March 16, 2017
Living cells change their signature with time
miRNA biotechnology developed at CiRA marks a cell's differentiation
by changes in fluorescence intensity
that traces intracellular signatures during the differentiation of iPS cells.
The differentiation process of iPS cells to a given cell type involves an extraordinary number of molecular events. To trace these changes, scientists depend on molecular signatures. The Hirohide Saito group has reported new biotechnology that detects the miRNA dynamics of the cell as a means to follow iPS cell fate. Collaboration with the Yoshinori Yoshida lab at CiRA has demonstrated how this technology could advance disease modeling and drug discovery.
The technology depends on miRNA, which Dr. Hideyuki Nakanishi, who led the study, describes as "molecular signatures that we can use to follow cell differentiation. Our system is a simple way to quantitatively and continuously monitor changes in the cellular state."
The lab had previously engineered miRNA switches to detect miRNA levels, but this technology was not effective at detecting miRNA dynamics, which can be used to describe the differentiation state of a cell.
"miRNA switches are good for transient monitoring of miRNA," said Nakanishi, but the short half-life of the switch is not suitable for "the differentiation of iPS cells [which] can take several days or weeks."
To observe miRNA dynamics as an iPS cell differentiated, Nakanishi extended the lifetime of their miRNA switch by developing a vector system based on common molecular biology tools (piggyBac transposon, episomal plasmid, etc.) in which the switch was inserted.
By transfecting their new biotechnology into iPS cells, the differentiation of an iPS cell to various cell lineages could be traced over two weeks. In contrast, without this vector system, once inside a cell, "miRNA switches are designed to last several hours and degrade within 1 day," said Nakanishi.
He added that, "We can increase the miRNA sensitivity by reducing access energy or by increasing copy number."
Scientists at CiRA have already found the new vectors useful. "The purification of cardiomyocytes was very high," said Dr. Kenji Miki, who is researching cell therapies against heart disease in the Yoshida lab. "We could separate cardiomyoctyes and iPSCs into different populations easily."
The exceptionally high purification is allowing researchers like Miki to more precisely identify differences between healthy and diseased cells and observe the effects of experimental drugs.
- Journal: Biomaterials
- Title: Monitoring and visualizing microRNA dynamics during live cell differentiation using microRNA-responsive non-viral reporter vectors
- Authors: Hideyuki Nakanishi, Kenji Miki, Kaoru R. Komatsu, Masayuki Umeda, Megumi Mochizuki, Azusa Inagaki, Yoshinori Yoshida, and Hirohide Saito
- Author Affiliation: Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan