Faculty Bibliography

Granulin epithelin precursor (GEP) has been implicated in development, tissue regeneration, tumorigenesis, and inflammation. Herein we report that GEP stimulates chondrocyte differentiation from mesenchymal stem cells in vitro and endochondral ossification ex vivo, and GEP-knockdown mice display skeleton defects. Similar to bone morphogenic protein (BMP) 2, application of the recombinant GEP accelerates rabbit cartilage repair in vivo. GEP is a key downstream molecule of BMP2, and it is required for BMP2-mediated chondrocyte differentiation. We also show that GEP activates chondrocyte differentiation through Erk1/2 signaling and that JunB transcription factor is one of key downstream molecules of GEP in chondrocyte differentiation. Collectively, these findings reveal a novel critical role of GEP growth factor in chondrocyte differentiation and the molecular events both in vivo and in vitro.

Purpose: Survival of bone implants depends on biological fixation, and prosthesis loosening can be catastrophic leading to replacement of prostheses. Inflammation and osteoclast-mediated bone resorption in response to wear particles near prostheses contribute to loosening. Because we have demonstrated that adenosine A_2Areceptor activation is anti-inflammatory and prevents osteoclast formation and function we hypothesized that adenosine A_2Areceptor agonists might prevent osteoclast-mediated bone resorption at the site of prosthesis wear in a calvarial model of wear particle-induced bone resorption. Methods: Eighteen C57Bl/6 mice age 6-8 weeks were anesthetized by intraperitoneal injection of ketamine and xylazine and a 1cm midline sagittal incision was made over the calvarium anterior to the line connecting both ears. Six animals received no particles (control), and 12 received 15 ml of polyethylene particle suspension. Of the 12 mice receiving particulate, 6 were injected subcutaneously at the surgical site with 20 ml of 10 muM CGS21680 (A_2Areceptor agonist), and 6 mice were injected with saline 0.9%, beginning immediately after incision closure and continuing every other day until sacrifice. Animals were sacrificed after 14 days and the calvaria were removed, fixed, and prepared for microCT and histological staining with TRAP. Results: Histologic examination of calvaria demonstrated lymphocytic infiltration in both particle-exposed groups. TRAP staining revealed a reduction in osteoclast differentiation after treatment with CGS21680. mCT showed pitting and increased porosity in both particle-exposed groups compared to controls, although in CGS21680-treated mice the reduction in cortical bone was significantly less than in the untreated particle-exposed mice (p<0.01). Control bone volume/trabecular volume was significantly greater (p<0.005) than in either particulate group, however, calvarial bone from CGS21680-treated mice had significantly greater mean bone volume than did the untreated group (p<0.0005). Trabecular thickness was significantly reduced in both CGS21680-treated and untreated particle-exposed groups as compared to control mice (p<0.05). Finally, digital morphometric analysis of microCTs reveals that CGS21680 significantly reduced the area of bone pitting compared to control particle-treated mice (p<0.05). (Table presented) Conclusions: Adenosine A_2Areceptor activation reduces inflammation and bone destruction due to prosthetic wear particles. This observation suggests that delivery of an adenosine A_2Aagonist in the cement may enhance orthopedic implant survival, delaying or eliminating the need for revision arthroplasty surgery

Id proteins play important roles in osteogenic differentiation; however, the molecular mechanism remains unknown. In this study, we established that inhibitor of differentiation (Id) proteins, including Id1, Id2, and Id3, associate with core binding factor alpha-1 (Cbfa1) to cause diminished transcription of the alkaline phosphatase (ALP) and osteocalcin (OCL) gene, leading to less ALP activity and osteocalcin (OCL) production. Id acts by inhibiting the sequence-specific binding of Cbfa1 to DNA and by decreasing the expression of Cbfa1 in cells undergoing osteogenic differentiation. p204, an interferon-inducible protein that interacts with both Cbfa1 and Id2, overcame the Id2-mediated inhibition of Cbfa1-induced ALP activity and OCL production. We show that 1) p204 disturbed the binding of Id2 to Cbfa1 and enabled Cbfa1 to bind to the promoters of its target genes and 2) that p204 promoted the translocation from nucleus to the cytoplasm and accelerated the degradation of Id2 by ubiquitin-proteasome pathway during osteogenesis. Nucleus export signal (NES) of p204 is required for the p204-enhanced cytoplasmic translocation and degradation of Id2, because a p204 mutant lacking NES lost these activities. Together, Cbfa1, p204, and Id proteins form a regulatory circuit and act in concert to regulate osteoblast differentiation

