Functional, radiographic, and histologic assessment of healing of autogenous osteochondral grafts and full-thickness cartilage defects in the talus of dogs.

A circular (5.5 mm diameter) full-thickness cartilage defect was created on the medial ridge of the talus in 12 skeletally mature dogs. In 6 dogs, the articular surface of the lesion was repaired, using an osteochondral graft obtained from the ipsilateral manus. The graft (digit I, first phalanx, distal articular surface and diaphysis) was contoured to obtain a press fit in the drilled talar recipient site. In 6 dogs, the lesion was not treated and healed by fibrous tissue replacement. Functional assessment (lameness, hock range of motion, joint stability, joint crepitus, and mid-femoral muscle circumference) was completed before surgery and at postoperative weeks 2 through 20. Radiographic assessment (periarticular soft tissue width, joint space width, osteophyte formation, and graft incorporation) was completed before surgery and at postoperative weeks 0, 6, 12, and 20. To facilitate histologic assessment, tissues were stained with toluidine blue and H&E. Histologic assessment of the articular surface on the surgically treated talus, ipsilateral tibia, and contralateral talus was completed, using a modification of the Mankin grading system. Subchondral bone was examined to assess graft viability and incorporation. Analysis of the ordinal data was completed, using a Mann-Whitney rank sum test. All dogs were fully weight bearing by postoperative week 7. Dogs without grafts had significantly (P = 0.036) better clinical function at postoperative week 6. Significant difference in functional assessment was not evident at postoperative week 20. Immediate postoperative radiographic assessment revealed significant (P = 0.005) difference between nongrafted and grafted groups. Significant difference was not observed at postoperative week 6, 12, or 20. All grafts appeared radiographically incorporated by postoperative week 12. All grafts restored joint surface congruity, whereas 3 of 6 nongrafted lesions had poor articular congruity. Of 6 grafts, 4 partially retained normal hyaline cartilage, resulting in significantly (P = 0.014) lower Mankin grades. Significant histologic differences between groups were not apparent when the apposing tibia and control talus were examined. Talar reconstruction by use of a phalangeal osteochondral graft is a viable surgical procedure. These data indicate that normal articular and subchondral architecture are more closely approximated by osteochondral reconstruction than by fibrous tissue repair.     

Resurfacing Canine Femoral Trochleoplasties with Free Autogenous Periosteal Grafts

Femoral trochleoplasties resurfaced with autogenous free periosteal grafts in six dogs were compared to femoral trochleoplasties performed without grafts in six dogs. The grafts produced cartilage as early as 4 weeks after surgery; however, it was fibrocartilage rather than normal hyaline articular cartilage. The trochleoplasties not resurfaced with periosteum also healed with fibrocartilage but not until 40 weeks postoperatively. The grafted stifles had less patellar cartilage damage when compared to trochleoplasties left to heal without resurfacing.     

Histomorphometric analysis of articular cartilage, zone of calcified cartilage, and subchondral bone plate in femoral heads from clinically normal dogs and dogs with moderate or severe osteoarthritis

OBJECTIVE: To quantify and compare the microscopic changes in articular cartilage (AC), zone of calcified cartilage (ZCC), and subchondral bone plate in femoral heads from clinically normal dogs and dogs with moderate or severe osteoarthritis. SAMPLE POPULATION: Femoral heads from clinically normal dogs (n = 16) and dogs with moderate (24) or severe (14) osteoarthritis. PROCEDURES: Femoral heads were allocated to 3 categories (normal, moderate, or severe osteoarthritis) on the basis of radiographic findings, macroscopic findings, and histologic grade determined by use of a modified Mankin scale. Equally spaced 2-mm sections were cut in each femoral head in a coronal or transverse plane. Thickness of the AC, ZCC, and subchondral bone plate was recorded. RESULTS: Mean thickness of AC was significantly greater in samples with moderate and severe osteoarthritis than those considered normal. Mean thickness of the ZCC was significantly greater in samples with moderate and severe osteoarthritis than those considered normal. Mean thickness of the subchondral bone plate in samples with severe osteoarthritis was significantly greater than those with moderate osteoarthritis and those considered normal. A significant decrease in AC thickness was detected in the proximomedial area of femoral heads with severe osteoarthritis, compared with those considered normal. CONCLUSIONS AND CLINICAL RELEVANCE: A cause and effect association between thickening of subchondral structures and thinning and loss of the overlying AC was not detected. Changes in AC were associated with changes in the subchondral bone plate, which is compatible with the theory of adaptation in response to altered load distribution.     

