Further, considering the feasibility of gene manipulations in iPSCs, other platelet products may bring forth novel therapeutic measures.
Topics: Animals; Blood Platelets; Blood Transfusion; Cell Differentiation; Humans; Induced Pluripotent Stem Cells; Megakaryocytes; Regenerative Medicine
PubMed: 33439272
DOI: 10.1007/s00018-020-03749-8 -
Journal of Thrombosis and Haemostasis :... Sep 2023Germline mutations in the ETV6 transcription factor gene are responsible for familial thrombocytopenia and leukemia predisposition syndrome. Although previous studies...
BACKGROUND
Germline mutations in the ETV6 transcription factor gene are responsible for familial thrombocytopenia and leukemia predisposition syndrome. Although previous studies have shown that ETV6 plays an important role in megakaryocyte (MK) maturation and platelet formation, the mechanisms by which ETV6 dysfunction promotes thrombocytopenia remain unclear.
OBJECTIVES
To decipher the transcriptional mechanisms and gene regulatory network linking ETV6 germline mutations and thrombocytopenia.
METHODS
Presuming that ETV6 mutations result in selective effects at a particular cell stage, we applied single-cell RNA sequencing to understand gene expression changes during megakaryopoiesis in peripheral CD34 cells from healthy controls and patients with ETV6-related thrombocytopenia.
RESULTS
Analysis of gene expression and regulon activity revealed distinct clusters partitioned into 7 major cell stages: hematopoietic stem/progenitor cells, common-myeloid progenitors (CMPs), MK-primed CMPs, granulocyte-monocyte progenitors, MK-erythroid progenitors (MEPs), progenitor MKs/mature MKs, and platelet-like particles. We observed a differentiation trajectory in which MEPs developed directly from hematopoietic stem/progenitor cells and bypassed the CMP stage. ETV6 deficiency led to the development of aberrant cells as early as the MEP stage, which intensified at the progenitor MK/mature MK stage, with a highly deregulated core "ribosome biogenesis" pathway. Indeed, increased translation levels have been documented in patient CD34-derived MKs with overexpression of ribosomal protein S6 and phosphorylated ribosomal protein S6 in both CD34-derived MKs and platelets. Treatment of patient MKs with the ribosomal biogenesis inhibitor CX-5461 resulted in an increase in platelet-like particles.
CONCLUSION
These findings provide novel insight into both megakaryopoiesis and the link among ETV6, translation, and platelet production.
Topics: Humans; Cell Differentiation; Megakaryocytes; Ribosomal Protein S6; Single-Cell Analysis; Thrombocytopenia; Thrombopoiesis; Antigens, CD34; ETS Translocation Variant 6 Protein
PubMed: 37085035
DOI: 10.1016/j.jtha.2023.04.007 -
British Journal of Haematology Oct 2023Immune thrombocytopenia (ITP) is a disorder characterized by low platelets due to increased clearance and decreased platelet production. While ITP has been characterized... (Review)
Review
Immune thrombocytopenia (ITP) is a disorder characterized by low platelets due to increased clearance and decreased platelet production. While ITP has been characterized as an acquired disorder of the adaptive immune system, the resulting platelet autoantibodies provide ancillary links to the innate immune system via antibody interaction with the complement system. Most autoantibodies in patients with ITP are of the IgG1 subclass, which can be potent activators of the classical complement pathway. Antibody-coated platelets can initiate complement activation via the classical pathway leading to both direct platelet destruction and enhanced clearance of C3b-coated platelets by complement receptors. Similar autoantibody interactions with bone marrow megakaryocytes can also result in complement injury and ineffective thrombopoiesis. The development of novel therapeutic complement inhibitors has revived interest in the role of complement in autoantibody-mediated disorders, such as ITP. A recent early-phase clinical trial of a classical complement pathway inhibitor has demonstrated efficacy in a subset of ITP patients refractory to conventional immune modulation. In this review, we will analyse the role of complement in refractory ITP.
Topics: Humans; Purpura, Thrombocytopenic, Idiopathic; Thrombocytopenia; Complement System Proteins; Blood Platelets; Autoantibodies
PubMed: 37735550
DOI: 10.1111/bjh.19070