A ring of five genes for reprogramming cells
Stem cells easily turn into pluripotent without the use of viruses
Dmitry Safin, CompulentaResearchers from the Stanford University School of Medicine (USA) have developed a sufficiently effective technology for cell transformation, in which mini-rings of DNA are used to transfer genetic material.
Since the proposed method is simple and relatively safe, the authors hope for its early official review and approval by the American Food and Drug Administration. "It will not be difficult for laboratories to switch to our technology," says study participant Michael Longaker. "And then we will be one step closer to creating a system of practical regenerative medicine."
In their experiments, scientists transformed human fat stem cells into induced pluripotent stem cells (iPSCs). The effectiveness of DNA mini-rings is due to the fact that they contain only four genes necessary for reprogramming cells and one gene of a green fluorescent protein, which allows evaluating the results of the experiment. Larger and more commonly used DNA fragments – plasmids – also contain bacterial DNA, which is fraught with conflicts with the body's natural defense system and reduces the likelihood of penetration into cells. In addition, the genes in the mini-rings show a higher level of expression.
When using mini-rings, the expression of green fluorescent protein was observed in 10.8% of all stem cells, and the effectiveness of plasmids reached only 2.7%. The researchers separated these ten percent of the cells and treated them with mini–rings twice more – on the fourth and sixth days of the experiment. After 14-16 days, the first clusters of cells resembling colonies of embryonic stem cells appeared, and some of them no longer expressed the fluorescent protein.
Such "non-fluorescent" clusters, according to scientists, demonstrated all the properties of iPSCs: they expressed the genes of embryonic stem cells, had the appropriate parameters of DNA methylation and could form teratomas (tumors consisting of tissues that are absent in the organs and areas of a healthy organism corresponding to their place of formation) when injected under the skin of mice. The authors also confirmed the safety of the technique, making sure that the mini-rings were not integrated into the DNA of stem cells.
The researchers managed to create a total of 22 new IPSC lines from fat stem cells and fibroblasts. The resulting efficiency of the technology was about 0.005%, which, of course, is lower than that of "viral" methods offering 0.01-0.05%, but higher than that of traditional plasmid methods.
It is worth noting that the popularity of potentially dangerous from a genetic point of view techniques using viruses is gradually falling.
The full version of the report (Fangjun Jia et al., A nonviral minicircle vector for deriving human iPS cells) will be published in the journal Nature Methods.
Prepared based on the materials of the Stanford University Medical School.
Portal "Eternal youth" http://vechnayamolodost.ru08.02.2010