Date: Monday, June 3, 2019
Session Time: 4:30pm-6:00pm
Presentation Time: 4:54pm-5:06pm
Location: Room 206
*Purpose: Exosomes are membrane vesicles released by cells into body fluids. Our laboratory demonstrated the presence of circulating exosomes with lung self-antigens (SAgs), Collagen V and K-α Tubulin, and donor HLA in lung transplant recipients (LTxR) undergoing rejection. Since respiratory viral infections (RVI) is a known risk factor for development of chronic rejection following LTx, the goal of this study is to determine whether RVI leads to induction of circulating exosomes with SAgs and to demonstrate the presence of viral DNA/RNA in exosomes isolated from LTxR with RVI.
*Methods: Exosomes were isolated using ultracentrifugation and purity was confirmed by sucrose cushion gradient. DNA and RNA were isolated using kits and quantified on the Nanodrop. Libraries were generated using Kapa Biosystem’s library kit. The raw Illumina 2x150bp pair-end reads were checked on FastQC and were aligned to the human and viral genome build from CHIPseeker Database. Validation was done using Abs and primers to respiratory syncytial virus, coronavirus, and rhinovirus. To determine the role of exosomes in inducing stress and DNA damage, we incubated airway epithelial cell line, KCC266, with exosomes from LTxR with RVI or stable.
*Results: Viral nucleic acid sequences were demonstrable in exosomes from LTxR with RVI. Comparing the sequences with human genome, we identified the presence of DNA sequences specific to defensins and GTPase pathways, 195 unique in LTxR with RVI specific to apoptotic cleavage, NMDA receptor activation and stable had 91 sequences specific to MAP kinase and cell death signaling. We also identified H. influenzae sequences in exosomes isolated from LTxR with influenza. Further, we demonstrated increases in proteins associated with endoplasmic reticulum stress, i.e, PERK, ATF4 and BiP in cells incubated with exosomes isolated from LTxR with RVI. However, significant increase in the levels of STING and IRF3 at the protein levels were not detected.
*Conclusions: Based on these preliminary results we conclude that RVI LTxR leads to induction of circulating exosomes having unique nucleic acid sequences of viral etiology suggesting that these nucleic acids of viral origin may have functional consequences including upregulation of stress markers.
To cite this abstract in AMA style:Bansal S, Bremner R, Smith M, Mohanakumar T. Circulating Exosomes Isolated from Human Lung Transplant Recipients with Respiratory Viral Infections Contain Nucleic Acids Specific to Virus and Can Lead to Activation of Stress Proteins [abstract]. Am J Transplant. 2019; 19 (suppl 3). https://atcmeetingabstracts.com/abstract/circulating-exosomes-isolated-from-human-lung-transplant-recipients-with-respiratory-viral-infections-contain-nucleic-acids-specific-to-virus-and-can-lead-to-activation-of-stress-proteins/. Accessed March 8, 2021.
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