Long-term complete chimerism and stable hematopoiesis in beagles after fetal liver hematopoietic stem cell transplantation

Karyotype analysis of bone marrow cells was performed from 6 Beagles that had received sex-mismatched fetal liver hematopoietic stem cell grafts 2.4 to 6.5 years earlier. Two dogs received dog leukocyte antigen-matched fetal liver cells and 4 received dog leukocyte antigen-mismatched cells. In each dog, all 25 evaluated metaphase spreads were of donor genotype. All dogs had normal hemograms and repopulation of bone marrow and no dog developed graft-vs-host disease.     

Allogeneic tooth transplantation in the dog

Allogeneic tooth transplantation was evaluated as a functional and aesthetic treatment for dental fracture in the dog. Of 7 dogs that received tooth transplants, 5 were research animals and 2 were clinical patients. Canine teeth were transplanted immediately after extraction in the research dogs. Endodontic therapy had been performed on the affected canine tooth of one clinical patient. The other clinical patient had bilateral maxillary canine fractures 2 months earlier. One of the research dog transplants failed at 3 weeks as a result of improper surgical technique. Four of the research dogs had a solid implant for 18 months, after which time the dogs were euthanatized serially. All transplanted teeth were anchored firmly into the alveoli, but were nonviable. Root resorption, with bone replacement, was first noticed at 24 months. The transplanted tooth in the first clinical patient remained functional for 3 months, after which time the tooth was fractured. The right canine transplant in the second clinical patient failed by 3 months, probably because of preexisting periapical inflammation. The left transplanted tooth remains stable at 38 months. It was concluded that allogeneic tooth transplantation may have merit as a rapid and inexpensive method for replacement of fractured teeth in the dog. Function is compromised gradually as a result of root resorption and ankylosis, with tooth fracture likely to occur after 2 years.     

Radiologic and gross anatomic evaluation of bone healing in the dog.

Bone healing associated with 3 techniques of midshaft femoral fracture fixation in 36 young adult Beagle-type dogs was evaluated in radiographic and gross pathoanatomic studies. A serrated transverse fracture was surgically and aseptically created on the midshaft of either the left or the right femur of each dog. The fixation devices used were intramedullary (IM) pin, IM pin and 1/2 Kirschner device, and tension bone plates. The radiographic evaluation was done in series. The first radiographs were taken during surgery. Radiographs were then taken immediately after surgery, to record the status of reduction, alignment, and fixation. Radiographs were taken at the 4th and 10th postoperative weeks, to monitor healing. All dogs were euthanatized at the 10th week. Both the normal femur and the healing femur were removed from all dogs, all soft tissue was removed from the bone, and each femur was examined grossly. Each type of fixation was associated with a distinct mode of osteogenesis. Bony union and clinical union (that stage in the healing process when fixation can be removed) were defined as a successful conclusion to each case. Malunion and nonunion were defined as unsuccessful conclusions. Delayed union was defined as neither successful nor unsuccessful. The IM pin cases had a 64.2% success rate and a 14.2% failure rate. The IM pin and 1/2 Kirschner cases had a 100% success rate. The bone plate cases were 91% successful, with 0% failure. A problem identified with IM pinning was axial rotation (6 of 14 or 42% of the cases). It was concluded that IM pins should not be used alone for the fixation of femoral fractures in adult dogs.     

Clinical and pathological findings in dogs following supralethal total body irradiation with and without infusion of autologous long-term marrow culture cells.

