Porcine Tendon Research Articles

Biomechanical comparison of double grasping repair versus cross-locked cruciate flexor tendon repair.

This study was conducted to compare the in vitro biomechanical properties of tensile strength and gap resistance of a double grasping loop (DGL) flexor tendon repair with the established four-strand cross-locked cruciate (CLC) flexor tendon repair, both with an interlocking horizontal mattress (IHM) epitendinous suture. The hypothesis is that the DGL-IHM method which utilizes two looped core sutures, grasping and locking loops, and a single intralesional knot will have greater strength and increased gap resistance than the CLC-IHM method. Forty porcine tendons were evenly assigned to either the DGL-IHM or CLC-IHM group. The tendon repair strength, 2-mm gap force and load to failure, was measured under a constant rate of distraction. The stiffness of tendon repair was calculated and the method of repair failure was analyzed. The CLC-IHM group exhibited a statistically significant greater resistance to gapping, a statistically significant higher load to 2-mm gapping (62.0N), and load to failure (99.7N) than the DGL-IHM group (37.1N and 75.1N, respectively). Ninety percent of CLC-IHM failures were a result of knot failure whereas 30% of the DGL-IHM group exhibited knot failure. This study demonstrates that the CLC-IHM flexor tendon repair method better resists gapping and has a greater tensile strength compared to the experimental DGL-IHM method. The authors believe that while the DGL-IHM provides double the number of sutures at the repair site per needle pass, this configuration does not adequately secure the loop suture to the tendon, resulting in a high percentage of suture pullout and inability to tolerate loads as high as those of the CLC-IHM group.

Evaluation of porcine xenograft in collateral ligament reconstruction in beagle dogs

This study evaluated the effectiveness of irradiated porcine tendon xenografts for lateral collateral ligament (LCL) reconstruction. Twenty healthy adult beagle dogs underwent LCL reconstruction using irradiated porcine tendons treated with poly-gamma-glutamic acid. Serological and histological assessments were performed to evaluate host immunological response at 3 and 12 months after surgery. The healing and functional integrity of the LCL reconstructions were assessed by mechanical testing and gait analysis. Histological assessment of the porcine xenografts showed gradual host cellular infiltration and graft collagen remodeling during the healing process. Porcine xenografts showed angiogenesis and no signs of inflammatory reaction. Additionally, biomechanical and gait evaluations supported graft functional integration with no differences between normal and porcine xenograft reconstruction at 12 months after surgery. Irradiated porcine xenografts showed greater cellular responses and healing properties in short- and long-term evaluations. Irradiated porcine tendons appear to be useful as xenografts for the reconstruction of damaged ligaments.

In Vitro Characteristics of Porcine Tendon Hydrogel for Tendon Regeneration

PURPOSE: Previous work has characterized the development of a human tendon hydrogel capable of improving mechanical strength following tendon injury.1 Animal tendon hydrogel has not yet been described, but would prove beneficial due to the cost and ethical concerns associated with the use of human cadaveric tendon. This study details the manufacture and assesses the biocompatibility of porcine tendon hydrogel seeded with human adipoderived stem cells (ASCs). METHODS AND MATERIALS: Porcine tendon was dissected from surrounding connective and muscle tissue and decellularized via 0.2% SDS and 0.2% SDS/EDTA wash solutions before lyophilization. Tendon was milled and reconstituted by previously described methods.2 Decellularization was confirmed by H and E staining, SYTO Green 11 nucleic acid dye, and DNeasy assay. The protein composition of milled tendon matrix before and after digestion was identified by mass spectrometry. Rheological properties were determined using an ARG2 rheometer. Biocompatibility was assessed by live/dead assay. The proliferation of human ASCs seeded in porcine and human hydrogel was measured by MTS assay. All experimental conditions were performed in triplicate. RESULTS: Histology of H and E and SYTO Green stained tendon showed no cells or nucleic acid, respectively, following SDS and EDTA washes. Using DNeasy assay, only 132 ng DNA per mg of tendon persisted after washes – a value similar to decellularized human tendon.3 Mass spectrometry showed that collagen composes 32.3% and 35.8% of milled porcine tendon before and after pepsin digestion, respectively (Table 1). Rheology demonstrated that porcine hydrogel maintains a fluid consistency over a range of temperatures, unlike human hydrogel, which tends to solidify. Live/Dead staining revealed that human ASCs survive in hydrogel ten days after seeding and retain spindle-like morphology. Cell proliferation was measured in 2% human hydrogel and 1% porcine hydrogel at 3 (O.D. 0.68 and 0.58, respectively; p = 0.051) and 5-day (O.D. 1.17 and 0.92, respectively; p = 0.05) time points (Figure 1).Table 1: Mass spectrometry of porcine tendon hydrogel before and after digestion.Figure 1: Proliferation of human ASCs in human and porcine tendon hydrogel.CONCLUSION: Following reconstitution and digestion, porcine hydrogel was capable of growing human ASCs for at least ten days. The minimal difference in proliferative capacity suggests that porcine tendon hydrogel may be an effective and viable alternative to human hydrogel for the enhancement of tendon healing.

