henhaobio
Sign in

bioRxivPathologyPreprint

Host response against SARS-CoV-2 infection is impacted by allosteric modulation of cholinergic and purinergic receptors

Victoire PerraudiD, Guilherme Dias de MeloiD, Marco CecchiniiD, Lauriane KergoatiD, Julien Lannoy, Jean‐Pierre ChangeuxiD, Hervé BourhyiD, Florence LarrousiD

PDFDOI

In-depth summary

Claude Opus 5.5 · xhigh effort · 8 Nov 2026 · from the full text of version 1 (26 Oct 2026) · CC BY 4.0

Ivermectin's protection in SARS-CoV-2-infected hamsters depended partly on both α7 nicotinic and P2X4 receptors; in co-cultures its effects were α7nAChR-independent, and P2X4 blockade prevented CXCL10 induction at 48 hpi.

Ivermectin improved clinical outcome in infected hamsters in earlier work, but its mechanism of action is still unknown. Here it serves as a pharmacological tool to test the roles of α7nAChR, P2X4 and P2X7 in host response, using cell cultures and hamsters.

Key results

  1. ShownIn Vero E6 cells, ivermectin and two other α7nAChR modulators (GAT-107, EVP-6124) protected cells and lowered the share of infected cells. Two α7nAChR antagonists did not abolish ivermectin's effect, so it is α7nAChR-independent in this model.

    Ivermectin 5 µM, GAT 25 µM and EVP 16 µM reduced cell death and infected-cell percentage versus untreated controls. Ivermectin's effect persisted with MLA (0.1-10 µM) or mecamylamine (8-32 µM). Similar protection was seen with Alpha, Beta and Delta variants. · Modulation of α7nAChRs is not sufficient to…; IVM exerts a protective effect against…

  2. ShownIn an A549-ACE2-TMPRSS2/THP-1 transwell co-culture, ivermectin raised intracellular ATP and lowered nanoluciferase release without changing viral RNA in THP-1 cells. It shifted THP-1 transcripts (CXCL10 up, CXCL5 down) at 72 hpi, and MLA did not change this pattern.

    Four independent experiments, Mann-Whitney tests. MLA partially reversed ivermectin's antiviral effect in A549-ACE2-TMPRSS2 but not THP-1 cells. IL-6 was unchanged and ISG20 was up at 72 hpi. · IVM exerts a protective effect against…

  3. SuggestedIn the co-culture time course, THP-1 cells responded to ivermectin first (24 hpi) and epithelial cells later (CXCL10 at 48 hpi). The P2X4 antagonist BX430 blocked ivermectin-induced CXCL10 at 48 hpi, while the P2X7 antagonist further lowered CXCL5 in THP-1.

    At 24 hpi in THP-1: CXCL10 up, CXCL5 and IL-6 down (p=0.0286 for each, Mann-Whitney). MLA did not alter the signature. BX430 also lowered IL-6 in THP-1 at 24 hpi and IL-6 and ISG20 in epithelial cells at 48 hpi (p=0.0286). · IVM-mediated immunomodulation reveals distinct…

  4. ShownIn hamsters, ivermectin's benefit on clinical score, olfactory dysfunction and lung pathology was abolished by α7nAChR antagonists. The P2X4 antagonist abolished the clinical and olfactory benefit but not the lung benefit.

    Mecamylamine and MLA abolished all three outcomes; BX430 abolished clinical score and olfaction only. Olfactory dysfunction: 11.1% with GAT-107 and 37.5% with ivermectin versus 68.8% in non-treated animals; EVP-6124 had no effect. Groups of n=8 or 9. · α7nAChR and P2X4 receptors mediate IVM-induced…; Figure legends

  5. ShownHamster lung RT-qPCR did not reproduce the in vitro pattern. Viral RNA did not differ between groups, and Il-6 rose with ivermectin instead of falling.

    No group differences in viral RNA, Cxcl10 or Cxcl5. Il-6 was higher with ivermectin than in controls (p=0.0286, Mann-Whitney). Isg20 was not significantly changed by ivermectin alone, but rose significantly with ivermectin plus BX430. · α7nAChR and P2X4 receptors mediate IVM-induced…; Discussion

Methods, data and reproducibility

Ivermectin, other α7nAChR modulators and receptor antagonists were tested in infected Vero E6 cells, in an epithelial/macrophage transwell co-culture, and in infected golden hamsters. Readouts were infection and viability imaging, ATP and nanoluciferase assays, RT-qPCR of selected genes, clinical score, a food-finding olfaction test and lung weight ratio.

