Radiologic, mechanical, and histologic evaluation of 2 glenoid prosthesis designs in a canine model.

Aseptic loosening of glenoid components is a common problem associated with total shoulder arthroplasty. A new glenoid design aimed at improving fixation outcomes was compared with conventional keeled glenoids in weight-bearing canine shoulders. Radiographic, histologic, and mechanical tests were performed at 3 postoperative intervals (0, 3, and 6 months). The uncemented pegged glenoid achieved bone ingrowth around the peg flanges in each case. This result was confirmed histologically and radiographically. Mechanical results indicated that mean fixation strength increases significantly between 0 and 3 months after surgery and remains strong through 6 months. In contrast, conventional keeled glenoids were found to have partial or complete radiolucent lines around the keel in each instance, and mechanical testing demonstrated that mean fixation strength weakens significantly between 0 and 3 months after surgery and remains weak through 6 months. These results show that stem design changes can improve implant fixation. A cementless fluted peg stem was superior to a conventional cemented keel design in achieving osseous integration and fixation in a weight-bearing animal model.

Bone morphogenetic protein but not transforming growth factor-beta enhances bone formation in canine diaphyseal nonunions implanted with a biodegradable composite polymer.

BACKGROUND: The purpose of the present study was to create an effective bone-graft substitute for the treatment of a diaphyseal nonunion. METHODS: A standardized nonunion was established in the midportion of the radial diaphysis in thirty mongrel dogs by creating a three-millimeter segmental bone defect (at least 2 percent of the total length of the bone). The nonunion was treated with implantation of a carrier comprised of poly(DL-lactic acid) and polyglycolic acid copolymer (50:50 polylactic acid-polyglycolic acid [PLG50]) containing canine purified bone morphogenetic protein (BMP) or recombinant human transforming growth factor-beta (TGF-beta1), or both, or the carrier without BMP or TGF-beta1. Five groups, consisting of six dogs each, were treated with implantation of the carrier alone, implantation of the carrier with fifteen milligrams of BMP, implantation of the carrier with 1.5 milligrams of BMP, implantation of the carrier with fifteen milligrams of BMP and ten nanograms of TGF-beta1, or implantation of the carrier with ten nanograms of TGF-beta1. At twelve weeks after implantation, the radii were examined radiographically and the sites of nonunion were examined histomorphometrically. RESULTS: We found that implantation of the polylactic acid-polyglycolic acid carrier alone or in combination with ten nanograms of TGF-beta1 failed to induce significant radiographic or histomorphometric evidence of healing at the site of the nonunion. The radii treated with the carrier enriched with either 1.5 or fifteen milligrams of BMP showed significantly increased periosteal and endosteal bone formation on histomorphometric (p < 0.05) and radiographic (p < 0.02) analysis. CONCLUSIONS: Bone formation in a persistent osseous defect that is similar to an ununited diaphyseal fracture is increased when species-specific BMP incorporated into a polylactic acid-polyglycolic acid carrier is implanted at the site of the nonunion. TGF-beta1 at a dose of ten nanograms per implant did not induce a similar degree of bone formation or potentiate the effect of BMP in this model. CLINICAL RELEVANCE: The biodegradable implant containing BMP that was used in the present study to treat diaphyseal nonunion is an effective bone-graft substitute.

Guided tissue fabrication from periosteum using preformed biodegradable polymer scaffolds.

A successful tissue engineering method for bone replacement would imitate natural bone graft by providing the essential elements for new bone formation using synthetic scaffolds, osteogenic cell populations, and bone induction factors. This is a study of the suitability of various formulations of poly(DL-lactic-co-glycolic acid) (PLGA) foams to provide a tissue conducting scaffold in an ovine model for bone flap fabrication. Three formulations were used of different copolymer ratio and molecular weight. Porous wafers of PLGA were stacked into rectangular chambers (volume 4 cm3) enclosed on five sides. Some chambers also contained autologous morcellized bone graft (MBG). The chambers were inserted with the open face adjacent to the cambium layer of the periosteum in rib beds of seven sheep and harvested after 8 weeks in vivo. Gross and histologic examination of the resulting tissue specimens demonstrated molded units of vascularized tissue generally conforming to the shape of the chambers and firmly attached to the periosteum. Polymer degradation appeared to occur by varying degrees based on polymer formulation. New bone formation was observed only in areas containing MBG. There was no evidence of significant inflammatory reaction or local tissue damage at 8 weeks. We conclude that a PLGA foam scaffold is (1) an efficient conductor of new tissue growth but not osteoinductive, (2) contributes to the shape of molded tissue, and (3) biocompatible when used in this model. Further studies are warranted to develop practical methods to deliver bone induction factors to the system to promote osseous tissue generation throughout the synthetic scaffold.

