Effects of Hydroxychloroquine and Azithromycin on iPSC-derived Cardiomyocytes: Considerations for the Treatment of COVID-19 Patients

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   ScreenIT

   Aug 21, 2021

   SciScore for 10.1101/2021.08.19.456950: (What is this?)

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

   Table 1: Rigor

   Ethics	IRB: The iPSC generation was approved by the Ethics Committee of the University Medical Center Göttingen (approval number: 21/1/11 and 10/9/15) and used following the approval guidelines.
   Sex as a biological variable	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   For staining, cells were incubated with the following primary antibodies: anti-α-actinin, clone EA-53 (1:500; mouse monoclonal, IgG1, Sigma-Aldrich, 7811), anti-Nav1.5 (1:200; rabbit polyclonal, Alomone Labs, ASC-005), and anti-Cx43, clone 2	anti-α-actinin suggested: None IgG1 suggested: (R and D Systems Cat# BAM7811, RRID:AB_10639875)
   Afterward, cells …

   SciScore for 10.1101/2021.08.19.456950: (What is this?)

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

   Table 1: Rigor

   Ethics	IRB: The iPSC generation was approved by the Ethics Committee of the University Medical Center Göttingen (approval number: 21/1/11 and 10/9/15) and used following the approval guidelines.
   Sex as a biological variable	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   For staining, cells were incubated with the following primary antibodies: anti-α-actinin, clone EA-53 (1:500; mouse monoclonal, IgG1, Sigma-Aldrich, 7811), anti-Nav1.5 (1:200; rabbit polyclonal, Alomone Labs, ASC-005), and anti-Cx43, clone 2	anti-α-actinin suggested: None IgG1 suggested: (R and D Systems Cat# BAM7811, RRID:AB_10639875)
   Afterward, cells were washed three times with PBS and incubated with the corresponding secondary antibodies (1:1000; anti-rabbit Alexa Fluor 488, Invitrogen,	anti-rabbit suggested: None
   Membranes were blocked in 5% milk in TBS-T for 30-45 min at room temperature and probed with anti-Cx43, clone 4E6.2 (1:1000; mouse monoclonal, Merck, MAB3067), anti-Nav1.5 (1:200; rabbit polyclonal, Alomone Labs, ASC-005), or anti-EEF2 (1:5000; rabbit polyclonal, Abcam, ab40812) at 4°C overnight, followed by incubation with horseradish peroxidase-conjugated secondary antibodies goat anti-mouse (1:10,000; Sigma Aldrich, A2304) or goat anti-rabbit (1:10,000; Cell Signaling, 7074S), respectively, for 1 hour at room temperature.	anti-Cx43 suggested: (SICGEN Cat# AB0015-200, RRID:AB_2333132) anti-Nav1.5 suggested: (Alomone Labs Cat# ASC-005, RRID:AB_2040001) anti-EEF2 suggested: (Abcam Cat# ab40812, RRID:AB_732082) horseradish peroxidase-conjugated secondary antibodies goat anti-mouse ( 1:10,000; Sigma Aldrich , A2304 ) suggested: None anti-mouse suggested: (Sigma-Aldrich Cat# A2304, RRID:AB_257993) goat anti-rabbit suggested: (Cell Signaling Technology Cat# 7074, RRID:AB_2099233)
   Software and Algorithms
   Sentences	Resources
   Sarcomere-length was determined manually using FIJI as described previously22.	FIJI suggested: (Fiji, RRID:SCR_002285)
   Several key parameters including conduction velocity (CV), corrected FPDC (corrected by Fridericia’s formula) and inter-beat interval were determined using AxIS Navigator, and further analyzed with AxIS Metric Plotting Tool (Axion BioSystems).	AxIS suggested: (AxIS , RRID:SCR_016308)
   Densitometry analyses of the immunoblots were performed using ImageJ software and the intensity of individual bands was normalized to EEF2.	ImageJ suggested: (ImageJ, RRID:SCR_003070)
   Statistical analysis was performed with GraphPad Prism 9.	GraphPad Prism 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: We detected the following sentences addressing limitations in the study:

   Study limitations: Aiming to gain mechanistic insights for the increased rates of cardiac complications observed for the combined treatment with AZM and HCQ, we characterized the consequences of the two drugs as well as their combination on the viability, structure and functionality of iPSC-CMs. Despite iPSC-CMs represent an important model system to study drug effects on the human heart, different aspects, including the immaturity of the cells and the lack of the multicellular environment limit, the predictive value of our findings. Furthermore, modeling the situation in patients with severe COVID-19 may require infection of iPSC-CMs with SARS-CoV-2 before drug treatment to model structural and functional abnormalities, which will make the execution of the study technically challenging.

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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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   Results from scite Reference Check: We found no unreliable references.

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   Version published to 10.1101/2021.08.19.456950v1 on bioRxiv

   Aug 19, 2021