Review 3: "Lipid droplets fuels SARS-CoV-2 replication and inflammatory response"

RR:C19 Evidence Scale rating by reviewer:

• Potentially informative. The main claims made are not strongly justified by the methods and data, but may yield some insight. The results and conclusions of the study may resemble those from the hypothetical ideal study, but there is substantial room for doubt. Decision-makers should consider this evidence only with a thorough understanding of its weaknesses, alongside other evidence and theory. Decision-makers should not consider this actionable, unless the weaknesses are clearly understood and there is other theory and evidence to further support it.

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Review:

The manuscript by Dias et al shows an increased in lipid droplets in monocytes from COVID-19 patients. The same phenomenon was also observed in human monocytes and other cell lines infected with SARS-CoV2 in vitro. This increase in lipid droplets is probably explained by increases in expression of a number of genes involved in lipid uptake and synthesis. It was hypothesized that the virus induces these changes to promote viral replication and that lipid droplets play an important in viral replication. Applying an inhibitor of fatty acid synthesis could reduce the number of lipid droplets in infected cell and the viral output as well as production of proinflammatory cytokines. The authors also tried to show co-localization between lipid droplets and viral antigens / dsRNA in infected cells.

Except for the co-localization, the data seem to be reliable and trustworthy and in agreement with previous publications on a number of other viruses, which induced lipid synthesis and relied on the increased availability of intracellular lipid for optimal viral replication. While there are some co-localized areas between the viral antigens and lipid droplets, many areas of viral antigen do not co-localized with lipid droplet. This has to be better described and explained. Because the viral antigen detection was done using serum, which might react to many antigens and positive areas might contain different antigens. This may explain the lack of co-localization of some antigen signal. The co-localization with dsRNA looks less convincing as only a few co-localized dots were shown. The description and interpretation should be more cautiously stated. It is possible that inhibition of DGAT simply reduce the viral replication by reducing availability of intracellular lipid. Availability of lipid and number of lipid droplets is related but may not be the same thing.

In addition, more information should be given on the COVID-19 patients, whose blood monocytes are shown in Fig. 1. How many patients were the data derived from, and what were their clinical conditions? Was there any correlation between the levels of lipid droplets and clinical severity?