Monitoring the maturation of the sarcomere network: a super-resolution microscopy-based approach

The in vitro generation of human cardiomyocytes derived from induced pluripotent stem cells (iPSC) is of great importance for cardiac disease modeling, drug-testing applications and for regenerative medicine. Despite the development of various cultivation strategies, a sufficiently high degree of ma...

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Vydáno v:Cellular and molecular life sciences : CMLS Ročník 79; číslo 3; s. 149
Hlavní autoři: Skorska, Anna, Johann, Lisa, Chabanovska, Oleksandra, Vasudevan, Praveen, Kussauer, Sophie, Hillemanns, Maximilian, Wolfien, Markus, Jonitz-Heincke, Anika, Wolkenhauer, Olaf, Bader, Rainer, Lang, Hermann, David, Robert, Lemcke, Heiko
Médium: Journal Article
Jazyk:angličtina
Vydáno: Cham Springer International Publishing 01.03.2022
Springer Nature B.V
Témata:
Actinin
Actinin - metabolism
adults
Animals
Biochemistry
Biomedical and Life Sciences
Biomedicine
Calcium - metabolism
Calcium Signaling - physiology
Cardiomyocytes
Cell Biology
Cell culture
Cell differentiation
Cell Differentiation - physiology
Cell morphology
cell structures
Cells, Cultured
Coronary artery disease
Cultivation
Cytology
Filaments
Fluorescence
Heart diseases
Humans
Image acquisition
Image analysis
Image processing
Induced Pluripotent Stem Cells - cytology
Induced Pluripotent Stem Cells - metabolism
Learning algorithms
Life Sciences
Machine Learning
Maturation
medicine
Mice
Microscopy
Microscopy, Fluorescence - methods
Muscle contraction
Muscle, Skeletal - cytology
Muscles
Myocardium - cytology
Neonates
Original
Original Article
Phenotype
Phenotypes
Pluripotency
quantitative analysis
Regenerative medicine
RNA - genetics
RNA - isolation & purification
sarcomeres
Sarcomeres - metabolism
Skeletal muscle
Stem cells
Tissue engineering
Workflow
Super resolution microscopy
Sarcomere network
Maturation
iPSC cardiomyocytes
ISSN:1420-682X, 1420-9071, 1420-9071
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Popis
Shrnutí:The in vitro generation of human cardiomyocytes derived from induced pluripotent stem cells (iPSC) is of great importance for cardiac disease modeling, drug-testing applications and for regenerative medicine. Despite the development of various cultivation strategies, a sufficiently high degree of maturation is still a decisive limiting factor for the successful application of these cardiac cells. The maturation process includes, among others, the proper formation of sarcomere structures, mediating the contraction of cardiomyocytes. To precisely monitor the maturation of the contractile machinery, we have established an imaging-based strategy that allows quantitative evaluation of important parameters, defining the quality of the sarcomere network. iPSC-derived cardiomyocytes were subjected to different culture conditions to improve sarcomere formation, including prolonged cultivation time and micro patterned surfaces. Fluorescent images of α-actinin were acquired using super-resolution microscopy. Subsequently, we determined cell morphology, sarcomere density, filament alignment, z-Disc thickness and sarcomere length of iPSC-derived cardiomyocytes. Cells from adult and neonatal heart tissue served as control. Our image analysis revealed a profound effect on sarcomere content and filament orientation when iPSC-derived cardiomyocytes were cultured on structured, line-shaped surfaces. Similarly, prolonged cultivation time had a beneficial effect on the structural maturation, leading to a more adult-like phenotype. Automatic evaluation of the sarcomere filaments by machine learning validated our data. Moreover, we successfully transferred this approach to skeletal muscle cells, showing an improved sarcomere formation cells over different differentiation periods. Overall, our image-based workflow can be used as a straight-forward tool to quantitatively estimate the structural maturation of contractile cells. As such, it can support the establishment of novel differentiation protocols to enhance sarcomere formation and maturity.
Bibliografie:ObjectType-Article-1
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ISSN:1420-682X
1420-9071
1420-9071
DOI:10.1007/s00018-022-04196-3