Enhanced matrix synthesis and in vitro formation of cartilage‐like tissue by genetically modified chondrocytes expressing BMP‐7

Abstract

Bone morphogenic protein-7 (BMP-7) supports ectopic cartilage and bone formation, is expressed in normal articular cartilage, and increases matrix synthesis in chondrocytes. Based on this knowledge, we hypothesized that an adenovirus (Ad) vector encoding human BMP-7 could be used to modify chondrocytes genetically to improve their capacity for cartilage repair. An adenovirus vector encoding BMP-7 (AdBMP-7) was constructed and its bioactivity confirmed by ectopic bone formation assay. AdBMP-7 modification of bovine chondrocytes induced expression of BMP-7 mRNA and bioactive protein, resulting in an increase in incorporation of 35SO; into proteoglycan, 3H-proline uptake into protein, and the expression of the cartilage-specific matrix genes, aggrecan and type I1 collagen. An in vitro model of chondrocyte transplantation was used to demonstrate the feasibility of using genetically modified chondrocytes to enhance formation of cartilage-like tissue. When transplanted onto cartilage explants and maintained in vitro for 3 weeks, chondrocytes modified with AdBMP-7 formed 1.9-fold thicker tissue than chondrocytes modified with a control vector (P < 0.001). This tissue was positive for type I1 collagen and proteoglycan but negative for type X collagen and demonstrated a cartilage-like morphology. These observations suggest that Ad-mediated transfer of BMP-7 gene to chondrocytes enhances the chondrocyte-specific matrix synthesis and their capacity to form cartilage-like tissue, thus representing a strategy that may improve cell-based cartilage repair.

Differences in matrix accumulation and hypertrophy in superficial and deep zone chondrocytes are controlled by bone morphogenetic protein

“…We and others have reported that exogenous BMPs also increase matrix synthesis by chondrocytes (Flechtenmacher et al, 1996;Hidaka et al, 2003;Hidaka et al, 2001;Huch et al, 1997;Sailor et al, 1996). In the current study, we have identified SZC as the sub-population that is particularly responsive, even to low doses of BMPs.…”

“…Several family members, including BMP7, are expressed in post-natal articular cartilage where they are important for cartilage ECM homeostasis (Chang et al, 1994;Chubinskaya and Kuettner, 2003;Chubinskaya et al, 2000;Rountree et al, 2004;Soder et al, 2005). Previously, we have reported increased matrix gene expression and matrix synthesis in chondrocytes overexpressing BMP7 (Hidaka et al, 2001). Furthermore, we showed that BMP7 overexpressing chondrocytes can be implanted into experimental cartilage defects to accelerate repair in vivo (Hidaka et al, 2003).…”

“…To elucidate the aspects of the chondrocyte phenotype that may be controlled by BMPs, SZC and DZC were genetically modified using adenovirus (Ad) vectors encoding BMP2 (AdBMP2) or BMP7 (AdBMP7) (Hidaka et al, 2001;Zhu et al, 2004) or Noggin (AdNoggin) (Lim et al, 2000), then maintained as pellet cultures for 7 days. As controls, cells were incubated with AdNull (encoding no transgene) or no virus (Naïve).…”

“…In particular, we and others have shown the efficacy of bone morphogenetic protein-7 (BMP7, also known as osteogenic protein-1) in increasing matrix production in chondrocytes (Flechtenmacher et al, 1996;Hidaka et al, 2001;Huch et al, 1997;Masuda et al, 2006;Merrihew et al, 2003). The BMPs are a family of growth factors whose expression is critical for skeletal development, including the formation of the joints Francis-West et al, 1999;Rosen, 2006).…”

Restoration of the extracellular matrix in human osteoarthritic articular cartilage by overexpression of the transcription factor SOX9

“…Direct application of rAAV-FLAG-hSOX9 to human normal and OA cartilage in situ mediated high levels of transgene expression that was distributed throughout the thickness of the explant cultures, probably due to the ability of these small vectors to penetrate the dense ECM of the cartilage (23,24). The manipulation of vectors such as rAAV might thus be desirable for introducing candidate genes directly into the targets as compared with other classes of vectors still used in ex vivo gene transfer protocols (5,12,45). While no effects on cell proliferation were seen, expression of the SOX9 gene cassette promoted a significant, dose-dependent increase in the proteoglycan and type II collagen content in OA cartilage to levels higher than or similar to those in the untreated normal cartilage and to depths relevant for clinical applications (24).…”