Histological characterization of soft‐embalmed porcine tendon and muscle (914.1)

Embalmed material is used in many fields because of its benefits over fresh materials; however, typical formalin‐fixation results in materials that are vastly different from their original state. Soft‐embalming has received recognition as a viable alternative to fresh or formalin‐fixed materials in education and research, especially in orthopedics. Soft‐embalmed tissue has mainly been characterized by gross features, with minimal investigation into the microscopic features of different types of soft‐embalmed tissues.Deep flexor muscle and tendon were harvested from porcine forelimbs. Samples were soft‐embalmed using one of three different methods: phenol‐based, phenoxyethanol‐based, and Thiel embalming. The samples were visualized using Masson’s trichrome staining immediately following embalming and three weeks later.All types of soft‐embalming displayed typical tendon structural characteristics at both time points. The staining of the tendons ranged from almost entirely blue, as would be expected, to almost entirely red, which was unexpected, depending on the treatment. All soft‐embalmed muscle samples exhibited typical colouring, but there were structural differences. At both time points, muscle of phenol‐based and phenoxyethanol‐based embalming generally exhibited normal muscle structure, while Thiel‐embalmed muscle appeared to have suffered degradation.This histological analysis is a step forward in understanding the effects of different soft‐embalming methods on different tissues, which can aid in determining their efficacy.Grant Funding Source: This research was supported in part by NSERC and CIHR

Biochemical characterisation and assessment of fibril-forming ability of collagens extracted from Bester sturgeon Huso huso × Acipenser ruthenus

Collagens purified from Bester sturgeon organs were characterised biochemically, and their fibril-forming abilities and fibril morphologies formed in vitro clarified. Yields of collagens were 2.1%, 11.9%, 0.4%, 18.1%, 0.4%, 0.8% and 0.03% (collagen dry weight/tissue wet weight) from scales, skin, muscle, swim bladder, digestive tract, notochord and snout cartilage, respectively. Using SDS–PAGE and amino acid composition analyses, collagens from scales, skin, muscle, the swim bladder and digestive tract were characterised as type I, and collagens from the notochord and snout cartilage as type II. Denaturation temperatures of the collagens, measured using circular dichroism, were 29.6, 26.8, 29.0, 32.9, 31.6 and 36.3°C in scales, skin, muscle, swim bladder, digestive tract, and notochord, respectively. For fibril formation, swim bladder and skin collagen showed a more rapid rate of increase in turbidity, a shorter time to attain the maximum turbidity, and formed thicker fibrils compared with porcine tendon type I collagen.

Optimal Parameters for Sutures Tied to a Post during Anterior Cruciate Ligament Reconstruction: Thread Numbers, Knot Numbers, Suture Techniques and Stitch Numbers: An Experimental Laboratory Study Using Porcine Tendon

We evaluated the conditions required for sutures tied to a post for tibial fixation during anterior cruciate ligament (ACL) reconstruction. Harvested porcine tendon was used as a graft material and nonabsorbable suture was used for sutures. Samples were tested for ultimate tensile load and elongation according to thread numbers, knot numbers, suture techniques and stitch numbers. As thread numbers were increased, ultimate tensile load was increased and elongation was decreased. However, more than 4 strands of threads provided the sufficient ultimate tensile load more than 454 N of normal ACL for daily activities. As knot numbers were increased, ultimate tensile load was increased, but elongation was decreased. In terms of failure mode, unraveling occurred 100% in 3 and 4 knots, 81.2% in 5 knots, 54.5% in 6 knots, and 0% in 7 knots. Suture techniques and stitch numbers didn’t significantly affect the ultimate tensile load and the elongation. For sutures tied to a post for tibial fixation in ACL reconstruction, more than 4 threads, more than 7 knots, and more than 4 stitches provide adequate ultimate tensile load and elongation.

