Use of Platelet-Rich Plasma Immediately After an Injury Did Not Improve Ligament Healing, and Increasing Platelet Concentrations Was Detrimental in an In Vivo Animal Model.

BACKGROUND: Limited information in basic science and clinical trials exists to determine if ligament healing may be accelerated with the use of biological adjuvants, such as platelet-rich plasma (PRP). However, there has been widespread acceptance of PRP for use in clinical practice, despite an inadequate understanding of its biological mechanism of action. PURPOSE: To determine whether a single dose of PRP could accelerate ligament healing and correspondingly improve histological characteristics and biomechanical properties when injected immediately postoperatively into the injured medial collateral ligament (MCL) of New Zealand White rabbits. STUDY DESIGN: Controlled laboratory study. METHODS: Eighty skeletally mature New Zealand White rabbits (160 knees) were used. The MCL was torn midbody to simulate a grade 3 tear. After an acute injury of the MCL, the administration of autologous PRP at 3 different platelet concentrations (0 million/uL, platelet-poor plasma [PPP]; 0.6 million/uL, 2 times the baseline [2× PRP]; and 1.2 million/uL, 4 times the baseline [4× PRP]) was performed and compared with a saline injection control in the contralateral knee. Histological analysis and a biomechanical endpoint characterization were utilized to assess ligamentous healing and compare it to a sham surgery group. RESULTS: The PPP ( P = .001) and 4× PRP ( P = .002) groups had a significantly lower collagen subscore than the sham surgery group. No other differences were observed among the treatment groups, including the vascularity subscore and overall ligament tissue maturity index score. Compared with saline-injected contralateral knees, the maximum load for PPP and 2× PRP was not significantly different ( P = .788 and .325, respectively). The maximum load and stiffness for knees treated with 4× PRP were significantly less than for the saline-treated contralateral knees ( P = .006 and .001, respectively). CONCLUSION: One single dose of PPP or 2× PRP at the time of injury did not improve ligament healing. In addition, 4× PRP negatively affected ligament strength and histological characteristics at 6 weeks after the injury. CLINICAL RELEVANCE: The current practice of treating knee ligament injuries with PRP may not improve healing at low doses of PRP. The decreased mechanical properties and histological appearance of the torn MCL suggest that high doses of PRP decrease the quality of repair tissue. Further in vivo studies are necessary to determine the dosing and timing of PRP administration after a ligament injury before the widespread use of PRP to treat ligament injuries is recommended.

Evaluation of a collagen-coated, resorbable fiber scaffold loaded with a peptide basic fibroblast growth factor mimetic in a sheep model of rotator cuff repair.

BACKGROUND: A new scaffold design combined with a peptide growth factor was tested prospectively for safety and for improved tendon healing in sheep. METHODS: The infraspinatus tendon was detached and then surgically repaired to the humerus using sutures and anchors in 50 adult sheep. The repairs in 40 of these sheep were reinforced with a scaffold containing F2A, a peptide mimetic of basic fibroblast growth factor. The sheep were examined after 8 or 26 weeks with magnetic resonance imaging, full necropsy, and histopathologic analysis. A second cohort of 30 sheep underwent surgical repair--20 with scaffolds containing F2A. The 30 shoulders were tested mechanically after 8 weeks. RESULTS: The scaffold and F2A showed no toxicity. Scaffold-repaired tendons were 31% thicker than surgically repaired controls (P = .037) at 8 weeks. There was more new bone formed at the tendon footprint in sheep treated with F2A. Surgically repaired tendons delaminated from the humerus across 14% of the footprint area. The extent of delamination decreased to 1.3% with increasing doses of F2A (P = .004). More of the repair tissue at the footprint was tendon-like in the peptide-treated sheep. On mechanical testing, only 7 shoulders tore at the repair site. The repairs in the other 23 shoulders were already stronger than the midsubstance tendon at 8 weeks. CONCLUSIONS: The new scaffold and peptide safely improved tendon healing.

