Recommendations of the OARSI FDA Osteoarthritis Devices Working Group.

Osteoarthritis (OA) is the most common type of arthritis and a major cause of chronic musculoskeletal pain and functional disability. While both pharmacologic and non-pharmacologic modalities are recommended in the management of OA, when patients with hip or knee OA do not obtain adequate pain relief and/or functional improvement, joint replacement surgery or other surgical interventions should be considered. Total joint arthroplasties are reliable and cost-effective treatments for patients with significant OA of the hip and knee. Evidence from cohort and observational studies has confirmed substantial improvements in pain relief with cumulative revision rates at 10 years following total hip (THA) and total knee arthroplasties (TKA) at 7% and 10%, respectively. Joint replacements have been used in most every synovial joint, although results for joints other than hip and knee replacement have not been as successful. The evolution of new device designs and surgical techniques highlights the need to better understand the risk to benefit ratio for different joint replacements and to identify the appropriate methodology for evaluating the efficacy and optimal outcomes of these new devices, designed to treat OA joints.

'Gelatinase-like' activity from articular chondrocytes in monolayer culture.

In addition to releasing collagenase and proteoglycanase activity, rabbit articular chondrocytes in monolayer culture released into the culture medium, latent, neutral enzyme activity which when activated by p-aminophenylmercuric acetate degraded fluorescein-labeled polymeric rat tail tendon Type I collagen and the tropocollagen TCA and TCB fragments of human Type II collagen into smaller peptides at 37 degrees C. Enzyme activity was abolished if p-aminophenylmercuric acetate-activated culture medium was preincubated with 1.10-phenanthroline, a metal chelator. Thus, articular chondrocytes in monolayer culture are capable of producing neutral proteinases which acting together can result in complete degradation of tendon and cartilage collagen to small peptides.

Osteoclast differentiation requires ascorbic acid.

Osteoclast differentiation assays are usually conducted in alpha minimal essential medium (alpha-MEM). We reasoned that determining which components of this media are critical for osteoclast differentiation might provide insight into the mechanisms that regulate osteoclast differentiation. This study demonstrates that ascorbic acid is the crucial component of alpha-MEM that stimulates differentiation of murine osteoclasts in cocultures with murine mesenchymal support cells. Thus, supplementation with ascorbic acid allows osteoclast differentiation to occur in basal MEM media as well as in RPMI-1640 and basal media Eagle (BME) media. The conclusion that osteoclast differentiation is stimulated by ascorbic acid was obtained whether osteoclast differentiation was induced by 1,25-dihydroxyvitamin D3 or parathyroid hormone, whether ST2 or CIMC-2 cells were used as mesenchymal support cells, and whether osteoclast precursors were obtained from spleen or bone marrow. Time course studies revealed that although ascorbic acid only modestly increases the rate at which osteoclast precursors begin to express tartrate-resistant acid phosphatase, it strongly increases the rate at which precursors fuse into mature, multinucleated cells. Moreover, ascorbic acid strongly increases the life span of both osteoclasts and their precursors. The increases in precursor formation, fusion, and life span induced by ascorbic acid are together responsible for the stimulation of osteoclast differentiation by ascorbic acid. Given the known effects of ascorbic acid on differentiation of mesenchymal cells, it may stimulate osteoclast differentiation indirectly by regulating the differentiation state of the mesenchymal cells that support osteoclast differentiation.

Adherent endotoxin on orthopedic wear particles stimulates cytokine production and osteoclast differentiation.

Aseptic loosening of orthopedic implants is thought to be caused primarily by osteoclast differentiation induced by bone resorptive cytokines produced in response to phagocytosis of implant-derived wear particles. This study examined whether adherent endotoxin on the wear particles is responsible for inducing osteoclast differentiation as well as production of interleukin-1beta (IL-1beta), IL-6, and tumor necrosis factor a (TNF-alpha). Removal of adherent endotoxin almost completely inhibited the responses to titanium (Ti) particles by both murine marrow cells and human peripheral blood monocytes. In vivo experiments showed that endotoxin removal reduced particle-induced osteolysis by 50-70%. Addition of lipopolysaccharide (LPS) to the "endotoxin-free" particles restored their ability to induce cytokine production and osteoclast differentiation in vitro. Moreover, marrow cells from mice that are hyporesponsive to endotoxin because of mutation of Toll-like receptor 4 induced significantly less cytokine production and osteoclast differentiation in response to Ti particles with adherent endotoxin than did marrow cells from normoresponsive mice. This mutation also resulted in significantly less particle-induced osteolysis in vivo. Taken together, these results show that adherent endotoxin is involved in many of the biological responses induced by orthopedic wear particles and should stimulate development of new approaches designed to reduce the activity of adherent endotoxin in patients with orthopedic implants.

