期刊
CELLULAR AND MOLECULAR BIOENGINEERING
卷 10, 期 1, 页码 54-62出版社
SPRINGER
DOI: 10.1007/s12195-016-0462-7
关键词
Confocal microscopy; Image analysis; Phenylephrine; Aging; Myofilament volume
资金
- NIH [R00 HL115123]
- Brown University Eccleston Fellowship
- Brown University Karen T. Romer Undergraduate Teaching and Research Award
Advances in the use of human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes for heart regeneration and in vitro disease models demand a greater understanding of how these cells grow and mature in 3-dimensional space. In this study, we developed an analysis methodology of single cardiomyocytes plated on 2D surfaces to assess their 3D myofilament volume and its z-height distribution, or shape, upon hypertrophic stimulation via phenylephrine (PE) treatment or long-term culture (aging). Cardiomyocytes were fixed and labeled with alpha-actinin for confocal microscopy imaging to obtain z-stacks for 3D myofilament volume analysis. In primary neonatal rat ventricular myocytes (NRVMs), area increased 72% with PE, while volume increased 31%. In hiPSC-cardiomyocytes, area increased 70% with PE and 4-fold with aging; however, volume increased significantly only with aging by 2.3-fold. Analysis of z-height myofilament volume distribution in hiPSC-cardiomyocytes revealed a shift from a fairly uniform distribution in control cells to a basally located volume in a more flat and spread morphology with PE and even more so with aging, a shape that was akin to all NRVMs analyzed. These results suggest that 2D area is not a sufficient measure of hiPSC-cardiomyocyte growth and maturation, and that changes in 3D volume and its distribution are essential for understanding hiPSC-cardiomyocyte biology for disease modeling and regenerative medicine applications.
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