SARS-CoV-2 Causes Severe Epithelial Inflammation and Barrier Dysfunction

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1. Version published to 10.1128/jvi.00110-21

   Apr 26, 2021
2. 

   ScreenIT

   Sep 9, 2020

   SciScore for 10.1101/2020.08.31.276725: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	not detected.
   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
   ) IgG monoclonal, primary antibodies and AlexaFluor® goat anti-mouse IgG polyclonal antibodies (Dianova; # 115-545-146).	anti-mouse IgG suggested: (Jackson ImmunoResearch Labs Cat# 115-545-146, RRID:AB_2307324)
   Rabbit anti-E-cadherin IgG monoclonal (CellSignaling; 3195S) or rabbit anti-VE-cadherin polyclonal, primary antibodies (CellSignaling; 2158S) and Cy5 goat anti-mouse IgG polyclonal antibodies (Dianova; #111-175-144) were used to detect cell borders of …

   SciScore for 10.1101/2020.08.31.276725: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	not detected.
   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
   ) IgG monoclonal, primary antibodies and AlexaFluor® goat anti-mouse IgG polyclonal antibodies (Dianova; # 115-545-146).	anti-mouse IgG suggested: (Jackson ImmunoResearch Labs Cat# 115-545-146, RRID:AB_2307324)
   Rabbit anti-E-cadherin IgG monoclonal (CellSignaling; 3195S) or rabbit anti-VE-cadherin polyclonal, primary antibodies (CellSignaling; 2158S) and Cy5 goat anti-mouse IgG polyclonal antibodies (Dianova; #111-175-144) were used to detect cell borders of Calu-3 or HUVEC cells on the membrane of the alveolus-on-a-chip model, respectively.	anti-E-cadherin IgG suggested: None anti-VE-cadherin suggested: (Cell Signaling Technology Cat# 2158, RRID:AB_2077970)
   For the detection of SARS-CoV-2 spike protein a rabbit polyclonal anti-SARS-CoV-2 spike S2 antibody (Sino Biological #40590-T62) was used.	anti-SARS-CoV-2 spike S2 suggested: (Thermo Fisher Scientific Cat# PA5-112048, RRID:AB_2866784)
   Experimental Models: Cell Lines
   Sentences	Resources
   For the cultivation of the human alveolus-on-a-chip model we used Calu-3 cells and macrophages at the epithelial side, and HUVECs at the endothelial side.	HUVECs suggested: None
   For infection of Vero-76 or Calu-3 cells, cells were washed with PBS and either left uninfected (mock) or infected with SARS-CoV-2 with a multiplicity of infection (MOI) of 1 for 120 min in medium (EMEM with HEPES modification and 5 mM L-Glutamine for Vero-76 cells and RPMI 1640 for Calu-3 cells) supplemented with 10 % FCS.	Vero-76 suggested: None Calu-3 suggested: None
   Rabbit anti-E-cadherin IgG monoclonal (CellSignaling; 3195S) or rabbit anti-VE-cadherin polyclonal, primary antibodies (CellSignaling; 2158S) and Cy5 goat anti-mouse IgG polyclonal antibodies (Dianova; #111-175-144) were used to detect cell borders of Calu-3 or HUVEC cells on the membrane of the alveolus-on-a-chip model, respectively.	HUVEC suggested: KCB Cat# KCB 200648YJ, RRID:CVCL_2959)
   Software and Algorithms
   Sentences	Resources
   The cDNA preparation was performed using the SuperScript IV (Thermofisher), followed by a multiplex PCR to generate overlapping 400 nt amplicons using version 3 of the primer set (https://github.com/artic-network/artic-ncov2019/tree/master/primer_schemes/nCoV-2019/V3).	Thermofisher suggested: (ThermoFisher; SL 8; Centrifuge, RRID:SCR_020809)
   Infection by SARS-CoV-2 was visualized using mouse anti-SARS-CoV-2 spike (GeneTex; #GTX632604	GeneTex suggested: (GeneTex, RRID:SCR_000069)
   Images were acquired using an Axio Observer.Z1 microscope (Zeiss) with Plan Apochromat 20x/0.8 objective (Zeiss), ApoTome.2 (Zeiss) and Axiocam 503 mono (Zeiss) and the software Zen 2.6 (blue edition; Zeiss).	Zen suggested: None
   Fiji V 1.52b (ImageJ) was used for further image processing, including Z-stack merging with maximum intensity projection and gamma correction.	Fiji suggested: (Fiji, RRID:SCR_002285) ImageJ suggested: (ImageJ, RRID:SCR_003070)
   Statistical analysis: Statistical analyses were performed using Prism 8 (GraphPad Software).	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:

   • No conflict of interest statement was detected. If there are no conflicts, we encourage authors to explicit state so.
   • 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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3. 

   Version published to 10.1101/2020.08.31.276725v1 on bioRxiv

   Sep 2, 2020