Fluorescence molecular tomography enables in vivo visualization and quantification of nonunion fracture repair induced by genetically engineered mesenchymal stem cells

Abstract

Fluorescence molecular tomography (FMT) is a novel tomographic near-infrared (NIR) imaging modality that enables 3D quantitative determination of fluorochrome distribution in tissues of live small animals at any depth. This study demonstrates a noninvasive, quantitative method of monitoring engineered bone remodeling via FMT. Murine mesenchymal stem cells overexpressing the osteogenic gene BMP2 (mMSCs-BMP2) were implanted into the thigh muscle and into a radial nonunion bone defect model in C3H/HeN mice. Real-time imaging of bone formation was performed following systemic administration of the fluorescent bisphosphonate imaging agent OsteoSense TM , an hydroxyapatite-directed bone-imaging probe. The mice underwent imaging on days 7, 14, and 21 postimplantation. New bone formation at the implantation sites was quantified using microcomputed tomography (micro-CT) imaging. A higher fluorescent signal occurred at the site of the mMSC-BMP2 implants than that found in controls. Micro-CT imaging revealed a mass of mature bone formed in the implantation sites on day 21, a finding also confirmed by histology. These findings highlight the effectiveness of FMT as a functional platform for molecular imaging in the field of bone regeneration and tissue engineering. ß

Near-infrared fluorescent probe traces bisphosphonate delivery and retention in vivo

“…FRFP has been used to visualize osteogenic labeling in bone-defect models, with high fluorescence concentrating in areas of high osteoblastic activity. (24,26) These studies corroborate earlier studies using radiolabeled bisphosphonates to demonstrate high drug delivery to sites of active bone regeneration during fracture repair (66) and histologic studies demonstrating unremodeled cartilage retained in fracture callus treated with bisphosphonate. (67) Similarly, FRFP, when injected into mice with osteolytic lesions resulting from tumor xenograft injections, localizes under osteoclasts at the tumor-bone interface.…”

“…Fluorescence molecular tomography (FMT) is less dependent on depth of signal than fluorescence reflectance imaging (69,70) and has been used to visualize and quantify FRFP and similar near-infrared probes in vivo within the skeleton. (26,71) Further advances will continue to widen options for using agents such as FRFP for in vivo imaging of biologic activity.…”

“…Recently, the functionalization of the bisphosphonate pamidronate with a near-infrared fluorophore has been developed and characterized. (24,25) This far-red fluorescent pamidronate (FRFP) has been used as a tool for monitoring bone turnover and repair (24)(25)(26) and extraskeletal calcification in vivo. (25,(27)(28)(29) Similar fluorescent bisphosphonate probes have been used to monitor bisphosphonate-osteoclast interactions in vitro.…”

Advances in multimodality molecular imaging of bone structure and function

“…In agreement, new mineral deposition as assessed with micro-CT had occurred in the non-union bone defect at 3 weeks in mice implanted with MSCs -BMP2, whereas in control mice, no bone formation was found. 75 Similarly, intervertebral discs injected with MSCs -BMP2 under a tetracycline-controlled expression system retained a high fluorescence signal up to 4 weeks post injection detected with BLI, and caused fusion of vertebral bodies as revealed with micro-CT. 76 …”

Comparative efficacy of dermal fibroblast-mediated and direct adenoviral bone morphogenetic protein-2 gene therapy for bone regeneration in an equine rib model

“…[1][2][3][4] Although direct BMP2 gene delivery is shown to promote bone regeneration, [5][6][7] the relative potency between gene-and cell-mediated BMP2 deliveries is unknown. Although embryonic and adult mesenchymal stem cells have been extensively studied for cell-mediated gene therapy, 8,9 other cell sources, such as dermal fibroblasts (DFbs) may be an alternative candidate because of their excellent reprogramming capacity. 10 In fact, DFbs have been differentiated into bone-forming cells by the transduction of osteogenic genes including BMP2, [11][12][13][14][15] and such genetically modified DFbs have been autologously transplanted to induce ectopic bone formation 14,15 and promote bone healing in rodent models.…”