Extracellular vesicles produced by bronchial epithelial cells in response to oxidative stress contain micro-RNAs associated with senescence

Background:

Cellular senescence is one of the hallmarks of COPD. The miR-34a and mir-570 are involved in the induction of cellular senescence by inhibiting anti-ageing sirtuins. Extracellular vesicles (EVs) play a key role in intercellular communication. Senescent cells produce high amounts of EVs and contain miRNA that can regulate cellular senescence. The role of EVs in spreading cell senescence remains unclear.

Aim:

To characterize extracellular vesicles released by epithelial cells in response to oxidative stress and to assess their effect on epithelial cells senescence.

Methods:

BEAS-2B bronchial epithelial cells were stimulated with H2O2 for 24 or 48h. Microvesicles and exosomes were isolated by differential ultracentrifugation of cell supernatant and characterised by Western blot and labelling membrane with the dye PKH67. The RNA content of EVs was determined by fluorescence and by qRT-PCR.

Results:

After 48h treatment, EVs are released by cells stimulated with medium or H2O2. The fluorescence intensity of pkh67 increased in response to H2O2 (n=2, from 2.6 fold for microvesicles, and 1.4 fold for exosomes) suggesting that epithelial cells release more EVs in response to oxidative stress. In response to H2O2, miR-34a and miR-570 expression is augmented only in exosomes.

Conclusion:

These data suggest that in response to oxidative stress, epithelial cells produced exosomes enriched in miRNA involved in cellular senescence. This may contribute to COPD pathophysiology by spreading senescence in adjacent epithelial and other cells.