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Culture and adenoviral infection of adult mouse cardiac myocytes: methods for cellular genetic physiology
Abstract
Rapid development of transgenic and gene-targeted mice and acute genetic manipulation via gene transfer vector systems have provided powerful tools for cardiovascular research. To facilitate the phenotyping of genetically engineered murine models at the cellular and subcellular levels and to implement acute gene transfer techniques in single mouse cardiomyocytes, we have modified and improved current enzymatic methods to isolate a high yield of high-quality adult mouse myocytes (5.3 ± 0.5 × 10 5 cells/left ventricle, 83.8 ± 2.5% rod shaped). We have also developed a technique to culture these isolated myocytes while maintaining their morphological integrity for 2–3 days. The high percentage of viable myocytes after 1 day in culture (72.5 ± 2.3%) permitted both physiological and biochemical characterization. The major functional aspects of these cells, including excitation-contraction coupling and receptor-mediated signaling, remained intact, but the contraction kinetics were significantly slowed. Furthermore, gene delivery via recombinant adenoviral infection was highly efficient and reproducible. In adult β 1/β 2-adrenergic receptor (AR) double-knockout mouse myocytes, adenovirus-directed expression of either β 1- or β 2-AR, which occurred in 100% of cells, rescued the functional response to β-AR agonist stimulation. These techniques will permit novel experimental settings for cellular genetic physiology.
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Cited by
Cardiac progenitor cells from adult myocardium: Homing, differentiation, and fusion after infarction
Oh, H., Bradfute, S. B., Gallardo, T. D., Nakamura, T., Gaussin, V., Mishina, Y., Pocius, J., Michael, L. H., Behringer, R. R., Garry, D. J., et al. (2003) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.2132126100 · Cited by 1,743
The role of autophagy in cardiomyocytes in the basal state and in response to hemodynamic stress
Nakai, A., Yamaguchi, O., Takeda, T., Higuchi, Y., Hikoso, S., Taniike, M., Omiya, S., Mizote, I., Matsumura, Y., Asahi, M., et al. (2007) · Nature Medicine · doi:10.1038/nm1574 · Cited by 1,520
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Ackers-Johnson, M., Li, P. Y., Holmes, A. P., O’Brien, S.-M., Pavlovic, D., & Foo, R. S. (2016) · Circulation Research · doi:10.1161/circresaha.116.309202 · Cited by 543
The long noncoding RNA Chaer defines an epigenetic checkpoint in cardiac hypertrophy
Wang, Z., Zhang, X.-J., Ji, Y.-X., Zhang, P., Deng, K.-Q., Gong, J., Ren, S., Wang, X., Chen, I., Wang, H., et al. (2016) · Nature Medicine · doi:10.1038/nm.4179 · Cited by 386
Metadata
- Authors
- Zhou, Y.-Y., Wang, S.-Q., Zhu, W.-Z., Chruscinski, A., Kobilka, B. K., Ziman, B., Wang, S., Lakatta, E. G., Cheng, H., & Xiao, R.-P.
- Venue
- American Journal of Physiology-Heart and Circulatory Physiology
- Publisher
- American Physiological Society
- Published
- 2000-07-01
- Volume
- 279
- Issue
- 1
- Pages
- H429-H436
- ISSN
- 0363-6135 (ISSN-L)
- 1522-1539
- OpenAlex
- W2114235073
- Cited by
- 280
- References
- 0
- FWCI
- 3.90
- Type
- journal article
- Language
- en
- Open access
- closed
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- Zhou, Y.-Y., Wang, S.-Q., Zhu, W.-Z., Chruscinski, A., Kobilka, B. K., Ziman, B., Wang, S., Lakatta, E. G., Cheng, H., & Xiao, R.-P. (2000). Culture and adenoviral infection of adult mouse cardiac myocytes: methods for cellular genetic physiology. American Journal of Physiology-Heart and Circulatory Physiology. https://doi.org/10.1152/ajpheart.2000.279.1.h429
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