Autogenous Cancellous Bone Grafts From the Sternum in Horses Comparison With Other Donor Sites and Results of Use in Orthopedic Surgery

Cancellous bone was collected from the fourth or fifth sternebra of six horses aged 12 to 36 months, and compared quantitatively and qualitatively with cancellous bone collected from the tuber coxa, proximal tibia, and rib at the time of necropsy, 28 to 49 days after surgery. Cancellous bone collected from the sternum was equivalent in amount and in microscopic appearance to that collected from the other three sites.
Cancellous bone also was collected from the sternum for use during clinical orthopedic surgery in 18 horses. Wound dehiscence occurring in two horses healed uneventfully by second intention. In each horse, an adequate amount of cancellous bone was obtained from one sternebra, with the exception of one horse where cancellous bone from two sternebra was used. Major complications with the donor sites were not encountered.     

Histologic features of the healing of bone graft donor sites in dogs

Healing of cancellous bone graft donor sites in the proximal tibial metaphysis of 12 healthy adult dogs was studied histologically. Cancellous bone was curetted from the metaphysis of the proximal end of the tibia, via a 1-cm diameter circular opening in the medial cortex. A hematoma and fibrovascular tissue filled the bone defect at 2 weeks. At 4 and 8 weeks, endosteal callus, composed initially of cartilage and woven bone and later of lamellar bone, filled the marrow cavity. At 12 weeks, the normal structural arrangement of lamellar bone and hematopoietic marrow was reestablished in the marrow cavity. The medial cortex defect was filled only with lamellar trabecular bone. It was concluded that, in adult dogs, a second cancellous bone graft could be collected from the proximal portion of the tibial metaphysis 12 weeks or more after an initial collection.     

Optimisation of bone marrow aspiration from the equine sternum for the safe recovery of mesenchymal stem cells

Reasons for performing study: Mesenchymal stem cell (MSC) therapy for orthopaedic disease is being used with increasing frequency; there is a need to define a safe, reliable and effective technique for the recovery of MSCs from the sternum of the horse. Objectives: To describe an optimised safe technique for obtaining bone marrow‐derived MSCs from the sternum of the Thoroughbred horse. Methods: The anatomical relationship of the sternum with the heart and internal anatomy was demonstrated in cadavers. Sternal anatomy was evaluated ultrasonographically and after midline sectioning. Sternebrae were examined histologically after aspiration to determine the effect of needle insertion. The quality of the aspirate was evaluated as the number of colony‐forming units from sequential and separately aspirated 5 ml aliquots and assessed for their multipotency using trilineage differentiation. Results: The optimal safe location for the needle was the 5th sternebra because it had a safe dorsoventral thickness and was cranial to the apex of the heart. This sternebra could be reliably identified ultrasonographically. Aspirates could also be obtained from the 4th and 6th sternebrae, although the former is between the front limbs and the latter closer to the heart. Minimal disruption of the internal bony architecture was seen after needle insertion through the thin outer cortex and the first 5 ml aliquot contained the greatest number of colony‐forming units of mesenchymal stem cells with trilineage capabilities. Conclusions: Accurate placement of a Jamshidi needle into the medullary cavity of the 4th–6th individual sternebrae is facilitated by the use of ultrasonography and enables aspiration of bone marrow reliably with minimal damage to the sternum and risk to the horse. Potential clinical relevance: Sternal marrow aspiration as described is a safe and reliable technique to obtain MSCs for orthopaedic cell‐based therapies.     

Cancellous Bone Graft Collection from the Tibia in Dogs

Autologous cancellous bone was collected from the metaphysis of the proximal end of the tibia in 30 medium and large dogs. Access to the metaphyseal cancellous bone was gained by making a medial opening in the cortex with a 5.6, 7.9, or 10 mm bone trephine. Sufficient bone was obtained from this site for grafting in each case. Four dogs had bilateral carpal arthrodesis that necessitated graft collection from both tibias. Complications arising from graft collection included wound disruption 5 days postoperatively (3/34 graft sites) and incomplete fracture of the tibial cortex created during the surgical procedure (1/34 graft sites). Collection of autologous cancellous bone from this sita in mature, medium and large dogs was found to be a safe procedure with a low incidence of donor site morbidity.     

