title: Ex vivo non-invasive assessment of cell viability and proliferation in bio-engineered whole organ constructs.
abstract: Decellularized organ scaffolds allow whole organ regeneration and study of cell behavior in three-dimensional culture conditions. Cell viability within the bio-engineered organ constructs is an essential parameter reflecting the performance of participating cells during long-term ex vivo culture, and is a prerequisite for further functional performance. Resazurin-based redox metabolic assays have been used to monitor cell viability in both two- and three-dimensional cell cultures. Here we developed a method for monitoring cell viability and proliferation in bio-engineered organ constructs using a resazurin perfusion assay. This method allows non-invasive, repetitive and rapid estimation of viable cell numbers during long-term ex vivo culture. As a proof-of-principle, we assessed the performance of two different endothelial sources and the impact of different perfusion programs on endothelial viability after re-endothelialization of decellularized lung scaffolds. The resazurin-based perfusion assay revealed changes in endothelial viability and proliferation during long-term ex vivo culture, which was consistent with histological assessment at different time points. Finally, we showed that this method could be used for assessment of proliferation and cytotoxicity after pharmacological treatment on a three-dimensional non-small cell lung cancer culture model.
authors.affiliation: Center for Regenerative Medicine, Massachusetts General Hospital, United States; Harvard Medical School, United States.
authors.forename: Xi
authors.initials: X
authors.lastname: Ren
authors.affiliation: Harvard Medical School, United States; Division of Thoracic Surgery, Department of Surgery, Massachusetts General Hospital, United States.
authors.forename: Luis F
authors.initials: LF
authors.lastname: Tapias
authors.affiliation: Center for Regenerative Medicine, Massachusetts General Hospital, United States; Harvard Medical School, United States.
authors.forename: Bernhard J
authors.initials: BJ
authors.lastname: Jank
authors.affiliation: Harvard Medical School, United States; Division of Thoracic Surgery, Department of Surgery, Massachusetts General Hospital, United States.
authors.forename: Douglas J
authors.initials: DJ
authors.lastname: Mathisen
authors.affiliation: Harvard Medical School, United States; Division of Thoracic Surgery, Department of Surgery, Massachusetts General Hospital, United States.
authors.forename: Michael
authors.initials: M
authors.lastname: Lanuti
authors.affiliation: Center for Regenerative Medicine, Massachusetts General Hospital, United States; Harvard Medical School, United States; Division of Thoracic Surgery, Department of Surgery, Massachusetts General Hospital, United States; Harvard Stem Cell Institute, United States. Electronic address: hott@mgh.harvard.edu.
authors.forename: Harald C
authors.initials: HC
authors.lastname: Ott
mesh_terms: D000818:Animals; D057005:Bioengineering / Q000379:methods; D002289:Carcinoma, Non-Small-Cell Lung / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D066026:Endothelial Progenitor Cells / Q000166:cytology; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D007202:Indicators and Reagents; D008168:Lung / Q000737:chemistry / Q000166:cytology* / Q000473:pathology / Q000502:physiology*; D008175:Lung Neoplasms / Q000473:pathology; D008297:Male; D009924:Organ Culture Techniques / Q000379:methods*; D010078:Oxazines; D010477:Perfusion / Q000379:methods; D017207:Rats, Sprague-Dawley; D012038:Regeneration; D023822:Tissue Engineering / Q000379:methods; D054457:Tissue Scaffolds / Q000737:chemistry*; D014966:Xanthenes
chemical_list: D007202:Indicators and Reagents; D010078:Oxazines; D014966:Xanthenes; C005843:resazurin
keywords: Decellularization; Organ regeneration; Perfusion; Resazurin; Tissue engineering; Viability
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