Novel formulation of fibroblast growth factor-2 in a hyaluronan gel accelerates fracture healing in nonhuman primates.

Recent advances in understanding the biology of fracture healing and the availability of specific macromolecules has resulted in the development of novel treatments for injuries to bone. Fibroblast growth factor-2 or basic fibroblast growth factor (4 mg/ml), a potent mitogen, and hyaluronan (20 mg/ml), an extracellular matrix component, were combined into a viscous gel formulation intended for direct, percutaneous injection into fresh fractures. In an experimental primate fracture model, a bilateral 1-mm-gap osteotomy was surgically created in the fibulae of baboons. A single direct administration of this hyaluronan/fibroblast growth factor-2 formulation to the defect site significantly promoted local fracture healing as evidenced by increased callus formation and mechanical strength. Radiographic analysis showed that the callus area was statistically significantly larger at the treated sites than at the untreated sites. Specimens treated with 0.1, 0.25, and 0.75 ml hyaluronan/fibroblast growth factor-2 demonstrated a 48, 50, and 34% greater average load at failure and an 82, 104, and 66% greater energy to failure than the untreated controls, respectively. By histologic analysis, the callus size, periosteal reaction, vascularity, and cellularity were consistently more pronounced in the treated osteotomies than in the untreated controls. These results suggest that hyaluronan/fibroblast growth factor-2 may provide a significant advance in the treatment of fractures.

Neoplastic odontogenic epithelial cells express bone sialoprotein.

Bone sialoprotein (BSP) is synthesized and secreted by bone-, dentine- and cementum-forming cells and has been implicated in de novo bone formation and mineralization. In this study, we used histological sections of odontogenic neoplasms and performed immunohistochemical and in situ hybridization analyses. In ameloblastoma, BSP mRNA signals were seen in the neoplastic epithelial cells forming nests, strips and islands. BSP deposition was also seen in the stellate reticulum of the tumour masses revealed by immunohistochemistry using human BSP antibodies. In calcifying epithelial odontogenic tumour, the calcified masses demonstrated positive immunoreactivity to the human BSP antibodies, and the hybridization signals for BSP were located in the cells near the calcified particles. In the calcifying odontogenic cyst, strong BSP signals were seen in cells surrounding the characteristic nests of ghost cells, which often calcify subsequently. BSP protein was also found in these cells by immunohistochemistry. The active expression of BSP in the epithelial elements of the odontogenic tumours of adult patients suggests the activation of this matrix protein gene in the neoplastic process, and that BSP may play an important role in tumour formation and differentiation with respect to pathological calcification.

Enamel epithelium expresses bone sialoprotein (BSP).

Bone sialoprotein (BSP) is a major non-collagenous extracellular matrix protein in bone and other mineralized connective tissues. BSP is synthesized and secreted by bone-, dentin- and cementum-forming cells. In this study we hypothesized that BSP may be also involved in enamel formation through its postulated role in matrix mineralization. In situ hybridization with cRNA probes for rat and hamster BSP, respectively, showed strong mRNA signals in ameloblasts actively synthesizing enamel matrix in developing incisors. However, no hybridization signals were observed at an earlier developmental stage when bell-shaped molar tooth germs were being formed. Immunohistochemical analysis of tooth tissues from transgenic mice harboring a 2.7 kb rat BSP promoter ligated to a luciferase reporter gene revealed strong staining for luciferase in the enamel epithelium of the developing tooth germ. Interestingly, BSP expression was also observed in epithelial cells of an ameloblastoma. The neoplastic epithelial nests and cords demonstrated strong mRNA signals to the human BSP probe while the connective tissue stroma showed only a background level of silver grains. Immunostaining also showed deposition of BSP by the odontogenic cells of the tumor. These results demonstrate that BSP is expressed by the enamel-forming epithelium of developing teeth, suggesting a possible role for BSP in enamel formation and its subsequent mineralization. Expression of the BSP gene in ameloblastomas is consistent with the expression of BSP by the enamel epithelium and also with the expression of BSP by neoplastic tissues, suggesting a possible role in tumorigenesis.

Repair of soft tissue to bone using a biodegradable suture anchor.

