Expandable Megakaryocyte Cell Lines Enable Clinically Applicable Generation of Platelets from Human Induced Pluripotent Stem Cells

Nakamura, Sou; Takayama, Nobuki; Hirata, Shinji; Seo, Hideya; Endô, Hiroshi; Ochi, Kenji; Fujita, Ken‐ichi; Koike, Tsuneaki; Harimoto, Kenichi; Dohda, Takeaki; Watanabe, Akira; Okita, Keisuke; Takahashi, Nobuyasu; Sawaguchi, Akira; Yamanaka, Shinya; Nakauchi, Hiromitsu; Nishimura, Satoshi; Eto, Koji

doi:10.1016/j.stem.2014.01.011

Cited by 271 publications

(343 citation statements)

References 40 publications

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“…18 The development of human embryonic stem cell cultures (hESC), 44,46 and more recently, hiPSC 47 (Feng Q et al, manuscript submitted 2014) offer a physiologically relevant and potentially unlimited source of progenitor cells that can be differentiated into human MKs in vitro to address the first quantitative roadblock. 48 Indeed, because PLTs are anucleate, PLT bioreactor-derived units could be irradiated prior to infusion, addressing concerns that cellular products derived from hESC or hiPSCs could be oncogenic or teratogenic. 49 Nevertheless, attempts to study the environmental drivers of PLT production have fallen short.…”

Section: Discussionmentioning

confidence: 99%

Platelet bioreactor-on-a-chip

Thon

Mažutis

et al. 2014

Blood

179

201

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• We have developed a biomimetic microfluidic platelet bioreactor that recapitulates bone marrow and blood vessel microenvironments.• Application of shear stress in this bioreactor triggers physiological proplatelet production, and platelet release.Platelet transfusions total >2.17 million apheresis-equivalent units per year in the United States and are derived entirely from human donors, despite clinically significant immunogenicity, associated risk of sepsis, and inventory shortages due to high demand and 5-day shelf life. To take advantage of known physiological drivers of thrombopoiesis, we have developed a microfluidic human platelet bioreactor that recapitulates bone marrow stiffness, extracellular matrix composition, micro-channel size, hemodynamic vascular shear stress, and endothelial cell contacts, and it supports high-resolution live-cell microscopy and quantification of platelet production. Physiological shear stresses triggered proplatelet initiation, reproduced ex vivo bone marrow proplatelet production, and generated functional platelets. Modeling human bone marrow composition and hemodynamics in vitro obviates risks associated with platelet procurement and storage to help meet growing transfusion needs. (Blood. 2014;124(12):1857-1867) IntroductionAlthough platelets (PLTs) play critical roles in hemostasis, 1 angiogenesis, 2 and innate immunity, 3 PLT production remains poorly understood. Consequently, PLT units are derived entirely from human donors, despite serious clinical concerns owing to their immunogenicity and associated risk of sepsis. 4 More than 2.17 million apheresisequivalent PLT units are transfused yearly in the United States 5,6 at a cost of .$1 billion per year. Although demand for PLT transfusions has increased markedly in the past decade, a near-static pool of donors and a 5-day PLT unit shelf life resulting from bacterial contamination 7 and storage-related PLT deterioration, 8 have resulted in significant PLT shortages. 9 Furthermore, artificial platelet substitutes have failed to replace physiological platelet products. 10 An efficient, donorindependent PLT bioreactor capable of generating clinically significant numbers of functional human PLTs is necessary to obviate risks associated with PLT procurement and storage, and help meet growing transfusion needs. In vivo, megakaryocytes (MKs) PLT progenitors sit outside blood vessels in the bone marrow (BM) and extend long, branching cellular structures designated proPLTs into the circulation from which PLTs are released. 11-15 Nearly 100% of human adult MKs must produce ;10 3 PLTs each to account for circulating PLT counts. 16 Although functional human PLTs were first grown in vitro in 1995, 17 to date only ;10% of human MKs initiate proPLT production in culture. This results in yields of 10 122 PLTs per CD34 1 cord blood-derived or embryonic stem cell-derived MK, 18 which are themselves of limited availability, constituting a significant bottleneck in the ex vivo production of a PLT transfusion unit. Although second-generation c...

