Dominique J. Griffon

Effect of chitosan scaffold microstructure on mesenchymal stem cell chondrogenesis.

Although numerous biomaterials have been investigated as scaffolds for cartilage tissue engineering, the effect of their microstructure on final construct characteristics remains unclear. The biocompatibility of chitosan and its similarity with glycosaminoglycans make it attractive as a scaffold for cartilage engineering. Our objective was to evaluate the effect of chitosan scaffold structure on mesenchymal stem cell proliferation and chondrogenesis. Chitosan fibrous scaffolds and chitosan sponges were seeded with mesenchymal stem cells in a chondrogenic medium. Constructs were analyzed 72 h after seeding via scanning electron microscopy (SEM), weight measurements and DNA quantification. Constructs were cultured for 10 or 21 days prior to confocal microscopy, SEM, histology, quantitative analysis (weight, DNA and glycosaminoglycan (GAG)), and quantitative real-time polymerase chain reaction. Mesenchymal stem cells maintained a viability above 90% on all chitosan scaffolds. The cell numbers in the constructs were similar at 72 h, 10 days and 21 days. However, matrix production was improved in chitosan fibrous constructs based on the GAG quantification and collagen II mRNA expression. Chondrogenesis on chitosan scaffolds is superior on microfibers compared to macroporous sponges.

Comparison of short- and long-term function and radiographic osteoarthrosis in dogs after postoperative physical rehabilitation and tibial plateau leveling osteotomy or lateral fabellar suture stabilization

OBJECTIVES To compare short- and long-term functional and radiographic outcome of cranial cruciate ligament (CrCL) injury in dogs treated with postoperative physical rehabilitation and either tibial plateau leveling osteotomy (TPLO) or lateral fabellar suture stabilization (LFS). STUDY DESIGN Prospective observational clinical study. ANIMALS Medium to large breed dogs with naturally occurring CrCL injury (n=65). METHODS Dogs with CrCL injury were treated with either TPLO or LFS and with identical physical rehabilitation regimes postoperatively. Limb peak vertical force (PVF) was measured preoperatively and at 3, 5, and 7 weeks, and 6 months and 24 months postoperatively. Stifles were radiographically assessed for osteoarthrosis (OA) preoperatively and 24 months postoperatively. RESULTS Thirty-five dogs had LFS and 30 dogs had TPLO. Radiographic OA scores were significantly increased at 24 months compared with preoperative scores in all dogs. Radiographic OA scores preoperatively and at 24 months were not significantly different between treatment groups. PVF was significantly increased from preoperative to 24 months among both treatment groups but not significantly different between treatment groups preoperatively or at 3, 5, 7 weeks, 6, or 24 months. CONCLUSION No significant difference in outcome as determined by ground reaction forces or radiographic OA scores were found between dogs with CrCL injury treated with LFS or TPLO. CLINICAL RELEVANCE LFS and TPLO remain good options for stabilizing stifles with CrCL injury with all dogs showing significant functional improvement. This study does not support the superiority of either surgical technique.

A Review of the Pathogenesis of Canine Cranial Cruciate Ligament Disease as a Basis for Future Preventive Strategies

Cranial cruciate ligament (CCL) deficiency is the leading cause of lameness of the canine stifle and has important consequences in terms of morbidity and cost associated with its management. In spite of this impact, development of preventive strategies remains in its infancy, largely because of gaps in our understanding of the complex and likely multifactorial origin of CCL deficiency. The purpose of this article is to provide a critical review of the literature related to the pathogenesis of CCL deficiency and place this evidence in the context of potential preventive measures. Trauma accounts for a minority of CCL ruptures in dogs, whereas progressive degeneration of the ligament has been attributed to a variety of factors that may be broadly classified as genetic, conformational, environmental, immune-mediated, and inflammatory. Genetic screening appears promising as a long-term option in selected breeds while immunomodulating therapies may be implemented in the nearer future to reduce the incidence of contralateral disease in dogs with unilateral CCL deficiency. Preventive modification of conformation factors may be applicable to most breeds predisposed to CCL deficiency but requires further investigations into the relative contribution of individual factors, as well as into the feasibility, and potential risks of large-scale implementation.Cranial cruciate ligament (CCL) deficiency is the leading cause of lameness of the canine stifle and has important consequences in terms of morbidity and cost associated with its management. In spite of this impact, development of preventive strategies remains in its infancy, largely because of gaps in our understanding of the complex and likely multifactorial origin of CCL deficiency. The purpose of this article is to provide a critical review of the literature related to the pathogenesis of CCL deficiency and place this evidence in the context of potential preventive measures. Trauma accounts for a minority of CCL ruptures in dogs, whereas progressive degeneration of the ligament has been attributed to a variety of factors that may be broadly classified as genetic, conformational, environmental, immune-mediated, and inflammatory. Genetic screening appears promising as a long-term option in selected breeds while immunomodulating therapies may be implemented in the nearer future to reduce the incidence of contralateral disease in dogs with unilateral CCL deficiency. Preventive modification of conformation factors may be applicable to most breeds predisposed to CCL deficiency but requires further investigations into the relative contribution of individual factors, as well as into the feasibility, and potential risks of large-scale implementation.

