Abstract
Recently, ex vivo three-dimensional organ culture systems have emerged to study the physiology and pathophysiology of human organs. These systems also have potential as a translational tool in tissue engineering; however, this potential is limited by our ability to longitudinally monitor the fate and action of cells used in regenerative therapies. Therefore, we investigated luciferase-mediated bioluminescence imaging (BLI) as a non-invasive technique to continuously monitor cellular behavior in ex vivo whole organ culture. Goat adipose-derived stem cells (gADSCs) were transduced with either Firefly luciferase (Fluc) or Gaussia luciferase (Gluc) reporter genes and injected in isolated goat intervertebral discs (IVD). Luciferase activity was monitored by BLI for at least seven days of culture. Additionally, possible confounders specific to avascular organ culture were investigated. Gluc imaging proved to be more suitable compared to Fluc in monitoring gADSCs in goat IVDs. We conclude that BLI is a promising tool to monitor spatial and temporal cellular behavior in ex vivo organ culture. Hence, ex vivo organ culture systems allow pre-screening and pre-validation of novel therapeutic concepts prior to in vivo large animal experimentation. Thereby, organ culture systems can reduce animal use, and improve the speed of innovation by overcoming technological, ethical and financial challenges.
Highlights
Non-invasive imaging technique which allows real-time in vivo monitoring of location and proliferation of luciferase-expressing cells[8,9]
The aim of the current study has been to evaluate the use of luciferase-mediated bioluminescence imaging for longitudinal monitoring of adipose derived mesenchymal stem cells injected in large mammal intervertebral discs (IVD), cultured in an ex vivo organ culture system
Expression of the fluorescent proteins in the U87 cells was increased compared to the goat adipose derived stem cells (gADSCs); longer exposure times were needed for a clear visualization of Cyan Fluorescent Protein (CFP) and mCherry in the gADSCs
Summary
Non-invasive imaging technique which allows real-time in vivo monitoring of location and proliferation of luciferase-expressing cells[8,9]. BLI has been shown to be highly correlated to the above mentioned cell viability assays (for both Fluc and Gluc) and is a well-established method for cell viability monitoring[10,11] These advantages highlight the potential of BLI for longitudinal evaluation of cellular therapies in ex vivo tissue culture[12]. We developed an ex vivo loaded disc organ culture system (LDCS), to study the physiology of intervertebral discs (IVD) and their response to mechanical loads[4,19] This system allows us to investigate the process of intervertebral disc degeneration, identified as an important etiological factor of low back pain[20,21]. The aim of the current study has been to evaluate the use of luciferase-mediated bioluminescence imaging for longitudinal monitoring of adipose derived mesenchymal stem cells injected in large mammal IVDs, cultured in an ex vivo organ culture system. Possible confounders specific to avascular organ culture were investigated
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