Development of in situ forming implants for controlled delivery of punicalagin.

Due to efficient drainage of the joint, the development of intra-articular depots for long-lasting drug release is a difficult challenge. Moreover, a disease-modifying osteoarthritis drug (DMOAD) that can effectively manage osteoarthritis has yet to be identified. The current study was undertaken to explore the potential of injectable, in situ forming implants to create depots that support the sustained release of punicalagin, a promising DMOAD. In vitro experiments demonstrated punicalagin's ability to suppress production of interleukin-1ß and prostaglandin E2, confirming its chondroprotective properties. Regarding the entrapment of punicalagin, it was demonstrated by LC-MS/MS to be stable within PLGA in situ forming implants for several weeks and capable of inhibiting collagenase upon release. In vitro punicalagin release kinetics were tunable through variation of solvent, PLGA lactide:glycolide ratio, and polymer concentration, and an optimized formulation supported release for approximately 90 days. The injection force of this formulation steadily increased with plunger advancement and higher rates of advancement were associated with greater forces. Although the optimal formulation was highly cytotoxic to primary chondrocytes if cells were exposed immediately or shortly after implant formation, upwards of 70 % survival was achieved when the implants were first allowed to undergo a 24-72 h period of phase inversion prior to cell exposure. This study demonstrates a PLGA-based in situ forming implant for the controlled release of punicalagin. With modification to address cytotoxicity, such an implant may be suitable as an intra-articular therapy for OA.

Laser microgrooving and resorbable blast texturing for enhanced surface function of titanium alloy for dental implant applications.

Long-term dental implant success is dependent on biocompatibility and osseointegration between the bone and the implant. Surface modifications such as laser-induced microgrooving which increase contact area can enhance osseointegration by establishing and directing a stable attachment between the implant surface and peri-implant bone. The objective of this study was to evaluate pre-osteoblast proliferation, morphology, and differentiation on titanium alloy (Ti64) surfaces-Laser-Lok© (LL), resorbable blast textured (RBT), and machined (M)-compared to tissue culture plastic (TCP) control. We hypothesized the LL surfaces would facilitate increased cellular alignment compared to all other groups, and LL and RBT surfaces would demonstrate enhanced proliferation and differentiation compared to M and TCP surfaces. Surface roughness was quantified using a surface profilometer, and water contact angle was measured to evaluate the hydrophilicity of the surfaces. Cellular function was assessed using quantitative viability and differentiation assays and image analyses, along with qualitative fluorescent (viability and cytoskeletal) imaging and scanning electron microscopy. No differences in surface roughness were observed between groups. Water contact angle indicated LL was the least hydrophilic surface, with RBT and M surfaces exhibiting greater hydrophilicity. Cell proliferation on day 2 was enhanced on both LL and RBT surfaces compared to M, and all three groups had higher cell numbers on day 2 compared to day 1. Cell orientation was driven by the geometry of the surface modification, as cells were more highly aligned on LL surfaces compared to TCP (on day 2) and RBT (on day 3). At day 21, cell proliferation was greater on LL, RBT, and TCP surfaces compared to M, though no differences in osteogenic differentiation were observed. Collectively, our results highlight the efficacy of laser microgrooved and resorbable blast textured surface modifications of Ti64 for enhancing cellular functions, which may facilitate improved osseointegration of dental implants.

Comparison of the Capacitance of a Cyclically Fatigued Stretch Sensor to a Non-Fatigued Stretch Sensor When Performing Static and Dynamic Foot-Ankle Motions.

Motion capture is the current gold standard for assessing movement of the human body, but laboratory settings do not always mimic the natural terrains and movements encountered by humans. To overcome such limitations, a smart sock that is equipped with stretch sensors is being developed to record movement data outside of the laboratory. For the smart sock stretch sensors to provide valuable feedback, the sensors should have durability of both materials and signal. To test the durability of the stretch sensors, the sensors were exposed to high-cycle fatigue testing with simultaneous capture of the capacitance. Following randomization, either the fatigued sensor or an unfatigued sensor was placed in the plantarflexion position on the smart sock, and participants were asked to complete the following static movements: dorsiflexion, inversion, eversion, and plantarflexion. Participants were then asked to complete gait trials. The sensor was then exchanged for either an unfatigued or fatigued plantarflexion sensor, depending upon which sensor the trials began with, and each trial was repeated by the participant using the opposite sensor. Results of the tests show that for both the static and dynamic movements, the capacitive output of the fatigued sensor was consistently higher than that of the unfatigued sensor suggesting that an upwards drift of the capacitance was occurring in the fatigued sensors. More research is needed to determine whether stretch sensors should be pre-stretched prior to data collection, and to also determine whether the drift stabilizes once the cyclic softening of the materials comprising the sensor has stabilized.

