Speaker
Description
Introduction
Patient specific, tissue engineered oesophagi represent a possible solution for conditions which currently lack effective therapeutic options, such as long-gap oesophageal atresia1. Histology, the current gold standard for tissue engineering (TE) protocol design, is destructive and 2D, disregarding volumetric information which are essential for confirming the regeneration of the appropriate organ architecture and subsequently function. X-ray phase contrast computed tomography (PC-CT), can generate novel volumetric density maps of organ architecture in a non-destructive manner2,3.
Methods
Laboratory-based edge-illumination PC-CT2 was integrated in an oesophageal TE protocol, imaging 50 piglet derived specimens. This included specimens of type: native (n=20), as extracted from the piglet, scaffold (n=20), native-derived cell-free constructs1, and tissue engineered (n=10), recellularised - in-vitro matured cell-seeded scaffolds4. A semi-automated, machine learning-based, image processing pipeline was developed for perfuming specimen characterisation5. This entailed 1) the volumetric visualisation and 2) quantitative, intra- and inter-sample comparison, based on oesophageal physical density and morphology. A new, comprehensive specimen evaluation was made possible by combing 1) and 2).
Results/Discussion
All oesophageal layers, classically visualised in 2D using histology, are assessed in 3D in the native oesophagi, their preservation is confirmed in the scaffold specimens, whilst an evident loss in layer architecture is observed in the cell-repopulated portion of the recellularised samples. One of the developed tools, the “radial density profile”, extracts density variation within PC-CT slices, and allows for assessing quantitatively cell-distribution throughout the volume of a recellularised sample in a non-destructive manner. Further to guiding the progression of the existing TE protocol, the imaging tools and developed pipeline demonstrated to be compatible with scale-up models, from piglet to pig derived oesophagi. PC-CT will play a fundamental role in in-vivo testing stages, as its non-destructive nature will eliminate the need for extracting the engineering construct for assessment, and will enable the evaluation of construct integration with the host.
Conclusions
The comprehensive construct evaluation, delivered by PC-CT in a non-destructive manner can unlock the great potential of the field of TE, ultimately allowing for the successful regeneration of organs with complex architecture and function.
Acknowledgements
Funding contributions for this work include: the National Research Facility for lab X-ray CT (NXCT) through EPSRC (EP/T02593X/1), the Engineering and Physical Sciences Research Council (EPSRC-EP/T005408/1), the Great Ormond Street Hospital for Children, the NHS Foundation Trust. A.U, the NIHR (NIHR-RP-2014-04-046), the Horizon 2020 grant INTENS 668294 and the OAK Foundation (W1095/OCAY-14-191). Author fundings include: the EPSRC (EP/L016478/1), which supported SS & AA. The Royal Academy of Engineering under their Chairs in Emerging Technologies scheme (CiET1819/2/78) supports AO. The Royal Academy of Engineering under the Research Fellowship Scheme, supports ME and CKH. MFMG is supported by The European Commission H2020 Marie Skłowdoska-Curie Action Individual Fellowship (843265 AmnioticID).
References
[1] Totonelli, G. et al. Surg. Int. 29, 87–95 (2013)
[2] Hagen, C. K. et al. Sci. Rep. 5, 18156 (2015)
[3] Hagen, C. K. et al. Opt. Express 22, 7989 (2014)
[4] Urbani, L. et al. Nat. Commun. 9, (2018)
[5] Savvidis, S. et al. Acta Biomaterialia (Accepted, January 2022)
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