Effect of low-temperature ethylene oxide and electron beam sterilization on the in vitro and in vivo function of reconstituted extracellular matrix-derived scaffolds.

Reconstituted extracellular matrix (ECM)-derived scaffolds are commonly utilized in preclinical tissue engineering studies as delivery vehicles for cells and growth factors. Translation into clinical use requires identifying a sterilization method that effectively removes bacteria but does not harm scaffold function. To determine effectiveness of sterilization and impact on ECM scaffold integrity and function, low-temperature ethylene oxide and 15 kGy electron beam irradiation techniques were evaluated. Scaffold sterility was assessed in accordance to United States Pharmacopeia Chapter 71. Scaffold matrix degradation was determined in vitro using enzymatic resistance tests and gel electrophoresis. Scaffold mechanics including elastic modulus, yield stress and collapse modulus were tested. Lastly, 14 Yorkshire pigs underwent ACL transection and bio-enhanced ACL repair using sterilized scaffolds. Histologic response of ligament, synovium, and lymph nodes was compared at 4, 6, and 8 weeks. Ethylene oxide as well as electron beam irradiation yielded sterile scaffolds. Scaffold resistance to enzymatic digestion and protein integrity slightly decreased after electron beam irradiation while ethylene oxide altered scaffold matrix. Scaffold elastic modulus and yield stress were increased after electron beam treatment, while collapse modulus was increased after ethylene oxide treatment. No significant changes in ACL dimensions, in vivo scaffold resorption rate, or histologic response of synovium, ligament, and lymph nodes with either terminal sterilization technique were detectable. In conclusion, this study identifies two methods to terminally sterilize an ECM scaffold. In vitro scaffold properties were slightly changed without significantly influencing the biologic responses of the surrounding tissues in vivo. This is a critical step toward translating new tissue engineering strategies to clinical trials.

Electron beam sterilization does not have a detrimental effect on the ability of extracellular matrix scaffolds to support in vivo ligament healing.

Extracellular matrix (ECM) scaffolds have been used to enhance anterior cruciate ligament (ACL) repair in large animal models. To translate this technology to clinical care, identifying a method which effectively sterilizes the material without significantly impairing in vivo function is desirable. Sixteen Yorkshire pigs underwent ACL transection and were randomly assigned to bridge-enhanced ACL repair-primary suture repair of the ACL with addition of autologous blood soaked ECM scaffold--with either (i) an aseptically processed ECM scaffold, or (ii) an electron beam irradiated ECM scaffold. Primary outcome measures included sterility of the scaffold and biomechanical properties of the scaffold itself and the repaired ligament at 8 weeks after surgery. Scaffolds treated with 15 kGy electron beam irradiation had no bacterial or fungal growth noted, while aseptically processed scaffolds had bacterial growth in all tested samples. The mean biomechanical properties of the scaffold and healing ligament were lower in the electron beam group; however, differences were not statistically significant. Electron beam irradiation was able to effectively sterilize the scaffolds. In addition, this technique had only a minimal impact on the in vivo function of the scaffolds when used for ligament healing in the porcine model.

Evaluation of human acellular dermis versus porcine acellular dermis in an in vivo model for incisional hernia repair.