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.     

Effects of gelatin sponge implantation in cancellous bone defects in dogs

Reports from oral surgeons suggested that packing bone defects with cellulose or collagen stimulated cancellous bone replacement. The study reported here was designed to evaluate a similar method for augmenting the remodeling of cancellous bone defects and graft donor sites. Gelatin sponge was implanted into bony defects created in the proximal humeral metaphysis of 5 adult Beagles after collection of cancellous bone. A similar defect created in the contralateral humerus was left unpacked. Four months after creation of the defect, the dogs were euthanatized, and the surgical sites were evaluated by use of correlated microradiographic and histologic examination. Unpacked sites had filled completely with loosely woven trabecular bone. Remodeling had not taken place in sites packed with gelatin sponge. Gelatin sponge did not elicit any reaction in the surrounding bone, but remained in situ and obstructed new bone formation. Gelatin sponge blocked rather than augmented cancellous bone replacement.     

Effects of autogenous cancellous bone on healing of homogenous cortical bone grafts

A comparative clinical and histological study was made of the effects of autogenous cancellous grafts on the rate of healing of homogenous cortical bone grafts in dogs: their right foreleg received homogenous bone grafts and their left foreleg received pre-drilled homogenous bone grafts filled with autogenous cancellous bone. Histologically, the use of homogenous cortical grafts with pre-drilled holes and autogenous cancellous bone appeared to accelerate reorganization and development of these grafts. This was seen particularly 1–2 months post-operatively and at nine months when the graft in the left leg was made up of spongy bone marrow; whereas the right leg graft still contained compact dead bone and the marrow was filled with fibrous tissue.     

In vitro comparison of equine cancellous bone graft donor sites and tibial periosteum as sources of viable osteoprogenitors

OBJECTIVE: To compare the osteogenic potential of cancellous bone of conventional graft sites with that of one nonconventional site (fourth coccygeal vertebra) and to investigate the tibial periosteum as a donor site with respect to osteogenic potential. STUDY DESIGN: In vitro osteogenic cell culture system. SAMPLE POPULATION: Eight adult horses. METHODS: Cancellous bone or tibial periosteum was aseptically collected and cut into bone chips or periosteal strips of 1 to 2 mm(3) for primary explant cultures. After 2 weeks, primary tissue cultures that yielded a population of osteogenic cells were counted and subcultured at 1 x 10(5) cells/35-mm dish in osteogenic media. After 7 to 10 days, subcultures were stained with Von Kossa (VK) to assess mineralized bone nodule formation. VK-positive bone nodules were counted as osteoprogenitors and compared among 3 donor sites, which provided consistent primary osteogenic cells (tuber coxae, fourth coccygeal vertebra, periosteum) using ANOVA (P <.05). RESULTS: Sternal and tibial bone yielded viable osteogenic cells from 25% and 50% of horses, respectively, whereas yields from tuber coxae, coccygeal vertebra, and periosteum were 75%, 100%, and 100%, respectively. Tuber coxae and periosteum had significantly greater numbers of osteoprogenitors compared with fourth coccygeal vertebra. CONCLUSIONS: Among the conventional donor sites, tuber coxae most consistently yielded viable osteogenic cells with an acceptable percentage of osteoprogenitors. Sternal and tibial sites were unreliable in providing osteogenic cells. Two new donor sites, the fourth coccygeal vertebra and tibial periosteum, were tissues with good osteogenic potential. CLINICAL RELEVANCE: When a source of transplantable viable osteoprogenitor cells is desired, use of the tuber coxae as a conventional donor site is warranted. Use of tibial periosteum or fourth coccygeal vertebra as reliable sources of transplantable osteoprogenitors should be considered.     

