scholarly journals Improving Human Induced Pluripotent Stem Cell-Derived Megakaryocyte Differentiation and Platelet Production

2021 ◽  
Vol 22 (15) ◽  
pp. 8224
Author(s):  
Linda Krisch ◽  
Gabriele Brachtl ◽  
Sarah Hochmann ◽  
André Cronemberger Andrade ◽  
Michaela Oeller ◽  
...  
Keyword(s):  
Pluripotent Stem Cells ◽  
Iterative Process ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Mesoderm Induction ◽  
Related Protein ◽  
Media Composition ◽  
Platelet Production ◽  
Induced Pluripotent ◽  
Related Proteins

Several protocols exist for generating megakaryocytes (MKs) and platelets from human induced pluripotent stem cells (hiPSCs) with limited efficiency. We observed previously that mesoderm induction improved endothelial and stromal differentiation. We, therefore, hypothesized that a protocol modification prior to hemogenic endothelial cell (HEC) differentiation will improve MK progenitor (MKP) production and increase platelet output. We further asked if basic media composition affects MK maturation. In an iterative process, we first compared two HEC induction protocols. We found significantly more HECs using the modified protocol including activin A and CHIR99021, resulting in significantly increased MKs. MKs released comparable platelet amounts irrespective of media conditions. In a final validation phase, we obtained five-fold more platelets per hiPSC with the modified protocol (235 ± 84) compared to standard conditions (51 ± 15; p < 0.0001). The regenerative potency of hiPSC-derived platelets was compared to adult donor-derived platelets by profiling angiogenesis-related protein expression. Nineteen of 24 angiogenesis-related proteins were expressed equally, lower or higher in hiPSC-derived compared to adult platelets. The hiPSC-platelet’s coagulation hyporeactivity compared to adult platelets was confirmed by thromboelastometry. Further stepwise improvement of hiPSC-platelet production will, thus, permit better identification of platelet-mediated regenerative mechanisms and facilitate manufacture of sufficient amounts of functional platelets for clinical application.

scholarly journals Induced Pluripotent Stem Cells for Duchenne Muscular Dystrophy Modeling and Therapy

Cells ◽  
2018 ◽  
Vol 7 (12) ◽  
pp. 253 ◽  
Author(s):  
Lubos Danisovic ◽  
Martina Culenova ◽  
Maria Csobonyeiova
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Cell Reprogramming ◽  
Progressive Degeneration ◽  
Cardiac Muscles ◽  
Dmd Gene ◽  
Induced Pluripotent

Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder, caused by mutation of the DMD gene which encodes the protein dystrophin. This dystrophin defect leads to the progressive degeneration of skeletal and cardiac muscles. Currently, there is no effective therapy for this disorder. However, the technology of cell reprogramming, with subsequent controlled differentiation to skeletal muscle cells or cardiomyocytes, may provide a unique tool for the study, modeling, and treatment of Duchenne muscular dystrophy. In the present review, we describe current methods of induced pluripotent stem cell generation and discuss their implications for the study, modeling, and development of cell-based therapies for Duchenne muscular dystrophy.

scholarly journals Major Histocompatibility Complex–Matched Arteries Have Similar Patency to Autologous Arteries in a Mauritian Cynomolgus Macaque Major Histocompatibility Complex–Defined Transplant Model

2019 ◽  
Vol 8 (15) ◽  
Author(s):  
John P. Maufort ◽  
Jacqueline S. Israel ◽  
Matthew E. Brown ◽  
Steve J. Kempton ◽  
Nicholas J. Albano ◽  
...  
Keyword(s):  
Major Histocompatibility Complex ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Small Diameter ◽  
Induced Pluripotent Stem Cell ◽  
Major Histocompatibility ◽  
Arterial Grafts ◽  
Histocompatibility Complex ◽  
Induced Pluripotent

Background Arterial bypass and interposition grafts are used routinely across multiple surgical subspecialties. Current options include both autologous and synthetic materials; however, each graft presents specific limitations. Engineering artificial small‐diameter arteries with vascular cells derived from induced pluripotent stem cells could provide a useful therapeutic solution. Banking induced pluripotent stem cells from rare individuals who are homozygous for human leukocyte antigen alleles has been proposed as a strategy to facilitate economy of scale while reducing the potential for rejection of induced pluripotent stem cell–derived transplanted tissues. Currently, there is no standardized model to study transplantation of small‐diameter arteries in major histocompatibility complex–defined backgrounds. Methods and Results In this study, we developed a limb‐sparing nonhuman primate model to study arterial allotransplantation in the absence of immunosuppression. Our model was used to compare degrees of major histocompatibility complex matching between arterial grafts and recipient animals with long‐term maintenance of patency and function. Unexpectedly, we (1) found that major histocompatibility complex partial haplomatched allografts perform as well as autologous control grafts; (2) detected little long‐term immune response in even completely major histocompatibility complex mismatched allografts; and (3) observed that arterial grafts become almost completely replaced over time with recipient cells. Conclusions Given these findings, induced pluripotent stem cell–derived tissue‐engineered blood vessels may prove to be promising and customizable grafts for future use by cardiac, vascular, and plastic surgeons.

