62.09 Using Bioluminescent Imaging for In-Vivo Monitoring of Implanted Human Intestinal Organoids

P. Obidike¹, C. Moneme¹, V. Ives De Silva², S. Moore², Y. Zhang¹, L.S. Cheng^{1  1}University of Virginia, Department Of Surgery, Charlottesville, VIRGINIA, USA ²University Of Cincinnati, Division Of Pediatric Gastroenterology, Cincinnati, OH, USA

Introduction:
Human intestinal organoids (HIOs) have emerged as a useful model to study the gastrointestinal tract. However, monitoring their successful implantation and activity in vivo thus far has required non-survival surgery. Here, we present a noninvasive approach to monitor the growth of HIOs implanted under the kidney capsule of mice using bioluminescent imaging.

Methods:
Human embryonic stem cell (hESC) H9 line was transduced with IVISbrite red F-luc-Puromycin lentiviral particles (RediFect) at a concentration of MOI=1. Cells were differentiated into mid/hindgut endodermal spheroids and matured into intestinal organoids according to previously published protocols. Maturation of spheroids was confirmed using immunofluorescent staining for intestinal cell types and the stability of the luciferase expression was confirmed by monitoring the HIOs in a luminometer for 5 days. HIOs were then surgically implanted under the left kidney capsule of 5 female and 5 male mice. Noninvasive imaging was performed weekly using an in vivo imaging system (IVIS). Before imaging, 150uL of IVISbrite D-Lucerfin, RediJect (30mg/ml) solution was administered via intraperitoneal injection. The bioluminescent signal was measured and quantified using the IVIS spectrum region of interest (ROI) tool and color scale.

Results:
Nine of 10 mice had bioluminescent signals at the implantation site two weeks after implantation. The region of interest per mouse varied weekly. The fold change in average radiance over time in female mice was notably greater than in male mice (2.0×10⁷ vs 1.0×10⁶, respectively). 

Conclusion:

The growth of HIOs implanted under the kidney capsule of mice can be monitored noninvasively using bioluminescent IVIS imaging. Further study may uncover sex differences in the rate of HIO growth in vivo.