The medial collateral ligaments of 18 New Zealand rabbits were surgically detached from bone. In one knee, the ligament was repaired using a biodegradable suture anchor composed of a co-polymer of lactic and glycolic acid. The contralateral medial collateral ligament was not repaired. Animals were sacrificed at 4, 8, and 12 weeks after the operation, and the knee that had the ligament repair was compared with the contralateral control knee. All knees were tested manually tested for stability to valgus stress and then prepared for histologic examination. Medial collateral ligaments repaired using the biodegradable suture anchor demonstrated stability to valgus stress and anatomic healing at the bone-tendon junction. Resorption of the implant was virtually complete by 12 weeks. All specimens demonstrated less inflammatory reaction to the suture anchor than to the attached Vicryl suture. This contrasts with the control group, which was grossly unstable and demonstrated scarring in this nonanatomic position. These results demonstrate efficacy of this particular material of biodegradable implant and justify further investigative efforts.

Markers for macrophage and osteoclast lineages in giant cell lesions of the oral cavity.

PURPOSE: Giant cell lesions of the oral cavity are a well recognized entity. However, the histogenesis of these lesions is still the subject of controversy, with support for both histiocyte/macrophage and osteoclast origins being found in the literature. This study evaluated a set of peripheral giant cell lesions (PGCLs) and central giant cell lesions (CGCLs) for characteristics of both cell types to address this dilemma. MATERIALS AND METHODS: Detection of histiocyte/macrophage characteristics was accomplished immunohistochemically by evaluating for markers specific for this cell type, namely alpha-1 -antichymotrypsin (1 -ACT) and factor XIIIa antibodies. Detection of osteoclast characteristics made use of the fact that osteoclasts possess a unique enzyme, tartrate-resistant acid phosphatase, which can be appreciated by histochemical procedures. RESULTS: A large percentage of the multinucleated cells stained with the 1-ACT (38.08% in PGCLs and 15.84% in CGCLs), while only isolated cells stained for factor XIIIa (1.20% PGCLs, 0.99% CGCLs). Isolated stromal cells also were stained. Virtually all multinucleated cells reacted with the tartrate-resistant acid phosphatase stain (99.26% PGCLs, 98.34% CGCLs), as did a number of the mononuclear stromal cells. CONCLUSIONS: This study supports the contention that GCLs of the oral cavity may arise from precursor cells related to the granulocyte/macrophage line, and may originate from mononuclear cells that express markers for both macrophages and osteoclasts.

Evaluation of the Palmaz stent in the trachea and mainstem bronchi of normal dogs.

OBJECTIVE: This study evaluates the efficacy of the Palmaz balloon expandable intraluminal stent in the trachea and mainstem bronchi of normal dogs. STUDY DESIGN: Effects of the stent were evaluated by physical examination, thoracic radiography, respiratory tract fluoroscopy and endoscopy, tracheal diameter measurement, postmortem examination, and airway histomorphometry. ANIMALS OR SAMPLE POPULATION: Ten normal beagle dogs. METHODS: Stent size was estimated from thoracic radiographs in awake dogs. Group I dogs (n = 4) had two stents placed: one in the thoracic trachea (TT) plus a randomly chosen mainstem bronchus (MB). Group II dogs (n = 3) had stents placed in the MB (one stent), TT (one stent), and mid- or proximal cervical trachea (CT) (one or two stents). Three dogs were used as sham-operated controls (group III). Temperature, pulse, respiration, and cough were measured twice daily. Dogs were evaluated at 21 and 49 to 56 days after stent placement, euthanatized, and tissues were collected for histomorphometric analysis of stent integration and epithelial pathology. RESULTS: Mean tracheal diameters of awake (10.5 +/- 1.7 mm) and anesthetized dogs before stent implantation (13.9 +/- 2.0 mm) were significantly different (P < .01). Complications associated with stent placement included acute pulmonary edema (n = 2), stent migration (n = 7), stent collapse (n = 4 CT, 2 TT, and 1 MB), and positive tracheal culture (n = 10). Group II dogs coughed more at rest, exercise, and with tracheal palpation than dogs in other groups (P < .01). Group I dogs coughed more at rest than group III dogs (P < .01). Stent integration ranged from 0 to 91.3%. Squamous metaplasia and epithelial ulceration associated with stents ranged from 0 to 57.5% and 0 to 32.7%, respectively. CONCLUSIONS: Determination of stent size should be based on measurements taken on anesthetized dogs because use of inappropriately sized stents may promote stent migration, squamous metaplasia, and/or ulceration. Epithelialization over stent struts may occur if the stent is closely associated with tracheal epithelium. CLINICAL RELEVANCE: Palmaz stents do not appear to be appropriate for placement in the CT of dogs; however, with technical modifications, application in the TT and MB may be feasible.