Evaluation of Electrospun PCL-PLGA for Sustained Delivery of Kartogenin.

In this study, kartogenin was incorporated into an electrospun blend of polycaprolactone and poly(lactic-co-glycolic acid) (1:1) to determine the feasibility of this system for sustained drug delivery. Kartogenin is a small-molecule drug that could enhance the outcome of microfracture, a cartilage restoration procedure, by selectively stimulating chondrogenic differentiation of endogenous bone marrow mesenchymal stem cells. Experimental results showed that kartogenin did not affect the electrospinnability of the polymer blend, and it had negligible effects on fiber morphology and scaffold mechanical properties. The loading efficiency of kartogenin into electrospun membranes was nearly 100%, and no evidence of chemical reaction between kartogenin and the polymers was detected by Fourier transform infrared spectroscopy. Analysis of the released drug using high-performance liquid chromatography-photodiode array detection indicated an abundance of kartogenin and only a small amount of its major hydrolysis product. Kartogenin displayed a typical biphasic release profile, with approximately 30% being released within 24 h followed by a much slower, constant rate of release up to 28 days. Although additional development is needed to tune the release kinetics and address issues common to electrospun scaffolds (e.g., high fiber density), the results of this study demonstrated that a scaffold electrospun from biodegradable synthetic polymers is a suitable kartogenin delivery vehicle.

Effects of short-duration treatment of cartilage with punicalagin and genipin and the implications for treatment of osteoarthritis.

Punicalagin (PA) not only binds type II collagen, but also blocks its MMP-13-mediated degradation, and genipin (GNP) is a collagen cross-linking agent. We hypothesized that these drugs could mitigate the loss of cartilage if administered in the early phase of osteoarthritis, and experiments were designed to provide proof-of-concept. Porcine cartilage was exposed to both drugs in a manner designed to simulate intra-articular (IA) injection. Based on penetration of PA into cartilage, the rate of drug diffusion was conservatively estimated at 2 µm per minute. GNP caused a measurable degree of cross-linking, increased compressive resistance and coefficient of friction, and substantially inhibited degradation by collagenase, but not by hyaluronidase. Pre-incubation of GNP with collagenase had no effect on enzymatic activity. PA did not cross-link collagen nor affect the mechanical properties of cartilage. It did, however, increase resistance to degradation by collagenase and hyaluronidase. Furthermore, it reacted with collagenase in solution and inhibited its subsequent enzymatic activity. Effects of PA and GNP were not additive. The chondroprotective effect of semi-weekly IA injections was investigated in the monoiodoacetate-induced model of OA in rats. Quantitative histology suggested that injection of PA decreased the amount of cartilage lost compared to saline-injected controls, and the addition of GNP made no difference. This study supports the notion that IA delivery of PA could mitigate OA-induced cartilage erosion.

Assessment of erythrocyte damage and in-line pressure changes associated with simulated transfusion of canine blood through microaggregate filters.

OBJECTIVE: To determine whether passage of whole blood through a microaggregate filter by use of a syringe pump would damage canine erythrocytes. SAMPLE: Blood samples obtained from 8 healthy client-owned dogs. PROCEDURES: Whole blood was passed through a standard microaggregate filter by use of a syringe pump at 3 standard administration rates (12.5, 25, and 50 mL/h). Prefilter and postfilter blood samples were collected at the beginning and end of a simulated transfusion. Variables measured at each time point included erythrocyte osmotic fragility, mean corpuscular fragility, RBC count, hemoglobin concentration, RBC distribution width, and RBC morphology. In-line pressure when blood passed through the microaggregate filter was measured continuously throughout the simulated transfusion. After the simulated transfusion was completed, filters were visually analyzed by use of scanning electron microscopy. RESULTS: Regardless of administration rate, there was no significant difference in mean corpuscular fragility, RBC count, hemoglobin concentration, or RBC distribution width between prefilter and postfilter samples. Additionally, there were no differences in in-line pressure during the simulated transfusion among administration rates. Echinocytes were the erythrocyte morphological abnormality most commonly observed at the end of the transfusion at administration rates of 12.5 and 25 mL/h. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that regardless of the administration rate, the microaggregate filter did not alter fragility of canine RBCs, but may have altered the morphology. It appeared that the microaggregate filter would not contribute to substantial RBC damage for transfusions performed with a syringe pump.

