1. Volume 19, Issue 1, Pages 53-59 (January 2011)
Targeted Gene-and-host Progenitor Cell Therapy for Nonunion Bone Fracture Repair 
Nadav Kimelman-Bleich, Gadi Pelled, Yoram Zilberman, Ilan Kallai, Olga Mizrahi, Wafa Tawackoli, Zulma Gazit, Dan Gazit 
Molecular Therapy 
Volume 19, Issue 1, Pages (January 2011)
DOI: /mt
Copyright © 2011 The American Society of Gene & Cell Therapy Terms and Conditions

2. Figure 1
Host progenitor cells populate the defect site. A 1.5-mm long defect was made in the mouse radius, and a collagen sponge was placed in the defect site. Ten days after surgery, the radius was harvested and stained using H&E. (a–c) Radial defect 10 days after surgery. Staining shows the presence of HPCs (arrowheads). HPC, host progenitor cells; H&E, hematoxylin and eosin; M, defect margin; U, ulna.
Molecular Therapy  , 53-59DOI: ( /mt )
Copyright © 2011 The American Society of Gene & Cell Therapy Terms and Conditions

3. Figure 2
Gene transfer efficiency and localization to HPCs. (a) BLI was used to monitor luciferase activity in bone defects following pLuc injection and electroporation. The x axis displays time postelectroporation; the y axis shows activity in RLUs. *P < 0.05, two-tailed t-test, n = 4. Note the representative pictures of one mouse. (b) Results of another BLI study performed in a representative mouse in order to verify gene delivery to HPCs. After they underwent the BLI study, mice were sacrificed and cells from explanted radii were isolated. (c) photomicrograph shows HPCs isolated from the radial explants. (d) Isolated HPCs were lyzed and mRNA was isolated. Using RT-PCR followed by real-time PCR, the presence of Luc expression in the isolated cells was verified (EP Luc). HPCs isolated from defects in which electroporation was not performed were used as a negative control (no EP). BLI, bioluminescence imaging; HPC, host progenitor cells; RLU, relative light unit; RT-PCR, reverse transcription PCR.
Molecular Therapy  , 53-59DOI: ( /mt )
Copyright © 2011 The American Society of Gene & Cell Therapy Terms and Conditions

4. Figure 3
Bone formation in the defect area. (a) Bone volume analysis: a comparison of newly formed bone in the radial defect in animals in the pBMP-9 with electroporation (BMP-9, six mice), pLuc with electroporation (Luc, four mice), and pBMP-9 without electroporation (no EP, four mice) groups and of segments of native radii having the same dimensions (native, six mice). *P < 0.01, two-tailed t-test. Note that bone formation in the defect site was significantly higher than in controls. No statistical significant difference was found between mice electroporated with pLuc and mice with pBMP-9 without electroporation. (b) µCT reconstruction comparison of defects treated with pLuc and electroporation (Luc), defects treated with pBMP-9 and electroporation (BMP-9), and defects treated with BMP-9 without electroporation (no EP). M, defect margin. Arrowhead indicates new bone formation in the defect. In the 3D images, orange regions denote new bone formation. (c) Histological sections containing newly formed bone. 1.5-mm long defects were made in mice radii, and a collagen sponge was placed in the defect site. Ten days postoperation, the radius was injected with either pBMP-9 or pLuc followed by electroporation. As a control, radii were injected with pBMP-9 but electroporation was not performed. Five weeks postelectroporation, the radii were harvested and stained using Masson's trichome. M, defect margin; U, ulna. Arrowheads indicate new bone formation; asterisks indicate soft tissue in the defect site. BMP, bone morphogenetic protein 9; µCT, microcomputed tomography.
Molecular Therapy  , 53-59DOI: ( /mt )
Copyright © 2011 The American Society of Gene & Cell Therapy Terms and Conditions

5. Figure 4
Quantitative analysis of structural parameters of induced bone formation in the defect area. (a–f) Newly formed bone in the radial defect in mice in the pBMP with electroporation group (BMP-9) was compared to segments of native radii having the same dimensions (native). *P < 0.05, two-tailed t-test, n = 6 in each group. The structural parameters include: (a) bone thickness (mm); (b) trabecular number (1/mm); (c) bone separation (mm); (d) connectivity density (1/mm3); (e) bone volume density (BV/TV, mm/mm); and (f) bone mineral density (mg HA/cm3). BMP-9, bone morphogenetic protein 9.
Molecular Therapy  , 53-59DOI: ( /mt )
Copyright © 2011 The American Society of Gene & Cell Therapy Terms and Conditions

6. Figure 5
Setup of the electroporation system. A 1.5-mm long defect was made in the mouse radius, and a collagen sponge was placed in the defect site. Ten days after surgery pLuc or pBMP-9 was injected into the defect site, and electroporation was immediately performed using needle electrodes. (a) Mouse ready for electroporation. The animal is mounted on the fluoroscope detector and needle electrodes are placed. (b) Locations of the electrodes and the needle used for plasmid injection are verified using the fluoroscope.
Molecular Therapy  , 53-59DOI: ( /mt )
Copyright © 2011 The American Society of Gene & Cell Therapy Terms and Conditions