scholarly journals Present and future challenges of induced pluripotent stem cells

2015 ◽  
Vol 370 (1680) ◽  
pp. 20140367 ◽  
Author(s):  
Mari Ohnuki ◽  
Kazutoshi Takahashi
Keyword(s):  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Cell Types ◽  
Disease Modelling ◽  
Future Perspectives ◽  
Differentiated Cells ◽  
Human Ipscs ◽  
Future Challenges ◽  
Induced Pluripotent

Growing old is our destiny. However, the mature differentiated cells making up our body can be rejuvenated to an embryo-like fate called pluripotency which is an ability to differentiate into all cell types by enforced expression of defined transcription factors. The discovery of this induced pluripotent stem cell (iPSC) technology has opened up unprecedented opportunities in regenerative medicine, disease modelling and drug discovery. In this review, we introduce the applications and future perspectives of human iPSCs and we also show how iPSC technology has evolved along the way.

scholarly journals In Situ Expansion, Differentiation, and Electromechanical Coupling of Human Cardiac Muscle in a 3D Bioprinted, Chambered Organoid

2020 ◽  
Vol 127 (2) ◽  
pp. 207-224 ◽  
Author(s):  
Molly E. Kupfer ◽  
Wei-Han Lin ◽  
Vasanth Ravikumar ◽  
Kaiyan Qiu ◽  
Lu Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Induced Pluripotent Stem Cells ◽  
Cardiac Muscle ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Pump Function ◽  
Induced Pluripotent

Rationale: One goal of cardiac tissue engineering is the generation of a living, human pump in vitro that could replace animal models and eventually serve as an in vivo therapeutic. Models that replicate the geometrically complex structure of the heart, harboring chambers and large vessels with soft biomaterials, can be achieved using 3-dimensional bioprinting. Yet, inclusion of contiguous, living muscle to support pump function has not been achieved. This is largely due to the challenge of attaining high densities of cardiomyocytes—a notoriously nonproliferative cell type. An alternative strategy is to print with human induced pluripotent stem cells, which can proliferate to high densities and fill tissue spaces, and subsequently differentiate them into cardiomyocytes in situ. Objective: To develop a bioink capable of promoting human induced pluripotent stem cell proliferation and cardiomyocyte differentiation to 3-dimensionally print electromechanically functional, chambered organoids composed of contiguous cardiac muscle. Methods and Results: We optimized a photo-crosslinkable formulation of native ECM (extracellular matrix) proteins and used this bioink to 3-dimensionally print human induced pluripotent stem cell–laden structures with 2 chambers and a vessel inlet and outlet. After human induced pluripotent stem cells proliferated to a sufficient density, we differentiated the cells within the structure and demonstrated function of the resultant human chambered muscle pump. Human chambered muscle pumps demonstrated macroscale beating and continuous action potential propagation with responsiveness to drugs and pacing. The connected chambers allowed for perfusion and enabled replication of pressure/volume relationships fundamental to the study of heart function and remodeling with health and disease. Conclusions: This advance represents a critical step toward generating macroscale tissues, akin to aggregate-based organoids, but with the critical advantage of harboring geometric structures essential to the pump function of cardiac muscle. Looking forward, human chambered organoids of this type might also serve as a test bed for cardiac medical devices and eventually lead to therapeutic tissue grafting.

scholarly journals Young at Heart: Pioneering Approaches to Model Nonischaemic Cardiomyopathy with Induced Pluripotent Stem Cells

2016 ◽  
Vol 2016 ◽  
pp. 1-15 ◽  
Author(s):  
Aoife Gowran ◽  
Marco Rasponi ◽  
Roberta Visone ◽  
Patrizia Nigro ◽  
Gianluca L. Perrucci ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Induced Pluripotent Stem Cells ◽  
Drug Screening ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Human Fibroblasts ◽  
Induced Pluripotent Stem Cell ◽  
Induced Pluripotent

A mere 9 years have passed since the revolutionary report describing the derivation of induced pluripotent stem cells from human fibroblasts and the first in-patient translational use of cells obtained from these stem cells has already been achieved. From the perspectives of clinicians and researchers alike, the promise of induced pluripotent stem cells is alluring if somewhat beguiling. It is now evident that this technology is nascent and many areas for refinement have been identified and need to be considered before induced pluripotent stem cells can be routinely used to stratify, treat and cure patients, and to faithfully model diseases for drug screening purposes. This review specifically addresses the pioneering approaches to improve induced pluripotent stem cell based models of nonischaemic cardiomyopathy.

scholarly journals A novel GNAS-mutated human induced pluripotent stem cell model for understanding GNAS-mutated tumors