Anatomic significance of magnetic resonance imaging findings in hip fracture.

This study was done to determine the histologic etiology for the linear black signal seen on magnetic resonance imaging at the site of proximal femoral fractures. No previous satisfactory explanation for this black line exists in the literature. To determine the pathologic etiology of this linear black signal, the magnetic resonance imaging appearances of osteotomies and fractures of proximal femora were correlated with gross and histologic appearances in 12 freshly frozen porcine hips. In phase 1, a unilateral femoral neck osteotomy was performed in 1 hip, and a sham operation was performed on the contralateral hip. The radiologist was able to easily determine the site and side of the osteotomy using magnetic resonance imaging. In phase 2, the remaining 8 femoral specimens were stressed using the 1125 Instron Materials Testing Machine in a compression mode. In the 8 specimens, 4 fractures were created that were visible on magnetic resonance imaging and plain radiographs; 2 fractures were apparent only on magnetic resonance images, and 2 specimens did not have fractures identified by plain radiographs or magnetic resonance images. Gross and microscopic analysis of the specimens confirmed the magnetic resonance imaging findings in all 8 cases (6 fractures and 2 nonfractures). Histologic evaluation showed trabecular impaction at the 6 fracture sites corresponding to the linear black signal on magnetic resonance images. No edema or hemorrhage was present in these postmortem specimens. The authors conclude that the low signal band seen with proximal femoral fractures is produced by impaction of the trabecular bone and is not the result of perifracture edema or hemorrhage.

Sublabral recess of the superior glenoid labrum: study of cadavers with conventional nonenhanced MR imaging, MR arthrography, anatomic dissection, and limited histologic examination.

PURPOSE: To evaluate size, location, and appearance of the sublabral recess of the superior glenoid labrum, which mimics a superior labral tear oriented in the anterior to posterior direction (SLAP) lesion with conventional magnetic resonance (MR) imaging, MR arthrography, gross dissection, and limited histologic evaluation. MATERIALS AND METHODS: Twenty-six cadaveric shoulder specimens that were freshly frozen were examined with nonenhanced MR imaging and with MR arthrography after intraarticular injection of dilute contrast material with gadolinium. In all specimens, the appearance of the superior glenolabral junction was categorized and was correlated to that seen at gross anatomic dissection. Histologic sections were obtained of two shoulders with large sublabral recesses in the plane of the MR sections. RESULTS: A sublabral recess was present in 19 (73%) shoulders and was deeper than 2 mm in 10 (39%). MR arthrography better demonstrated the sublabral recess than nonenhanced MR imaging. In 16 of 19 shoulders, the sublabral recess was located in the most anterior section obtained through the superior labrum. There was no definite correlation between subject age and sex and glenolabral junction type. At histologic examination of the two shoulders, a synovial lining of the sublabral recess was seen and there was no evidence of fibrosis to suggest a traumatic cause. CONCLUSION: A sublabral recess is common in cadaveric shoulders and has an appearance similar to that of published examples of SLAP lesions. Histologic findings were consistent with normal anatomic structure rather than with acquired, posttraumatic lesions.

Osteoblast migration on poly(alpha-hydroxy esters).

We investigated the migration of rat calvaria osteoblast populations on poly(alpha-hydroxy ester) films for up to 14 days to determine effects of substrate composition and culture conditions on the migratory characteristics of osteoblasts. Initial osteoblast culture conditions included cell colonies formed by seeding a high (84,000 cells/cm(2)) or low (42,000 cells/cm(2)) density of isolated osteoblasts on the polymer films, and bone tissue cultures formed by plating bone chips directly on the substrates. High density osteoblast colonies cultured and allowed to migrate and proliferate radially on 85:15 poly(DL-lactic-co-glycolic acid) (PLGA) films, 75:25 PLGA films, and tissue culture polystyrene controls demonstrated that the copolymer ratio in the polymer films did not affect the rate of increase in substrate surface area (or culture area) covered by the growing cell colony. However, the rate of increase in culture area was dependent on the initial osteoblast seeding density. Initial cell colonies formed with a lower osteoblast seeding density on 75:25 PLGA resulted in a lower rate of increase in culture area, specifically 4.9 +/- 0.3 mm(2)/day, versus 14.1 +/- 0.7 mm(2)/day for colonies seeded with a higher density of cells on the same polymer films. The proliferation rate for osteoblasts in the high and low density seeded osteoblast colonies did not differ, whereas the proliferation rate for the osteoblasts arising from the bone chips was lower than either of these isolated cell colonies. Confocal and light microscopy revealed that the osteoblast migration occurred as a monolayer of individual osteoblasts and not a calcified tissue front. These results demonstrated that cell seeding conditions strongly affect the rates of osteoblast migration and proliferation on biodegradable poly(alpha-hydroxy esters). (c) 1996 John Wiley & Sons, Inc.