Prediction of the elastic strain limit of tendons

The elastic strain limit (ESL) of tendons is the point where maximum elastic modulus is reached, after which micro-damage starts. Study of damage progression in tendons under repetitive (fatigue) loading requires a priori knowledge about ESL. In this study, we propose three different approaches for predicting ESL. First, one single value is assumed to represent the ESL of all tendon specimens. Second, different extrapolation curves are used for extrapolating the initial part of the stress–strain curve. Third, a method based on comparing the shape of the initial part of the stress–strain curve of specimens with a database of stress–strain curves is used. A large number of porcine tendon explants (97) were tested to examine the above-mentioned approaches. The variants of the third approach yielded significantly (p<0.05) smaller error values as compared to the other approaches. The mean absolute percentage error of the best-performing variant of the shape-based comparison was between 8.14±6.44% and 9.96±9.99% depending on the size of the initial part of the stress–strain curves. Interspecies generalizability of the best performing method was also studied by applying it for prediction of the ESL of horse tendons. The ESL of horse tendons was predicted with mean absolute percentage errors ranging between 10.53±7.6% and 19.16±14.31% depending on the size of the initial part of the stress–strain curves and the type of normalization. The results of this study suggest that both ESL and the shape of stress–strain curves may be highly different between different individuals and different anatomical locations.

Measurement of Three-Dimensional Anisotropic Diffusion by Multiphoton Fluorescence Recovery after Photobleaching

The multiphoton fluorescence recovery after photobleaching (MP-FRAP) technique has been developed to measure the three-dimensional (3D) solute diffusion within biological systems. However, current 3D MP-FRAP models are based on isotropic diffusion and spatial domain analysis. The 3D anisotropic diffusion and frequency domain analysis for MP-FRAP measurements are rarely studied. In this study, a new technique is demonstrated for the quantitative and non-destructive determination of 3D anisotropic solute diffusion tensors within biological fibrosis tissues by multiphoton photobleaching and spatial Fourier analysis (SFA). Compared to the spatial domain analysis based MP-FRAP techniques, this SFA-based method has the capability for determining the 3D anisotropic diffusion tensors as well as the flexibility for satisfying initial and boundary conditions. First, a close-form solution of the 3D anisotropic diffusion equation is derived by solely using SFA. Next, this new method is validated by computer-simulated MP-FRAP experiments with pre-defined 3D anisotropic diffusion tensors as well as experimental diffusion measurements of FITC-Dextran (FD) molecules in aqueous glycerol solutions. Finally, this MP-FRAP technique is applied to the measurement of 3D anisotropic diffusion tensors of FD molecules within porcine tendon tissues. This study provides a new tool for complete determination of 3D anisotropic solute diffusion tensor in biological tissues.

Quantitative ultrasound method for assessing stress–strain properties and the cross-sectional area of Achilles tendon

The Achilles tendon is one of the most commonly observed tendons injured with a variety of causes, such as trauma, overuse and degeneration, in the human body. Rupture and tendinosis are relatively common for this strong tendon. Stress–strain properties and shape change are important biomechanical properties of the tendon to assess surgical repair or healing progress. Currently, there are rather limited non-invasive methods available for precisely quantifying the in vivo biomechanical properties of the tendons. The aim of this study was to apply quantitative ultrasound (QUS) methods, including ultrasonic attenuation and speed of sound (SOS), to investigate porcine tendons in different stress–strain conditions. In order to find a reliable method to evaluate the change of tendon shape, ultrasound measurement was also utilized for measuring tendon thickness and compared with the change in tendon cross-sectional area under different stress. A total of 15 porcine tendons of hind trotters were examined. The test results show that the attenuation and broadband ultrasound attenuation decreased and the SOS increased by a smaller magnitude as the uniaxial loading of the stress–strain upon tendons increased. Furthermore, the tendon thickness measured with the ultrasound method was significantly correlated with tendon cross-sectional area (Pearson coefficient = 0.86). These results also indicate that attenuation of QUS and ultrasonic thickness measurement are reliable and potential parameters for assessing biomechanical properties of tendons. Further investigations are needed to warrant the application of the proposed method in a clinical setting.

