SARS-CoV-2-associated ssRNAs activate inflammation and immunity via TLR7/8

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   ScreenIT

   Apr 17, 2021

   SciScore for 10.1101/2021.04.15.439839: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement	IRB: All mouse experiments were carried out in accordance with guidelines prescribed by the Ethics Committee for the use of laboratory animals for research purposes at the University of Verona and by the Italian Ministry of Health.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Flow cytometry: Human and mouse DCs were stained with the following antibodies from Miltenyi Biotec or as specified: Vioblue-conjugated anti-human CD86 (clone FM95, Miltenyi Biotec), PE-conjugated anti-human CD83 (clone REA714), …

   SciScore for 10.1101/2021.04.15.439839: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement	IRB: All mouse experiments were carried out in accordance with guidelines prescribed by the Ethics Committee for the use of laboratory animals for research purposes at the University of Verona and by the Italian Ministry of Health.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Flow cytometry: Human and mouse DCs were stained with the following antibodies from Miltenyi Biotec or as specified: Vioblue-conjugated anti-human CD86 (clone FM95, Miltenyi Biotec), PE-conjugated anti-human CD83 (clone REA714), FITC-conjugated anti-human BDCA2 (clone AC144), APC-conjugated anti-human CCR7 (clone REA546), VioGreen-conjugated anti-mouse CD45 (clone REA737), VioBlue or FITC-conjugated anti-mouse MHCII (clone REA564), PerCP-Vio 700-conjugated anti-mouse CD11c (clone REA754), PE-conjugated anti-mouse SiglecH (clone 551.3D3), PE-Vio 615-conjugated anti-mouse CD11b (clone REA592), VioBlue-conjugated anti-mouse CD8a (cloneREA601), PE-Vio 770-conjugated anti-mouse B220 (clone RA3-6B2), PE-conjugated anti-mouse CD40 (clone REA965), FITC-conjugated anti-mouse CD40 (clone HM40-3, Biolegend) and APC-CY7-conjugated anti-mouse CD86 (clone GL-1, Biolegend).	anti-human CD86 suggested: None anti-human CD83 suggested: (BD Biosciences Cat# 334098, RRID:AB_2074438) anti-human BDCA2 suggested: (Miltenyi Biotec Cat# 130-090-510, RRID:AB_244167) anti-human CCR7 suggested: None anti-mouse CD45 suggested: None anti-mouse MHCII suggested: None anti-mouse CD11c suggested: None anti-mouse SiglecH suggested: (Creative Diagnostics Cat# CABT-35771RM, RRID:AB_2356706) anti-mouse CD11b suggested: (BD Biosciences Cat# 559971, RRID:AB_10053179) anti-mouse CD8a suggested: None anti-mouse B220 suggested: None anti-mouse CD40 suggested: None anti-mouse CD86 suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   RAW264.7 cells were purchased from American Type Culture Collection and cultured in DMEM complemented with 10% FBS.	RAW264.7 suggested: None
   NF-κB luciferase reporter assay: TLR-specific activation assays were performed using human HEK293 cells expressing luciferase under control of the NF-κB promoter and stably transfected with human TLR7 and TLR8 as previously described (21).	HEK293 suggested: CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045)
   HEK293-transfected cells were maintained in DMEM supplemented with 10% FBS and specific antibiotics were added.	HEK293-transfected suggested: None
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Sex and age matched C57Bl6/J mice were obtained by Charles River Laboratories and housed in the specific pathogen-free animal facility of the Department of Medicine, University of Verona.	C57Bl6/J suggested: None
   MyD88−/− mice were kindly provided by S. Akira (Osaka University, Osaka, Japan).	MyD88−/− suggested: None
   Software and Algorithms
   Sentences	Resources
   These sequences were checked for uniqueness with BLAST in the database RefSeq Genome Database (refseq_genomes) within the RNA viruses (taxid: 2559587).	BLAST suggested: (BLASTX, RRID:SCR_001653)
   Samples were read on a MACSQuant Analyzer (Miltenyi Biotec) and analysed with FlowJo (Tree Star Inc.).	FlowJo suggested: (FlowJo, RRID:SCR_008520)
   The SsoAdvanced Universal SYBR Green Supermix (Bio-Rad Laboratories) was used according to the manufacturer’s instructions.	Bio-Rad Laboratories suggested: (Bio-Rad Laboratories, RRID:SCR_008426)
   Statistical analysis: Statistical significance among the experimental groups was determined using paired or unpaired Student’s t test or one-way ANOVA with Dunnet’s post-hoc test (GraphPad Prism 7, GraphPad Software) as indicated in each figure legend.	GraphPad suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

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   Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

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   Results from JetFighter: We did not find any issues relating to colormaps.

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   Results from rtransparent:

   • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
   • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
   • No protocol registration statement was detected.

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2. 

   Version published to 10.1101/2021.04.15.439839 on bioRxiv

   Apr 15, 2021