Identification of RNA content of CHO-derived extracellular vesicles from a production process.

Recent studies have unveiled the unique roles of extracellular vesicles (EVs) in various cellular processes including protein degradation, transport, and intercellular communication. However, the EVs of Chinese hamster ovary (CHO) cells, the workhorse of biologics manufacturing, have not been well-characterized despite their significant roles in protein production. Herein, we successfully isolated CHO EVs from CHO fed-batch cultures and identified their messenger RNA (mRNA) and micro RNA (miRNA) contents through next-generation sequencing. We found that mRNAs corresponding to oxidative phosphorylation were highly enriched in microvesicles (large EVs) but absent in exosomes (small EVs). We also found that both large EVs and small EVs had enriched mRNA species corresponding to key signaling pathways for cell proliferation, survival, and growth, including the TGFβ and PI3K/Akt pathways. In addition, the enrichment of miR-196a-5p in both small EVs and large EVs suggests an anti-apoptotic and proliferative function for EVs through intercellular communication. The identification of these mRNAs and miRNAs associated with cell growth and survival sheds light on the potential role of extracellular vesicles in the context of biologics manufacturing and may help further optimize CHO biologics production.