Morphometric characteristics of the pelvic limbs of Labrador Retrievers with and without cranial cruciate ligament deficiency

OBJECTIVE-To evaluate skeletal characteristics of pelvic limbs with and without cranial cruciate ligament (CCL) deficiency in Labrador Retrievers. ANIMALS-30 adult purebred Labrador Retrievers. PROCEDURES-Pelvic limbs (n = 28) of 14 dogs without CCL deficiency were classified as control limbs, whereas the limbs of 16 dogs with CCL deficiency were considered affected by (18 limbs) or predisposed to (10 contralateral limbs of dogs with 1 affected limb) CCL deficiency. Skeletal characteristics were evaluated via physical examination, radiography, and computed tomography. Radiographic and computed tomographic variables were compared among limb groups by use of a mixed-model ANOVA. RESULTS-The tibial plateau slope was steeper in CCL-deficient limbs but not in predisposed limbs, compared with the slope in control limbs. The angle between diaphyseal and proximal tibial axes was increased in both CCL-deficient and predisposed limbs. The relative width of the proximal portion of the tibia and the inclination of the patellar ligament did not differ among limb groups. The overall and distal femoral anteversion angles were greater in CCL-deficient and predisposed limbs, whereas the femoral condyle trochanteric angle was decreased in those limb groups, compared with findings in control limbs. CONCLUSIONS AND CLINICAL RELEVANCE-Cranial angulation of the proximal portion of the tibia, excessive steepness of the tibial plateau, and distal femoral torsion appeared more likely to be associated with CCL deficiency than femoral angulation, tibial torsion, intercondylar notch stenosis, and increased inclination of the patellar ligament.

The effect of type II collagen coating of chitosan fibrous scaffolds on mesenchymal stem cell adhesion and chondrogenesis

The biocompatibility of chitosan and its similarity to glycosaminoglycans (GAG) make it attractive for cartilage tissue engineering. We have previously reported improved chondrogenesis but limited cell adhesion on chitosan scaffolds. Our objectives were to produce chitosan scaffolds coated with different densities of type II collagen and to evaluate the effect of this coating on mesenchymal stem cell (MSC) adhesion and chondrogenesis. Chitosan fibrous scaffolds were obtained by a wet spinning method and coated with type II collagen at two different densities. A polyglycolic acid mesh served as a reference group. The scaffolds were characterized by Fourier-transform infrared spectroscopy, scanning electron microscopy (SEM), transmission electron microscopy (TEM) and type II collagen content. Constructs were analyzed after MSCs seeding via live/dead assay, weight and DNA evaluations, SEM, and TEM. Constructs were cultured in chondrogenic medium for 21 days prior to quantitative analysis (weight, DNA, and GAG), SEM, TEM, histology, immunohistochemistry, and quantitative real time polymerase chain reaction. The cell attachment and distribution after seeding correlated with the density of type II collagen. The cell number, the matrix production, and the expression of genes specific for chondrogenesis were improved after culture in collagen coated chitosan constructs. These findings encourage the use of type II collagen for coating chitosan scaffolds to improve MSCs adhesion and chondrogenesis, and confirm the importance of biomimetic scaffolds for tissue engineering.