Incisional hernias commonly occur following abdominal wall surgery. Human acellular dermal matrices (HADM) are widely used in abdominal wall defect repair. Xenograft acellular dermal matrices, particularly those made from porcine tissues (PADM), have recently experienced increased usage. The purpose of this study was to compare the effectiveness of HADM and PADM in the repair of incisional abdominal wall hernias in a rabbit model. A review from earlier work of differences between human allograft acellular dermal matrices (HADM) and porcine xenograft acellular dermal matrices (PADM) demonstrated significant differences (P < 0.05) in mechanical properties: Tensile strength 15.7 MPa vs. 7.7 MPa for HADM and PADM, respectively. Cellular (fibroblast) infiltration was significantly greater for HADM vs. PADM (Armour). The HADM exhibited a more natural, less degraded collagen by electrophoresis as compared to PADM. The rabbit model surgically established an incisional hernia, which was repaired with one of the two acellular dermal matrices 3 weeks after the creation of the abdominal hernia. The animals were euthanized at 4 and 20 weeks and the wounds evaluated. Tissue ingrowth into the implant was significantly faster for the HADM as compared to PADM, 54 vs. 16% at 4 weeks, and 58 vs. 20% for HADM and PADM, respectively at 20 weeks. The original, induced hernia defect (6 cm(2)) was healed to a greater extent for HADM vs. PADM: 2.7 cm(2) unremodeled area for PADM vs. 1.0 cm² for HADM at 20 weeks. The inherent uniformity of tissue ingrowth and remodeling over time was very different for the HADM relative to the PADM. No differences were observed at the 4-week end point. However, the 20-week data exhibited a statistically different level of variability in the remodeling rate with the mean standard deviation of 0.96 for HADM as contrasted to a mean standard deviation of 2.69 for PADM. This was significant with P < 0.05 using a one tail F test for the inherent variability of the standard deviation. No significant differences between the PADM and HADM for adhesion, inflammation, fibrous tissue or neovascularization were noted.

Abdominal ventral hernia repair with current biological prostheses: an experimental large animal model.

Biologic prostheses have emerged to address the limitations of synthetic materials for ventral hernia repairs; however, they lack experimental comparative data. Fifteen swine were randomly assigned to 1 of 3 bioprosthetic groups (DermaMatrix, AlloDerm, and Permacol) after creation of a full thickness ventral fascial defect. At 15 weeks, host incorporation, hernia recurrence, adhesion formation, neovascularization, inflammation, and biomechanical properties were assessed. No animals had hernia recurrence or eventration. DermaMatrix and Alloderm implants demonstrated more adhesions, greater inflammatory infiltration, and more longitudinal laxity, but near identical neovascularization and tensile strength to Permacol. We found that porcine acellular dermal products (Permacol) contain following essential properties of an ideal ventral hernia repair material: low inflammation, less elastin and stretch, lower adhesion rates and cost, and more contracture. The addition of lower cost xenogeneic acellular dermal products to the repertoire of available acellular dermal products demonstrates promise, but requires long-term clinical studies to verify advantages and efficacy.

Homeostatic role of transforming growth factor-beta in the oral cavity and esophagus of mice and its expression by mast cells in these tissues.

Transforming growth factor-beta (TGF-beta) is a pleiotropic growth factor; its overexpression has been implicated in many diseases, making it a desirable target for therapeutic neutralization. In initial safety studies, mice were chronically treated (three times per week) with high doses (50 mg/kg) of a murine, pan-neutralizing, anti-TGF-beta antibody. Nine weeks after the initiation of treatment, a subset of mice exhibited weight loss that was concurrent with decreased food intake. Histopathology revealed a unique, nonneoplastic cystic epithelial hyperplasia and tongue inflammation, as well as dental dysplasia and epithelial hyperplasia and inflammation of both the gingiva and esophagus. In an effort to determine the cause of this site-specific pathology, we examined TGF-beta expression in these tissues and saliva under normal conditions. By immunostaining, we found higher expression levels of active TGF-beta1 and TGF-beta3 in normal tongue and esophageal submucosa compared with gut mucosal tissues, as well as detectable TGF-beta1 in normal saliva by Western blot analysis. Interestingly, mast cells within the tongue, esophagus, and skin co-localized predominantly with the TGF-beta1 expressed in these tissues. Our findings demonstrate a novel and restricted pathology in oral and esophageal tissues of mice chronically treated with anti-TGF-beta that is associated with basal TGF-beta expression in saliva and by mast cells within these tissues. These studies illustrate a previously unappreciated biological role of TGF-beta in maintaining homeostasis within both oral and esophageal tissues.