Characterization of cells with osteogenic potential from human marrow.

Studies using animal tissue suggest that bone marrow contains cells with the potential to differentiate into cartilage and bone. We report the extension of these studies to include human marrow. Bone marrow from male and female donors of various ages was obtained either from the femoral head or as aspirates from the iliac crest, and introduced into culture. Culture-adherent cells were expanded, subcultured, and then tested for bone and cartilage differentiation potential utilizing two different in vivo assays in nude mice. One assay involved subcutaneous implantation of porous calcium phosphate ceramics loaded with cultured, marrow-derived, mesenchymal cells; the other involved peritoneal implantation of diffusion chambers, also inoculated with cultured, marrow-derived, mesenchymal cells. Histological evaluation showed bone formation in ceramics implanted with cultured, marrow-derived, mesenchymal cells originating from both the femoral head and the iliac crest. Immunocytochemical analysis indicates that the bone is derived from the implanted human cells and not from the cells of the rodent host. No cartilage was observed in any of these ceramic grafts. In contrast, aliquots from the same preparations of cultured, marrow-derived, mesenchymal cells failed to form bone or cartilage in diffusion chambers. These data suggest that human marrow contains cells with osteogenic potential, which can be enriched and expanded in culture. Our findings also suggest that subcutaneous implantation of these cells in porous calcium phosphate ceramics may be a more sensitive in vivo assay than diffusion chambers for measuring their osteogenic lineage potential.

Bone mass and comparative rates of bone resorption and formation of fibular autografts: comparison of vascular and nonvascular grafts in dogs.

The metabolic fate of whole grafts that were either vascularized or nonvascularized were compared. This study was designed to quantify and correlate changes in bone resorption, formation, and mass in orthotopic, stably fixed, weight-bearing autografts. The grafts were 8-cm segments of the fibula that were internally fixed. Fibula segments subjected to sham operations, nonvascularized autografts, and vascularized autografts were studied in 16 dogs at three months after surgery. Three months prior to surgery the dogs were labeled repeatedly over two months with 3H-tetracycline and 3H-proline. Metabolic turnover of whole grafts was evaluated by quantifying loss of 3H-tetracycline for measuring postoperative resorption of bone mineral and loss of 3H-collagen for resorption of bone collagen. Net changes in bone dry weight, calcium, and collagen per whole grafts were obtained to determine differential changes in the mineral and matrix mass. The difference in change between bone resorption and bone mass was used to determine the amount of new bone formation that had replaced the resorbed bone. Vascularized autografts lost more mass (12%), and had more bone resorption (40%) and more bone formation (28%) than sham operated and unoperated fibulas. Nonvascularized grafts lost much more bone mass (48%) because resorption was large (61%) and formation was relatively small (13%). More new bone was formed in vascularized autografts than in nonvascularized autografts. During the incorporation of bone grafts, resorption is an early and rapid process, whereas formation is a late and slow process.(ABSTRACT TRUNCATED AT 250 WORDS)

Bone and cartilage formation in diffusion chambers by subcultured cells derived from the periosteum.

Periosteal cells were enzymatically isolated from the tibiae of young chicks, introduced into cell culture, allowed to reach confluence, and subcultured. The freshly isolated or subcultured cells were loaded into diffusion chambers and implanted into the peritoneal cavity of athymic mice to test their osteo-chondrogenic potential in a contained in vivo location. Freshly isolated periosteal cells formed both bone and cartilage tissue in such test chambers, but with a relatively low incidence. In contrast, cultured periosteal cells consistently gave rise to bone and cartilage even after 10 population doublings. With further passages of cells, the osteo-chondrogenic potential diminished substantially, until complete loss of expressivity at 16 population doublings or longer. Cultured muscle fibroblasts, when loaded into diffusion chambers under identical conditions to those of cultured periosteal cells, formed neither bone nor cartilage. These observations suggest that periosteal cells of young chicks contain subsets of progenitor cells or mesenchymal stem cells which possess the potential to differentiate into osteoblasts or chondrocytes, and this potential is retained after enzymatic isolation and for several population doublings in culture.