Comparative Structure and Morphology of the Goat Sternum in West African Dwarf and Danish Landrace Breeds

The structure and morphology of the sternum from 33 West African dwarf (WAD) and sixteen Danish Landrace breed goats were studied radiographically. In young kids of the Danish Landrace breed (DL), the number of sternal elements (Sternebrae) was six or seven with the fifth or the last but one being double in some animals due to bilateral bodies. All the sternal elements in the WAD breed appeared to have each a single locus of ossification and the number of sternal elements varied between 5, 6 or 7 at birth. In those goats with 5 or 6 sternal segments, bilateral indentations were observed on the left and right side on either the last or the last two segments. These indentations were considered to indicate a prenatal cranial coalescence of the last segments leading to the reduced number of sternebra. The structure and morphology of the sternum in neonatal kids for the two breeds appears to be basically different. 8 sternal and 5 asternal ribs were observed in animals of both breeds. The cartilaginous attachments of the ribs on the sternal segments were variable. A floating rib was also observed in one of the WAD goats. The comparative structure and morphology of other ruminant sternum is also discussed.     

Comparison of bone healing,as assessed by computed tomography,following tibial tuberosity advancement in dogs with and without autogenous cancellous bone grafts

Abstract

AIMS: To objectively compare measures of bone healing, using computed tomography (CT) in dogs following bilateral tibial tuberosity advancement (TTA), between tibiae treated with and without autogenous cancellous bone grafts.

METHODS: Ten dogs with bilateral cranial cruciate ligament disease requiring surgical stabilisation were prospectively recruited to undergo single-session bilateral TTA, with only one, randomly assigned, tibia receiving bone graft in the osteotomy deficit. Bone healing at the osteotomy site was assessed using CT performed 38–70 days post-operatively. CT images were evaluated using both objective measurements of osseous bridging and subjective evaluation by six radiologists. Repeated measures ANOVA was used to compare the objective outcomes between the grafted and non-grafted tibiae.

RESULTS: The mean percentage of the osteotomy deficit bridged at the lateral cortex was greater in grafted (77.6, SD 35.2%) compared to non-grafted (63.0, SD 36.5%) tibiae (p=0.001), but did not differ at the medial cortex (p=0.1). The mean minimum callus width was greater in grafted (7.2, SD 3.3 mm) compared to non-grafted (3.6, SD 2.9 mm) tibiae (p<0.001). There was no difference in mean attenuation (measured in Hounsfield units) of the callus between grafted and non-grafted tibiae (p=0.5). The grafted tibia was deemed to have superior bone healing in 50/60 subjective assessments made by radiologists.

CONCLUSIONS: Superior osseous bridging was detected by CT analysis following TTA using autogenous cancellous bone grafts compared with no graft. This was shown by greater bridging percentage at the lateral cortex and formation of a broader callus. Qualitative assessments made by six radiologists also supported the conclusion that bone healing was improved by use of autogenous cancellous bone graft. CT was a useful method for assessing evidence of bone healing following TTA.

CLINICAL RELEVANCE: These findings justify the application of autogenous cancellous bone graft to augment healing following TTA in dogs.     

