Intravital imaging of DSS-induced cecal mucosal damage in GFP-transgenic mice using two-photon microscopy.
BACKGROUND
Two-photon laser-scanning microscopy (TPLSM) is a powerful diagnostic tool for real-time, high-resolution structural imaging. However, obtaining high-quality in vivo TPLSM images of intra-abdominal organs remains technically challenging.


METHODS
An organ-stabilizing system was applied to high-quality TPLSM imaging. Real-time imaging of visceral organs, such as the liver, spleen, kidney and intestine, of transgenic green fluorescent protein (GFP) mice was performed in vivo using TPLSM. The bacterial translocation model using dextran sodium sulfate (DSS)-induced colitis was also investigated in prepared GFP mice following simple surgery. This allowed the capture of morphological real images using in vivo TPLSM. Immunohistochemical analysis of ZO-1 was performed to support the morphological findings of TPLSM.


CONCLUSIONS
We established an organ-stabilizing system to evaluate the real-time imaging of visceral organs in actin-GFP transgenic mice using in vivo TPLSM. DSS-induced colitis showed irregularity of crypt architecture, disappearance of crypts, inflammatory cell infiltration and increased rolling of white blood cells along the vasculature. In addition, the intercellular distance of mucosal cells in the crypt and vascular endothelial cells in the intestinal wall was increased in the intestinal mucosa during DSS colitis. In DSS colitis, there was remarkable loss of mucosal and vascular endothelial ZO-1 expression, as could be seen by a decrease in ZO-1 staining. In conclusion, our observations suggested the possibility that our TPLSM imaging system can be used to clarify the pathophysiological changes in various diseases using longitudinal studies of microscopic changes in the same animal over long periods of time.
Toiyama
Yuji
Y
Department of Gastrointestinal and Pediatric Surgery, Division of Reparative Medicine, Institute of Life Sciences, Mie University Graduate School of Medicine, Tsu, Mie, Japan. ytoi0725@clin.medic.mie-u.ac.jp
Mizoguchi
Akira
A
Okugawa
Yoshinaga
Y
Koike
Yuhki
Y
Morimoto
Yuhki
Y
Araki
Toshimitsu
T
Uchida
Keiichi
K
Tanaka
Koji
K
Nakashima
Hisako
H
Hibi
Mayumi
M
Kimura
Kazushi
K
Inoue
Yasuhiro
Y
Miki
Chikao
C
Kusunoki
Masato
M
D000818:Animals; D018988:Bacterial Translocation / Q000187:drug effects / Q000502:physiology*; D002432:Cecum / Q000187:drug effects / Q000473:pathology*; D003092:Colitis / Q000139:chemically induced / Q000473:pathology*; D016264:Dextran Sulfate; D004926:Escherichia coli / Q000502:physiology*; D005240:Feasibility Studies; D049452:Green Fluorescent Proteins; D007413:Intestinal Mucosa / Q000187:drug effects / Q000473:pathology*; D007668:Kidney / Q000382:microbiology / Q000473:pathology; D008099:Liver / Q000382:microbiology / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D018613:Microscopy, Confocal / Q000379:methods*; D013154:Spleen / Q000382:microbiology / Q000473:pathology
D049452:Green Fluorescent Proteins; D016264:Dextran Sulfate