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.     

Glycosaminoglycan and collagen distribution in the developing human vitreous

BACKGROUND: We determined the distribution of glycosaminoglycans and collagens in the developing human vitreous. METHODS: Eighty human eyes from 5 gestational weeks to 2 postnatal years of age were used. Glycosaminoglycan components were determined by enzyme digestion with hyaluronidase or chondroitinase AC and ABC and immunohistochemistry for chondroitin, chondroitin-4-sulfate, chondroitin-6-sulfate, and dermatan sulfate. Collagen distribution was determined by immunohistochemistry for types I, II, and III collagens. RESULTS: Enzyme digestion showed that throughout development hyaluronic acid is the main glycosaminoglycan in the vitreous and in the extraocular space at 5-7 gestational weeks. Both areas were filled with mesenchymal cells. Immunohistochemistry showed chondroitin-6-sulfate in the vitreous between 6 and 40 gestational weeks, and chondroitin-4-sulfate between 12 and 40 gestational weeks. Hyaluronic acid and chondroitin sulfate appeared in the retina and around the hyaloid vessels at 12-40 weeks. Immunohistochemistry showed type III collagen in the vitreous and around the mesenchymal cells at 5-7 weeks that was replaced by type II collagen after 8 weeks. CONCLUSIONS: Hyaluronic acid is the major glycosaminoglycan in the vitreous throughout development, except for the transient appearance of chondroitin sulfate at 6-40 gestational weeks. Type III is the main collagen in the early developing vitreous that converts to type II collagen at 8 weeks. The primary and secondary vitreous has the same components as these macromolecules. These vitreous glycosamino-glycans and collagens seem to be produced by mesenchymal cells at an early stage and by the retina and hyaloid vessels during middle and late development.     

Comparison of surgically attached and non-attached repair of the rat Achilles tendon-bone interface. Cellular organization and type X collagen expression

The effects of surgical repair versus non-repair on cell morphology and type X collagen expression were investigated using a rat model of Achilles tendon avulsion. The animals were divided into four groups. In Group 1, tendon was reattached to the original attachment site by suturing through a drill hole in the calcaneus; in Group II, tendon was not reattached and a drill hole was not made; in Group III, tendon was not reattached but a drill hole was made; and the animals in Group IV were sham operated. In Group I (tendon reattached), at 2 weeks postoperatively, many hypertrophic chondrocytes appeared at the reattachment site adjacent to bone and type X collagen was detected immunologically both in the cells and in the extracellular matrix. After 4 weeks, the cells at the original site of attachment were arranged in rows along the newly formed tendon fibers and were stained with type X collagen antibody. By contrast, when tendon was not reattached (Groups II and III), a gap between the original attachment site and the tendon stump was observed through the entire postoperative period. At 8 weeks, the original attachment site was covered by fibrocartilaginous tissue and tendon became attached to the calcaneal fibrocartilage area, which is proximal to the original attachment site. Type X collagen was detected in the cells which were adjacent to bone. In Group IV (sham operation), there were no changes in histology or type X collagen distribution, either at the attachment site or in tendon and bone, compared with the non-operated control rats. These results suggest that surgical reattachment of tendon to the original site is important to help reorganize cells during the repair process. Type X collagen was identified immunohistochemically in the cells adjacent to bone in all the groups, suggesting that it may play a role in maintaining distinct areas of calcified and non-calcified fibrocartilage.     

Morphology and matrix composition during early tendon to bone healing

This study outlines the early morphologic phenomenon of tendon to bone healing in the rabbit model. Twelve skeletally mature, male New Zealand White rabbits received transplantation of the hallucis longus tendon into a 2-mm calcaneal bone tunnel. The morphologic characteristics of the healing tendon to bone interface were evaluated at 1, 2, 4, and 6 weeks after surgery by the use of conventional histology and immunohistochemical localization of collagen Types I, II, and III. Histologic analysis illustrated progressive maturation and reorganization of the tendon to bone interface with subsequent development of tissue collagen fiber continuity between the tendon and bone. Initially, diffuse immunolocalization of all three collagen types was observed within the scar tissue filling the space between the tendon and bone. During a 6-week period, reorganization of the scar tissue into an interface occurred, similar to an indirect insertion. Although a definitive fibrocartilage region did not form, Type II collagen was localized at the remodeling insertion site throughout the first 6 weeks of repair. In addition, Type III collagen fibers, resembling Sharpey's fibers, were noted to span this interface. The characterization of the insertion between tendon and bone is important to the understanding of healing in commonly used orthopaedic grafting procedures, such as anterior cruciate ligament reconstructions.     