Future directions for research and treatment of osteoarthritis.

Future directions in the research and treatment of osteoarthritis (OA) will be based on the emerging picture of pathophysiological events that govern the initiation and progression of OA. The fundamental event resulting in the destruction of articular cartilage in OA arises from an imbalance between anabolic and catabolic pathways. The extracellular matrix (ECM) of cartilage is degraded by matrix metalloproteinases (MMPs) induced by cytokines. Cytokines also blunt chondrocyte compensatory synthesis pathways required to restore the integrity of the degraded ECM. Inhibition of the MMPs, their activators, and cytokines that induce MMP gene up-regulation would appear to be fertile targets for drug development in the treatment of OA. Restoration of damaged articular surfaces via tissue engineering strategies which could employ chondroprogenitor cells in biomatrices appropriate for transplantation to cartilage surfaces appears feasible. A reduction in cytokine-mediated up-regulation of MMP gene expression as well as augmentation of cartilage ECM biosynthesis may also be possible by employing the principles of gene transfer using suitable vectors that establish long-term stable expression of genes which suppress MMPs while at the same time supporting cartilage ECM biosynthesis.

Groundwater pollution by nitrates from livestock wastes.

Utilization of wastes from livestock complexes for irrigation involves the danger of groundwater pollution by nitrates. In order to prevent and minimize pollution, it is necessary to apply geological-hydrogeological evidence and concepts to the situation of wastewater irrigation for the purposes of studying natural groundwater protectiveness and predicting changes in groundwater quality as a result of infiltrating wastes. The procedure of protectiveness evaluation and quality prediction is described. With groundwater pollution by nitrate nitrogen, the concentration of ammonium nitrogen noticeably increases. One of the reasons for this change is the process of denitrification due to changes in the hydrogeochemical conditions in a layer. At representative field sites, it is necessary to collect systematic stationary observations of the concentrations of nitrogenous compounds in groundwater and changes in redox conditions and temperature.

Osteogenic potential of culture-expanded rat marrow cells as assayed in vivo with porous calcium phosphate ceramic.

It has been established that, when whole marrow is introduced into porous calcium phosphate ceramic, bone forms on the walls of the pores. To extend earlier studies, bone marrow cells derived from the femora of inbred rats were introduced into tissue culture and the adherent cells cultivated, mitotically expanded, passaged, harvested, placed in small cubes of porous calcium phosphate ceramics and grafted into subcutaneous sites of syngeneic rats. Marrow-derived, cultured mesenchymal cells introduced into ceramics showed strong osteogenic potential, with bone forming in the pore regions of ceramics as early as 2 wk after implantation. Osteogenesis could be observed after the eighteenth passage. With increasing passage number, the initiation of osteogenesis and the apparent rate of bone formation declined and the course of osteogenesis was delayed. In the future, it may be possible to culture marrow cells as a source for reparative cells for implantation back into autologous in vivo sites.

Proteoglycan composition and biosynthesis by human osteochondrophytic spurs of the femoral head.

The composition and biosynthesis of sulphated proteoglycan by human osteophytes and cells derived from them by explanation were studied. In organ-culture, the newly synthesized proteoglycan aggregated poorly with hyaluronic acid but was roughly compared to the proteoglycan aggregates derived from osteofemoral cartilage. Other characteristics of adult hyaline cartilage were preserved (i.e., high ratio of chondroitin-6-sulphate to chondroitin-4-sulphate, large hydrodynamic size and long chondroitin sulphate chains). Moderately high levels of keratan-sulphate (15-17% of the total glycosaminoglycans) were found and large keratan sulphate chains (Mr greater than 7000) confirmed by use of monoclonal antibody 1/20/5D4. Cells generated from the explants synthesized both hydrodynamically large proteoglycans and small proteoglycans. The most buoyant fraction of a CsCl density gradient (A4) contained considerable amounts of dermatan sulphate (5.2-24.4%). The Kav of the A4 fraction was 0.56 on Sepharose CL-2B and contained large glycosaminoglycan chains (Kav, 0.36 on Sepharose CL-6B). The measurements were similar to those obtained separately on osteofemoral head cartilage. These studies showed that the cartilaginous cap of human osteophytes has the capacity to synthesize the entire repertoire of sulphated proteoglycans of mature hyaline cartilage.

