Molecular imaging of the skeleton: quantitative real-time bioluminescence monitoring gene expression in bone repair and development.
Monitoring gene expression in vivo, noninvasively, is a critical issue in effective gene therapy systems. To date, there are no adequate molecular imaging techniques, which quantitatively monitor gene expression in vivo in skeletal development and repair. The aim of this study was to monitor gene expression in skeletal development and repair, using a real-time molecular imaging system, which quantitatively and noninvasively detects bioluminescence in vivo. Our experimental model consisted of transgenic mice harboring the luciferase marker gene under the regulation of the human osteocalcin (hOC) promoter. A new light detection cooled charge coupled device (CCCD) camera was applied to monitor luciferase expression. In vitro, mesenchymal stem cells (MSCs) isolated from bone marrow of transgenic mice exhibited hOC promoter regulation, detected by luciferase expression that correlated with their osteogenic differentiation. During development from 1 week to 1.5 years, transgenic mice exhibited transgene expression in a wide spectrum of skeletal organs, including calvaria, vertebra, tail, and limbs, reaching a peak at 1 week in most of the skeletal organs. In two skeletal repair models, bone fracture and marrow ablation, the noninvasive CCCD system revealed a peak of luciferase expression at 6 days postsurgery. All quantitative, noninvasive, real-time CCCD measurements correlated with a luciferase biochemical assay and luciferase immunohistochemistry, which demonstrated luciferase expression in hypertrophic chondrocytes and trabecular osteoblasts. Our studies show for the first time (1) the CCCD detection system is a reliable quantitative gene detection tool for the skeleton in vivo, (2) expression of luciferase regulated by the hOC promoter is significantly decreased with age in most skeletal sites, and (3) the dynamics of hOC regulation during mice skeletal development and repair in real time, quantitatively and noninvasively.
Iris
Bar
B
Skeletal Biotech Laboratory, Hebrew University, Hadassah Medical Center, Jerusalem, Israel.
Zilberman
Yoram
Y
Zeira
Eveline
E
Galun
Eithan
E
Honigman
Alik
A
Turgeman
Gadi
G
Clemens
Thomas
T
Gazit
Zulma
Z
Gazit
Dan
D
D000818:Animals; D001483:Base Sequence; D001846:Bone Development; D016723:Bone Remodeling; D017931:DNA Primers; D015870:Gene Expression; D006801:Humans; D008156:Luciferases; D008163:Luminescent Measurements; D051379:Mice; D008822:Mice, Transgenic; D015675:Osteocalcin / Q000235:genetics
D017931:DNA Primers; D015675:Osteocalcin; D008156:Luciferases