Shawn A. Hunter

Using Functional Tissue Engineering and Bioreactors to Mechanically Stimulate Tissue-Engineered Constructs

Bioreactors precondition tissue-engineered constructs (TECs) to improve integrity and hopefully repair. In this paper, we use functional tissue engineering to suggest criteria for preconditioning TECs. Bioreactors should (1) control environment during mechanical stimulation; (2) stimulate multiple constructs with identical or individual waveforms; (3) deliver precise displacements, including those that mimic in vivo activities of daily living (ADLs); and (4) adjust displacement patterns based on reaction loads and biological activity. We apply these criteria to three bioreactors. We have placed a pneumatic stimulator in a conventional incubator and stretched four constructs in each of five silicone dishes. We have also programmed displacement-limited stimuli that replicate frequencies and peak in vivo patellar tendon (PT) strains. Cellular activity can be monitored from spent media. However, our design prevents direct TEC force measurement. We have improved TEC stiffness as well as PT repair stiffness and shown correlations between the two. We have also designed an incubator to fit within each of two electromagnetic stimulators. Each incubator provides cell viability like a commercial incubator. Multiple constructs are stimulated with precise displacements that can mimic ADL strain patterns and record individual forces. Future bioreactors could be further improved by controlling and measuring TEC displacements and forces to create more functional tissues for surgeons and their patients.

Biomechanical and biologic effects of meniscus stabilization using triglycidyl amine.

The susceptibility of meniscus allografts to enzymatic degradation may be reduced through tissue stabilization. We have previously reported on an epoxide-based crosslinker, triglycidyl amine (TGA), which can be used alone or with a bisphosphonate (MABP) to stabilize heterograft heart valves and reduce their pathologic calcification. Our objective was to evaluate the effects of TGA and TGA-MABP pretreatment on an orthopedic allograft involving meniscus crosslinking, degradation, calcification, and compressive properties. Ovine menisci treated with TGA or TGA-MABP for up to seven days and glutaraldehyde crosslinked controls were examined in vitro for degree of crosslinking, resistance to degradation by collagenase, and material property changes. Likewise treated menisci were implanted in rats for eight weeks and examined for calcium content and biomechanical changes. TGA treatment for three days significantly reduced collagen loss by 88% and increased thermal denaturation temperatures (Ts) above 80 degrees C versus Ts of 70 degrees C or less for non-crosslinked meniscus. In vitro, TGA and TGA-MABP significantly increased aggregate modulus by 19% and 32% compared to native controls, respectively. TGA decreased permeability by 53% while TGA-MABP increased it by 303%. In vivo, TGA significantly reduced explant calcification by 42% compared to glutaraldehyde, and including MABP reduced it by 90%. Analyses revealed that TGA and TGA-MABP stabilized menisci had significantly lower modulus and permeability values than glutaraldehyde controls by at least 28% and 86%, respectively. It is concluded that TGA crosslinking of meniscus increases resistance to both collagenase degradation and pathologic calcification, while demonstrating comparable or improved biomechanical properties versus glutaraldehyde controls.

Demineralization Removes Residual Alendronate in Allograft Bone Procured From Donors With a History of Bisphosphonate Use

BACKGROUND Bisphosphonate-associated osteonecrosis (BON) of the jaw is a growing concern in the dental community, but the possible presence of residual bisphosphonates in demineralized allograft bone from bisphosphonate-using tissue donors and the clinical implications of using such bone are unclear. The objectives of this study are to determine whether alendronate remained in demineralized bone matrix (DBM) procured from donors with a documented history of oral bisphosphonate use and to examine whether the demineralization process removes alendronate from allograft bone. METHODS A gas chromatography?mass spectrometry method was developed and validated to quantify residual alendronate in allograft bone. Alendronate levels in DBM procured from tissue donors with a history of oral bisphosphonate use were compared to alendronate levels in DBM procured from donors without a history of bisphosphonate use. In addition, mineralized and demineralized bone was soaked in alendronate at concentrations of 0.002, 2.0, and 2,000 ng/mg bone and analyzed to examine the effect of the demineralization process. RESULTS Residual alendronate was not detected in the DBM from either group, nor was it detected in any of the DBM samples soaked in alendronate solutions. Soaked mineralized bone contained measureable alendronate, but the substance was removed by demineralization. CONCLUSIONS The demineralization process effectively removed residual alendronate from allograft bone. These results may relieve anxieties regarding the use of DBM in dental patients because it is unlikely to trigger BON of the jaw.