scholarly journals Machine learning-assisted high-content analysis of pluripotent stem cell-derived embryos in vitro

2021 ◽  
Author(s):  
Jianying Guo ◽  
Peizhe Wang ◽  
Berna Sozen ◽  
Hui Qiu ◽  
Yonglin Zhu ◽  
...  
Keyword(s):  
Machine Learning ◽  
Content Analysis ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
High Content Analysis

Abstract 12279: Novel and Reliable Method to Screen for Drug Cardiac Toxicity Using Human Induced Pluripotent Stem Cell Derived Cardiomyocytes (hiPS-CM)

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Tamer M Mohamed ◽  
Bernhard Ellinger ◽  
Vera Stankovikj ◽  
Adriana Wiesinger ◽  
Kaomei Guan ◽  
...  
Keyword(s):  
Content Analysis ◽  
Stem Cell ◽  
Confocal Microscopy ◽  
Pluripotent Stem Cell ◽  
Transmembrane Potential ◽  
Cell Phenotype ◽  
Plasma Membrane Permeability ◽  
Prediction Rate ◽  
High Content Analysis ◽  
Induced Pluripotent

Unexpected cardiotoxicity underlies high rates of attrition during drug development, posing a multi-billion dollar burden on the pharmaceutical industry. Over reliance on the use of animals and materials derived from animals in preclinical assays to predict the cardiotoxic effect of new drugs in humans has contributed to this problem. The drug responses in human cardiomyocytes compared to animal-derived primary cardiomyocytes (CM) for in vitro assays is not always the same. Here, we describe a combination of human pluripotent stem cell-derived cardiomyocytes (hiPS-CM) and high content analysis confocal microscopy as a potential solution for this major setback in drug development. Initially human skin fibroblasts were reprogrammed into human induced pluripotent stem cells (hiPSCs) by lentiviral transduction and utilization of the following combination of transcription factors: Oct3/4, Sox2, c-Myc and Klf4. The human pluripotent stem cell phenotype of the generated hIPSCs was confirmed. hiPSCs were differentiated into cardiomyocytes and the cardiac cell phenotype was confirmed by immunofluorescence and RT-PCR analysis of cardiac markers i.e. αMHC, cTNT, NKX2.5 and connexin 43. Seventeen known cardiotoxic compounds, as well as controls, were applied to the cells in 384 well format at a dose of 10μM for 48 hours. Then hiPSC-CM underwent confocal microscopy high content analysis to simultaneously evaluate the cell mitochondrial transmembrane potential (using TMRM dye), and plasma membrane permeability (using TOTO-3 dye). All known cardiotoxins showed a significant decrease in mitochondrial transmembrane potential ranging from 74% to 95% and an increase in plasma membrane permeability ranging from 67-327 fold in comparison to the controls. These results showed 100% prediction rate of cardiotoxicity of known cardiotoxins by hiPS-CM. This was compared to only 12% general cytotoxicity prediction rate when these compounds tested on A549 and ACHN cancer cell lines. In conclusion, combining two state of the art technologies 1) hiPSC-CM and 2) confocal microscopy high content analysis, we were able to provide a reliable high throughput method to assess cardiotoxicity of compounds.

Use of 3D culture systems to generate human induced pluripotent stem cell-derived [beta]-cells in vitro

2018 ◽  
Author(s):  
Fantuzzi Federica ◽  
Toivonen Sanna ◽  
Schiavo Andrea Alex ◽  
Pachera Nathalie ◽  
Rajaei Bahareh ◽  
...  
Keyword(s):  
Stem Cell ◽  
Beta Cells ◽  
Pluripotent Stem Cell ◽  
3D Culture ◽  
Induced Pluripotent Stem Cell ◽  
Culture Systems ◽  
Induced Pluripotent

In vitro pharmacologic testing using human induced pluripotent stem cell-derived cardiomyocytes

2009 ◽  
Vol 385 (4) ◽  
pp. 497-502 ◽  
Author(s):  
Tomofumi Tanaka ◽  
Shugo Tohyama ◽  
Mitsushige Murata ◽  
Fumimasa Nomura ◽  
Tomoyuki Kaneko ◽  
...  
Keyword(s):  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Induced Pluripotent

scholarly journals Persistence of intramyocardially transplanted murine induced pluripotent stem cell-derived cardiomyocytes from different developmental stages

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Gabriel Peinkofer ◽  
Martina Maass ◽  
Kurt Pfannkuche ◽  
Agapios Sachinidis ◽  
Stephan Baldus ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Stem Cell ◽  
Cell Adhesion ◽  
Developmental Stage ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Induced Pluripotent

Abstract Background Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) are regarded as promising cell type for cardiac cell replacement therapy, but it is not known whether the developmental stage influences their persistence and functional integration in the host tissue, which are crucial for a long-term therapeutic benefit. To investigate this, we first tested the cell adhesion capability of murine iPSC-CM in vitro at three different time points during the differentiation process and then examined cell persistence and quality of electrical integration in the infarcted myocardium in vivo. Methods To test cell adhesion capabilities in vitro, iPSC-CM were seeded on fibronectin-coated cell culture dishes and decellularized ventricular extracellular matrix (ECM) scaffolds. After fixed periods of time, stably attached cells were quantified. For in vivo experiments, murine iPSC-CM expressing enhanced green fluorescent protein was injected into infarcted hearts of adult mice. After 6–7 days, viable ventricular tissue slices were prepared to enable action potential (AP) recordings in transplanted iPSC-CM and surrounding host cardiomyocytes. Afterwards, slices were lysed, and genomic DNA was prepared, which was then used for quantitative real-time PCR to evaluate grafted iPSC-CM count. Results The in vitro results indicated differences in cell adhesion capabilities between day 14, day 16, and day 18 iPSC-CM with day 14 iPSC-CM showing the largest number of attached cells on ECM scaffolds. After intramyocardial injection, day 14 iPSC-CM showed a significant higher cell count compared to day 16 iPSC-CM. AP measurements revealed no significant difference in the quality of electrical integration and only minor differences in AP properties between d14 and d16 iPSC-CM. Conclusion The results of the present study demonstrate that the developmental stage at the time of transplantation is crucial for the persistence of transplanted iPSC-CM. iPSC-CM at day 14 of differentiation showed the highest persistence after transplantation in vivo, which may be explained by a higher capability to adhere to the extracellular matrix.

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