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Next Generation Sequencing of SARS-CoV-2 infected lung epithelial cells derived from iPSC

ABSTRACT: We present evidence on how a lung epithelial cell culture system based on human induced pluripotent stem cells (hiPSC) could emerge as a relevant and sensitive platform for modelling SARS-CoV-2 infection. In order to develop a human model system, we differentiated hiPSC into lung epithelial lineage (airway and alveolar cells) through a three-step protocol. De novo generated cells were authenticated by spatiotemporal expression of stage specific markers including SOX2, FOXJ1 and CC10 in proximal cells and SOX9, FOXP2, SFTPC, MUC5AC, and CC10 in distal cells. Further we showed upregulation of ACE2 and TMPRSS2, the two proteins responsible for SARS-CoV-2 binding and invasion into the host lung in our iPSC-derived proximal and distal lung epithelial cells. In order to find out whether our in-house created lung cell culture system was permissive to SARS-CoV-2 infection, two viral doses representing two multiplicities of infections (MOIs) was tested. Plaque assay, RT-PCR, and ICC results demonstrated that the virus was able to enter and replicate inside these cells in a reproducible manner. Further, transcriptomics (RNA seq) data uncovered a robust epithelial cell-specific response to virus infection including perturbation of metabolic processes, disruption in alveolar and airway maturation program, in addition to the well-known upregulation of antiviral defense and immune responses.

ORGANISM(S): Homo sapiens

PROVIDER: GSE190193 | GEO | 2022/03/07

REPOSITORIES: GEO

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Project description:The COVID-19 pandemic remains a global health crisis, yet, the immunopathological mechanisms driving the development of severe disease remain poorly defined. Here, we utilize a rhesus macaque (RM) model of SARS-CoV-2 infection to delineate perturbations in the innate immune system during acute infection using an integrated systems analysis. We found that SARS-CoV-2 initiated a rapid infiltration (two days post infection) of plasmacytoid dendritic cells into the lower airway, commensurate with IFNA production, natural killer cell activation, and induction of interferon-stimulated genes. At this early interval, we also observed a significant increase of blood CD14-CD16+ monocytes. To dissect the contribution of lung myeloid subsets to airway inflammation, we generated a novel compendium of RM-specific lung macrophage gene expression using a combination of sc-RNA-Seq data and bulk RNA-Seq of purified populations under steady state conditions. Using these tools, we generated a longitudinal sc-RNA-seq dataset of airway cells in SARS-CoV-2-infected RMs. We identified that SARS-CoV-2 infection elicited a rapid recruitment of two subsets of macrophages into the airway: a C206+MRC1- population resembling murine interstitial macrophages, and a TREM2+ population consistent with CCR2+ infiltrating monocytes, into the alveolar space. These subsets were the predominant source of inflammatory cytokines, accounting for ~75% of IL6 and TNF production, and >90% of IL10 production, whereas the contribution of CD206+MRC+ alveolar macrophages was significantly lower. Treatment of SARS-CoV-2 infected RMs with baricitinib (Olumiant®), a novel JAK1/2 inhibitor that recently received Emergency Use Authorization for the treatment of hospitalized COVID-19 patients, was remarkably effective in eliminating the influx of infiltrating, non-alveolar macrophages in the alveolar space, with a concomitant reduction of inflammatory cytokines. This study has delineated the major subsets of lung macrophages driving inflammatory and anti-inflammatory cytokine production within the alveolar space during SARS-CoV-2 infection.

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Next Generation Sequencing of SARS-CoV-2 infected lung epithelial cells derived from iPSC

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