Biochemical, biophysical, and immunological characterization of respiratory secretions in severe SARS-CoV-2 infections
Michael J. Kratochvil, Gernot Kaber, Sally Demirdjian, Pamela C. Cai, Elizabeth B. Burgener, Nadine Nagy, Graham L. Barlow, Medeea Popescu, Mark R. Nicolls, Michael G. Ozawa, Donald P. Regula, Ana E. Pacheco-Navarro, Samuel Yang, Vinicio A. de Jesus Perez, Harry Karmouty-Quintana, Andrew M. Peters, Bihong Zhao, Maximilian L. Buja, Pamela Y. Johnson, Robert B. Vernon, Thomas N. Wight, Stanford COVID-19 Biobank Study Group, Carlos E. Milla, Angela J. Rogers, Andrew J. Spakowitz, Sarah C. Heilshorn, Paul L. Bollyky
Michael J. Kratochvil, Gernot Kaber, Sally Demirdjian, Pamela C. Cai, Elizabeth B. Burgener, Nadine Nagy, Graham L. Barlow, Medeea Popescu, Mark R. Nicolls, Michael G. Ozawa, Donald P. Regula, Ana E. Pacheco-Navarro, Samuel Yang, Vinicio A. de Jesus Perez, Harry Karmouty-Quintana, Andrew M. Peters, Bihong Zhao, Maximilian L. Buja, Pamela Y. Johnson, Robert B. Vernon, Thomas N. Wight, Stanford COVID-19 Biobank Study Group, Carlos E. Milla, Angela J. Rogers, Andrew J. Spakowitz, Sarah C. Heilshorn, Paul L. Bollyky
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Research Article COVID-19 Pulmonology

Biochemical, biophysical, and immunological characterization of respiratory secretions in severe SARS-CoV-2 infections

Abstract

Thick, viscous respiratory secretions are a major pathogenic feature of COVID-19, but the composition and physical properties of these secretions are poorly understood. We characterized the composition and rheological properties (i.e., resistance to flow) of respiratory secretions collected from intubated COVID-19 patients. We found the percentages of solids and protein content were greatly elevated in COVID-19 compared with heathy control samples and closely resembled levels seen in cystic fibrosis, a genetic disease known for thick, tenacious respiratory secretions. DNA and hyaluronan (HA) were major components of respiratory secretions in COVID-19 and were likewise abundant in cadaveric lung tissues from these patients. COVID-19 secretions exhibited heterogeneous rheological behaviors, with thicker samples showing increased sensitivity to DNase and hyaluronidase treatment. In histologic sections from these same patients, we observed increased accumulation of HA and the hyaladherin versican but reduced tumor necrosis factor–stimulated gene-6 staining, consistent with the inflammatory nature of these secretions. Finally, we observed diminished type I interferon and enhanced inflammatory cytokines in these secretions. Overall, our studies indicated that increases in HA and DNA in COVID-19 respiratory secretion samples correlated with enhanced inflammatory burden and suggested that DNA and HA may be viable therapeutic targets in COVID-19 infection.

Authors

Michael J. Kratochvil, Gernot Kaber, Sally Demirdjian, Pamela C. Cai, Elizabeth B. Burgener, Nadine Nagy, Graham L. Barlow, Medeea Popescu, Mark R. Nicolls, Michael G. Ozawa, Donald P. Regula, Ana E. Pacheco-Navarro, Samuel Yang, Vinicio A. de Jesus Perez, Harry Karmouty-Quintana, Andrew M. Peters, Bihong Zhao, Maximilian L. Buja, Pamela Y. Johnson, Robert B. Vernon, Thomas N. Wight, Stanford COVID-19 Biobank Study Group, Carlos E. Milla, Angela J. Rogers, Andrew J. Spakowitz, Sarah C. Heilshorn, Paul L. Bollyky

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Figure 2

COVID-19 human lung sections have high levels of HA.

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COVID-19 human lung sections have high levels of HA. (A) Quantification ...
(A) Quantification of HA in respiratory secretion samples. COVID-19 (n = 8), CF (n = 4), and healthy (n = 7) respiratory secretion samples. One-way ANOVA with Tukey’s multiple comparisons tests; **P < 0.01, ****P < 0.0001. (B) Representative chromatogram of HA molecular weight (MW). Solid traces are the averages of COVID-19, CF, and healthy respiratory secretion samples. The dotted trace is the chromatogram of standard loaded with HA of known MWs, as indicated on graph. The dashed trace is representative of a commercially available 100 kDa MW HA. (C) The bar graph represents the percentage low–molecular weight HA in respiratory secretion samples: healthy (n = 6) and COVID-19 (n = 8). We chose 250 kDa as the cutoff to define low–molecular weight HA. Unpaired t test with Welch’s correction; **P < 0.005. (D–I) Representative histologic cadaveric lung sections from donors with COVID-19, donors with CF, and healthy donors, both with (D–F) and without (G–I) HAdase treatment. Nuclei are stained in blue, and HA binding proteins (HABPs) are stained in brown. (J–L) Enlarged sections from G–I, respectively. Scale bars: 800 μm (D–I), 400 μm (J–L).