key: cord-312558-w2vjuidm authors: Gambardella, J.; Sardu, C.; Morelli, M. B.; Messina, V.; Castellanos, V.; Marfella, R.; Maggi, P.; Paolisso, G.; Wang, X.; Santulli, G. title: Exosomal microRNAs Drive Thrombosis in COVID-19 date: 2020-06-18 journal: nan DOI: 10.1101/2020.06.16.20133256 sha: doc_id: 312558 cord_uid: w2vjuidm Thrombotic and thromboembolic complications have been shown to play a critical role in the clinical outcome of COVID-19. Emerging evidence has shown that exosomal miRNAs are functionally involved in a number of physiologic and pathologic processes. However, neither exosomes nor miRNAs have been hitherto investigated in COVID-19. To test the hypothesis that exosomal miRNAs are a key determinant of thrombosis in COVID-19, we enrolled patients positive for COVID-19. Circulating exosomes were isolated from equal amounts of serum and levels of exosomal miRNAs were quantified. We divided our population in two groups based on the serum level of D-dimer on admission. Strikingly, we found that exosomal miR-424 was significantly upregulated whereas exosomal miR-103a, miR-145, and miR-885 were significantly downregulated in patients in the high D-dimer group compared to patients in the low D-Dimer group (p<0.0001). . CC-BY-NC 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 18, 2020. MicroRNAs (miRNAs) are small non-coding RNAs that enhance mRNA degradation and/or inhibit protein translation (5, 6) . Emerging evidence has shown that exosomal miRNAs are functionally involved in a number of physiologic and pathologic processes (5, 7, 8) . However, neither exosomes nor miRNAs have been hitherto investigated in COVID-19. To test the hypothesis that exosomal miRNAs are a key determinant of thrombosis in COVID-19, we enrolled 26 patients positive for COVID-19 admitted to the Infectious Disease Departments of University of Naples "Vanvitelli" and San Sebastiano Caserta Hospital (Italy). The study was conducted according to the Declaration of Helsinki principles and approved by the local Ethical Committee. Written informed consent was obtained from all participants. Circulating exosomes were isolated from equal amounts of serum, as previously described and validated by our group (9,10) and levels of exosomal miRNAs were quantified by RT-qPCR, normalizing values to spiked cel-miR-39 (10). We divided our population in two groups based on the serum level of D-dimer on admission, using a previously published cut-off of 3 μg/ml (4). . CC-BY-NC 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 18, 2020. Strikingly, we found a that exosomal miR-424 was significantly upregulated whereas exosomal miR-103a, miR-145, and miR-885 were significantly downregulated (Figure 1, A-D) in patients in the high D-dimer group compared to patients in the low D-Dimer group (p<0.0001). Regression analysis confirmed these findings (Figure 1, E-H) . Mechanistically, Tissue Factor has been identified as a direct target of miR-145, while miR-885 has been shown to target the von Willebrand Factor (5, 11) . Equally important, miR-424 has been associated with hypercoagulability whereas low levels of miR-103a have been observed in deep vein thrombosis (5, 12) , although precise mechanisms have not been fully defined for these miRNAs. To our knowledge, this is the first study showing a functional contribution of exosomal noncoding RNA in COVID19. Limitations of our study include the relatively small population and the fact that we did not determine the exact source of exosomes; nevertheless, since endothelial dysfunction has been shown to be a prominent feature of COVID-19 and to contribute to the pro-thrombotic and pro-inflammatory state of the vasculature (2), we speculate that a main source could be represented by endothelial cells, which express these miRNAs in normal conditions (5). . CC-BY-NC 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 18, 2020. . https://doi.org/10.1101/2020.06.16.20133256 doi: medRxiv preprint Conflict of interest: none. . CC-BY-NC 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted June 18, 2020. . https://doi.org/10.1101/2020.06.16.20133256 doi: medRxiv preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted June 18, 2020. . https://doi.org/10.1101/2020.06.16.20133256 doi: medRxiv preprint Hypertension, Thrombosis, Kidney Failure, and Diabetes: Is COVID-19 an Endothelial Disease? A Comprehensive Evaluation of Clinical and Basic Evidence COVID-19 and Thrombotic or Thromboembolic Disease: Implications for Prevention, Antithrombotic Therapy, and Follow-Up: JACC State-of-the-Art Review Is Endothelial Dysfunction the Concealed Cornerstone of COVID-19 COVID-19-associated hyperviscosity: a link between inflammation and thrombophilia? 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