Periodontitis in the baboon: a potential model for human disease.

Advances in periodontics with respect to disease activity, microbiology and immunology have demonstrated the multifactorial nature of periodontal diseases. This serves to underscore the need for an ideal animal model for periodontal research. Non-human primates are most similar to man in comparison to other animal models. The baboon is an Old World monkey that has infrequently been used in periodontal research. Periodontal exams were accomplished on 116 baboons (Papio anubis, P. cynocephalus) ages 5 to 30 years with one baboon year being roughly equivalent to 3 to 4 human years. The study population consisted of 29 males and 87 females. Clinical parameters including probing depth, attachment level, mobility, plaque index and gingival index were collected. Radiographs were taken on 25 animals and correlated to clinical findings. Results showed a significant increase in mean probing depth and mean attachment level with age (p = 0.0001). Disease prevalence and severity were not significantly different between genders. Mobility was uncommon; however, the prevalence and severity of furcation involvement increased with age. Radiographs suggested horizontal and isolated vertical bone loss. Plaque and gingival indices were at sustained high levels for all age groups and showed a statistically significant increase with age. Some baboons were found to develop a naturally-occurring periodontitis that increased in severity with age. This primate may be a suitable model for studies in human periodontal disease.

Analysis of transforming growth factor beta and other cytokines in autoimmune exocrinopathy (Sjögren's syndrome).

Cytokines play a major role in tissue destruction caused by autoimmune dysregulation. In Sjögren's syndrome (SS) patients, salivary glands are the target organs for autoimmune tissue damage. In the present study, reverse transcriptase-polymerase chain reaction (RT-PCR) was used to look for cytokine mRNA expressed in SS salivary glands. Focus score was used to determine the severity of the lesions. Cytokine production in supernatants of the salivary gland cell culture was measured by enzyme-linked immunosorbent assay (ELISA). Immunohistochemical staining was used to identify the local presence of transforming growth factor beta (TGF-beta). Interleukin (IL)-2, IL-6, and IL-10 mRNA were expressed in moderate to severe SS salivary gland lesions. TGF-beta mRNA was constitutively expressed in normal and SS salivary glands. In SS salivary gland cell cultures, IL-6 and IL-10 proteins were produced. TGF-beta production was reduced in high focus score SS glands. Normal and minimally involved SS salivary gland ductal epithelium and acinar cells were found to produce TGF-beta by immunostaining. In conclusion, an excess production of IL-2, IL-6, and IL-10 and a reduced production of the immunosuppressive cytokine, TGF-beta, may be responsible for the progression of the salivary gland lesion in SS. Specific immunotherapy can now be designed based on mechanisms to correct this cytokine imbalance and benefit patients with autoimmune diseases, such as SS.

Localization of cytokines in cholesteatoma tissue.

Acquired cholesteatoma is associated with an intense inflammatory reaction with resultant tissue and bone destruction. Cytokines are molecules released by inflammatory cells at the site of infection and are potent mediators of inflammation and the immune response. Five cytokines, tumor necrosis factor-alpha, transforming growth factor-beta 1 and 2, and interleukin-1 and 6, were immunolocalized in human cholesteatoma epithelium and subepithelial stroma, with greater intensity of staining compared with noninflamed external auditory canal skin. Increased interleukin-6 activity in cholesteatoma epithelium and stroma correlated significantly with the presence of ossicular and bony erosion and granulation tissue noted intraoperatively. Transforming growth factor-beta 2 activity in cholesteatoma epithelium correlated significantly with bony erosion at surgery. Additionally, transforming growth factor-beta 1 activity in cholesteatoma epithelium correlated significantly with increased length of disease. Tumor necrosis factor-alpha, interleukin-1, and interleukin-6 appear to be involved in the inflammation and resultant remodeling associated with cholesteatoma. We hypothesized a protective function of transforming growth factor-beta 1 and 2 in the presence of cholesteatoma. The antiinflammatory and osteoclast and keratinocyte inhibitory actions of the transforming growth factor-beta s could potentially slow the proliferation and resultant tissue destructiveness associated with cholesteatoma.