THE EFFECTS of TYPE i & II IFNs DISCLOSE A ROLE for DOPAMINE in SARS-CoV-2 INFECTION

ABSTRACT

Background: Recent studies highlight the dynamic nature of virus-host interaction during SARS-CoV-2 infection, raising intriguing questions about the role and timing of interferon (IFN) responses. In fact, SARS-CoV-2 delays/antagonizes Type-I, and to a definitely lesser extent, Type II-IFNs. While paving the way for potential antiviral therapies based on immune activation, the molecular mechanisms linking different IFN pathways to SARS-CoV-2 susceptibility remain to be elucidated. The present study investigates the role of Type-I &-II IFNs in SARS-CoV-2 replication in human lung cells, with a focus on molecular pathways related with innate and adaptive immunity. Methods: Human lung carcinoma cells (CaLu3) were pretreated with IFN-α,-β or-γ (from 1 to 1000 U/mL), O.N. Cells were infected with SARS-CoV-2 (MOI 0.05) for 3h, and IFNs were added during infection. In another set of experiments, IFNs were added only p.i. Supernatants were harvested at 24 and 48h p.i. to assess viral replication by RT-qPCR, and to quantify the levels of cytokines/chemokines through Multiplex assay. At 48h post-infection, cells were collected and RNA was retrotranscribed to investigate a variety of transcriptional targets. Cell viability was assessed by MTT. Results are presented as the average of the relative expression units to the GAPDH gene, calculated by the 2-ΔΔCt equation. Statistical analyses were performed through the Student t-test. Results: Pretreatment with both Type-I &-II IFNs dramatically reduces SARS-CoV-2 replication in the absence of cell toxicity. Such an effect is maintained, though at a lower magnitude, when IFNs are added only p.i. The antireplicative effects of Type-I &-II IFNs are associated with both convergent and divergent mechanisms. Both Types decrease the expression and/or protein levels of most pro-inflammatory mediators while augmenting anti-inflammatory and anti-apoptotic factors. Surprisingly, IFN-γ shows the strongest effect in potentiating antiviral effectors besides boosting adaptive immunity pathways. Remarkably, a convergent effect of both IFN Types is observed upon the expression of genes associated with DA activity, including DA receptors (D1-D5) and the DA transporter (DAT), which are dramatically altered by SARS-CoV-2. Conclusion: Both Type-I &-II IFNs halt SARS-CoV-2 replication by acting through complementary mechanisms. Their effects also disclose a potential role for DA activity, and neuromodulators in general, in host immunity during SARS-CoV-2 infection in pulmonary cells.