We developed a canine model for autologous bone marrow transplantation (AuBMT) with long-term marrow culture (LTMC) cells. Marrow was harvested from nine normal dogs. Harvests from dogs 2-7 were placed into 21 day LTMC. Cells in LTMC from dogs 4-7 were labelled with the neomycin phosphotransferase gene neo. Dogs were given 60Co total body irradiation (TBI) and then infused with LTMC cells: dog 1 received 500 cGy TBI and 2.08 x 10(8)/kg uncultured marrow cells. Dogs 2-7 received 600-800 cGy TBI and 0.07-0.45 x 10(8)/kg LTMC cells. Dogs 8 and 9 received 600 and 800 cGy TBI, respectively, but no infusion of marrow or LTMC cells. For all dogs, profound myelosuppression developed during week 1 and pyrexia developed during week 2. Enrofloxacin was given from one day before TBI until a peripheral neutrophil count > 1.0 x 10(9)/L was achieved, which eliminated Escherichia coli from feces. Dogs 1, 2 and 5-9 also received gentamicin and/or combination beta-lactam antibiotics. Numerous platelet transfusions were needed to control hemorrhages in all dogs except dog 1. Dog 1 achieved neutrophils > 1.0 x 10(9)/L on day 15, while dogs 2 and 5-9 achieved this count on days 33-48. Dogs 3 and 4 died on days 17 and 18, respectively, of beta-hemolytic streptococcal sepsis and hemorrhage, with no evidence of hematopoiesis at necropsy. The marker gene, neo, was documented in lymphoid and myeloid cells of dogs 5-7 up to 21 months post-AuBMT. Our studies indicate that dogs can recover following supralethal TBI and can survive the delayed engraftment associated with AuBMT using LTMC cells, if they receive intensive platelet and antimicrobial therapy. Used prophylactically for such therapy, enrofloxacin achieved selective intestinal decontamination, but did not prevent sepsis when used as the sole antimicrobial agent during myelosuppression. Furthermore, our studies indicate that infused LTMC cells, at the above doses, can contribute to hematopoietic recovery, but are not essential for recovery following TBI, and do not shorten the period of prolonged profound myelosuppression induced by TBI.     

Evaluation of dirlotapide for sustained weight loss in overweight Labrador retrievers

The effects of dirlotapide on body weight (BW) reduction were investigated in overweight Labradors in two parallel-design studies. Study A involved 42 dogs randomized to 0.0, 0.025, 0.05, 0.1, 0.2 or 0.4 mg dirlotapide/kg/day orally for 4 weeks. Study B involved 72 dogs randomized to nine treatments: placebo (24 weeks); dirlotapide (24 weeks) followed by placebo (28 weeks); or dirlotapide (52 weeks); on diets containing 5%, 10% or 15% fat. Dirlotapide dose (initially 0.1 mg/kg) was adjusted monthly during 24-week weight-loss and subsequent 28-week weight-stabilization phases. Food was offered above maintenance energy requirements (MER× 1.1–1.2) based on initial BW. Body composition (body fat, lean tissue and bone mineral content) was monitored using dual-energy X-ray absorptiometry. After treatment, dogs that had received dirlotapide for 52 weeks were fed 90% of quantity consumed at week 52. In study A, BW and food intake decreased asymptotically with dose: mean weekly weight loss exceeded 1% at 0.1–0.4 mg/kg. In study B, dirlotapide resulted in significant mean weekly weight loss (>0.8%) and decreased food intake over 24 weeks compared with placebo ( P  = 0.0001) for all diets. Food restriction minimized post-treatment weight rebound. Dirlotapide administered daily to dogs for up to 52 weeks was clinically safe and resulted in sustained weight reduction.     