Effects of short- and long-term administration of nonsteroidal anti-inflammatory drugs on osteotomy healing in dogs.

OBJECTIVE: To determine the influence of short-term administration of carprofen on bone healing in dogs. STUDY DESIGN: Randomized controlled experimental study. ANIMALS: Eighteen purpose-bred sexually mature hound dogs. METHODS: Tibial osteotomies were performed, and dogs were divided into three groups: no carprofen (n = 6), 2-week administration of carprofen at 2.2 mg/kg twice daily (n = 6), and 8-week administration of carprofen at 2.2 mg/kg twice daily (n = 5). Bone healing was evaluated radiographically at 4 and 8 weeks postoperatively. Postmortem, fracture healing was assessed via biomechanical testing (three-point bending), histological cartilage:callus ratio, and bone mineral density (BMD) with quantitative computed tomography. RESULTS: No biomechanical difference was detected between dogs that received no carprofen and those that received 2 weeks of carprofen or between those that received 2 weeks vs 8 weeks of carprofen. Stiffness (P = .035) and maximum stress (P = .042) were higher in dogs that received no carprofen than in those that received 8 weeks of carprofen. Radiographic healing did not differ between dogs without carprofen and those with 2-week administration of carprofen (P = .9923). However, tibias of dogs without carprofen and those with 2-week administration of carprofen were more healed compared with those in the 8-week-carprofen group at 4 and 8 weeks after surgery (P = .0011). No treatment effect was detected by cartilage:callus ratio or BMD. CONCLUSION: Long-term administration of carprofen had a negative effect on bone healing compared with short-term or no administration of carprofen. CLINICAL SIGNIFICANCE: Nonsteroidal anti-inflammatory drugs should be used cautiously in dogs at risk for delayed bone healing, and administration should be discontinued beyond the perioperative period in dogs with fractures or osteotomies.

Influence of Microtextured Implant Surfaces on Peri-implantitis and Its Treatment: A Preclinical Trial.

PURPOSE: The prevalence of peri-implantitis has increased significantly, forcing clinicians to search for ways to prevent it. Laser-microtextured surfaces promote soft tissue attachment and provide a tight seal around implants. Hence, the aim of this study was to examine the clinical, radiographic, and histologic features of ligature-induced peri-implantitis, as well as the effect of surgical treatment of these induced peri-implantitis lesions on laser-microtextured implants in a controlled animal model. MATERIALS AND METHODS: Six mini-pigs (three males/three females) received 6 implants each (3 resorbable blast textured [RBT] implants and 3 laser-microtextured [LM] implants) in mandibular premolar sites, for a total of 36 implants. Two groups were identified based on the time point of sample analysis. After osseointegration was achieved, metal wire ligatures were placed and left for 12 weeks. Group 1 samples were then obtained, and group 2 samples received rescue therapy following a guided bone regeneration (GBR) protocol. Sample collection in group 2 was completed 12 weeks after the samples were submerged and treated. All samples were analyzed histologically and measurements were taken. RESULTS: Four implants (three RBT, one LM) were lost at early time points because of implant instability. Interimplant distances and soft tissue thicknesses varied subtly between groups. More notable was the mean (± standard error of the mean) crestal bone loss (group 1: 1.860 ± 1.618 mm [LM] and 2.440 ± 2.691 mm [RBT]; group 2: 2.04 ± 1.613 mm [LM] and 3.00 ± 2.196 mm [RBT]) (P < .05), as demonstrated by a paired t test. Histologic pocket depth was also greater at RBT sites than at LM sites (4.448 ± 2.839 mm and 4.121 ± 2.251 mm, respectively, in group 1; and 3.537 ± 2.719 mm and 2.339 ± 1.852 mm, respectively [P < .005] in group 2). CONCLUSION: LM implants had less crestal bone loss and shallower histologic pocket depth compared with their RBT counterparts. Also, LM implants had higher bone fill when a rescue therapy (GBR) was performed.