Mechanical Properties of Revision ACL Reconstruction

The aim of this study is to evaluate the mechanical properties of a revision anterior cruciate ligament (ACL) reconstruction after redrilling the original tibial bone tunnel through a retained composite screw compared with initial soft tissue graft fixation. A total of 24 porcine tendons were fixed to porcine tibial tunnels with a 10 × 35 mm composite interference screw. Following the pullout test, a revision tunnel was drilled through the first interference screw and a second graft was fixed in the bone tunnel using a larger composite screw (11 × 35 mm). Following insertion of the revision screw, the graft was reloaded as described for the primary reconstruction. Load versus displacement data were recorded for each test. There were no significant differences between the primary and revision reconstruction constructs for yield load (p = 0.62), linear stiffness (p = 0.18), maximum failure load (p = 0.57), and yield displacement (p = 0.46). These results indicate that the mechanical properties of tibial fixation for ACL reconstruction with a composite screw following a revision provide similar fixation compared with initial reconstruction in this model. Revising a failed composite ACL construct by means of overdrilling and reinstrumenting may provide fixation equivalent to the initial reconstruction.

Imaging monitored loosening of dense fibrous tissues using high-intensity pulsed ultrasound

Pulsed high-intensity focused ultrasound (HIFU) is proposed as a new alternative treatment for contracture of dense fibrous tissue. It is hypothesized that the pulsed-HIFU can release the contracted tissues by attenuating tensile stiffness along the fiber axis, and that the stiffness reduction can be quantitatively monitored by change of B-mode images. Fresh porcine tendons and ligaments were adapted to an ex vivo model and insonated with pulsed-HIFU for durations ranging from 5 to 30 min. The pulse length was 91 µs with a repetition frequency of 500 Hz, and the peak rarefactional pressure was 6.36 MPa. The corresponding average intensities were kept around 1606 W cm−2 for ISPPA and 72.3 W cm−2 for ISPTA. B-mode images of the tissues were acquired before and after pulsed-HIFU exposure, and the changes in speckle intensity and organization were analyzed. The tensile stiffness of the HIFU-exposed tissues along the longitudinal axis was examined using a stretching machine. Histology examinations were performed by optical and transmission electron microscopy. Pulsed-HIFU exposure significantly decreased the tensile stiffness of the ligaments and tendons. The intensity and organization of tissue speckles in the exposed region were also decreased. The speckle changes correlated well with the degree of stiffness alteration. Histology examinations revealed that pulsed-HIFU exposure probably damages tissues via a cavitation-mediated mechanism. Our results suggest that pulsed-HIFU with a low duty factor is a promising tool for developing new treatment strategies for orthopedic disorders.

Human hamstring tenocytes survive when seeded into a decellularized porcine Achilles tendon extracellular matrix

Tendon ruptures and defects remain major orthopaedic challenges. Tendon healing is a time-consuming process, which results in scar tissue with an altered biomechanical competence. Using a xenogeneic tendon extracellular matrix (ECM) as a natural scaffold, which can be reseeded with autologous human tenocytes, might be a promising approach to reconstruct damaged tendons. For this purpose, the porcine Achilles (AS) tendons serving as a scaffold were histologically characterized in comparison to human cell donor tendons. AS tendons were decellularized and then reseeded with primary human hamstring tenocytes using cell centrifuging, rotating culture and cell injection techniques. Vitality testing, histology and glycosaminoglycan/DNA quantifications were performed to document the success of tendon reseeding. Porcine AS tendons were characterized by a higher cell and sulfated glycosaminoglycan content than human cell donor tendons. Complete decellularization could be achieved, but led to a wash out of sulfated glycosaminoglycans. Nevertheless, porcine tendon could be recellularized with vital human tenocytes. The recellularization led to a slight increase in cell number compared to the native tendon and some glycosaminoglycan recovery. This study indicates that porcine tendon can be de- and recellularized using adult human tenocytes. Future work should optimize cell distribution within the recellularized tendon ECM and consider tendon- and donor species-dependent differences.

