IDF21-0230 Anti-inflammatory activity of glyoxalase-1 inducer supports further evaluation for prevention of COVID-19

Abstract

Background: Pre-clinical and clinical studies suggest an essential role of proteases, plasma membrane-associated transmembrane protease serine 2 (TMPRSS2) and furin, in virion fusion with lung alveolar epithelial cells for viral infectivity of coronavirus SARS-CoV-2. Angiotensin converting enzyme-2 (ACE2) and glucose regulated protein-78 (GRP78) are considered to be receptors for cell surface binding of the spike protein of SARS-CoV-2. Risk of progression of associated COVID-19 disease to severe symptoms and risk of mortality is linked to increased levels of inflammatory mediators, including monocyte chemoattract protein-1 (MCP-1) and interleukin-8 (IL8), which are also likely risk factors for severe, life-threatening disease. Increased MCP-1 and IL8 in obese subjects and patients with diabetes may contribute to poor clinical outcomes when infected with coronavirus. In a double blind placebo-controlled clinical crossover study in overweight and obese subjects, treatment with Glo1 inducer, trans-resveratrol and hesperetin combination (tRES-HESP), once daily by oral capsule dietary supplement for 8 weeks, decreased expression of MCP-1, IL8, the receptor for advanced glycation endproducts (RAGE) and cyclo-oxygenase-2 (COX-2) in peripheral blood mononuclear cells (PBMCs). Placebo had no effect. We hypothesized that tRES-HESP may decrease protease and inflammatory gene expression linked to SARS-CoV-2 infection and COVID-19 progression. If so, the anti-inflammatory activity of Glo1 inducer in PBMCs found in clinical trial may also be evidenced in human small airway alveolar epithelial cells in primary culture. Aim: The aim of this project was to assess the anti-inflammatory effect of tRES-HESP on human alveolar epithelial cells in primary culture, assessing effects on protease and inflammatory gene expression considered essential for SARS-CoV-2 infectivity and progression of COVID-19 severity. Method: Human small airway epithelial cells (SAECs) were purchased from Lonza. SAECs were cultured in Small Airway Growth Media with supplements, according to the supplier’s instructions. SAECs (10,000 cells/cm2) were cultured with and without 5 µM tRES-HESP and the effect of basal gene expression studied after 72 h for mRNA (qRT-PCR) and protein (Western blotting); and significance of effect assessed by Student’s t-test, n = 3 or 4. The concentration of glucose in the medium was 6 mM. Results: In SAEC primary cultures, tRES-HESP decreased basal protein levels of TMPRSS2 (- 58%; P<0.001) and furin protease (- 43%; P<0.01) after 72 h. tRES-HESP also decreased mRNA of TMPRSS2 (- 62%, P<0.01), GRP78 (- 67%, P<0.001), COX-2 (-34%, P<0.001) and RAGE mRNA (- 56%, P<.001) and decreased basal secretion of MCP-1 and IL8 (-29% and - 25%, P<0.001). Discussion: tRES-HESP decreased basal expression of proteases essential for SARS-CoV-2 infection, GRP78 and secretion of inflammatory mediator risk predictors of severe COVID-19. These effects are likely due to activation of Nrf2 by tRES-HESP and decrease of unfolded protein response activation by suppression of dicarbonyl stress. These outcomes suggest previous anti-inflammatory response of tRES-HESP in overweight and obese subjects found in PBMCs may also occur concomitantly in the lung. TMPRSS2 is an antioxidant response element-linked gene down regulated by activation of Nrf2. These anti-proteolytic and anti-inflammatory responses of tRES-HESP suggest it is suitable for further evaluation for prevention of COVID-19, particularly in obesity and diabetes.