This study clearly shows the benefits of combined PTH and BMP-2 treatment on ectopic and orthotopic bone formation in a microsphere/scaffold composite for bone tissue engineering. Although an anabolic effect on the rest of the skeleton was seen, the 10
μg PTH/kg/day treatment had no significant effect on local bone regeneration in unfilled and empty scaffold-filled defects. Implantation of BMP-2-loaded scaffolds resulted in a significant increase of BMD and bone volume in both sites as opposed to the empty scaffolds. The combination of BMP-2-containing scaffolds with daily PTH injections resulted in a further increase of both ectopic and orthotopic bone volumes.
The exact cellular mechanism underlying the anabolic effect of PTH treatment is not fully understood. Histological observations have shown that intermittently administered PTH stimulates new bone formation on existing surfaces by increasing the osteoblast number.5,19–21
Previous studies have suggested that this increase in osteoblast number is not dependent on the proliferation of osteoprogenitor cells and osteoblastogenesis, but the result of PTH effects on existing cells.19,20,22
Likely target cells for short-term PTH effects are bone-lining cells, which are thought to be inactive osteoblasts. These lining cells may undergo hypertrophy and resume matrix synthesis in response to PTH treatment.5,19,21
Another proposed method for the continued effect of long-term PTH treatments is the inhibition of osteoblast apoptosis.5,20
The absence of bone formation in the ectopically implanted empty scaffolds was anticipated, as the synthetic polymers are not osteoinductive. Since PTH is expected to act upon cells committed to the osteoblastic lineage, it was not surprising that PTH was not able to affect ectopic bone formation in these empty scaffolds.23
Although PTH cannot induce bone formation from uncommitted cells, it can enhance bone formation once it has been initiated. Previous bone fracture, distraction, and conduction chamber models have shown a strong dose-related effect of intermittent PTH administration on local bone regeneration.24–27
Despite bone induction at the edges of the unfilled or empty scaffold filled defects and the positive PTH effects on preexistent bone, no significant differences were found between these untreated and PTH-treated defects. The discrepancy between previous studies and these results may be due to the more challenging critical-sized defect model and low PTH dose used in this study.
In contrast to PTH, BMP-2 plays an essential role in the commitment and differentiation of mesenchymal stem cells toward the osteoblastic lineage. Its high osteoinductive potential is clearly demonstrated by its ability to induce bone formation in an ectopic implantation site.28,29
The BMP-induced bone formation occurs through endochondral (through a cartilage intermediate) and/or intramembranous (direct) ossification and results in woven bone, which is later remodeled into normal bone.1,28
In our study, after implantation of the microsphere/PPF/gelatin delivery vehicle, clear signs of endochondral and intramembranous ossification were also seen at the ectopic or orthotopic location.
Despite the nonsignificant effect of PTH alone, PTH significantly enhanced the BMP-2-induced bone formation at both ectopic and orthotopic locations. This clearly shows that the different mechanisms of action of BMP-2 and PTH enhance each other when used as a combination therapy. Whereas BMP-2 induces bone formation by committing mesenchymal stem cells toward the osteoblastic lineage, PTH could act upon these committed cells to prolong the matrix-synthesizing function. Further, the antiapoptotic effect of PTH could also have enhanced bone formation by expanding the osteoblast life span by counteracting the possible apoptotic effects of BMP-2 on osteoblasts.20,30
Unfortunately, further histological analysis (e.g., counting of the osteoblast number) of the mechanism underlying the anabolic effect in the combination therapy was impossible due to the woven aspect of the newly formed bone.
Compared to previous studies, the ectopic results obtained in our study corresponded with previous findings of PTH-mediated enhancement of ectopic BMP-induced bone formation in a collagen sponge.8,9
In contrast to the clearly enhanced ectopic effect, no significant differences could be observed between a PTH/BMP-7 combination and PTH alone in a previous study.31
This nonsignificant effect might be caused by their use of a less challenging partial thickness metaphysic defect model. Since the spontaneous regeneration response was capable of healing the nontreated defects as well, no significant effect of BMP-7 could be shown compared to the control group. Consequently, PTH enhanced both spontaneous and BMP-7-assisted healing response and no significant differences in bone formation were observed between the PTH/BMP-7 combination and PTH alone. In the critical-sized defect model in this study, the normal bone regeneration response resulted in limited amounts of bone formation at the defect edges, which was not significantly enhanced by PTH. However, bone regeneration was significantly stimulated by the BMP-2-releasing implants and PTH could act upon this to synergistically enhance the healing response.
In conclusion, this study clearly shows that BMP-2-induced osteogenesis can be enhanced by intermittent administration of PTH. Although PTH alone did not significantly improve bone formation, it can be beneficial for local bone regeneration in combination with BMP-2. On the basis of this study, PTH/BMP-2 combination therapy could be considered for the restoration of large bone defects in orthopedic surgery. However, due to the physiological and anatomical differences between humans and different animal species for bone regeneration, future studies in more relevant large animal models and clinical trials will be needed to maximize the clinical efficacy of PTH/BMP-2 combination therapies.32,33