Blocking vascular endothelial growth factor with soluble Flt-1 improves the chondrogenic potential of mouse skeletal muscle-derived stem cells.
OBJECTIVE
To investigate the effect of vascular endothelial growth factor (VEGF) stimulation and the effect of blocking VEGF with its antagonist, soluble Flt-1 (sFlt-1), on chondrogenesis, using muscle-derived stem cells (MDSCs) isolated from mouse skeletal muscle.


METHODS
The direct effect of VEGF on the in vitro chondrogenic ability of mouse MDSCs was tested using a pellet culture system, followed by real-time quantitative polymerase chain reaction (PCR) and histologic analyses. Next, the effect of VEGF on chondrogenesis within the synovial joint was tested, using genetically engineered MDSCs implanted into rat osteochondral defects. In this model, MDSCs transduced with a retroviral vector to express bone morphogenetic protein 4 (BMP-4) were coimplanted with MDSCs transduced to express either VEGF or sFlt-1 (a VEGF antagonist) to provide a gain- and loss-of-function experimental design. Histologic scoring was used to compare cartilage formation among the treatment groups.


RESULTS
Hyaline-like cartilage matrix production was observed in both VEGF-treated and VEGF-blocked (sFlt-1-treated) pellet cultures, but quantitative PCR revealed that sFlt-1 treatment improved the expression of chondrogenic genes in MDSCs that were stimulated to undergo chondrogenic differentiation with BMP-4 and transforming growth factor beta3 (TGFbeta3). In vivo testing of articular cartilage repair showed that VEGF-transduced MDSCs caused an arthritic change in the knee joint, and sFlt-1 improved the MDSC-mediated repair of articular cartilage, compared with BMP-4 alone.


CONCLUSIONS
Soluble Flt-1 gene therapy improved the BMP-4- and TGFbeta3-induced chondrogenic gene expression of MDSCs in vitro and improved the persistence of articular cartilage repair by preventing vascularization and bone invasion into the repaired articular cartilage.
Kubo
Seiji
S
Children's Hospital of Pittsburgh, and University of Pittsburgh, Pittsburgh, Pennsylvania 15213-2582, USA.
Cooper
Gregory M
GM
Matsumoto
Tomoyuki
T
Phillippi
Julie A
JA
Corsi
Karin A
KA
Usas
Arvydas
A
Li
Guangheng
G
Fu
Freddie H
FH
Huard
Johnny
J
D000818:Animals; D055415:Bone Morphogenetic Protein 4 / Q000235:genetics; D002358:Cartilage, Articular / Q000166:cytology; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000502:physiology*; D015316:Genetic Therapy / Q000379:methods*; D051457:Hyaline Cartilage / Q000166:cytology; D007202:Indicators and Reagents; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018482:Muscle, Skeletal / Q000166:cytology; D020370:Osteoarthritis, Knee / Q000473:pathology / Q000503:physiopathology / Q000628:therapy*; D010619:Phenazines; D051381:Rats; D011923:Rats, Nude; D012995:Solubility; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000502:physiology; D014161:Transduction, Genetic; D042461:Vascular Endothelial Growth Factor A / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics* / Q000378:metabolism
C525809:BMP4 protein, human; D055415:Bone Morphogenetic Protein 4; D007202:Indicators and Reagents; D010619:Phenazines; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; C501163:Flt1 protein, mouse; D040281:Vascular Endothelial Growth Factor Receptor-1; C009195:safranine T