Engineered pluripotent mesenchymal cells integrate and differentiate in regenerating bone: a novel cell‐mediated gene therapy
D Gazit, G Turgeman, P Kelley, E Wang… - The journal of gene …, 1999 - Wiley Online Library
D Gazit, G Turgeman, P Kelley, E Wang, M Jalenak, Y Zilberman, I Moutsatsos
The journal of gene medicine, 1999•Wiley Online LibraryBackground Among the approximately 6.5 million fractures suffered in the United States
every year, about 15% are difficult to heal. As yet, for most of these difficult cases there is no
effective therapy. We have developed a mouse radial segmental defect as a model
experimental system for testing the capacity of Genetically Engineered Pluripotent
Mesenchymal Cells (GEPMC, C3H10T1/2 clone expressing rhBMP‐2), for gene delivery,
engraftment, and induction of bone growth in regenerating bone. Methods Transfected …
every year, about 15% are difficult to heal. As yet, for most of these difficult cases there is no
effective therapy. We have developed a mouse radial segmental defect as a model
experimental system for testing the capacity of Genetically Engineered Pluripotent
Mesenchymal Cells (GEPMC, C3H10T1/2 clone expressing rhBMP‐2), for gene delivery,
engraftment, and induction of bone growth in regenerating bone. Methods Transfected …
Background
Among the approximately 6.5 million fractures suffered in the United States every year, about 15% are difficult to heal. As yet, for most of these difficult cases there is no effective therapy. We have developed a mouse radial segmental defect as a model experimental system for testing the capacity of Genetically Engineered Pluripotent Mesenchymal Cells (GEPMC, C3H10T1/2 clone expressing rhBMP‐2), for gene delivery, engraftment, and induction of bone growth in regenerating bone.
Methods
Transfected GEPMC expressing rhBMP‐2 were further infected with a vector carrying the lacZ gene, that encodes for β‐galactosidase (β‐gal). In vitro levels of rhBMP‐2 expression and function were confirmed by immunohistochemistry, and bioassay. Differentiation was assayed using alkaline phosphatase staining. GEPMC were transplanted in vivo into a radial segmental defect. The main control groups included lacZ clones of WT‐C3H10T1/2‐LacZ, and CHO‐rhBMP‐2 cells. New bone formation was measured quantitatively via fluorescent labeling, X‐ray analysis and histomorphometry. Engrafted mesenchymal cells were localized in vivo by β‐gal expression, and double immunofluorescence.
Results
In vitro, GEPMC expressed rhBMP‐2, β‐gal and spontaneously differentiated into osteogenic cells expressing alkaline phosphatase. Detection of transplanted cells revealed engrafted cells that had differentiated into osteoblasts and co‐expressed β‐gal and rhBMP‐2. Analysis of new bone formation revealed that at fout to eight week post‐transplantation, GEPMS significantly enhanced segmental defect repair.
Conclusions
Our study shows that cell‐mediated gene transfer can be utilized for growth factor delivery to signaling receptors of transplanted cells (autocrine effect) and host mesenchymal cells (paracrine effect) suggesting the ability of GEPMC to engraft, differentiate, and stimulate bone growth. We suggest that our approach should lead to the designing of mesenchymal stem cell based gene therapy strategies for bone lesions as well as other tissues. Copyright © 1999 John Wiley & Sons, Ltd.
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