Sex differences in cardio-pulmonary pathology of SARS-CoV2 infected and Trypanosoma cruzi co-infected mice

  1. Dhanya Dhanyalayam
  2. Kezia Lizardo
  3. Neelam Oswal
  4. Hariprasad Thangavel
  5. Enriko Dolgov
  6. David S. Perlin
  7. Jyothi F. Nagajyothi

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Abstract

Background

Coronavirus disease-2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2; CoV2) is a deadly contagious infectious disease. For those who survived COVID-19, post-COVID cardiac damage poses a major threat for the progression of cardiomyopathy and heart failure. Currently, the number of COVID-related cases and deaths are increasing in Latin America, where a major COVID comorbidity is Chagas’ heart disease (caused by the parasite Trypanosoma cruzi ). Here, we investigated the effect of T. cruzi infection on the pathogenesis and severity of CoV2 infection and, conversely, the effect of CoV2 infection on heart pathology during coinfection.

Methodology/findings

We used transgenic human angiotensin-converting enzyme 2 (huACE2) mice infected with CoV2, T. cruzi , or coinfected with both in this study. Our study shows for the first time that white adipose tissue (WAT) serves as a reservoir for CoV2 and the persistence of CoV2 in WAT alters adipose tissue morphology and adipocyte physiology. Our data demonstrate a correlation between the loss of fat cells and the pulmonary adipogenic signaling (via adiponectin isomers) and pathology in CoV2 infection. The viral load in the lungs is inversely proportional to the viral load in WAT, which differs between male and female mice. Our findings also suggest that adiponectin-PPAR signaling may differently regulate Chagas cardiomyopathy in coinfected males and females.

Conclusion

We conclude that adipogenic signaling may play important roles in cardio-pulmonary pathogenesis during CoV2 infection and T. cruzi coinfection. The levels of adiponectin isomers differ between male and female mice during CoV2 infection and coinfection with T. cruzi , which may differently regulate inflammation, viral load, and pathology in the lungs of both the sexes. Our findings are in line with other clinical observations that reported that males are more susceptible to COVID-19 than females and suffer greater pulmonary damage.

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  1. ScreenIT

    SciScore for 10.1101/2021.09.18.460895: (What is this?)