Suitability of EGCG as a Means of Stabilizing a Porcine Osteochondral Xenograft.

As a non-crosslinked osteochondral xenograft would be mechanically inferior to native cartilage and vulnerable to premature degradation, we seek a safe and effective method of xenograft stabilization. The purpose of this study was to evaluate the capacity for epigallocatechin gallate (EGCG) to stabilize a decellularized porcine osteochondral xenograft through collagen crosslinking. Our objectives were to assess the effects of EGCG on the degree of crosslinking, mechanical properties, collagenase resistance, cytotoxicity, and in vitro biocompatibility. EGCG is a green tea polyphenol that acts as a collagen crosslinker. Porcine osteochondral plugs were decellularized and then crosslinked by soaking in EGCG. The degree of crosslinking, cartilage compressive stiffness, cartilage-bone interface strength, coefficient of friction, and residual mass after collagenase exposure all increased with an increasing EGCG concentration. With the exception of the coefficient of friction, EGCG treatment could restore mechanical properties to levels equal to, or exceeding those, of native cartilage. EGCG treatment profoundly increased the enzymatic resistance, and 1% EGCG provided protection equivalent to 1% glutaraldehyde. EGCG up to 0.5 mM was essentially not cytotoxic to chondrocytes embedded in alginate, and autologous chondrocytes attached to decellularized, EGCG-fixed cartilage were all viable five days after seeding. Results demonstrate that EGCG has many beneficial effects on a decellularized osteochondral xenograft, and may be suitable for use in stabilizing such a graft prior to implantation for the repair of a defect.

A biomechanical comparison of conventional dynamic compression plates and string-of-pearls™ locking plates using cantilever bending in a canine Ilial fracture model.

BACKGROUND: Fracture of the ilium is common orthopedic injury that often requires surgical stabilization in canine patients. Of the various methods of surgical stabilization available, application of a lateral bone plate to the ilium is the most common method of fixation. Many plating options are available, each having its own advantages and disadvantages. The purpose of this study was to evaluate the biomechanical properties of a 3.5 mm String-of-Pearls™ plate and a 3.5 mm dynamic compression plate in a cadaveric canine ilial fracture model. Hemipelves were tested in cantilever bending to failure and construct stiffness, yield load, displacement at yield, ultimate load, and mode of failure were compared. RESULTS: The mean stiffness of dynamic compression plate (116 ± 47 N/mm) and String-of-Pearls™ plate (107 ± 18 N/mm) constructs, mean yield load of dynamic compression plate (793 ± 333 N) and String-of-Pearls™ plate (860 ± 207 N) constructs, mean displacement at yield of dynamic compression plate (8.6 ± 3.0 mm) and String-of-Pearls™ plate (10.2 ± 2.8 mm) constructs, and ultimate load at failure of dynamic compression plate (936 ± 320 N) and String-of-Pearls™ plate (939 ± 191 N) constructs were not significantly different. No differences were found between constructs with respect to mode of failure. CONCLUSIONS: No significant biomechanical differences were found between String-of-Pearls™ plate and dynamic compression plate constructs in this simplified cadaveric canine ilial fracture model.

Evaluation of genipin for stabilization of decellularized porcine cartilage.

We speculate that an acellular osteochondral xenograft may be a good alternative to allografts for repair of focal articular cartilage lesions. In order to make a xenograft resistant to enzymatic degradation and to prevent a chronic immune response it may be beneficial to stabilize it through crosslinking. The concept is analogous to treatment of porcine bioprosthetic heart valves with glutaraldehyde. The purpose of this study was to evaluate genipin, a natural crosslinking agent with low cytotoxicity, for stabilization of decellularized cartilage. Porcine articular cartilage discs were decellularized in SDS and nucleases and then crosslinked in genipin. The utility of genipin was determined from its effects on degree of crosslinking, mechanical properties, dimensional stability, enzymatic resistance, and in vitro biocompatibility. Degree of crosslinking, compressive moduli, and collagenase resistance varied over a wide range depending on genipin concentration. The equilibrium compressive modulus could be increased from approximately 50% to more than 120% that of native cartilage, and the time to complete degradation by collagenase could be extended from less than 12 h to more than 15 days. Radial shrinkage of approximately 4% was observed at a genipin concentration of 0.1% wt/vol, and cartilage coefficient of friction against glass increased in a concentration-dependent manner. Autologous chondrocytes displayed little difference in viability or their ability to attach and spread over the surface of genipin-fixed cartilage compared to non-crosslinked cartilage during 6 weeks of culture. These results indicate that genipin may be efficacious for stabilization of a decellularized porcine osteochondral xenograft. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1949-1957, 2017.