Early pathophysiologic feature of arthropathy in juvenile dogs induced by ofloxacin, a quinolone antimicrobial agent

Arthropathy in dogs induced by ofloxacin, a quinolone antimicrobial agent, was pathophysiologically investigated. In the in vivo studies, ofloxacin was administered orally once or twice at 20 mg/kg/day to male juvenile (3-month-old, n=3) or adult (36-month-old, n=2) dogs, and the humeral and femoral heads were examined pathologically. Unlike adult dogs, fluid-filled vesicles were macroscopically observed on the articular surfaces of one juvenile dog 24 hours after a single treatment with ofloxacin. These lesions were seen in all juvenile dogs by twice dosing. Microscopically, fissures or cavity formations in the middle zone of the articular cartilage were noted only in juvenile dogs. Furthermore, the cartilage matrix from the abnormal area to the articular surface showed a decreased safranin-O staining intensity, suggesting proteoglycan depletion. Ultrastructurally, chondrocytes in the middle zone of juvenile dogs displayed dilatation of the cisternae in the rough endoplasmic reticulum as an initial hallmark. In the in vitro studies, chondrocytes isolated from the articular cartilage of naive juvenile dogs were exposed to ofloxacin at 6.3-100 microg/ml for 24 hours. Although no changes were noted in the deoxyribonucleic acid synthesis, protein synthesis, or proteoglycan release at concentrations of up to 100 microg/ml, the proteoglycan synthesis was evidently decreased in a dose-dependent manner from 12.5 microg/ml. The results obtained suggest that the inhibitory action of ofloxacin on proteoglycan syntheses in the chondrocytes may largely contribute to the early morphologic features in the articular cartilage of the juvenile dog.     

Continent Jejunal Reservoir (Kock Pouch) for Urinary Diversion in Dogs

Urinary diversion by transplantation of both ureters into a reservoir constructed from 60 cm of jejunum was performed in six clinically normal dogs. The reservoir was connected to a stoma on the abdominal wall through a valve constructed from a short segment of intussuscepted bowel to provide continence. The reservoir was emptied three times daily by catheterization. One dog did not survive the surgical procedure and two dogs died within 1 week of surgical complications. The postoperative capacity of the reservoir in surviving dogs was 100 to 200 mL; it increased during the first month to 600 to 750 mL. Continence was excellent in two dogs. One dog was continent until month 2, when partial slippage of the intussusception occurred. A reversible hyponatremic, hypochloremic, hypokalemic metabolic acidosis developed. Bacteriuria was not responsive to systemic or local instillation of antibiotics. Ascending pyelonephritis, hydroureter, and hydronephrosis occurred in two dogs. Struvite urinary calculi formed in one dog. The procedure as described would be unsuitable for clinical use in dogs.     

Long-term outcome of distal femoral osteotomy for treatment of combined distal femoral varus and medial patellar luxation: 12 cases (1999-2004)

OBJECTIVE: To determine long-term outcome of distal femoral osteotomy as a component of treatment for distal femoral varus and medial patellar luxation in large-breed dogs. DESIGN:Retrospective case series. ANIMALS: 12 dogs (16 stifle joints). PROCEDURES: Medical records and radiographs were reviewed to identify large-breed dogs with medial patellar luxation (grade > or = 2) and femoral varus angle > or = 12 degrees treated with distal femoral osteotomy, with a minimum follow-up (by a veterinarian) of 18 months. Signalment, weight, medial patellar luxation and lameness grade, pre- and postoperative femoral varus angle, surgical technique, time to radiographic bone union, and complications were recorded. Follow-up with owners via questionnaire was performed > 18 months after surgery. RESULTS: 16 corrective distal femoral osteotomies were performed with ancillary medial patellar luxation procedures in 12 dogs; 4 dogs had staged bilateral procedures. Mean +/- SD preoperative and postoperative femoral varus angles were 16.3 +/- 4.3 degrees and 3.9 +/- 2.5 degrees , respectively. Mean +/- SD time to radiographic union of the distal femoral osteotomy was 52.6 +/- 13 days. One dog had Kirschner wire migration from the tibial tuberosity. Patellar luxation was not detected after surgery in any dog. Mean +/- SD follow-up by a veterinarian was 1,335 +/- 410 days and by use of an owner questionnaire was 1,497 +/- 464 days. All 10 variables of owner-observed patient comfort and function were significantly improved. CONCLUSIONS AND CLINICAL RELEVANCE: Distal femoral osteotomy in combination with traditional treatment provided predictable osteotomy healing, patellar stabilization, and long-term improvement in patient comfort and function when used to treat combined distal femoral varus and medial patellar luxation in large-breed dogs.     

