In vivo MR Imaging
MR imaging was performed on a 9.4T Bruker horizontal bore scanner (Billerica,
MA) equipped with a home-built RF transmit and receive 3 x 4-cm elliptical surface coil
and using ParaVision 3.0 Software. T1 maps were acquired using a RARE inversion
recovery sequence with the following parameters: TE = 7.253ms, TR = 10,000ms, TI =
0.001, 200, 400, 800, 1600, 3200, and 6400ms. FOV = 25.6 x 25.6mm2, spatial resolution
= 0.2 x 0.2mm.pixel-1, matrix size = 128 x 128, slice thickness = 1mm, and a total
imaging time of 16min 5sec.
For quantitative analysis of T1 relaxation, T1 color-coded maps were constructed
using Marevisi 3.5 software (Institute for Biodiagnostics, National Research Council,
Canada). The pancreas was manually segmented and subjected to region-of-interest
(ROI) analysis for the determination of pancreas-associated T1 relaxation times, as
previously described (1).
At the same time points, T1 weighted images of the same mice were obtained
using an MSME sequence with the following parameters: TE = 7.841ms, TR = 100ms,
FOV = 25.6 x 25.6mm2, spatial resolution = 0.2 x 0.2mm.pixel-1, matrix size = 128 x
128, slice thickness = 1mm, and a total imaging time of 3min 24sec.
In addition, 3D-angiograms were obtained using a gradient echo imaging sequence
comprising first-order flow compensation (GEFC). The following imaging parameters
were utilized: TE = 7.16ms, TR = 40ms, FOV = 51.2 x 25.6 x 25.6mm3, spatial resolution
= 0.4 x 0.4 x 0.4mm.pixel-1, matrix size = 128 x 64 x 64, slice thickness = 25.6mm, and a
total imaging time of 2min 46sec.
Finally, high-resolution angiograms were obtained at 1 and 40 hrs after injection
of PGC-GdDTPA-F with the following imaging parameters: TE = 3.016ms, TR = 20ms,
FOV = 19.2 x 25.6 x 12.8mm3, Spatial resolution = 0.1 x 0.1 x 0.1mm.pixel-1, matrix size
= 192 x 256 x 128, slice thickness = 12.8mm, and a total imaging time of 43min 41sec
440ms. Image reconstruction was performed using a maximum intensity projection (MIP)
algorithm.
PGC-GdDTPA-F localization in the pancreas by histology.
PGC-GdDTPA-F was synthesized to carry a fluorescent label (FITC). Pancreata
from experimental and control animals were removed immediately after imaging, snap
frozen, and cut into 5 µm sections. Tissue sections were fixed for 20 min in ice-cold
acetone, washed with PBS and mounted with a DAPI ((4’,6 diamidino-2-phenylindole))containing mounting medium (Vectashield, Vector Laboratories, Burlingame, CA) for
nuclear staining. Additional tissue sections were blocked in a solution of 10% goat serum
in PBS and stained for insulin. Sections were incubated with a rabbit anti-mouse insulin
primary antibody (1: 25 dilution, Santa Cruz Biotechnology, Santa Cruz, CA), followed
by a texas red-labeled goat anti-rabbit secondary IgG (1:100 dilution, Santa Cruz
Biotechnology, Santa Cruz, CA) and mounted with a DAPI mounting medium.
Fluorescence microscopy was performed in three channels (FITC for PGC-GdDTPA-F,
Texas red for insulin, and DAPI for nucleus) using a Nikon Eclipse 50i fluorescence
microscope equipped with an appropriate filter set (Chroma Technology Corporation,
Rockingham, VT). Images were acquired using a CCD camera with near infrared
sensitivity (SPOT 7.4 Slider RTKE; Diagnostic Instruments, Sterling Heights, MI) and
analyzed using SPOT 4.0 advanced version software (Diagnostic Instruments, Sterling
Heights, MI).
Consecutive sections were stained with hematoxylin & eosin (H&E) and analyzed
by light microscopy.
1. Grimm J, Potthast A, Wunder A, Moore A: Magnetic resonance imaging of the
pancreas and pancreatic tumors in a mouse orthotopic model of human cancer. Int J
Cancer 106:806-811, 2003