Biomechanical Testing and Histologic Examination of Intradermal Skin Closure in Dogs Using Barbed Suture Device and Non-Barbed Monofilament Suture.

OBJECTIVE: To compare the biomechanical strength and histologic features of 3-0 Glycomer™ 631 barbed suture (V-LOC™ 90 Absorbable Wound Closure Device, Covidien, Mansfield, MA) to non-barbed 3-0 Glycomer™ 631 suture (Biosyn™, Covidien) for intradermal skin wound closure in the dog. STUDY DESIGN: Randomized, factorial, in vivo. ANIMALS: Eighteen purpose-bred, mature male, and female hound dogs. METHODS: Eighteen adult hound dogs were randomly assigned to 1 of 3 groups designated by postoperative day of assessment. Six skin incisions were made along the dorsum in the thoracolumbar region of each dog with an equal number (n=3) randomly assigned to closure with barbed or non-barbed suture. Six dogs were euthanatized on postoperative days 3, 10, and 14, respectively. Two additional incisions were made on each dog after euthanasia for baseline data (Day 0). The skin incision specimens were harvested for biomechanical testing and histologic evaluation. RESULTS: Non-barbed closure had significantly higher maximum load at failure (P<.001) and stiffness (P<.001) than barbed closure regardless of day. The average tissue reaction score was significantly higher for barbed closure (P=.008), regardless of day. Suturing time for barbed closures was significantly shorter. There was no significant difference in frequency of complications between closures. CONCLUSION: Barbed Glycomer™ 631 closures had a significantly lower maximum load at failure and stiffness, and higher average tissue reaction scores, but showed no difference in short term outcome for intradermal closure of dorsally located skin incisions in dogs.

Pullout strength of monocortical and bicortical screws in metaphyseal and diaphyseal regions of the canine humerus.

OBJECTIVE: Monocortical screws are commonly employed in locking plate fixation, but specific recommendations for their placement are lacking and use of short monocortical screws in metaphyseal bone may be contraindicated. Objectives of this study were to evaluate axial pullout strength of two different lengths of monocortical screws placed in various regions of the canine humerus compared to bicortical screws, and to derive cortical thickness and bone density values for those regions using quantitative computed tomography analysis (QCT). METHODS: The QCT analysis was performed on 36 cadaveric canine humeri for six regions of interest (ROI). A bicortical, short monocortical, or 50% transcortical 3.5 mm screw was implanted in each ROI and axial pullout testing was performed. RESULTS: Bicortical screws were stronger than monocortical screws in all ROI except the lateral epicondylar crest. Short monocortical metaphyseal screws were weaker than those placed in other regions. The 50% transcortical screws were stronger than the short monocortical screws in the condyle. A linear relationship between screw length and pullout strength was observed. CLINICAL SIGNIFICANCE: Cortical thickness and bone density measurements were obtained from multiple regions of the canine humerus using QCT. Use of short monocortical screws may contribute to failure of locking plate fixation of humeral fractures, especially when placed in the condyle. When bicortical screw placement is not possible, maximizing monocortical screw length may optimize fixation stability for distal humeral fractures.

Comparison of natural crosslinking agents for the stabilization of xenogenic articular cartilage.

