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Blood–brain barrier injury and neuroinflammation induced by SARS-CoV-2 in a lung–brain microphysiological system

Nature Biomedical Engineering (2023)Cite this article

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An Author Correction to this article was published on 31 October 2023

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Abstract

In some patients, COVID-19 can trigger neurological symptoms with unclear pathogenesis. Here we describe a microphysiological system integrating alveolus and blood–brain barrier (BBB) tissue chips that recapitulates neuropathogenesis associated with infection by SARS-CoV-2. Direct exposure of the BBB chip to SARS-CoV-2 caused mild changes to the BBB, and infusion of medium from the infected alveolus chip led to more severe injuries on the BBB chip, including endothelial dysfunction, pericyte detachment and neuroinflammation. Transcriptomic analyses indicated downregulated expression of the actin cytoskeleton in brain endothelium and upregulated expression of inflammatory genes in glial cells. We also observed early cerebral microvascular damage following lung infection with a low viral load in the brains of transgenic mice expressing human angiotensin-converting enzyme 2. Our findings suggest that systemic inflammation is probably contributing to neuropathogenesis following SARS-CoV-2 infection, and that direct viral neural invasion might not be a prerequisite for this neuropathogenesis. Lung–brain microphysiological systems should aid the further understanding of the systemic effects and neurological complications of viral infection.

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Fig. 1: An integrated lung–brain MPS to understand the neuropathogenesis of viral infection.
Fig. 2: Direct exposure of SARS-CoV-2 on the individual BBB chip.
Fig. 3: Indirect impact of SARS-CoV-2 on the BBB after lung infection on the lung–brain MPS.
Fig. 4: Transcriptional analysis of brain endothelial cells and glial cells on the lung–brain MPS.
Fig. 5: Cerebral microvascular injury in SARS-CoV-2-infected mouse models.

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Data availability

The main data supporting the results in this study are available within the paper and its Supplementary Information. The raw RNA-seq data are available from the Sequence Read Archive via the accession number PRJNA764053. All data generated or analysed during the study are available from the corresponding authors on reasonable request. Source data are provided with this paper.

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Acknowledgements

We thank A. P. Xiang (Sun Yat-Sen University) and W. Li (Sun Yat-Sen University) for kindly providing the hPSC-PCs. We thank X. Cheng (Shanghai Institute of Biochemistry and Cell Biology, CAS) for kindly providing the lentivirus–GFP vector. We thank J. Han (Kunming Institute of Zoology, CAS) for helping with blood drawing. This research was supported by the National Key R&D Program of China (number 2022YFA1104700), Strategic Priority Research Program of the CAS (number XDB29050301), National Nature Science Foundation of China (numbers 32101163 and 31971373) and Yunnan Key Research and Development Program (number 202003AD150009).

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Author notes

  1. These authors contributed equally: Peng Wang, Lin Jin.

Authors and Affiliations

  1. Division of Biotechnology, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China

    Peng Wang, Min Zhang, Yunsong Wu, Yaqiong Guo, Wenwen Chen, Yaqing Wang & Jianhua Qin

  2. University of Science and Technology of China, Hefei, China

    Peng Wang & Jianhua Qin

  3. Suzhou Institute for Advanced Research, University of Science and Technology of China, Suzhou, China

    Peng Wang & Jianhua Qin

  4. Key Laboratory of Animal Models and Human Disease Mechanisms of the Chinese Academy of Sciences–Key Laboratory of Bioactive Peptides of Yunnan Province, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China

    Lin Jin, Zilei Duan, Chaoming Wang, Zhiyi Liao & Ren Lai

  5. University of Chinese Academy of Sciences, Beijing, China

    Min Zhang, Yunsong Wu & Jianhua Qin

  6. Core Technology Facility of Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China

    Yingqi Guo

  7. Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA, USA

    Luke P. Lee

  8. Department of Bioengineering, Department of Electrical Engineering and Computer Science, University of California, Berkeley, Berkeley, CA, USA

    Luke P. Lee

  9. Institute of Quantum Biophysics, Department of Biophysics, Sungkyunkwan University, Suwon, Korea

    Luke P. Lee

  10. Beijing Institute for Stem Cell and Regenerative Medicine, Chinese Academy of Sciences, Beijing, China

    Jianhua Qin

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Contributions

J.Q. and P.W. conceived the study. P.W. and L.J. performed the experiments and analysed results. L.J., Z.D., C.W. and Z.L. performed the SARS-CoV-2 infection in the BSL-3 laboratory. M.Z., Y. Wu, W.C., Yaqiong Guo and Y. Wang performed chip fabrication. Yingqi Guo prepared the electron microscopy samples. P.W., J.Q., L.P.L. and R.L. wrote and revised the paper.