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Section: Discussionmentioning

confidence: 99%

Platelet bioreactor-on-a-chip

Thon

Mažutis

et al. 2014

Blood

179

201

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“…It will be difficult to fulfil this necessity only with human volunteer donors (32), therefore, it is desirable to establish alternative PLT sources. To solve the problem of PLT availability, we and others have established different protocols to generate functional PLTs in vitro (9,19,(33)(34)(35). Nevertheless, PLT transfusion is associated with risks and adverse effects.…”

Section: Hla-universal Mks Generate Plts In a Refractoriness Mouse Modelmentioning

confidence: 99%

Generation of HLA-Universal iPSC-Derived Megakaryocytes and Platelets for Survival Under Refractoriness Conditions

Börger

Eicke

Wolf

et al. 2016

Mol Med

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Platelet (PLT) transfusion is indispensable to maintain homeostasis in thrombocytopenic patients. However, PLT transfusion refractoriness is a common life-threatening condition observed in multitransfused patients. The most frequent immune cause for PLT transfusion refractoriness is the presence of alloantibodies specific for human leukocyte antigen (HLA) class I epitopes. Here, we have silenced the expression of HLA class I to generate a stable HLA-universal induced pluripotent stem cell (iPSC) line that can be used as a renewable cell source for the generation of low immunogenic cell products. The expression of HLA class I was silenced by up to 82% and remained stable during iPSC cultivation. In this study, we have focused on the generation of megakaryocytes (MK) and PLTs from a HLA-universal iPSC source under feeder-and xeno-free conditions. On d 19, differentiation rates of MKs and PLTs with means of 58% and 76% were observed, respectively. HLA-universal iPSC-derived MKs showed polyploidy with DNA contents higher than 4n and formed proPLTs. Importantly, differentiated MKs remained silenced for HLA class I expression. HLA-universal MKs produced functional PLTs. Notably, iPSC-derived HLA-universal MKs were capable to escape antibody-mediated complement-and cellular-dependent cytotoxicity. Furthermore, HLA-universal MKs were able to produce PLTs after in vivo transfusion in a mouse model indicating that they might be used as an alternative to PLT transfusion. Thus, in vitro produced low immunogenic MKs and PLTs may become an alternative to PLT donation in PLT-based therapies and an important component in the management of severe alloimmunized patients.

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“…"Just -in -time delivery" is a hallmark of Japanese inventory logistics, but our typical 30 -40 minute delay between a clinician's request and a product's arrival is unduly long when there is critical bleeding or hemorrhagic stroke risk. The prospect of PLTs derived from induced Pluripotent Stem (iPS) cells is much anticipated 25) . As for other agents to reverse effects of APAs, recombinant activated factor VII (rVIIa) and DDAVP have been recommended in European and other guidelines 26 -28) , but with little supporting evidence.…”

Section: Discussionmentioning

confidence: 99%

Intracranial hemorrhage and platelet transfusion after administration of anti-platelets agents: Fukushima Prefecture

Suzuki

Sato

Sakuma

et al. 2016

FJMS

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We conducted a case series study to assess intracerebral hemorrhage (ICH) in the context of antiplatelets agents (APAs) and platelet (PLT) transfusion in Fukushima Prefecture. This study included patients who were newly diagnosed with ICH between January 2008 and June 2014 in the neurosurgical hospitals of Fukushima Prefecture. Four of ten neurosurgical hospitals responded to our questionnaire. Of 287 ICH patients, 51 (20.6%) were on APA therapy, of whom PLT transfusion was given to only one persistently bleeding patient who was on dual anti -platelet therapy. In a follow -up survey, 30 out of 51 ICH patients on APA therapy, average age 75 years, were analyzed, of whom 21 (70%) were male. The predominant underlying disease was diabetes mellitus. It is interesting to note that peripheral artery disease and aortic aneurysm were among the indications for APAs. ICH was mainly observed supratentorially. Hematoma enlargement was observed in 13 (44.8%) cases. By day 7, 3 patients (10%) had died from complications of ICH. In this study, we show that ICH during APA therapy matched what was observed in Kanagawa Prefecture. Whether or not a national survey differs, we anticipate greater statistical validity and an opportunity to improve patient outcomes in Japan and around the world.

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Expandable Megakaryocyte Cell Lines Enable Clinically Applicable Generation of Platelets from Human Induced Pluripotent Stem Cells

Cited by 271 publications

References 40 publications

Platelet bioreactor-on-a-chip

Platelet bioreactor-on-a-chip

Generation of HLA-Universal iPSC-Derived Megakaryocytes and Platelets for Survival Under Refractoriness Conditions

Intracranial hemorrhage and platelet transfusion after administration of anti-platelets agents: Fukushima Prefecture

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