Human flexor tendon tissue engineering: decellularization of human flexor tendons reduces immunogenicity in vivo.
BACKGROUND
In mutilating hand injuries, tissue engineered tendon grafts may provide a reconstructive solution. We have previously described a method to decellularize cadaveric human flexor tendons while preserving mechanical properties and biocompatibility. The purpose of this study is to evaluate the immunogenicity and strength of these grafts when implanted into an immunocompetent rat model.


METHODS
Cadaveric human flexor tendons were divided into two groups. Group 1 was untreated, and Group 2 was decellularized by treatment with sodium dodecyl sulfate (SDS), ethylenediaminetetraacetic acid (EDTA), and peracetic acid (PAA). Both groups were then analyzed for the presence of major histocompatibility complexes by immunohistochemistry (IHC). Pair-matched tendons from each group were then placed into the dorsal subcutaneous tissue and anchored to the spinal ligaments of Wistar rats for 2 or 4 weeks, and harvested. The infiltration of B-cells and macrophages was determined using IHC. The explants where then subjected to mechanical testing to determine the ultimate tensile stress (UTS) and elastic modulus (EM). Statistical analysis was performed using a paired Student's t-test.


RESULTS
The decellularization protocol successfully removed cells and MHC-1 complexes. At 2 weeks after implantation, there was increased infiltration of B-cells in Group 1 (untreated) compared with Group 2 (acellular), both in the capsule and tendon substance. There was improved ultimate tensile stress (UTS, 42.7 ± 8.3 vs. 22.8 ± 7.8 MPa, p<0.05) and EM (830.2 ± 206.7 vs. 421.2 ± 171.3 MPa, p<0.05) in tendons that were decellularized. At 4 weeks, there was continued B-cell infiltration in Group 1 (untreated) compared with Group 2 (acellular). There was no appreciable difference in macrophage infiltration at both time points. At 4 weeks Group 2 (acellular) demonstrated persistently greater UTS (40.5 ± 9.1 vs. 14.6 ± 4.2 MPa, p<0.05) and EM (454.05 ± 101.5 vs. 204.6 ± 91.3 MPa, p<0.05) compared with Group 1 (untreated).


CONCLUSIONS
Human flexor tendons that were decellularized with SDS, EDTA, and PAA resulted in removal of cellular antigens and a decreased immune response when placed into Wistar rats. These grafts showed better mechanical properties at 2 and 4 weeks when compared with control tendons. Decellularization is an important step toward the use of tissue engineered flexor tendons in upper extremity reconstruction.
Section of Plastic Surgery, VA Palo Alto Health Care System and Division of Plastic Surgery, Stanford University Medical Center, Stanford, California 94305, USA.
Shyam S
SS
Raghavan
Colin Y L
CY
Woon
Armin
A
Kraus
Kai
K
Megerle
Matthew S S
MS
Choi
Brian C
BC
Pridgen
Hung
H
Pham
James
J
Chang
D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D011817:Rabbits; D017208:Rats, Wistar; D013710:Tendons / Q000166:cytology* / Q000276:immunology*; D023822:Tissue Engineering / Q000379:methods*