En faceparametric imaging of tissue birefringence using polarization-sensitive optical coherence tomography

A technique for generating en face parametric images of tissue birefringence from scans acquired using a fiber-based polarization-sensitive optical coherence tomography (PS-OCT) system utilizing only a single-incident polarization state is presented. The value of birefringence is calculated for each A-scan in the PS-OCT volume using a quadrature demodulation and phase unwrapping algorithm. The algorithm additionally uses weighted spatial averaging and weighted least squares regression to account for the variation in phase accuracies due to varying OCT signal-to-noise-ratio. The utility of this technique is demonstrated using a model of thermally induced damage in porcine tendon and validated against histology. The resulting en face images of tissue birefringence are more useful than conventional PS-OCT B-scans in assessing the severity of tissue damage and in localizing the spatial extent of damage.

Use of a Silicon Tendon Model in an Orthopaedic Skills Course

The use of animal tendons for teaching suture techniques is well established on surgical courses. While providing an authentic simulation of human tendons, these may pose a number of potential problems including cultural issues, health risks and difficulties maintaining the quality of specimens throughout the course. These were evident at a recent Royal College of surgeons orthopaedic skills course in Dubai, with the use of porcine tendons clearly unacceptable to Moslem participants. After trials with alternatives, a silicon-based model was selected and found to be highly satisfactory.

Quantification of collagen fiber organization using three-dimensional Fourier transform-second-harmonic generation imaging

We present three-dimensional Fourier transform-second-harmonic generation (3D FT-SHG) imaging, a generalization of the previously reported two-dimensional FT-SHG, to quantify collagen fiber organization from 3D image stacks of biological tissues. The current implementation calculates 3D preferred orientation of a region of interest, and classifies regions of interest based on orientation anisotropy and average voxel intensity. Presented are some example applications of the technique which reveal the layered structure of collagen fibers in porcine sclera, and estimates the cut angle of porcine tendon tissues. This technique shows promising potential for studying biological tissues that contain fibrillar structures in 3D.

Comparison of modified Kessler and Yotsumoto-Dona suture: A biomechanical study on porcine tendons

There is a need for a strong suture technique that allows early active mobilisation after repair of flexor tendons, but the best method has not yet been found. The aim of this study was to compare the modified Kessler suture biomechanically with a newer, two–strand suture. Eighteen porcine tendons were cut and repaired according to either the grasping modified Kessler suture or the combined side-locking loop technique (Yotsumoto) and interlocking horizontal mattress suture (Dona). The specimens were tested linearly to failure. The 2 mm gap force, yield force, ultimate force, stiffness, energy to yield, and energy to failure were all significantly higher (p value = 0.005, 0.003, <0.001, 0.001, 0.004, and 0.001, respectively) in the Yotsumoto-Dona group (median values (IQR): 30.9 (28.1-39.5) N, 82.7 (64.9-114.1) N, 82.7 (76.6-114.1) N, 12.5 (10-14.5) N/mm, 0.45 (0.2-0.5) J, and 0.45 (0.35-0.5) J) than in the modified Kessler group (25.8 (12.2-28.1) N, 35 (24.6-54.4) N, 50.9 (34.4-55.1) N, 7 (5.8-91) N/mm, 0.09 (0.06-0.18) J, and 0.21 (0.18-0.28) J). All Yotsumoto-Dona specimens had a yield force exceeding 35 N, while in the Kessler group only four did. The early yielding rate was 6/9 and 2/9 in the modified Kessler and the Yotsumoto-Dona groups, respectively (p = 0.15). Most of the core sutures failed by breakage, but three Yotsumoto knots loosened. All the simple running and six of the Dona epitendinous sutures failed predominantly by pulling-out, and by breakage at the intersections in three of the latter. The relatively easy two-strand Yotsumoto-Dona suture is likely to withstand the loads of active finger flexion, whereas the modified Kessler suture is probably not.

