Preconditioning allows engraftment of mouse and human embryonic lung cells, enabling lung repair in mice

Abstract

Repair of injured lungs represents a longstanding therapeutic challenge. We show that human and mouse embryonic lung tissue from the canalicular stage of development (20–22 weeks of gestation for humans, and embryonic day 15–16 (E15–E16) for mouse) are enriched with progenitors residing in distinct niches. On the basis of the marked analogy to progenitor niches in bone marrow (BM), we attempted strategies similar to BM transplantation, employing sublethal radiation to vacate lung progenitor niches and to reduce stem cell competition. Intravenous infusion of a single cell suspension of canalicular lung tissue from GFP-marked mice or human fetal donors into naphthalene-injured and irradiated syngeneic or SCID mice, respectively, induced marked long-term lung chimerism. Donor type structures or 'patches' contained epithelial, mesenchymal and endothelial cells. Transplantation of differentially labeled E16 mouse lung cells indicated that these patches were probably of clonal origin from the donor. Recipients of the single cell suspension transplant exhibited marked improvement in lung compliance and tissue damping reflecting the energy dissipation in the lung tissues. Our study provides proof of concept for lung reconstitution by canalicular-stage human lung cells after preconditioning of the pulmonary niche.

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Figure 1: Histological comparison between embryonic lung implant and adult human lung, and comparison of conditioning protocols.
Figure 2: Engraftment of mouse GFP+ embryonic precursor lung cells following different conditioning protocols.
Figure 3: Morphometric analysis of the lung area occupied by the engrafted patches and their clonal origin.
Figure 4: Integration of donor-derived cells in epithelial lung compartment and functional repair of injured lungs.
Figure 5: Colony-forming efficiency and donor type lung chimerism after fractionation of E16 mouse lung cells.
Figure 6: Lung chimerism in NOD-SCID mice after transplantation of human embryonic lung (HEL) cells.

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Acknowledgements

The authors thank Z. Kam (Weizmann Institute) and O. Golani (Weizmann Institute) for excellent assistance with the colocalization analysis and R. Rotkopf (Weizmann Institute) for his critical support with the statistical analyses. This study was supported in part by a grant from the Israel Science Foundation (Legacy Bio-Med, 71035803 to Y.R. and M.W.) and by a research grant from Roberto and Renata Ruhman (Y.R.).

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Contributions

C.R. designed, performed and organized most of the experiments; analyzed and interpreted the data; and cowrote the manuscript. E.S. analyzed and interpreted the data and participated in discussions. A.A. and Y.Z.K. assisted in performing experiments and analyzed the data. Y.Y. participated in data analysis and discussions. M.A. performed, analyzed and interpreted lung function tests. I.E.B. performed and analyzed micro-CT studies. O.T. and G.S. performed and analyzed two-photon microscopy studies. H.B.-H., D.S., Z.V., O.S. and S.E. recruited the individuals scheduled for termination of pregnancy and obtained agreement and written informed consent to donate the fetal tissues for the studies. E.F., D.S. and M.W. participated in data analysis and discussions. N.B. performed, analyzed and interpreted lung function tests. W.E.F. and D.H. participated in data evaluation and critical reading of the manuscript. C.H.-K., I.M.K. and E.B.-L. assisted in experiments. Y.R. designed, coordinated and conducted the study, including analysis and interpretation of data, and cowrote the manuscript.

Corresponding author

Correspondence to Yair Reisner.

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Y.R. holds shares and serves as a consultant to Cell Source, Inc., which has acquired an exclusive option to license intellectual property arising from this publication.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8 & Supplementary Table 1 (PDF 34454 kb)

Progenitor cell niches in 20 week HEL, assessed by spinning-disc confocal microscopy.

Staining of thick slice for CK5 (red), nestin (green), blood vessels (blue). (MOV 7423 kb)

Chimeric lung 6 weeks after transplantation.

(a) Real-time evaluation of the donor-derived endogenous GFP (green) and host autofluorescence (blue). (b) 3D reconstructed structure of the same chimeric lung region 6 weeks after transplantation. Donor-derived endogenous GFP (green) and host autofluorescence (blue). (MOV 17872 kb)

Real-time chimerism of vascular and parenchymal compartments after i.v. injection of red quantum dots (QD 655), assessed by 2-photon microscopy.

Donor-derived endogenous GFP (green) and blood vessels (red). (MOV 3483 kb)

Chimeric lung 16 weeks after transplantation, assessed by two-photon microscopy.

Donor-derived endogenous GFP (green) and host autofluorescence (blue). (MOV 2285 kb)

Monochromic red and green patches in the chimeric lung, assessed by two-photon microscopy.

Donor-derived endogenous GFP (green) and endogenous tdTomato (red) and host autofluorescence (blue). (MOV 8087 kb)

Supplementary Video 6

Transplantation of E16 MEL cells from GFP donors into TdTomato recipients. Stained slice assessed by spinning disc confocal microscopy. Donor-derived GFP (green) stained with anti-GFP antibody and endogenous tdTomato of host (red). (MOV 7467 kb)

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Rosen, C., Shezen, E., Aronovich, A. et al. Preconditioning allows engraftment of mouse and human embryonic lung cells, enabling lung repair in mice. Nat Med 21, 869–879 (2015). https://doi.org/10.1038/nm.3889

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