305 - The Hyaluronan Receptor RHAMM is Critical for Toll-Like Receptor 7- and SARS-CoV-2-Mediated Increases in Lung Cytokines and Inflammation
Saturday, April 23, 2022
3:30 PM – 6:00 PM US MT
Poster Number: 305 Publication Number: 305.215
Rashmin C. Savani, UT Southwestern Medical Center, Dallas, TX, United States; Jie Liao, UT southwestern medical center, Plano, TX, United States; Christopher L. Longoria, University of Texas Southwestern Medical School, Dallas, TX, United States; Naeun Cheong, University of Texas Southwestern Medical School, Dallas, TX, United States; Elias Coutavas, Duke University School of Medicine, Durham, NC, United States; So-Jin Kim, Duke University School of Medicine, Durham, NC, United States; Patty J. Lee, Duke University School of Medicine, Durham, NC, United States; Len Luyt, Western University, London, ON, Canada; Eva Turley, The University of Western Ontario - Schulich School of Medicine & Dentistry, London, ON, Canada; Stavros Garantziotis, NIH, Reserach Triangle Park, NC, United States
Professor of Pediatrics UT Southwestern Medical Center Dallas, Texas, United States
Background: SARS-CoV-2 causes COVID-19, a viral pneumonia that includes increased cytokines, inflammation and acute respiratory failure. The glycosaminoglycan hyaluronan (HA) and the HA synthases (HASs) are elevated in patients with COVID-19. Activation of the host sensor for SARS-CoV-2, TLR7, increases interferons, interleukin 6 (IL6) and interleukin-1 beta (IL1ß) via Interferon Regulatory Factors (IRFs), NFkB and the NLRP3 inflammasome. We have previously implicated the Receptor for HA-Mediated Motility (RHAMM) in lung inflammation after injury and RHAMM-based peptides block injury-induced inflammation and fibrosis.
Objective: To determine if TLR7 activation increases TLRs, cytokines, RHAMM, HA and HASs and causes inflammation, and if RHAMM-based peptide abrogates these responses.
Design/Methods: We developed a COVID-19 model using intratracheal (IT) administration of the TLR7 agonist SM324405 in C57Bl/6 mice. Lung gene expression was determined by qRT-PCR. Lung inflammation was measured by myeloperoxidase (MPO) and N-acetyl glucosaminidase (NAG) activities for neutrophils and macrophages respectively, and by flow cytometry. HA content and IL1ß protein concentration were measured using ELISAs. RHAMM-based peptide was given subcutaneously once at the time of injury. IT SM324405 was also given to Rhamm knockout mice. Rhamm mRNA expression was measured in the lungs of K18-hACE2 receptor transgenic mice six days after intranasal SARS-CoV-2.
Results: IT SM324405 resulted in a 3 and 10-fold increase in lung neutrophils and macrophages with maximum accumulation at 72 hours. Steady state mRNA expression of Tlr2 (4-fold), Tlr4 (30-fold), Tlr7 (75-fold) and Nlrp3 (5-fold) was observed four hours after IT SM324405. There was an 8-fold increase in lung IL1ß mRNA at 4 hours and IL1ß protein at 8 hours. Expression of Rhamm, Has1, Has2 and Has3 as well as Ifng and IL6 were increased at 4 hours. Plasma HA was maximal at 16 hours and lung HA was maximal at 72 hours. Subcutaneous treatment with a RHAMM-based peptide resulted in decreased caspase 1 activity, plasma and lung HA content and IL1ß protein. This peptide also inhibited lung neutrophil and macrophage accumulation. Administration of IT SM324405 to Rhamm-/- mice did not result in increased neutrophil or macrophage accumulation. Importantly, K18-hACE2 receptor transgenic mice had a 15-fold increase in Rhamm mRNA expression six days after intranasal SARS-CoV-2.Conclusion(s): RHAMM and HA are mechanistically involved in lung inflammation seen in COVID-19. We speculate that RHAMM-based peptide may be an important therapeutic approach to abrogate the cytokine storm in COVID-19.