Noninvasive investigation of microRNAs (miRNAs) expression, which is deeply related to biological phenomena such as stem cell differentiation, in culture soup is particularly useful for monitoring of stem cell differentiation without phototoxicity of living cells, especially when cell morphologies remain unchanged during differentiation. However, real-time detection of miRNA in culture soup is not recommended because of insufficient miRNA amounts in culture soup. In this study, a convenient method is introduced for real-time assessing intracellular miRNA in culture soup by using lipovesicular miRNA beacon (Lipo-mB) and mechanical stimulus-mediated exocytosis. Pipetting-harvest of culture soup induces exocytosis-secretion of fluorescence signal of Lipo-mB from cytoplasm into culture soup. To demonstrate this method, Lipo-mB is applied for monitoring of adipogenesis by analyzing the expression levels of various intracellular miRNAs, which are related to adipogenesis regulators. The fluorescence intensity profile of the culture soup is correlated with the quantitative reverse-transcription-polymerase chain reaction data and absorbance of Oil Red O staining. These results demonstrate that Lipo-mB can successfully monitor stem cell differentiation by sensing changes in miRNA expression from culture soup of living cells. Lipo-mB can be further developed as an accurate sensing system for analyzing subtle differences in genotype, even when changes in phenotype cannot be observed.
Bibliographical noteFunding Information:
This research was supported by the Bio & Medical Technology Development Program of the National Research Foundation (NRF) funded by the Korean government (MEST) (NRF-2012M3A9C6050077, and NRF-2012M3A9C6050332), NRF-2015M3A7B6027963 and the NRF-2013 Global Ph.D. Fellowship Program (NRF-2013H1A2A1034907). This research was supported by Nano.Material Technology Development Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (Grant number 2017M3A7B4041798).
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All Science Journal Classification (ASJC) codes
- Biomedical Engineering
- Pharmaceutical Science