Effect of young exosomes injected in aged mice

Abstract

Introduction: Exosomes, nanosized extracellular vesicles, are known to circulate through the blood stream to transfer molecular signals from tissue to tissue.

Methods: To determine whether exosomes affect aging in animals, we primarily identified the changes in exosomal miRNA contents during the aging process. In exosomes from 12-month-old mice, mmu-miR-126-5p and mmu-miR-466c-5p levels were decreased and mmu-miR-184-3p and mmu-miR-200b-5p levels were increased significantly compared with those of 3-month-old mice. Their levels in exosomes were partially correlated with those in tissues: levels of only mmu-miR-126-5p and mmu-miR-466c-5p in lungs and/or liver were decreased, but those of mmu-miR-184-3p and mmu-miR-200b-5p in tissues did not coincide with those of exosomes.

Results and discussion: In the aged tissues injected with young exosomes isolated from serum, mmu-miR-126b-5p levels were reversed in the lungs and liver. Expression changes in aging-associated molecules in young exosome-injected mice were obvious: p16^Ink4A, MTOR, and IGF1R were significantly downregulated in the lungs and/or liver of old mice. In addition, telomerase-related genes such as Men1, Mre11a, Tep1, Terf2, Tert, and Tnks were significantly upregulated in the liver of old mice after injection of young exosomes.

Conclusion: These results indicate that exosomes from young mice could reverse the expression pattern of aging-associated molecules in aged mice. Eventually, exosomes may be used as a novel approach for the treatment and diagnosis of aging animals.

Keywords: biomarker; exosome; injection; molecular therapy; reverse aging; telomerase.

Figure 1

Characterization of the isolated exosomes by transmission electron microscopy (A), dynamic light scattering (B), and Western blotting for exosome-specific biomarkers (C).

Figure 2

Expression of miRNAs determined as differentially expressed miRNAs in exosomes, using droplet digital PCR. Notes: The relative expression of miRNAs at each age was determined in comparison with that of 5S rRNA. The statistical significance of expression changes at 8 and 12 months of 126b-5p, 184-3p, and 200b-5p were determined by comparison with those of 3 months, but the significance for miR-466c-5p was compared to that of 8 months. ^*P<0.05; ^**P<0.01.

Figure 3

Expression of differentially expressed miRNAs in tissues, determined using droplet digital PCR. Notes: (A) Expression in the lungs; (B) Expression in the liver. The statistical significance was determined by comparison with the expression at 3 months. ^*P<0.05.

Figure 4

Distribution of exosomes after injection of exogenous exosomes labeled with an exosome-specific dye (ExoGlow™-RNA, System Biosciences, Inc.). Notes: (A) Whole-body imaging for 3 days after exosome administration; (B) ex vivo imaging of exosome distribution in mouse tissues; (C) distribution of exosomes in the section of liver, lungs, and kidney observed using a confocal microscope.

Figure 5

Expression of differentially expressed miRNAs in lungs (A) and liver (B) after the injection of exosomes from 3-month-old to 18-month-old mice. Note: ^*P<0.05.

Figure 6

Expression of aging-associated biomarkers following the injection of exosomes from 3-month-old to 18-month-old mice. Notes: Following the analysis of protein expression by Western blotting (A), relative quantity of protein expression was determined by comparison with ACTB expression (B) and GAPDH expression (C). ^†P<0.1; ^*P<0.05; ^**P<0.01.

Figure 7

Expression of telomerase-related genes in lungs (A) and liver (B) after the injection of exosomes from 3-month-old to 18-month-old mice. Notes: ^†P<0.1; ^*P<0.05.