Evaluation of a free vascularized medial tibial bone graft in dogs

OBJECTIVE: To develop a free vascularized tibial bone graft based on the periosteal saphenous blood supply. STUDY DESIGN: Preliminary anatomic study of medial tibial blood supply. In vivo comparison of a vascularized and avascular tibial bone graft. ANIMALS: Nine canine cadavers; 14 healthy adult dogs that weighed 25 to 32 kg. METHODS: An anatomic study of the vascular supply of the medial aspect of the tibia was performed using the Spalteholz technique. A bone graft consisting of the medial aspect of the tibia was transferred to a mandibular defect as a vascularized graft in 7 dogs and as an avascular graft in 7 dogs. Bone scans were performed to evaluate graft perfusion. Radiographic evaluation of the mandibles and tibias was performed. The dogs were killed after 60 days, five mandibles from each group were examined histologically, and two from each group were evaluated using the Spalteholz technique. RESULTS: The saphenous vascular pedicle provides vascular perfusion to the medial tibial cortex. Bone scans and radiographic evaluations were consistent with viable bone in the vascularized grafts, and nonviable bone in the avascular grafts. Histological examination revealed live, healing bone in vascular grafts and necrotic bone in avascular grafts. Spalteholz evaluation revealed many small arborizing vessels in the vascular grafts and no organized vasculature in the avascular grafts. CONCLUSIONS: The vascularized medial tibial cortical bone graft survived and proceeded to bony union in the mandibular body defect more readily than the avascular graft in this experimental model. CLINICAL RELEVANCE: A vascularized medial tibial bone graft is a suitable free graft for use in reconstructing bone defects in dogs.     

Healing of Canine Autogenous Cancellous Bone Graft Donor Sites

Autogenous cancellous bone was procured from the proximal ends of one humerus and the contralateral tibia of eight adult mixed-breed dogs. Greater weights of bone were consistently harvested from the humerus than from the tibia. Restoration of cancellous bone was more rapid and complete in the humerus than in the tibia. The tibia harvest site filled primarily with fibrous tissue rather than with cancellous bone.     

Sternal bone biopsy in standing horses

OBJECTIVE: To develop a technique for sternal bone biopsy in standing horses. STUDY DESIGN: Experimental study. ANIMALS: Five adult horses. METHODS: Horses were restrained in a standing stocks and sedated. The sternal biopsy site, identified by ultrasonographic examination, was clipped and prepared for aseptic surgery and infiltrated with local anesthetic. An electric bone biopsy drill (Osteocore; Institut Straumann AG, CH-4437, Waldenburg, Switzerland)), which yielded 4-mm-diameter bone specimens, was used to obtain sternal biopsies through a small skin incision. Sections (7 microm) of the bone specimens were stained with toluidine blue and Goldner's green trichrome and observed microscopically to determine suitability for histological and histomorphometric evaluation. RESULTS: The most suitable sternal biopsy site was at the 4th or 5th sternebra. The surgical procedure was easy to perform and well tolerated by the horses, and adequate samples were obtained on the first attempt. The only complications were incisional edema in all horses and wound drainage in 1 horse. CONCLUSIONS: Sternal bone biopsy may be successfully performed in standing horses, and the technique described in this report yields architecturally intact bone specimens. CLINICAL RELEVANCE: The sternum is an accessible site for cancellous bone biopsy specimens in standing horses.     

Vascularization of cancellous chip bone grafts

Fresh autogenous, frozen allogeneic, and commercially prepared xenogeneic cancellous chip bone grafts were placed into plug holes in the tibia and into surgically simulated nonunion defects in the ulna of dogs. The microvascular and correlated histologic reactions were studied at given times up to 12 weeks after the graft implantation. Marked and characteristic differences were noted among the 3 graft materials. Fresh autogenous chips were extensively vascularized by 1 week, and subsequent microscopic healing was rapid. Allogeneic chips were incorporated into all defects, but vascularization and histologic healing rates were slower than those with the fresh autogenous graft material. The xenogeneic graft was poorly incorporated. A foreign body reaction was seen histologically, and corresponding vascularization of the graft was slow and incomplete.     

Combination of a Corticai Allograft With a Cancellous Autograft in the Canine Tibia

This study was undertaken to determine what difference, if any, occurred when autogenous cancellous bone was placed in the medullary canal of a fresh, 4-cm cortical allograft compared to a fresh cortical allograft alone. Twelve mature dogs were used. A tubular cortical graft was placed in the midshaft of both tibias. The right tibia received the combination graft while the left tibia received a fresh cortical allograft only. Study periods ranged from one week to 17 weeks. The progress of the grafts was assessed by technetium-99m radionuclide bone scans, radiographs, gross and histological examination, and tetracycline labeling. Results indicate that autogenous cancellous bone placed in the medullary canal of a fresh cortical allograft was beneficial to the ingrowth of new blood vessels and incorporation of the cortical graft.     

