In Cell Metabolism, researchers have described how a microRNA (miRNA) derived from exosomes generated by human embryonic stem cells (hESCs) restores function and fights senescence in cell cultures and mice.
Looking for a better senomorphic
This study begins with a discussion of cellular senescence and its role in aging, focusing on the main approaches to dealing with it: senolytics, which kill senescent cells, and senomorphics, which transform them. Senolytics may have side effects because their number goes up with advanced age [1], and some of them are necessary for life. Even such techniques as directly affecting the SASP may have adverse impacts on the immune system [2].
While senomorphics appear to be a viable strategy, they are still in their infancy. One potential strategy involves exosomes: messenger particles that are regularly sent by cells. Previous work with hESC-derived exosomes (hESC-Exos) has found that their contents are instrumental in rejuvenating multiple tissues [3]. Therefore, these researchers decided to examine them as a senomorphic, discovering which of their many components is best at restoring senescent cells.
Cells regained the power to proliferate
The first experiment was a basic test of hESC-Exos on IMR-90 cells, a line of human fibroblasts. After 30 divisions, this cell line remains youthful, but at 50, it has reached replicative senescence. However, treating those senescent cells with hESC-Exos almost entirely reversed their senescent biomarkers, downregulating SASP-related genes such as those for inflammatory interleukins, restoring cellular proliferation, and sharply decreasing the senescence biomarker SA-β-gal. Other genes related to senescence were inhibited, while those relating to proliferation were enhanced.
These findings were recapitulated at the single-cell level. First, the researchers created a population of human cells that were modified to express the fluorescence protein along with the senescence marker p21. Then, they drove these cells senescent by exposing them to doxycycline, after which they exposed some of them to hESC-Exos.
Compared to an untreated control group, the treated cells had substantially less p21-related fluorescence and less SA-β-gal, and some of them regained the ability to proliferate. Just like with the replicatively senescent cells, hESC-Exos diminished senescence-related gene expression and enhanced proliferation-related expression instead.
Mice regained youthful attributes
From the 20th month to the 30th month of life, wild-type mice on a normal diet were injected with hESC-Exos. Compared to a control group, the exosome-treated mice performed better on both fixed and accelerating rotarod tests, had higher body weight, and retained normal bladder activity. In the Morris water maze test, they both found the platform more quickly and remembered its location more accurately. They retained their hair color and skin smoothness as well.
Biomarkers of senescence, just as in the cellular culture, were substantially reduced in the treated mice. Inflammatory factors, such as cytokines and TNF-α, were also substantially reduced in their circulation, and γ-H2AX, a marker of genomic damage, was notably reduced. Just as in cellular cultures, the mice’s biomarkers of cellular proliferation were improved. Overall, the researchers found the treated mice to be substantially rejuvenated as a whole.
Finding the key molecule
The researchers hypothesized that much of this rejuvenative power can be distilled down to the exosomes’ individual components. They selected one of them, miR-302b, which is abundantly found in hESC-Exos and is documented to play a role in cellular proliferation [4]. However, it had remained untested against aging.
This particular miRNA was found to directly regulate expression of the Cdkn1a gene, which is related to cellular senescence. Exposing IMR-90 cells to miR-302b recapitulated the effects of hESC-Exos, reducing senescence and promoting proliferation.
Encouraged, the researchers then turned to mice. This time, they injected 25- to 30-month-old mice with artificially transfected exosomes containing miR-302b. They found that this approach recapitulated the results found in hESC-Exos as well, reducing inflammatory factors and SA-β-gal, substantially improving the results of rotarod and Morris water maze tests, and restoring cellular proliferation.
Llifespan itself was also improved by this approach. The median lifespan of the mice treated with miR-302b was 137 days greater than that of the control group. While the difference was not significant, the effect seemed to be stronger in males.
While hESC-Exos and miR-302b were not compared directly, they appear to be largely similar in terms of their effects. Still, this is a cell and mouse study, and further work needs to be done to determine if this approach is safe for clinical use. It is also not known which of these approaches will be the most scalable and suitable for mass production.
Lifterature
[1] Huang, W., Hickson, L. J., Eirin, A., Kirkland, J. L., & Lerman, L. O. (2022). Cellular senescence: the good, the bad and the unknown. Nature Reviews Nephrology, 18(10), 611-627.
[2] Zhang, L., Pitcher, L. E., Yousefzadeh, M. J., Niedernhofer, L. J., Robbins, P. D., & Zhu, Y. (2022). Cellular senescence: a key therapeutic target in aging and diseases. The Journal of Clinical Investigation, 132(15).
[3] Bi, Y., Qiao, X., Liu, Q., Song, S., Zhu, K., Qiu, X., … & Ji, G. (2022). Systemic proteomics and miRNA profile analysis of exosomes derived from human pluripotent stem cells. Stem Cell Research & Therapy, 13(1), 449.
[4] Subramanyam, D., Lamouille, S., Judson, R. L., Liu, J. Y., Bucay, N., Derynck, R., & Blelloch, R. (2011). Multiple targets of miR-302 and miR-372 promote reprogramming of human fibroblasts to induced pluripotent stem cells. Nature biotechnology, 29(5), 443-448.