Meniscus regeneration in a rabbit partial meniscectomy model.

Meniscectomy is known to be associated with osteoarthrosis of the knee. The purpose of this study was to compare the natural and augmented repair of menisci in the knees of New Zealand White rabbits. To create a partial defect in the medial meniscus, we used an experimental model that has been well characterized and extensively used in the study of osteoarthrosis and articular cartilage repair. The defect was left untreated or treated with one of the following: a periosteal autograft, a type I collagen sponge, or the same sponge loaded with autologous, bone marrow-derived, cultured mesenchymal stem cells. The natural repair was always incomplete and degenerative changes within these joints were progressive. The periosteal autograft underwent differentiation into a bone and hyaline cartilage composite that was ineffectual as a meniscus and accelerated the degenerative changes in those joints when compared to natural repair controls. There was evidence of a consistent sequence of events in the transformation of the periosteal grafts to a core of cartilage that underwent endochondral ossification. In the last two groups, the collagen sponge functioned as a scaffold that resulted in more abundant repair tissue. The collagen sponge alone supported a largely fibrous repair process. The cultured mesenchymal stem cells were observed to augment the repair process in some specimens to include fibrocartilage histologically similar to normal meniscus. Degenerative changes were present in both of these groups, which indicates that the biomechanical function of the meniscus was not restored, or an irreversible osteoarthrosis cascade was initiated during the repair period. Based on these preliminary studies, further investigation of cell-based meniscus regeneration appears to be warranted.

Repair of large full-thickness articular cartilage defects with allograft articular chondrocytes embedded in a collagen gel.

Full-thickness articular cartilage defects are a major clinical problem; however, presently there is no treatment available to regeneratively repair these lesions. The current therapeutic approach is to drill the base of the defect to expose the subchondral bone with its cells and growth factors. This usually results in a repair tissue of fibrocartilage that functions poorly in the loaded joint environment. The use of phenotypically appropriate chondrocytes embedded in a collagen gel delivery vehicle may provide a method that could be used to repair full-thickness articular cartilage defects with functionally satisfactory hyaline cartilage. Allograft articular chondrocytes embedded in a type I collagen gel were transplanted into large (6 x 3 x 3 mm), full-thickness articular cartilage defects in condylar and patellar weight-bearing surfaces to develop clinically applicable methods to repair articular cartilage defects. Chondrocytes were isolated from the articular cartilage of 4-week-old New Zealand rabbits and embedded in type I collagen gels. This composite was transplanted into a full-thickness defect on the medial femoral condyle and patellar groove of adolescent host rabbits. The repair cartilage was assessed histologically by a semiquantitative scoring system and biomechanically with a microindentation technique of specimens 4-48 weeks after chondrocyte transplantation. Defects in both locations were repaired with histologically apparent hyaline cartilage observed from as early as 4 weeks until 48 weeks after transplantation. The repair cartilage in the medial femoral condyle was more irregular than in the patellar groove, but in all other respects was similar. The grafted tissue did not remodel and differentiate into the morphological zones seen in normal articular cartilage. No tidemark or subchondral bony plate formed even 48 weeks after transplantation. Biomechanically, the repaired cartilage demonstrated indentation values similar to normal articular cartilage 12 weeks after transplantation and remained the same 48 weeks after transplantation. By contrast, the control (i.e., empty) defects healed with tissue that exhibited very poor metachromatic staining and exhibited very high indentation values. Incomplete bonding of the repair tissue to the normal cartilage was seen, and the surface was significantly irregular with major discontinuities. These observations provide the basis for considering the use of allograft articular chondrocytes to repair articular cartilage defects in the weight-bearing regions of the knee.

Tissue-engineered fabrication of an osteochondral composite graft using rat bone marrow-derived mesenchymal stem cells.