Treatment of Canine Nasal Aspergillosis with Enilconazole

Twenty-four dogs with nasal aspergillosis were treated with enilconazole (10 mg/kg bid for 7–14 days) administered topically through tubes surgically implanted into the nasal chambers. Aspergillosis was eliminated in 19 dogs over a median follow-up period of 18 months. Another dog died, but at necropsy there was no evidence of causative fungus. Two of the four dogs that were not cured had infection of periorbital soft tissues. An additional seven dogs received 6 weeks ketoconazole (5 mg/kg bid PO) and enilconazole therapy topically. Six of these dogs were disease-free over a median follow-up period of 35 months. The seventh dog responded to repeated treatment with enilconazole. Twenty-six of the 29 dogs (90%)without extranasal aspergillosis were cured.     

Effect of chronic lead exposure on the canine bone marrow

The effect of chronic oral led acetate administration on canine bone marrow was studied. Two dogs (group 1) were used as controls, 4 dogs (group 2) were given 2 mg of lead/kg of body weight daily, and 4 dogs (group 3) were given 5 mg of lead/kg daily. After a 7-day stabilizaion period, lead dosing was conducted for 91 days (13 weeks), after which half of each group was treated with calcium ethylenediaminetetraacetic acid. All dogs were then observed for another 28 days (4 weeks). Blood lead values and bone marrow cellular changes were monitored once a week during the 126 days (18 weeks) of study. Lead-dosed dogs had lower weight gains than the controls. Clinical signs of toxicosis were observed after 6 weeks in one dog in group 3. Anorexia, body weight loss, CNS depression, muscular weakness, and trembling were seen. Blood lead concentrations increased in all group 2 and 3 dogs. Lead caused increases in bone marrow segmented neutrophils and myeloid series cells, and increased myeloid:erythroid ratios. Blood lead concentrations and myeloid:erythroid ratios decreased after cessation of lead administration.     

Early Experience With the Use of an Interlocking Nail for the Repair of Canine Femoral Shaft Fractures

Static locked nailing was used to repair fractures of the femoral diaphysis in 15 dogs. The implant consists of a 5, 6, or 7 mm diameter stainless steel rod made up of two parts: the body of the nail, in which there are 13 threaded holes, and a piece without holes that contains a slot for anchoring the jig. Good limb function was obtained after less than 3 weeks in 12 dogs and these dogs remained sound throughout the study. Radiographic examination revealed fracture healing in 11 of the dogs, between 8 and 16 weeks after surgery. One dog was not returned for follow-up evaluation until 22 weeks after surgery; complete healing was apparent from radiographic examination. The remaining three dogs had to have additional operations, one because of lameness caused by excessive length of the distal screws, one because of a nonunion, and the third because of a sequestrum. Fracture healing in these dogs was observed at 18, 21, and 24 weeks respectively. Loosening of one screw and angulation of the bone occurred in one dog. These complications had no adverse effects on clinical outcome.     