A Biomechanical Comparison of 3 Loop Suture Materials in a 6-Strand Flexor Tendon Repair Technique

The braided polyblend (FiberWire) suture is recognized for its superiority in tensile strength in flexor tendon repair. The purpose of this study was to compare the biomechanical performance of 3 loop-suture materials used in a locking 6-strand flexor tendon repair configuration: braided polyblend (FiberLoop 4-0), cable nylon (Supramid Extra II 4-0), and braided polyester (Tendo-Loop 4-0). We hypothesized that, using this technique, the braided polyblend suture would give superior tensile strength compared with the other 2 suture materials. We divided 30 fresh porcine flexor tendons transversely and repaired each with 1 of the 3 suture materials using a modified Lim-Tsai 6-strand suture technique. We loaded the repaired tendons to failure using a materials testing machine and collected data on the mechanism of failure, ultimate tensile strength, gap strength, and stiffness. Failure mechanisms for the repaired specimens were as follows: the braided polyblend had 50% suture breakage and 50% suture pullout; the cable nylon had 100% suture breakage; and the braided polyester had 80% suture breakage and 20% suture pullout. Specimens repaired with the braided polyblend suture had the highest mean ultimate tensile strength (97 N; standard deviation, 22) and the highest mean gap force (35 N; standard deviation, 7). This study supports the findings of previous studies showing superior strength of the braided polyblend suture. We were able to achieve up to 124 N in ultimate tensile strength and 48 N of gap force with this suture in porcine tendons. This gives greater confidence in starting immediate controlled passive or active rehabilitation after repair of flexor tendon injuries.

Comparison of modified Kessler tendon suture at different levels in the human flexor digitorum profundus tendon and porcine flexors and porcine extensors: an experimental biomechanical study

This study compared the biomechanical behaviour of repairs in the human flexor digitorum profundus tendon in zones I, II and III with repairs of different segments of the porcine flexor tendon of the second digit and the extensor digiti quarti proprius tendon, in order to assess the validity of porcine tendons as models for human flexor tendon repairs. These porcine tendons were selected after comparing their size with the human flexor digitorum profundus tendon. The tendon repairs were done in three segments of each porcine tendon and repairs in the human tendons were done in zones I,II and III. Ten tendons in each group yielded a total of 90 specimens. A modified Kessler repair was done with 3-0 coated braided polyester suture and subjected to uniaxial tensile testing. In human flexor tendons, the ultimate force was higher in zones I and II than in zone III. The porcine flexor digitorum profundus tendon from the second digit and the proximal segment of the extensor digiti quarti proprius tendon behaved similarly to the human flexor tendon in zone III and can be considered as surrogates for the human flexor tendon.

White light interferometric detection of unpolarized light for complete Stokesmetric optical coherence tomography

Optical coherence tomography (OCT) relies on interference between a polarized reference and the target reflection. Thus, it has generally been impossible to detect any unpolarized part in the signal. Here, we demonstrate a scheme that overcomes this limitation. Using a combination of heterodyning and filtering, we realize a polarization-sensitive OCT system capable of measuring the full Stokes vector, including the depolarized part. Based on such a system, we perform full Stokesmetric imaging of different layers in a porcine tendon sample. The complete 4 × 4 backscattering Muellermetric images of one layer are acquired and investigated.

Characterization of bionanocomposite scaffolds comprised of amine‐functionalized gold nanoparticles and silicon carbide nanowires crosslinked to an acellular porcine tendon

As one of the most common proteins found in the human body, collagen is regarded as biocompatible and has many properties making it ideal for soft-tissue repair applications. However, collagen matrices fabricated from purified forms of collagen are notoriously weak and easily degraded by the body. The extracellular matrix of many tissues including human dermis, porcine dermis, and porcine small intestine submucosa are often utilized instead, and several of these scaffolds are crosslinked. Crosslinking has been shown to improve the mechanical properties of collagenous tissues and increase their resistance to degradation. In this study we investigated two novel "bionanocomposite" materials in which either gold nanoparticles or silicon carbide nanowires were crosslinked to a porcine tendon. Scanning electron micrographs confirmed that the nanomaterials were successfully crosslinked to the tissues. A collagenase assay, tensile testing, flow cytometry, and bioreactor studies were also performed to further characterize the properties of these novel materials. The results of these studies indicated that crosslinking porcine diaphragm tissues with nanomaterials resulted in scaffolds with improved resistance to enzymatic degradation and appropriate biocompatibility characteristics, thus warranting further study of these materials for soft tissue repair and tissue engineering applications.