This study tested the tissue engineering hypothesis that construction of an osteochondral composite graft could be accomplished using multipotent progenitor cells and phenotype-specific biomaterials. Rat bone marrow-derived mesenchymal stem cells (MSCs) were culture-expanded and separately stimulated with transforming growth factor-beta1 (TGF-beta1) for chondrogenic differentiation or with an osteogenic supplement (OS). MSCs exposed to TGF-beta1 were loaded into a sponge composed of a hyaluronan derivative (HYAF-11) for the construction of the cartilage component of the composite graft, and MSCs exposed to OS were loaded into a porous calcium phosphate ceramic component for bone formation. Cell-loaded HYAFF-11 sponge and ceramic were joined together with fibrin sealant, Tisseel, to form a composite osteochondral graft, which was then implanted into a subcutaneous pocket in syngeneic rats. Specimens were harvested at 3 and 6 weeks after implantation, examined with histology for morphologic features, and stained immunohistochemically for type I, II, and X collagen. The two-component composite graft remained as an integrated unit after in vivo implantation and histologic processing. Fibrocartilage was observed in the sponge, and bone was detected in the ceramic component. Observations with polarized light indicated continuity of collagen fibers between the ceramic and HYAFF-11 components in the 6-week specimens. Type I collagen was identified in the neo-tissue in both sponge and ceramic, and type II collagen in the fibrocartilage, especially the pericellular matrix of cells in the sponge. These data suggest that the construction of a tissue-engineered composite osteochondral graft is possible with MSCs and different biomaterials and bioactive factors that support either chondrogenic or osteogenic differentiation.

High variability in rabbit bone marrow-derived mesenchymal cell preparations.

The rabbit has been extensively used for preclinical models, especially in orthopedic applications. One of the more troubling features of this model is the high interindividual variability that is encountered and that requires a careful experimental design with sufficient sample size to make judgments valid. We have processed 241 individual preparations of rabbit bone marrow-derived mesenchymal progenitor cells (MPCs) over the last 3 years and have kept detailed records of the performance of these cells in various assays. This communication details the lack of correlation between the analyzed parameters. Bone marrow was harvested from 4-month-old rabbits; the cells were centrifuged, resuspended, and cultured. When cells reached 80% of confluence, they were removed from the plates with trypsin and assayed for their osteo- and chondrogenic potential. The average yield of the 241 individual MPC preparations exhibited a coefficient of variation of 77. An in vivo implantation assay with porous calcium phosphate ceramic cubes exhibited scores with a coefficient of variation of 65. Lastly, an in vitro assay of alkaline phosphatase enzyme activity exhibited the most variability with a coefficient of variation of 132. All of the cell preparations tested in an in vitro aggregate culture assay underwent chondrogenic differentiation. No relationships between any of these parameters were found. The variability of the results within the different assays is interpreted to be the result of the heterogeneity of the preparations. The lack of correlation between the parameters studied shows the importance of the conditions intrinsic to the different assays. These results serve to emphasize that any experimental design involving rabbit progenitor cells must include a sufficiently large sample size to allow statistically significant and rigorous conclusions.

Demonstration of HLA and ABH antigens in fresh and frozen human menisci by immunohistochemistry.

The expression of HLA and ABH antigens in fresh, frozen, and twice-frozen normal human meniscal tissue was evaluated with use of immunohistochemistry. A generalized expression of Class-I and Class-II HLA antigens was found in the endothelial and synovial cells of all three forms of tissue. Fibrochondrocytes were Class-I positive and Class-II negative. ABH antigens were expressed by endothelial cells in relationship to the blood group of the patient. Freezing preserved most of the HLA and ABH molecular structure. Although the normal meniscus consists of a relatively few chondrocytes embedded in an extracellular matrix, it also contains Class-II and ABH-positive endothelial cells and Class-II-positive synovial cells. These antigens are present at the moment of transplantation and could evoke an immune response in the host that would modulate the results of meniscal allografting.

S-100 protein immunostaining identifies cells expressing a chondrocytic phenotype during articular cartilage repair.

The healing of articular surface defects has been studied with conventional histology, which relies on the staining of the extracellular matrix to identify the phenotype of the cells present. A chondrospecific cellular marker would be useful. S-100 protein has been found in all chondroid tissues studied, and we evaluated its usefulness in the study of articular cartilage repair. Full-thickness rabbit femoral condylar defects were made, and the specimens were studied at serial time intervals. S-100 protein staining positively showed chondroid cells in the 7- and 14-day specimens, which were not identifiable by conventional techniques. At 30 and 60 days, an S-100 positive band of cells separated a deep safranin-O positive hypertrophic layer from a fibrocellular surface layer. At 120 days, the presence of S-100 protein identified cells with chondrogenic potential, and the lack of S-100 protein in other cells embedded in conventionally stained matrix suggested that these cells were no longer of a chondroid phenotype. The presence of S-100 protein-identified chondroid cells early in the repair process when the cells had not begun to synthesize conventionally stainable matrix and the lack of S-100 protein in cells late in the repair positively identified a phenotypic change earlier than conventional histology.