Osteochondral xenografts are potentially inexpensive, widely available alternatives to fresh allografts. However, antigen removal from xenogenic cartilage may damage the extracellular matrix and reduce compressive stiffness. Non-crosslinked xenogenic cartilage may also undergo rapid enzymatic degradation in vivo. We hypothesized that natural crosslinking agents could be used in place of glutaraldehyde to improve the mechanical properties and enzymatic resistance of decellularized cartilage. This study compared the effects of genipin (GNP), proanthocyanidin (PA), and epigallocatechin gallate (EGCG), on the physical and mechanical properties of decellularized porcine cartilage. Glutaraldehyde (GA) served as a positive control. Porcine articular cartilage discs were decellularized in 2% sodium dodecyl sulfate and DNase I followed by fixation in 0.25% GNP, 0.25% PA, 0.25% EGCG, or 2.5% GA. Decellularization decreased DNA by 15% and GAG by 35%. For natural crosslinkers, the average degree of crosslinking ranged from approximately 50% (EGCG) to 78% (GNP), as compared to 83% for the GA control. Among the natural crosslinkers, only GNP significantly affected the disc diameter, and shrinkage was under 2%. GA fixation had no significant effect on disc diameter. Decellularization decreased aggregate modulus; GA and GNP, but not EGCG and PA, were able to restore it to its original level. GNP, PA, and GA conferred a similar, almost complete resistance to collagenase degradation. EGCG also conferred substantial resistance but to a lesser degree. Overall, the data support our hypothesis and suggest that natural crosslinkers may be suitable alternatives to glutaraldehyde for stabilization of decellularized cartilage. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1037-1046, 2016.

Effect of platelet-rich plasma on chondrogenic differentiation in three-dimensional culture.

Platelet-rich plasma (PRP) may have the potential to enhance articular cartilage regeneration through release of growth factors including transforming growth factor isoforms. The purpose of this study was to investigate the potential for PRP to stimulate chondrogenic differentiation in three-dimensional PRP hydrogel constructs. Allogenic PRP was prepared using a double centrifugation protocol which resulted in a platelet concentration approximately 250% above baseline. Canine marrow stromal cells were encapsulated at 6.8×10(6) cells/ml in either 2% sodium alginate or in a 3:1 mixture of freshly prepared PRP and 2% alginate. PRP and alginate beads were cultured in chemically defined chondrogenic medium with and without 10 ng/ml TGF-ß3. PRP cultures were additionally supplemented with frozen-thawed PRP. In the absence of TGF-ß3, PRP had a mild stimulatory effect on cell proliferation. PRP did not stimulate cell proliferation in the presence of TGF-ß3. Cells exposed to TGF-ß3 accumulated significantly more GAG/DNA than those which were not, but there was not a statistically significant difference between alginate and PRP. Total collagen content was greater in PRP than in alginate, regardless of TGF-ß3. Chondrogenesis in PRP was qualitatively and spatially different than that which occurred in conventional alginate beads and was characterized by isolated centers of intense chondrogenesis. Overall the results demonstrate that PRP alone weakly promotes chondroinduction of marrow stromal cells, and the effect is greatly augmented by TGF-ß3.

Attachment, proliferation, and chondroinduction of mesenchymal stem cells on porous chitosan-calcium phosphate scaffolds.

Symptomatic osteochondral lesions occur frequently, but relatively few treatment options are currently available. The purpose of this study was to conduct a preliminary investigation into a new tissue engineering approach to osteochondral regeneration. The concept is a biphasic construct consisting of a porous, osteoconductive chitosan-calcium phosphate scaffold supporting a layer of neocartilage formed by marrow-derived mesenchymal stem cells. Two experiments were conducted to assess the feasibility of this approach. The first experiment characterized the attachment efficiency and proliferation of primary human marrow-derived mesenchymal stem cells seeded relatively sparely onto the scaffold's surface. The second experiment compared two different methods of creating a biphasic construct using a much higher density of primary porcine marrow stromal cells. About 40% of the sparsely seeded human cells attached and proliferated rapidly. Constructs formed by one of the two experimental techniques exhibited a layer of cartilaginous tissue which only partially covered the scaffold's surface due to inadequate adhesion between the cells and the scaffold. This study demonstrates some potential for the approach to yield an implantable biphasic construct, but further development is required to improve cell-scaffold adhesion.

Structural and biomechanical characterizations of porcine myocardial extracellular matrix.