Arthroscopic mosaic arthroplasty in the equine third carpal bone

OBJECTIVE--To investigate survival and function of autogenous heterotopic osteochondral grafts in a site where injuries are common. STUDY DESIGN--Three osteochondral grafts were harvested arthroscopically from the femoropatellar joint and transplanted to the third carpal bone (C(3)). Nine months later, histologic, histomorphometric, and biochemical comparisons were made between the transplanted grafts in C(3) and tissue adjacent to the recipient site, the opposing radial carpal bone (C(r)), the donor site in the femoropatellar joint, and the sham-operated contralateral C(3). ANIMALS--One mixed-breed pony and 5 Standardbred horses aged 3 to 8 years old. METHODS--Using instruments modified for equine use, four 4.5-mm-diameter osteochondral grafts were harvested arthroscopically from the distal aspect of the lateral trochlea of the right femur and inserted into the radial facet of the right third carpal bone. The fourth graft was kept as a donor-site control sample. Three months later, regular exercise was started and at 6 months, repeat arthroscopy was conducted to evaluate healing. The horses were euthanatized 9 months after transplantation, and comparisons were made between the grafts, opposing radial carpal bone, and contralateral third carpal bone. The assessment criteria included paravital staining, a modified Mankin scoring system, and biochemical analyses for collagen type, total collagen content, and sulfated glycosaminoglycan concentration. RESULTS--All horses were sound 21 days' postoperatively. At 6 months, all 18 grafts were intact but somewhat soft and opaque compared with surrounding carpal cartilage. Nine months' postoperatively, the bony portions of the grafts were well integrated with the recipient sites, but 6 grafts had histologic evidence of cartilage degeneration. From biochemical analysis of grafts, there was little or no new repair tissue invading the experimental sites, but sulfated glycosaminoglycan (proteoglycan) loss from the transplanted cartilage was marked. CONCLUSIONS--Heterotopic transfer of osteochondral grafts from the distal aspect of the lateral femoral trochlea to the third carpal bone is feasible with minor modifications of human mosaic arthroplasty instruments. The bony portion of the osteochondral grafts was quickly remodeled to provide subchondral support to the transplanted articular cartilage. The loss of proteoglycan from the transplanted cartilage indicates that the grafts might have been injured during harvesting or insertion, or, more likely, did not remodel to meet the demands of a new biomechanical environment. CLINICAL RELEVANCE--These findings suggest that arthroscopic resurfacing of focal osteoarticular defects will not be successful in the long term unless donor and recipient sites can be matched with respect to cartilage thickness, biochemical constituents, and physical properties. Mosaic arthroplasty may be indicated in selected cases in which no other options exist to create a confluent cartilage-covered surface.     

Complications of articular lag screw fixation of femoral capital epiphyseal separations

Four femoral capital epiphyseal separations in three immature dogs were repaired using an articular lag screw procedure. Two millimetre cortical screws were inserted from the articular surface across the fracture site in lag fashion. The screw heads were countersunk below the surface of the articular cartilage to prevent abrasion of the apposing articular surface. Marked osteolysis of the femoral neck was noted between four and eight weeks postoperatively in all cases, and excision of the femoral head and neck was carried out in two cases at six and eight weeks postoperatively. It is believed that the surgery failed because of iatrogenic vascular insult and the development of coxofemoral instability.     

Histomorphometric analysis of the proximal portion of the femur in healthy dogs

OBJECTIVE: To describe the cancellous bone architecture of the head and neck of the femur in healthy dogs by use of automated histomorphometry techniques in conjunction with histologic grading of articular cartilage. ANIMALS: 30 mature male dogs with healthy coxo-femoral joints PROCEDURE: Dogs were 1.5 to 4 years old and weighed 27 to 37 kg. Computer images of fine-detail radiographs of 100-microm-thick coronal and transverse plane sections of the head and neck of the femur (14 dogs) were analyzed by use of histomorphometry software. Statistical comparisons among histomorphometric indices of 4 regions were performed. Histologic preparations of coronal and transverse plane sections of femoral head articular cartilage (16 dogs) were graded. Median grades for lateral, medial, cranial, and caudal halves of the femoral head articular cartilage were determined. RESULTS: Bone volume/total volume, trabecular thickness and number, and bone surface/total volume were significantly higher in the femoral head than in the femoral neck. Anisotropy (trabecular alignment) and trabecular separation were significantly higher in the femoral neck than in the femoral head. Anisotropy was significantly higher in the caudal half of the femoral neck than in the cranial half. Cartilage had histologic grades indicating health without significant differences among lateral, medial, cranial, and caudal halves of femoral head cartilage. CONCLUSIONS AND CLINICAL RELEVANCE: A predictable cancellous architecture in the head and neck of the femur is associated with healthy cartilage.     

Measurement of the elastic properties of the cancellous bone in the femoral head of the dog

The degenerative wear and pathologic damage of the joints are reasons for total endoprotheses in man as well as in dogs. The main problem is the aseptic loosening of the protheses. By usig the finite-element-method, the total endoprothesis is designed with new features, with the purpose of preventing loosening and being better adapted to load transmission. In order to simulate the femur of the dog for the numerical analysis, a material law is developed. By taking into account the anisotropy and the local density of the cancellous bone in the femoral head, the young's modules are experimentally determined. The measurements are performed by ultrasonic methods on femoral heads of euthanised dogs. The results show planar isotropic cancellous bone.     