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

    Table 1: Rigor

    EthicsIACUC: Biosafety: All aspects of this study were approved by the Institutional Animal Care and Use and Institutional Biosafety Committee of Center for Discovery and Innovation of Hackensack University Medical Centre (IACUC 282) and adhere to the National Research Council guidelines.
    Sex as a biological variableBoth male and female mice (N=16) were intraperitoneally (i.p.) infected with 103 trypomastigotes at 6 weeks of age.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Adiponectin-specific mouse monoclonal antibody (#ab22554, Abcam), AdipoR1-specific rabbit polyclonal antibody (#ab70362, Abcam), AdipoR2-specific rabbit polyclonal antibody (#ABT12, Sigma-Aldrich), PPARα-specific rabbit polyclonal antibody (#PA1-822A, Thermo Fisher Scientific), PPARγ-specific rabbit polyclonal antibody (#2492, Cell Signaling Technology), pAMPK-specific rabbit monoclonal antibody (#2535S, Cell Signaling Technology), Cytochrome C-specific rabbit monoclonal antibody (#4280S, Cell Signaling Technology), Superoxide dismutase 1-specific mouse monoclonal antibody (#4266S, Cell Signaling Technology), Hexokinase 2-specific rabbit monoclonal antibody (#2867S, Cell Signaling Technology), β1 adrenergic receptor-specific rabbit polyclonal antibody (#12271S, Cell Signaling Technology), F4/80-specific rat monoclonal antibody (#NB 600-404, Novus Biologicals), TNFα-specific rabbit polyclonal antibody (#ab6671, Abcam), pHSL (Ser563)-specific rabbit monoclonal antibody(#4139, Cell Signaling Technology), ATGL-specific rabbit monoclonal antibody(#30A4, Cell Signaling Technology, Perilipin-specific rabbit monoclonal antibody (#D1D8, Cell Signaling Technology), IFNγ-specific rabbit monoclonal antibody (#EPR1108, Abcam), CD4-specific rabbit polyclonal antibody (#NBP1-19371, Novus biologicals), CD8-specific rabbit polyclonal antibody(#NBP2-29475, Novus biologicals), T-cadherin-specific rabbit polyclonal antibody (#ABT121, Millipore), FABP4-specific rabbit monoclonal antibody (#3544, Cell Signaling Technology), IL6-specific mouse monoclonal antibody (#66146-1-lg, Proteintech), IL10-specific rabbit polyclonal antibody (#20850-1-AP, Proteintech), BNIP3-specific rabbit monoclonal antibody (#44060, Cell Signaling Technology), Caspase 3-specific rabbit polyclonal antibody (#9662, Cell Signaling Technology) were used as primary antibodies.
    antibody(#4139
    suggested: (Cell Signaling Technology Cat# 3544, RRID:AB_2278257)
    antibody(#30A4, Cell Signaling Technology, Perilipin-specific
    suggested: None
    CD4-specific
    suggested: None
    CD8-specific
    suggested: None
    antibody(#NBP2-29475
    suggested: (Novus Cat# NBP1-44060, RRID:AB_2212858)
    IL10-specific
    suggested: None
    antibody (#9662, Cell Signaling Technology)
    suggested: (Cell Signaling Technology Cat# 9662, RRID:AB_331439)
    Horseradish peroxidase (HRP)-conjugated anti-mouse immunoglobulin (#7076, Cell Signaling Technology) or HRP-conjugated anti-rabbit immunoglobulin (#7074, Cell Signaling Technology) antibody was used to detect specific protein bands (as shown in the figure legends) using a chemiluminescence system.
    anti-mouse immunoglobulin (#7076, Cell Signaling Technology)
    suggested: (Cell Signaling Technology Cat# 7076, RRID:AB_330924)
    anti-rabbit immunoglobulin (#7074, Cell Signaling Technology)
    suggested: (Cell Signaling Technology Cat# 7074, RRID:AB_2099233)
    β-actin-specific rabbit monoclonal antibody (#4970S, Cell Signaling Technology) or Guanosine nucleotide dissociation inhibitor (GDI) (#71-0300, Invitrogen) were used as protein loading controls.
    β-actin-specific rabbit monoclonal antibody (#4970S, Cell Signaling Technology)
    suggested: None
    rabbit monoclonal antibody (#4970S, Cell Signaling Technology)
    suggested: (Cell Signaling Technology Cat# 4970, RRID:AB_2223172)
    antibody
    suggested: (Cell Signaling Technology Cat# 4970, RRID:AB_2223172)
    Guanosine nucleotide dissociation inhibitor (GDI
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    The Brazil strain of T. cruzi was maintained by passage in C3H/HeJ mice (Jackson Laboratories, Bar Harbor, ME).
    C3H/HeJ
    suggested: RRID:IMSR_JAX:000659)
    Software and Algorithms
    SentencesResources
    Adiponectin-specific mouse monoclonal antibody (#ab22554, Abcam), AdipoR1-specific rabbit polyclonal antibody (#ab70362, Abcam), AdipoR2-specific rabbit polyclonal antibody (#ABT12, Sigma-Aldrich), PPARα-specific rabbit polyclonal antibody (#PA1-822A, Thermo Fisher Scientific), PPARγ-specific rabbit polyclonal antibody (#2492, Cell Signaling Technology), pAMPK-specific rabbit monoclonal antibody (#2535S, Cell Signaling Technology), Cytochrome C-specific rabbit monoclonal antibody (#4280S, Cell Signaling Technology), Superoxide dismutase 1-specific mouse monoclonal antibody (#4266S, Cell Signaling Technology), Hexokinase 2-specific rabbit monoclonal antibody (#2867S, Cell Signaling Technology), β1 adrenergic receptor-specific rabbit polyclonal antibody (#12271S, Cell Signaling Technology), F4/80-specific rat monoclonal antibody (#NB 600-404, Novus Biologicals), TNFα-specific rabbit polyclonal antibody (#ab6671, Abcam), pHSL (Ser563)-specific rabbit monoclonal antibody(#4139, Cell Signaling Technology), ATGL-specific rabbit monoclonal antibody(#30A4, Cell Signaling Technology, Perilipin-specific rabbit monoclonal antibody (#D1D8, Cell Signaling Technology), IFNγ-specific rabbit monoclonal antibody (#EPR1108, Abcam), CD4-specific rabbit polyclonal antibody (#NBP1-19371, Novus biologicals), CD8-specific rabbit polyclonal antibody(#NBP2-29475, Novus biologicals), T-cadherin-specific rabbit polyclonal antibody (#ABT121, Millipore), FABP4-specific rabbit monoclonal antibody (#3544, Cell Signaling Technology), IL6-specific mouse monoclonal antibody (#66146-1-lg, Proteintech), IL10-specific rabbit polyclonal antibody (#20850-1-AP, Proteintech), BNIP3-specific rabbit monoclonal antibody (#44060, Cell Signaling Technology), Caspase 3-specific rabbit polyclonal antibody (#9662, Cell Signaling Technology) were used as primary antibodies.
    Cell Signaling Technology
    suggested: (Cell Signaling Technology, RRID:SCR_004431)
    Data were pooled, and statistical analysis was performed using a Student’s t-test (Microsoft Excel) as appropriate and significance differences were determined as p values between < 0.05 and <0.001 as appropriate.
    Microsoft Excel
    suggested: (Microsoft Excel, RRID:SCR_016137)

    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).

    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.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

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

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

    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.

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

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2021.09.18.460895v1 on bioRxiv