Tumor Biology ◽  
2020 ◽  
Vol 42 (9) ◽  
pp. 101042832096258
Author(s):  
Katsuhito Watanabe ◽  
Takashi Nakamura ◽  
Shoko Onodera ◽  
Akiko Saito ◽  
Takahiko Shibahara ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Epithelial Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Cell Model ◽  
Induced Pluripotent Stem Cell ◽  
Immunohistochemical Analysis ◽  
Induced Pluripotent

A missense mutation of the guanine nucleotide binding protein alpha stimulating activity polypeptide 1 ( GNAS) gene, typically Arg201Cys or Arg201His (R201H/R201C), leads to constitutive activation of the Gsα-cyclic AMP (cAMP) signaling pathway that causes several diseases. However, no germline mutations of GNAS have been identified to date, likely due to their lethality, and no robust human cell models have been generated. Therefore, the aim of this study was to generate GNAS-mutated disease-specific induced pluripotent stem cells as a model for these diseases. We then analyzed the functionality of this induced pluripotent stem cell model and differentiated epithelial cells. We generated disease-specific induced pluripotent stem cells by introducing a mutation in GNAS with the clustered regularly interspaced short palindromic repeats (CRISPR) nickase method, which has lower off-target effects than the conventional CRISPR/Cas9 method. We designed the target vector to contain the R201H mutation in GNAS, which was transfected into human control induced pluripotent stem cells (Nips-B2) by electroporation. We confirmed the establishment of GNASR201H -mutated ( GNASR201H/+) induced pluripotent stem cells that exhibited a pluripotent stem cell phenotype. We analyzed the effect of the mutation on cAMP production, and further generated teratomas for immunohistochemical analysis of the luminal epithelial structure. GNAS-mutated induced pluripotent stem cells showed significantly higher levels of intracellular cAMP, which remained elevated state for a long time upon hormonal stimulation with parathyroid hormone or adrenocorticotropic hormone. Immunohistochemical analysis revealed that several mucins, including MUC1, 2, and MUC5AC, are expressed in cytokeratin 18 (CK18)-positive epithelial cells. However, we found few CK18-positive cells in mutated induced pluripotent stem cell–derived teratoma tissues, and reduced MUCINs expression in mutated epithelial cells. There was no difference in CDX2 expression; however, mutated epithelial cells were positive for CEA and CA19-9 expression. GNASR201H-mutated induced pluripotent stem cells and GNASR201H-mutated epithelial cells have distinct phenotypic and differentiation characteristics. We successfully established GNASR201H-mutated human induced pluripotent stem cells with increased cAMP production. Considering the differentiation potential of induced pluripotent stem cells, these cells will be useful as a model for elucidating the pathological mechanisms of GNAS-mutated diseases.

scholarly journals Robust differentiation of human pluripotent stem cells into endothelial cells via temporal modulation of ETV2 with modified mRNA

2020 ◽  
Vol 6 (30) ◽  
pp. eaba7606 ◽  
Author(s):  
Kai Wang ◽  
Ruei-Zeng Lin ◽  
Xuechong Hong ◽  
Alex H. Ng ◽  
Chin Nien Lee ◽  
...  
Keyword(s):  
Stem Cells ◽  
Endothelial Cells ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
High Efficiency ◽  
Induced Pluripotent Stem Cell ◽  
Vascular Networks ◽  
Temporal Modulation ◽  
Induced Pluripotent

Human induced pluripotent stem cell (h-iPSC)–derived endothelial cells (h-iECs) have become a valuable tool in regenerative medicine. However, current differentiation protocols remain inefficient and lack reliability. Here, we describe a method for rapid, consistent, and highly efficient generation of h-iECs. The protocol entails the delivery of modified mRNA encoding the transcription factor ETV2 at the intermediate mesodermal stage of differentiation. This approach reproducibly differentiated 13 diverse h-iPSC lines into h-iECs with exceedingly high efficiency. In contrast, standard differentiation methods that relied on endogenous ETV2 were inefficient and notably inconsistent. Our h-iECs were functionally competent in many respects, including the ability to form perfused vascular networks in vivo. Timely activation of ETV2 was critical, and bypassing the mesodermal stage produced putative h-iECs with reduced expansion potential and inability to form functional vessels. Our protocol has broad applications and could reliably provide an unlimited number of h-iECs for vascular therapies.

Functional coculture of sympathetic neurons and cardiomyocytes derived from human-induced pluripotent stem cells

2020 ◽  
Vol 319 (5) ◽  
pp. H927-H937
Author(s):  
Annika Winbo ◽  
Suganeya Ramanan ◽  
Emily Eugster ◽  
Stefan Jovinge ◽  
Jonathan R. Skinner ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Pluripotent Stem Cell ◽  
Sympathetic Neurons ◽  
Induced Pluripotent Stem Cell ◽  
Induced Pluripotent

We present data on a functional coculture between human-induced pluripotent stem cell-derived sympathetic neurons and cardiomyocytes. Moreover, this study adds significantly to the available data on the electrophysiological function of human-induced pluripotent stem cell-derived sympathetic neurons.

Sign in / Sign up

Export Citation Format

Share Document