  • Vero E6 cell infections, imaged on a high-content system (3 independent experiments with 10 technical replicates); Wuhan, Alpha, Beta and Delta isolates · n = 3 independent experiments, 10 technical replicates · GISAID EPI_ISL_735391 (Alpha), EPI_ISL_964916 (Beta), EPI_ISL_2029113 (Delta)
  • Transwell co-culture of A549-ACE2-TMPRSS2 and differentiated THP-1 cells, with ATP, nanoluciferase and RT-qPCR readouts at 24, 48 and 72 hpi · human (A549 and THP-1 are described as human cell lines) · n = 4 independent experiments, technical duplicates or triplicates
  • Male golden Syrian hamsters infected intranasally with SARS-CoV-2 Wuhan; clinical score, olfaction test and lung weight ratio; lung RT-qPCR at 4 dpi · golden Syrian hamster (Mesocricetus auratus) · n = n=8 or 9 per group
Methods & tools
Opera Phenix High-Content Screening System, Wuhan_nLuc recombinant reporter virus, Nicotinic Acetylcholine Receptor Allosteric Ligand Library (ACRALL), ATPlite, QuantStudio 6 Flex, 2^-ΔΔCt method, GraphPad Prism 10, Kruskal-Wallis with Dunn's test, Mann-Whitney test
Reagents & environment
GraphPad Prism version 10; QuantStudio Real-time PCR System software v.2.7.0. Primer list in Supplementary Table S1.
Reproducibility gaps
Data will be made available on request. No code or repository is given.
Limitations (5)
  • The P2X7 antagonist could not be tested in hamsters because of toxicity, so P2X7's in vivo role is untested. · stated by the authors
  • In vitro and in vivo results diverged: MLA did not alter the transcriptomic signature in cells but abolished protection in hamsters, and lung transcripts did not fully match the co-culture pattern. The authors attribute this to the simple two-cell transwell system. · stated by the authors
  • Randomization and blinding were not possible in the animal experiments because of the pre-defined housing in separate isolators. · stated by the authors
  • Receptor roles are inferred from pharmacological antagonists alone, without genetic knockout or knockdown; co-culture and hamster experiments used one dose of each. Readout is RT-qPCR of four host genes, with small groups (four independent experiments in vitro, n=8 or 9 per hamster group). · noted in this summary
  • The proposed sequence of events (early P2X4-driven macrophage CXCL10, later α7nAChR-dependent tissue protection) is a model built from these data and is not directly tested. · noted in this summary

Read this if you study host-directed or immunomodulatory approaches to COVID-19 and want data on cholinergic versus purinergic receptor contributions in cell and hamster models.

Citations
0

Abstract

The COVID-19 pandemic has caused nearly 8 million deaths and a broad spectrum of clinical manifestations driven by dysregulated host inflammation, notably involving key signalling proteins IL-6, CXCL10, and CXCL5. We previously demonstrated that ivermectin (IVM) improves clinical outcomes and reduces the IL-6/IL-10 ratio in the lungs of SARS-CoV-2-infected hamsters. Here, combining high-throughput confocal microscopy and transcriptomics in human cell co-cultures and a hamster model, we dissected how IVM modulates host cells responses via 7nAChR and purinergic receptors. In vitro, IVM acted independently of 7nAChR, instead triggering a P2X4-dependent drop in intracellular ATP, rapidly upregulating CXCL10 in macrophages, and suppressing CXCL5 and IL-6. In vivo, however, 7nAChR antagonists abolished IVMs clinical benefit, as did the P2X4 blocker BX430; moreover, whereas the orthosteric agonist EVP-6124 was ineffective, the positive allosteric modulators (PAMs) IVM and GAT-107 reduced lung pathology and anosmia. Thus, IVM operates through a dynamic, context-dependent interplay: P2X4 drives early innate responses in cellular systems including macrophages, whereas the situation in vivo is more complex with 7nAChR being essential for tissue protection. These data position selective PAMs and the CXCL10/CXCL5 axis as promising targets for host-directed therapy in COVID-19.

Reproducibility ledger

Nobody has reported trying to reproduce this paper yet. Did you replicate an experiment or run the code? Your report saves the next lab a week.

Sign in to file a reproducibility report.

Related papers

Similarity Map
SelectedSame fieldCross-field
THISMelo '20Rocha '21Lecuyer '23Guo '20Resende '25Kazungu '26
76% semantic matchImmunology
G. D. de Melo, F. Lazarini, F. Larrous+8 more·bioRxiv·Nov 21, 2020