Current treatments for damaged articular cartilage (i.e., shaving the articular surface, perforation or abrasion of the subchondral bone, and resurfacing with periosteal and perichondrial resurfacing) often produce fibrocartilage, or hyaline-appearing repair that is not sustained over time (Henche 1967, Ligament and Articular Cartilage Injuries. Springer-Verlag, New York, NY, pp. 157-164; Insall 1974, Clin. Orthop. 101: 61-67; Mitchell and Shepard 1976, J. Bone Joint Surg. [Am.] 58: 230-233; O'Driscoll et al. 1986, J. Bone Joint Surg. [Am.] 68: 1017-1035; 1989, Trans. Orthop. Res. Soc. 14: 145; Kim et al. 1991, J. Bone Joint Surg. [Am.] 73: 1301-1315). Autologous chondrocyte transplantation, although promising, requires two surgeries, has site-dependent and patient age limitations, and has unknown long-term donor site morbidity (Brittberg et al. 1994, N Engl. J. Med. 331: 889-895; Minas 2003, Orthopedics 26: 945-947; Peterson et al. 2003, J. Bone Joint Surg. Am. 85-A(Suppl. 2): S17-S24). Osteochondral allografts remain a widely used method of articular resurfacing to delay arthritic progression. The present study compared the histological response to four types of osteochondral implants in a rabbit model: autograft, frozen, freeze-dried, and fresh implants. Specimens implanted in the femoral groove were harvested at 6 and 12 weeks. Results showed similar restoration of the joint surface regardless of implant type, with a trend toward better repair at the later timepoint. As has been observed in other studies (Frenkel et al. 1997, J. Bone Joint Surg. 79B: 281-286; Toolan et al. 1998, J. Biomed. Mater. Res. 41: 244-250), each group in this study had at least one specimen in which a healthy-appearing surface on the implant was not well-integrated with host tissues. Although the differences were not statistically significant, freeze-dried implants at both timepoints had the best histological scores. The osteochondral grafts tested successfully restored the gross joint surface and congruity. At 12 weeks, no significant differences were observed between the various allografts and autologous osteochondral grafts

OBJECTIVE: To investigate whether two different multiphasic implants could initiate and sustain repair of osteochondral defects in rabbits. The implants address the malleable properties of cartilage while also addressing the rigid characteristics of subchondral bone. DESIGN: The bone region of both devices consisted of D, D-L, L-polylactic acid invested with hyaluronan (HY). The cartilage region of the first device was a polyelectrolytic complex (PEC) hydrogel of HY and chitosan. In the second device the cartilage region consisted of type I collagen scaffold. Eighteen rabbits were implanted bilaterally with a device, or underwent defect creation with no implant. At 24 weeks, regenerated tissues were evaluated grossly, histologically and via immunostaining for type II collagen. RESULTS: PEC devices induced a significantly better repair than untreated shams. Collagen devices resulted in a quality of repair close to that of the PEC group, although its mean repair score (19.0+/-4.2) did not differ significantly from that of the PEC group (20.4+/-3.7) or the shams (16.5+/-6.3). The percentage of hyaline-appearing cartilage in the repair was highest with collagen implants, while the degree of bonding of repair to the host, structural integrity of the neocartilage, and reconstitution of the subchondral bone was greatest with PEC devices. Cartilage in both device-treated sites stained positive for type II collagen and GAG. CONCLUSIONS: Both implants are capable of maintaining hyaline-appearing tissue at 24 weeks. The physicochemical region between the cartilage and bone compartments makes these devices well suited for delivery of different growth factors or drugs in each compartment, or different doses of the same factor. It also renders these devices excellent vehicles for chondrocyte or stem cell transplantation