The effects of curcumin derivatives on SARS-CoV-2 spike S1 protein induced oxidative stress on macrophage cells

Abstract

The corona virus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has emerged as a global health crisis that claimed almost 7 million lives and a total of more than 700 million cases to date with new variants still being reported in various countries. Currently, there is no specific treatment for COVID-19 patients and the sporadic emergence of new variants make vaccine development a delayed response towards management of future outbreaks. Infection of host cells via interaction with SARS-CoV-2 spike proteins is characterised by hyper-inflammation of the innate immune response leading to overproduction of reactive oxygen species (ROS) by macrophage. The increased ROS production ultimately overwhelms the antioxidant pool and promote elevated oxidative stress levels, converting mild symptoms to severe COVID-19 outcomes. Curcumin has shown potent antioxidant and anti-inflammatory properties in various pathological conditions. Thus, this study investigated the antioxidant potential of curcumin derivatives in ameliorating SARS-CoV-2 spike protein-induced oxidative stress using macrophage cells as the innate immune model system. The cytotoxicity effect of curcumin derivatives was assessed by using two viability assays, namely the colorimetric MTT and propidium iodide (PI) fluorescence assays on Raw 264.7 macrophage cells. The curcumin derivatives were shown to exert no cytotoxic effect on Raw 264.7 cells at low doses (1.25-0.625 mM). The Annexin-V/ PI assay was employed to evaluate the mode of cell death induced by these curcumin derivatives. The results obtained showed that the derivatives at 5 mM induce apoptosis as a mode cell death. The production of IL-6 pro-inflammatory cytokines was examined using ELISA from supernatant after stimulation with 100 ng/ml SARS-CoV-2 spike S1 and treated with 1 mM curcumin derivatives. Treatment of SARS-CoV-2 spike S1-stimulated Raw 264.7 macrophages with curcumin resulted in lower production of IL-6 compared to untreated SARS-CoV-2 spike S1 stimulated control cells. The oxidative stress muse kit and DAF-2 DA were used to determine the levels of reactive oxygen species (ROS) and nitric oxide (NO) produced by Raw 264.7 cells post SARS-CoV-2 spike S1 stimulation and curcumin 13 derivative treatment. Treatment with 1 mM curcumin derivatives reduced the production of ROS and NO. The Western blot assay was used to measure the expression levels of Nrf2, IκΒ-α and NF-κB inflammatory proteins. Western blot protein expression assay demonstrated that these curcumin derivatives upregulate the expression of Nrf2 and IκΒ-α while downregulating that of NF-κB. Collectively, this study suggests that curcumin derivatives possess remarkable anti-inflammatory and antioxidant properties, making them a potential therapeutic treatment option to explore for the management of the SARS-Cov-2 induced oxidative stress and hyper-inflammation associated with COVID-19 infection.