Heterotopic osteogenesis in porous ceramics induced by marrow cells.

When untreated porous calcium phosphate ceramics were transplanted into subcutaneous (s.c.) or intramuscular (i.m.) sites, fibrovascular tissue grew in the pore region without evidence of bone formation. However, when these same ceramics were combined with syngeneic marrow cells, osteogenesis was observed inside the pore region of the implanted ceramic. The osteogenesis began on the surface of the pore region at approximately 3 weeks postimplantation by a process of intramembranous bone formation, with the de novo bone tissue observed directly interfacing with the ceramic surface. Infrequently, small isolated areas showed cartilage formation with no noticeable endochondral ossification. At 4 weeks postimplantation of the ceramic with marrow cells, the osteogenesis in the ceramic accompanied an observed increase in compressive strength, rigidity, and energy absorption of the ceramic. These results suggest that a combination of porous ceramics and marrow cells may be useful for clinical problems requiring osseous reconstruction.

Culture-expanded human periosteal-derived cells exhibit osteochondral potential in vivo.

Periosteal cells were enzymatically liberated from human rib periostea obtained from autopsies of 37 donors with an age distribution ranging from 25 weeks of gestation to 88 years old. These cells were introduced into cell culture and subcultured when they reached confluence. After subculture, the adherent periosteal-derived cells showed a nondescript, fibroblast-like morphology in cell culture. The cells from various passages of each donor were tested for in vivo osteochondrogenic potential with three different assay methods in athymic mice: (a) inoculation assay--the cells were directly inoculated into a subcutaneous site, (b) porous ceramics assay--the cells were combined with porous calcium phosphate ceramics, and this composite graft was implanted into a subcutaneous site, and (c) diffusion chamber assay--the cells were loaded into diffusion chambers and cultured in the peritoneal cavity. Frozen-preserved and recultured periosteal-derived cells were also assayed in the same way. In cases of donors younger than 19 years old, cultured, periosteal-derived cells from up to several passages consistently formed bone and/or cartilage in each of the three assays. Frozen-preserved and recultured cells from these donors also formed bone and/or cartilage after introduction into the three in vivo assays. In cases of donors older than 22 years of age, cultured, periosteal-derived cells formed neither bone nor cartilage in vivo. Cultured muscle fibroblasts from some of the same donors did not form bone or cartilage when assayed in vivo under identical conditions. These results suggest that periosteal cells with osteochondrogenic potentials can be liberated from the periosteum of a rib of human donors up to a certain age. Importantly, this potential is retained after enzymatic liberation, cell culture, subculturing, and freeze preservation. The present results suggest that culture-expanded human periosteal-derived cells from young donors may be useful in the repair of skeletal defects to foster cell-mediated regeneration of skeletal tissues, and that this methodology can be used to elucidate cellular, molecular, and genetic disorders in various metabolic bone diseases and skeletal dysplasias.

Histomorphometric analysis of the repair of a segmental diaphyseal defect with ceramic and titanium fibermetal implants: effects of bone marrow.

We used a rat femoral diaphyseal defect/implant model to quantify the ingrowth of bone, cartilage, and fibrous connective tissue in a comparative study of woven sintered titanium fibermetal and porous hydroxyapatite/tricalcium phosphate ceramic implanted with and without the addition of syngeneic bone marrow cells. The patterns of tissue growth into the implants were analyzed with respect to time, type of implant, and the presence or absence of syngeneic marrow. Significantly more bone was found in ceramic implants than in fibermetal implants, with the addition of syngeneic marrow than without it, and at 4 months than at 2 months. Significantly more bone was found at both time periods in ceramic implants with bone marrow than in any other combination studied. We hypothesize that these findings resulted from interactions between the implanted material and its surroundings, specifically its ability to serve as a substratum for cell attachment, and cells in and around the defect, whether surgically implanted or arising from the soft-tissue bed.