Extracellular matrix (ECM) of myocardium plays an important role to maintain a multilayered helical architecture of cardiomyocytes. In this study, we have characterized the structural and biomechanical properties of porcine myocardial ECM. Fresh myocardium were decellularized in a rotating bioreactor using 0.1 % sodium dodecyl sulfate solution. Masson's trichrome staining and SEM demonstrated the removal of cells and preservation of the interconnected 3D cardiomyocyte lacunae. Movat's pentachrome staining showed the preservation of cardiac elastin ultrastructure and vascular elastin distribution/alignment. DNA assay result confirmed a 98.59 % reduction in DNA content; the acellular myocardial scaffolds were found completely lack of staining for the porcine α-Gal antigen; and the accelerating enzymatic degradation assessment showed a constant degradation rate. Tensile and shear properties of the acellular myocardial scaffolds were also evaluated. Our observations showed that the acellular myocardial ECM possessed important traits of biodegradable scaffolds, indicating the potentials in cardiac regeneration and whole heart tissue engineering.

A comparative biomechanical analysis of term fetal membranes in human and domestic species.

OBJECTIVE: The purpose of this study was to biomechanically characterize and compare human, porcine, equine, and ovine fetal membranes. STUDY DESIGN: Noncontact metrology was used for topographic analyses. Uniaxial tensile testing was performed to resolve specific biomechanical values. Puncture force and radial stresses were determined with biaxial puncture testing. Microstructure and surface tortuosity were analyzed histologically. RESULTS: Equine and human membranes sustained larger magnitude loading, but ovine and porcine membranes exhibited stronger material properties. Biaxial puncture validated uniaxial results; human and equine groups accommodated the largest loads but lowest stresses. Equine membranes were mostly vascularized; tortuosity was highest in porcine membranes. Species' gestation length was correlated positively with membrane thickness. CONCLUSION: The anatomy of placentation and length of species gestation show distinct relationships to membrane biomechanics. Unlike other species, human fetal membranes do not compensate for structural weakness with a thicker membrane. This finding may explain the high incidence of preterm premature rupture of membranes in humans.

Effect of a mechanical stimulation bioreactor on tissue engineered, scaffold-free cartilage.

Achieving sufficient functional properties prior to implantation remains a significant challenge for the development of tissue engineered cartilage. Many studies have shown chondrocytes respond well to various mechanical stimuli, resulting in the development of bioreactors capable of transmitting forces to articular cartilage in vitro. In this study, we describe the production of sizeable, tissue engineered cartilage using a novel scaffold-free approach, and determine the effect of perfusion and mechanical stimulation from a C9-x Cartigen bioreactor on the properties of the tissue engineered cartilage. We created sizable tissue engineered cartilage from porcine chondrocytes using a scaffold-free approach by centrifuging a high-density chondrocyte cell-suspension onto an agarose layer in a 50 mL tube. The gross and histological appearances, biochemical content, and mechanical properties of constructs cultured in the bioreactor for 4 weeks were compared to constructs cultured statically. Mechanical properties were determined from unconfined uniaxial compression tests. Constructs cultured in the bioreactor exhibited an increase in total GAG content, equilibrium compressive modulus, and dynamic modulus versus static constructs. Our study demonstrates the C9-x CartiGen bioreactor is able to enhance the biomechanical and biochemical properties of scaffold-free tissue engineered cartilage; however, no additional enhancement was seen between loaded and perfused groups.

The effect of stifle angle on cranial tibial translation following tibial plateau leveling osteotomy: an in vitro experimental analysis.

This study was designed to determine the ability of tibial plateau leveling osteotomy (TPLO) to eliminate cranial tibial translation (CTT) through a loaded range of motion. Twenty-four large-breed canine cadaver limbs were compared. Each limb was placed in a custom designed jig at 120° of stifle extension under an axial load of 20% body weight. A force of approximately 10 N/s mimiced the action of the quadriceps muscle and allowed the limb to move from 120° to maximal extension. Positional data were acquired using electromagnetic motion-tracking sensors. Each limb was tested under normal, cranial cruciate ligament (CrCL)-deficient, and TPLO-treated conditions. Cranial tibial translation significantly increased after transection of the CrCL. The TPLO failed to normalize CTT within the CrCL deficient stifle; however, values trended towards intact values throughout the range of motion. The TPLO was more effective at higher angles of flexion. These altered biomechanics may help explain the continued progression of osteoarthritis in TPLO repaired stifles. This loaded model may serve as a method for future evaluation of other surgical techniques.