Proximodistal Alignment of the Canine Patella: Radiographic Evaluation and Association with Medial and Lateral Patellar Luxation

OBJECTIVES To evaluate the contribution of proximodistal alignment of the patella to patellar luxation, and to evaluate the structures contributing to proximodistal alignment of the patella relative to the femoral trochlea. STUDY DESIGN Retrospective study using a convenience sample. ANIMALS Medium to giant breed dogs (n=106). METHODS Medical records and stifle radiographs of 106 dogs were reviewed. Radiographic measurements evaluated the proximodistal alignment of the patella with respect to the femoral trochlea, distal aspect of the femur, and proximal aspect of the tibia. Measurements were compared between dogs with clinically normal stifles (controls; n=51 dogs, 66 stifles), and dogs with a clinical diagnosis of medial patellar luxation (MPL, n=46 dogs, 65 stifles) or lateral patellar luxation (LPL, n=9 dogs, 11 stifles) using ANOVA. RESULTS In dogs with MPL, the ratio of patellar ligament length (PLL) to patellar length (PL) was increased, as was the ratio of the distance from the proximal aspect of the patella to the femoral condyle (A) to PL (P<.0001). Dogs with LPL had a decreased A:PL (P=.003) and an increased ratio of the proximal tibial length (PTL) to distal tibial width (DTW; P=.009). CONCLUSIONS MPL is associated with a relatively long patellar ligament and patella alta in medium to giant breed dogs. LPL is associated with a relatively long proximal tibia and patella baja. Values for PLL:PL>2.06 and A:PL>2.03 are suggestive of the presence of patella alta, whereas a value for A:PL<1.92 is suggestive of patella baja. CLINICAL RELEVANCE Measurements of both PLL:PL and A:PL are recommended in dogs with patellar luxation, and surgical correction should be considered in those with abnormal values.

Comparison of lateral fabellar suture and tibial plateau leveling osteotomy techniques for treatment of dogs with cranial cruciate ligament disease

OBJECTIVE To compare 1-year outcomes after lateral fabellar suture stabilization (LFS) and tibial plateau leveling osteotomy (TPLO) for the treatment of dogs with cranial cruciate ligament disease. DESIGN Randomized blinded controlled clinical trial. ANIMALS 80 dogs with naturally occurring unilateral cranial cruciate ligament disease. PROCEDURES All dogs were randomly assigned to undergo LFS (n = 40) or TPLO (40). Clinical data collected included age, weight, body condition score, history information, stifle joint instability, radiographic findings, surgical findings, and complications. Outcome measures were determined prior to surgery and at 6 and 12 weeks and 6 and 12 months after surgery, including values of pressure platform gait analysis variables, Canine Brief Pain Inventory scores, owner satisfaction ratings, thigh circumference, and stifle joint goniometry values. RESULTS Signalment and data for possible confounding variables were similar between groups. Peak vertical force of affected hind limbs at a walk and trot was 5% to 11% higher for dogs in the TPLO group versus those in the LFS group during the 12 months after surgery. Canine Brief Pain Inventory, goniometry, and thigh circumference results indicated dogs in both groups improved after surgery, but significant differences between groups were not detected. Owner satisfaction ratings at 12 months after surgery were significantly different between groups; 93% and 75% of owners of dogs in the TPLO and LFS groups indicated a satisfaction score ≥ 9 (scale, 1 to 10), respectively. CONCLUSIONS AND CLINICAL RELEVANCE Kinematic and owner satisfaction results indicated dogs that underwent TPLO had better outcomes than those that underwent LFS.

Inverse Dynamics Analysis of the Pelvic Limbs in Labrador Retrievers With and Without Cranial Cruciate Ligament Disease

OBJECTIVES To quantify net joint moments, joint powers, and joint reaction forces (JRF) across the hock, stifle, and hip joints in Labrador Retrievers with and without cranial cruciate ligament (CCL) disease. To investigate differences in joint mechanics between normal, CCL deficient, and contralateral pelvic limbs. STUDY DESIGN Cross-sectional clinical study. ANIMALS Clinically normal (n=14) and CCL-deficient (n=9) Labrador Retrievers. METHODS Each limb was analyzed separately and classified as normal, CCL-deficient or sound contralateral limb. Kinematic, ground reaction force (GRF), and morphometric data were combined in an inverse dynamic approach to compute hock, stifle and hip net moments, powers and JRF, while trotting. RESULTS Vertical and braking GRF and JRF were significantly decreased in CCL-deficient limbs. In affected limbs, extensor moments at the hock and hip, flexor moment at the stifle and power in all 3 joints were less than normal. Kinetics also identified a greater joint moment and power of the contralateral limbs compared with normal, particularly of the stifle extensor muscles group. CONCLUSION Lameness resulting from CCL disease affected predominantly reaction forces during the braking phase and the extension during push-off. A greater contribution of the contralateral limbs to propel the dog forward was identified. CLINICAL RELEVANCE Reductions in joint motion, loads, and muscle contraction were interpreted as modifications adopted to reduce or avoid painful mobilization of the injured stifle joint. The increased joint moment and power of the contralateral compensating side may correlate with the lameness observed, but also with the predisposition of contralateral limbs to CCL deficiency in dogs.

