Molly M. Stevens, Robert P. Marini, Dirk Schaefer, Joshua Aronson, Robert Langer, and V. Prasad Shastril
PNAS (2005) 102: 11450-114555.
Currently, the repair of major bone defects, such as those associated with tumor resections and spinal fusions, requires bone grafts. Typically, autologous bone grafts are harvested from the iliac crest. However, the harvesting procedure is frequently accompanied by pain and morbidity. To address this issue, the authors tested an in vivo bioreactor designed to produce autologous bone without requiring the administration of growth factors or cell transplantation.
The bone bioreactor “space” was created in the tibia of rabbits by injecting a biocompatible gel composed of calcium and alginate between the periosteum and long bone. The maturation of bone within the bioreactor was observed over 12 weeks. The engineered bone was harvested from the reactor and then transplanted into tibial defects to determine how well it would integrate with existing bone.
The authors observed that after 6 weeks, the engineered bone demonstrated all the histological and structural characteristics of native lamellar bone. Also, they determined that both the compressive strength and ultimate strength of the engineered bone was within the range for compact bone. Moreover, histological and radiographical tests revealed that after 6 weeks, transplanted neo-bone integrated with native bone within a tibial defect.
The authors also demonstrated that by adding the angiogenic factor, Suramin, to the gel matrix, cartilage formation instead of bone formation occurred within the bioreactor. However, no quantitative analysis on the cartilage was performed.
The authors have established that in vivo engineering of autologous bone without the need for growth factors or transplanted cells is possible. Their approach may potentially lead to the generation of large quantities of autologous bone at an easily accessible surgical site. The study may have important implications for bone banking and bone transplantations.
13 comments:
Did they mention what kind of pain is associated with the removal of the bone from the reactor?
Also, what kind of defects did they use on the tibial bones? Did they use the same kind each time or did they vary the type of defect?
The authors did not test for pain in their study. However, they did briefly mention that no apparent post-operative morbidity was observed. Also, the authors noted that the presence of a demarcation line between engineered bone and native bone allowed for harvest of the bone without compromising the native skeletal structure. It is possible that this may lead to less pain and morbidity at the harvest site.
To create the tibial defects, the researchers created a 7x3x1.5-mm defect using a dental drill and dental burr. The defects were not varied.
Yes, the authors performed a control in which a defect was filled with Gelfoam. Radiographs of the defects were taken before and after the surgery and every two weeks for six weeks. No bone formation was observed within the defect.
Was the Suramin just injected into the middle of the gel matrix? If it was added as a "top layer" to the matrix do you think it would induce cartilage formation on the outside of the bone graft as well and possibly help in osteoarthritic patients?
Did they also test to see if the cells from the engineered bone were secreting a comparable amount of ECM to that of native cells?
Suramin was incorporated into the matrix solution as either a "free" or liposome-encapsulated form.
You make a very interesting point; inducing cartilage formation outside of the engineered bone would be novel. However, the presence of Suramin, which inhibits angiogenesis, may interfere with bone formation, which requires vascularization.
Also, immunostaining of the bone tissue indicated that both osteonectin and collagen type I were present. However, quantitative amounts were not determined.
Did they say why they chose the tibia among all other bones? Perhaps different bones have different conductive and inductive properties, and even show a marked difference in the type of bone they produce after injecting the same biocompateble gel. These properties could be mimicked in vitro for further bone applications..?
Why did the authors purposefully not include growth factors and transplanted cells?--was it to insure that the process can be used in mass production?
It seems to me that growth factors and transplanted cells can be added to this bioreactor to make the process produce even better results.
Did they have to regularly inject calcium or Suramin into the gel? Over the period of the experiment, it seems diffusion through the gel might have an effect. If they had to regularly inject materials, how practical would this bioreactor be?
I'm slightly confused- the point of the bone bioreactor seems to be to avoid the pain and potential morbidity associated with the way bone grafts are harvested currently. But from what I've read, it seems that the bone bioreactor is essentially a way of engineering bone but is still within the body, so the bone still needs to be harvested from an individual before being transplanted. What, then, makes this better than the current method of harvesting bone grafts?
Also, you mentioned that the authors tested the compressive strength and ultimate strength of the engineered bone- did they test any other mechanical properties? Bone seems to be a complex tissue in the sense that, different bones (just like different muscles) undergo different cyclic loads and different types of stress depending on their location in the body. Where in the body is this engineered bone designed for?
Todd : I believe the authors chose the tibia because the site is surgically easily accessible with less chance for complications. Additional studies involving different types of bone may prove to be useful
Willie : I think you are correct. The addition of growth factors may produce better results. However, the ability to reproduce synthesized bone that is structurally similar to native bone without requiring any additional factors would be an excellent commercial selling point.
Jay : The authors did not have to inject any additional factors after the creation of the bioreactor. Controlled release of Suramin was produced through the use of liposome capsules.
Jennifer : The authors noted that the presence of a demarcation line between engineered bone and native bone allowed for harvest of the bone without compromising the native skeletal structure. It is possible that this may lead to less pain and morbidity at the harvest site. To answer your next question, the authors did not perform any additional mechanical tests. As you suggest, more specific testing may be necessary in order to determine ideal locations for the engineered bone.
So is the eventual goal to be able to implant these bioreactors in the patients that require bone grafts? Can I get a sense of how much material was produced in the 6 weeks this experiment was run? I believe that may be the crucial factorv - the efficiency of the procfess.
Congrats Al. You win for most bizarre use of rabbit tibias. I'd be curious to know if there are any complications in the procedure and the dangers of damaging the healthy tissue surrounding the tibia when removing the engineered bone. Also how does the harvested bone from the tibia compare to the grafts that could be taken from the iliac crest?
Concord: Yes, that's the eventual goal. In fact, the authors performed tests on human cadavers to determine whether or not a bioreactor could be created in human tibia (see supplemental info).
To answer your next question, the investigators were able to harvest approximately 162 mm^3 of bone from a 200 mm^3 bioreactor space.
To compare these numbers on a human scale: about 4-6 cc of bone is required for an interbody fusion procedure. Assuming a human = 70kg and a rabbit = 2kg, this would roughly correspond to 170 mm^3 of bone harvested from a rabbit.
Zach: No serious complications appear to be associated with the harvesting procedure. The "border" between the bioreactor space and the engineered bone seems to facilitate removal of the bone. The authors did not compare bone harvested from the bioreactor to bone harvested from the iliac crest. However, this comparison would be important in future experiments.
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