Evaluation of the proximal portion of the femur as an autogenous cancellous bone donor site in dogs.

The proximal portion of the femur was evaluated as a source of autogenous cancellous bone in dogs. Bilateral oval cortical defects were created in the lateral subtrochanteric area of the femur in 16 dogs. Cancellous bone was removed and the weight recorded. Cancellous bone was similarly harvested from the proximal portion of the humerus in 7 of these dogs. Subtrochanteric femoral defects in 11 dogs were randomly assigned to receive cancellous bone graft obtained from the femur (n = 4) or the humerus (n = 7). Subtrochanteric defects in 5 dogs were not grafted. Radiographic assessment of subtrochanteric defects was performed at 4-week intervals, and histologic assessment at 4, 8, 16, and 24 weeks after surgery. Nongrafted donor sites healed by ingrowth of trabecular bone during the first 12 weeks after surgery. By week 24, the lateral cortical wall had reformed, but remodeling was incomplete. Donor sites grafted with cancellous bone healed similarly, but with more rapid healing and more complete remodeling evident by week 24. Although the mean weight of cancellous bone harvested from the proximal portion of the femur (0.82 +/- 0.22 g) was significantly (P less than 0.05) less than that harvested from the proximal portion of the humerus (1.38 +/- 0.29 g), there was no qualitative histologic or radiographic difference in bony healing of grafted defects. We determined that the proximal portion of the femur can be safely used to provide moderate amounts of cancellous bone, and that a second bone graft can be collected from the same subtrochanteric donor site after 12 weeks.     

Isolation and characterization of pediatric canine bone marrow CD34+ cells

Historically, the dog has been a valuable model for bone marrow transplantation studies, with many of the advances achieved in the dog being directly transferable to human clinical bone marrow transplantation protocols. In addition, dogs are also a source of many well-characterized homologues of human genetic diseases, making them an ideal large animal model in which to evaluate gene therapy protocols. It is generally accepted that progenitor cells for many human hematopoietic cell lineages reside in the CD34+ fraction of cells from bone marrow, cord blood, or peripheral blood. In addition, CD34+ cells are the current targets for human gene therapy of diseases involving the hematopoietic system. In this study, we have isolated and characterized highly enriched populations of canine CD34+ cells isolated from dogs 1 week to 3 months of age. Bone marrow isolated from 2- to 3-week-old dogs contained up to 18% CD34+ cells and this high percentage dropped sharply with age. In in vitro 6-day liquid suspension cultures, CD34+ cells harvested from 3-week-old dogs expanded almost two times more than those from 3-month-old dogs and the cells from younger dogs were also more responsive to human Flt-3 ligand (Flt3L). In culture, the percent and number of CD34+ cells from both ages of dogs dropped sharply between 2 and 4 days, although the number of CD34+ cells at day 6 of culture was higher for cells harvested from the younger dogs. CD34+ cells harvested from both ages of dogs had similar enrichment and depletion values in CFU-GM methylcellulose assays. Canine CD34+/Rho123lo cells expressed c-kit mRNA while the CD34+/Rhohi cells did not. When transplanted to a sub-lethally irradiated recipient, CD34+ cells from 1- to 3-week-old dogs gave rise to both myeloid and lymphoid lineages in the periphery. This study demonstrates that canine CD34+ bone marrow cells have similar in vitro and in vivo characteristics as human CD34+ cells. In addition, ontogeny-related functional differences reported for human CD34+ cells appear to exist in the dog as well, suggesting pediatric CD34+ cells may be better targets for gene transfer than adult bone marrow. The demonstration of similarities between canine and human CD34+ cells enhances the dog as a large, preclinical model to evaluate strategies for improving bone marrow transplantation protocols, for gene therapy protocols that target CD34+ cells, and to study the engraftment potential of various cell populations that may contain hematopoietic progenitor cell activity.