Optimized gene transduction in human lung organoids: A high-efficiency method for advanced research applications
Human induced pluripotent stem cell (iPSC)-derived lung organoids, engineered to carry targeted genes, offer a robust platform for investigating mechanistic insights in lung research. Although lentiviral vectors (LVVs) are highly effective for stable expression due to their integrative properties, achieving efficient transduction in human iPSC-derived lung organoids poses a significant technical challenge, likely due to the complex structure of these organoids. In this study, we optimized a method to enhance LVV transduction efficiency by physically disrupting the organoids to increase surface area, followed by spinoculation to apply shear force during cell dissociation. This approach, combined with the use of an optimized culture medium, significantly improved transduction efficiency. The success of this method was validated at both the gene and protein levels using single-cell RNA sequencing (scRNA-seq) and various cellular and molecular assays. Our optimized transduction protocol may provide a valuable tool for investigating specific cellular and molecular mechanisms in development and disease models using human iPSCs-derived lung organoids.
Authors
- Ori D. Rotstein (ORCID: https://orcid.org/0009-0004-4388-5352)
- Giovanna Pellecchia (ORCID: https://orcid.org/0000-0003-4747-3473)
- Yuchong Li (ORCID: https://orcid.org/0000-0003-2462-3109)
- Jady Liang (ORCID: https://orcid.org/0000-0001-9228-4407)
- Bhooma Thiruvahindrapuram (ORCID: https://orcid.org/0000-0003-1128-008X)
- Jasmin Khateeb (ORCID: https://orcid.org/0000-0002-4575-2199)
- Arthur S. Slutsky (ORCID: https://orcid.org/0000-0002-6063-3876)
- Julie Khang
- Thenuka Thanabalasingam
- Haibo Zhang (ORCID: https://orcid.org/0000-0002-1714-3038)
- Ya‐Wen Chen (ORCID: https://orcid.org/0000-0003-2382-8444)
- Mirjana Jerkic
Institutions
- St. Michael's Hospital (CA)
- Technion – Israel Institute of Technology (IL)
- University of Toronto (CA)
- Muscular Dystrophy Canada (CA)
- Rambam Health Care Campus (IL)
- Hospital for Sick Children (CA)
- St Michael’s Hospital (IE)
- First Affiliated Hospital of Guangzhou Medical University (CN)
- Rappaport Family Institute for Research in the Medical Sciences (IL)
- Public Health Ontario (CA)
- SickKids Foundation (CA)
- Ontario Tobacco Research Unit (CA)
- State Key Laboratory of Respiratory Disease (CN)
- Unity Health Toronto
- Guangzhou Medical University (CN)
- Icahn School of Medicine at Mount Sinai (US)
Publication Details
- Journal
- Communications Biology
- Published
- 2025-02-03
- DOI
- https://doi.org/10.1038/s42003-025-07461-w
- Citations
- 5
- Primary Topic
- Energy Harvesting in Wireless Networks
- Type
- article
- Field-Weighted Citation Impact
- 3.65
Funders
- University of Toronto
- Canadian Institutes of Health Research