Corresponding authors

Correspondence to Ren Lai, Luke P. Lee or Jianhua Qin.

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Nature Biomedical Engineering thanks Jacquelyn A. Brown, Seung-Woo Cho, John Wikswo and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Extended data

Extended Data Fig. 1 SARS-CoV-2 infection caused severe alveolar injury and inflammatory responses on the alveolus chip.

a, Confocal micrographs of alveolar epithelial cells immunostained for E-cadherin (green) and SFTPC (red) on Day 4 following SARS-CoV-2 infection (n = 3). b, Confocal micrographs of pulmonary microvascular endothelial cells immunostained for VE-cadherin (green) on Day 4 following SARS-CoV-2 infection (n = 3). c, Confocal micrographs of alveolar epithelial cells and pulmonary microvascular endothelial cells immunostained for Ki67 (red) on Day 4 following SARS-CoV-2 infection (n = 4). d, Quantification of Ki67+ cells on the mock- and SARS-CoV-2-infected chips based on c. Data are presented as the mean ± SEM and were analyzed using an unpaired Student’s t-test (***: P < 0.001). e, Multiplex cytokine assays showing 13 cytokine concentrations in culture supernatants from the upper epithelial channel (-U) and lower vascular channel (-L) of mock-, or SARS-CoV-2-infected alveolus chips on Day 4 post-infection (n = 3). Data are presented as the mean ± SEM and were analyzed using an unpaired Student’s t-test (***: P < 0.001).

Extended Data Fig. 2 Characterizing immune responses of glial cells and immune cells on the lung–brain MPS.

a, Confocal micrographs of brain endothelial cells immunostained for VE-cadherin and ZO-1 without astrocytes (Δ astrocytes) or without microglia (Δ microglia) following conditioned medium treatment on Day 4 (n = 4). b, Quantification of endothelial cell density based on a. Data are presented as the mean ± SEM and were analyzed using a one-way ANOVA with the Bonferroni post hoc test. c, Confocal micrographs of microglia immunostained for CD206 and F4/80 following conditioned medium treatment on Day 4 (n = 3). d, Quantification of CD206 and F4/80 fluorescence intensity for control and SARS-CoV-2 groups based on c. Data are presented as the mean ± SEM and were analyzed using an unpaired Student’s t-test (***: P < 0.001). e, Fluorescent micrographs of PBMCs (pre-stained with Cell Tracker Red dye) attached to the endothelium following conditioned medium treatment on Day 4 (n = 4). f, Quantification of the density of attached PBMCs based on e. Data are presented as the mean ± SEM and were analyzed using an unpaired Student’s t-test.

Extended Data Fig. 3 SARS-CoV-2 caused injuries of brain pericytes on the lung–brain MPS indirectly.

a, Cell morphology and expression of pericyte markers (NG2, CD13 and PDGFRβ) of hPSC-PCs were revealed by bright-field imaging and immunostaining imaging, respectively (n = 3). b, Schematic description of the BBB chip by co-culturing brain endothelial cells, astrocytes, pericytes and microglia. c, A side view of confocal immunofluorescent image showing BBB interface, which was formed by co-culture of human endothelial cells (ZO-1 staining, red), astrocytes (GFAP staining, red) and hPSC-PCs (labeled by GFP, green) under fluid flow conditions for 3 days. The white dotted line indicates the porous membrane. d, Confocal micrographs showing the co-culture of human astrocytes (GFAP staining, red) and hPSC-PCs (labeled by GFP) on the BBB chip (n = 3). e, Confocal micrographs of hPSC-PCs (labeled by GFP) immunostained for viral Spike (red) following conditioned medium treatment (n = 4). f, Quantification of the ratio of hPSC-PCs (green) coverage based on e. Data are presented as the mean ± SEM and were analyzed using an unpaired Student’s t-test. g, Quantification of the hPSC-PCs density based on e. Data are presented as the mean ± SEM and were analyzed using an unpaired Student’s t-test.

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Wang, P., Jin, L., Zhang, M. et al. Blood–brain barrier injury and neuroinflammation induced by SARS-CoV-2 in a lung–brain microphysiological system. Nat. Biomed. Eng (2023). https://doi.org/10.1038/s41551-023-01054-w

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