Morphometric Study of the Equine Navicular Bone: Age-Related Changes and Influence of Exercise

Navicular bones from the four limbs of 95 horses, classified in 9 categories, were studied. The effects of age on navicular bone morphometry and histomorphometry were estimated, after adjustment of the data to even out the effects of front and rear limbs, morphometrical type, sex, weight, and size.All the external measurements of the navicular bone decreased significantly with increasing age. From the histomorphometrical data, cortical bone volume decreased with age in most horses, whereas cancellous bone volume and, in particular, the marrow spaces increased. The increase in the cancellous bone volume could have resulted from tunnelling of the internal part of the cortex, which converted it progressively into a porous trabecular-like structure. Trabecular bone volume also decreased with age and the trabecular lattice changed dramatically to become disconnected in aged horses. These observations corresponded closely to those reported for ageing of the skeletal system in humans.However, in sporting horses, the navicular cortical bone volume increased with age and the cancellous bone volume decreased. Exercise appeared to have decreased bone resorption and increased bone formation at the endocortical junction. The cancellous bone architecture was also improved, in that the trabecular lattice and trabecular bone volume remained unchanged in aged sporting horses. Our findings confirmed that exercise may be good practice to prevent age-related bone loss.     

Bone morphological analyses in Spontaneously Diabetic Torii (SDT) fatty rats

The Spontaneously Diabetic Torii (SDT) fatty rat, a model for obese type 2 diabetes, shows bone quantitative abnormalities, namely low bone mineral density (BMD). The objective of this study was to evaluate bone morphological changes, in particular identifying the bone qualitative abnormalities, in the SDT fatty rat. Male SDT fatty rats showed increases in total trabecular area and trabecular number and decreases in trabecular thickness in cancellous bones of the proximal tibia, indicating trabecular miniaturization. The SDT fatty rat is useful for investigation of pathophysiological changes in bone quality in diabetic osteoporosis.     

Effect of hyperbaric oxygen treatment on incorporation of an autogenous cancellous bone graft in a nonunion diaphyseal ulnar defect in cats

OBJECTIVE: To determine whether hyperbaric oxygen treatment (HBOT) would affect incorporation of an autogenous cancellous bone graft in diaphyseal ulnar defects in cats. ANIMALS: 12 mature cats. PROCEDURE: Bilateral nonunion diaphyseal ulnar defects were created in each cat. An autogenous cancellous bone graft was implanted in 1 ulnar defect in each cat, with the contralateral ulnar defect serving as a nongrafted specimen. Six cats were treated by use of hyperbaric oxygen at 2 atmospheres absolute for 90 minutes once daily for 14 days, and 6 cats were not treated (control group). Bone labeling was performed, using fluorochrome markers. Cats were euthanatized 5 weeks after implanting, and barium sulfate was infused to evaluate vascularization of grafts. Ulnas were evaluated by use of radiography, microangiography, histologic examination, and histomorphometric examination. RESULTS: Radiographic scores did not differ between treatment groups. Microangiographic appearance of grafted defects was similar between groups, with all having adequate vascularization. Differences were not observed between treated and nontreated groups in the overall histologic appearance of decalcified samples of tissue in grafted defects. Mean distance between fluorescent labels was significantly greater in cats given HBOT than in nontreated cats. Median percentage of bone formation in grafted defects was significantly greater in cats given HBOT. CONCLUSIONS: Hyperbaric oxygen treatment increased the distance between fluorescent labels and percentage of bone formation when incorporating autogenous cancellous bone grafts in induced nonunion diaphyseal ulnar defects in cats, but HBOT did not affect revascularization, radiographic appearance, or qualitative histologic appearance of the grafts.     

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.