Double‐Blind Placebo‐Controlled Trial of Adjuvant Pamidronate with Palliative Radiotherapy and Intravenous Doxorubicin for Canine Appendicular Osteosarcoma Bone Pain

BACKGROUND Canine osteosarcoma (OSA) causes focal malignant osteolysis leading to severe pain. Despite the documented efficacy of radiotherapy or IV aminobisphosphonates for managing cancer bone pain, their potential combined therapeutic value has not been reported in OSA-bearing dogs. HYPOTHESIS Pamidronate combined with standardized palliative therapy will improve pain control and bone biologic effects in OSA-bearing dogs. ANIMALS Fifty dogs with appendicular OSA treated with standardized palliative therapy and either pamidronate or sterile saline. METHODS Randomized, prospective, double-blinded, placebo-controlled study. Treatment responses for dogs receiving standardized palliative therapy with (n = 26) or without (n = 24) adjuvant pamidronate were serially evaluated for changes in subjective pain scores, urine N-telopeptide (NTx) excretion, primary tumor relative bone mineral density (rBMD), and computerized pressure platform gait analysis. RESULTS Median duration of subjective pain relief for dogs treated with adjuvant pamidronate or placebo was 76 and 75 days, respectively (P= .39). Forty percent (20/50; pamidronate [11/26] and placebo [9/24]) of dogs experienced durable analgesia, defined by pain alleviation > or =112 days. For patients achieving durable pain control, dogs treated with pamidronate achieved greater reductions in NTx excretion and larger increases in rBMD compared with placebo controls. Changes in peak vertical force assessed by computerized pressure platform gait analysis correlated with pain alleviation in OSA-bearing dogs. CONCLUSIONS AND CLINICAL IMPORTANCE Combining pamidronate with standardized palliative therapy is safe, but does not clearly improve pain alleviation. However, in dogs achieving durable pain control, adjuvant pamidronate appears to decrease focal bone resorption in the local tumor microenvironment.

Hyaluronic acid and chondrogenesis of murine bone marrow mesenchymal stem cells in chitosan sponges

OBJECTIVE To establish the dose-dependent effects of high-molecular-weight hyaluronic acid (HA) supplementation on chondrogenesis by mesenchymal stem cells (MSCs) cultured on chitosan sponges and to determine the extent to which MSC matrix production (chondrogenesis) can be influenced by incorporation of high-molecular-weight HA into chitosan scaffolds. SAMPLE POPULATION Murine MSCs derived from a multipotent bone marrow stromal precursor. PROCEDURES MSCs were seeded on chitosan and chitosan-HA scaffolds in chondrogenic medium with various HA concentrations. Scanning electron microscopy, fluorescence microscopy (viability assay), and DNA quantification were used to assess cell attachment, distribution, and viability 48 hours after seeding. Constructs were cultured for 3 weeks prior to evaluation of cell distribution and chondrogenic differentiation via histologic evaluation and quantification of DNA, glycosaminoglycan, and collagen II. RESULTS 48 hours after MSC seeding, cell viability and DNA content were similar among groups. Three weeks after seeding, HA supplementation of the culture medium improved matrix production in a dose-dependent manner, as indicated by matrix glycosaminoglycan and collagen II concentrations. The scaffold composition, however, had no significant effect on matrix production. CONCLUSIONS AND CLINICAL RELEVANCE High-molecular-weight HA supplementation in culture medium had a dose-dependent effect on matrix production and thus chondrogenic differentiation of MSCs cultured on chitosan sponges. The addition of HA in the surrounding fluid during chondrogenesis should improve cartilage production and may be useful for producing engineered cartilage tissues.