Topography after repair of full-thickness corneal laceration

PURPOSE: To characterize corneal topography after repair of full-thickness corneal laceration. SETTING: Ophthalmic emergency room serving as a trauma referral center. METHODS: Twenty-two eyes with full-thickness corneal lacerations were prospectively studied after standardized surgical repair. Computerized videokeratography was done 2 and 14 weeks after surgery, with the latter measurement corresponding to 6 to 8 weeks after all sutures were removed. Fellow uninjured eyes served as the control group. RESULTS: Twenty eyes (91%) had a significant reduction in topographic distortion after suture removal. Mean corneal astigmatism, measured by simulated keratometry, was 10.70 diopters (D) +/- 5.90 D (SD) with sutures in place and 2.25 +/- 4.90 D after their removal (P < .005). Eighteen patients (82%) had 2.00 D or less of corneal astigmatism 6 to 8 weeks after all sutures were removed. The final distribution of topographic patterns was bow tie (50%), spherical/oval (36%), and irregular (14%). There was no significant correlation between laceration configuration (curvilinear, jagged, branched wound margins) and final topography. Lacerations that passed within 2.0 mm of the line of sight, however, were significantly more likely to have more than 2.00 D of final astigmatism. Mean central corneal power was 42.40 +/- 3.20 D in the injured eyes and 42.40 +/- 2.40 D in the uninjured fellow eyes. CONCLUSION: Although high astigmatism is frequently produced by corneal sutures used to repair full-thickness lacerations, the cornea has a substantial topographic memory that results in a marked normalization of contour after suture removal.     

Reparative dentin formation in rat molars after direct pulp capping with growth factors

Growth factors involved in normal wound healing may promote tissue repair when applied as a direct pulp capping medication. A minimal pulp exposure was made in rat molars, a pulp capping medication was placed, and the cavity was sealed. Epidermal growth factor, basic fibroblast growth factor, insulin-like growth factor II, platelet-derived growth factor-BB, and transforming growth factor-beta 1 (TGF-beta 1)--each absorbed onto a sterile collagen membrane (BioMend; Calcitek, Carlsbad, CA)--were used separately as pulpal medicaments. Dycal, unimpregnated collagen membrane, and no medication were used as controls. Eight samples from each treatment regimen were collected 2 and 3 weeks after surgery. Pulpal soft and hard tissue responses were graded. Data were analyzed by one-way ANOVA and Tukey-Kramer tests. No significant differences were detected after 2 wk. Pulp treated with TGF-beta 1 showed significantly improved soft and hard tissue healing at week 3, compared with the procedure control. We conclude that TGF-beta 1 as a pulp-capping medication enhances reparative dentin formation in rat molars.     

Determination of substance P in human nasal lavage fluid

Depending on the cell type studied, the involvement of type II transglutaminase (TGase) has been proposed in almost any event of the cell life such as differentiation, apoptosis, growth, aging, cell morphology and adhesion, metastatic capacity or extracellular matrix stabilization. In order to define the field(s) where this enzyme may be implicated in chondrocytes, type II TGase expression was studied in chondrocytes at different passages which differentiated state was modulated by retinoic acid, dihydrocytochalasin B or staurosporin. Results showed that (i) type II TGase expression is not incompatible with type II collagen expression, a main marker of chondrocyte differentiation (ii) type II TGase expression is higher when cells are in the exponential phase of growth than when growth arrested (iii) a high type II TGase expression does not imply that cells are apoptotic although cell apoptosis correlates with increased type II TGase expression (iv) non-adherent cells do not express type II TGase whereas adherent cells do whatever their differentiation state as assessed by type II collagen synthesis. These results suggest that, in articular chondrocytes, type II TGase is specifically implicated in the cell adhesion capacity.     

Immunohistochemical characterization of collagen in liver cirrhosis (author's transl)

Using indirect immunofluorescence technique, 21 cases of hepatic cirrhosis of differing etiology were studied with type-specific antibodies to collagen type I, II, and III. In all cases the fibrous septa and portal tracts showed an increase in type III collagen. No fluorescence could be observed with antibodies to collagen type I and II. Thus, biochemical studies are supported which show, in addition to type III collagen, a new, as yet undescribed type of collagen in liver cirrhosis that is similar to type I collagen electronmicroscopically, but differs from type I collagen biochemically and immunologically. No correlation between the etiology of cirrhosis and the pattern of different collagen types could be found. The origin of different collagen types in liver cirrhosis is briefly discussed.     

Effect of insulin-like growth factor-1 on zygomatic arch bone regeneration: a preliminary histological and histometric study

A number of physicians have attempted to pharmacologically manipulate the healing of bony fractures with a variety of agents such as growth hormone, thyroxine, chondroitin sulfate, and parathyroid hormone. Thus far, results from these experiments have been inconclusive. Previous research dealing with insulin-like growth factors has centered on cultures of osteoblast-like cells and has demonstrated a stimulatory effect on bone collagen synthesis, which may in fact play a critical role in the process of bone formation itself. The purpose of this investigation was to examine the effects of a genetically engineered growth factor, insulin growth factor type I, on midfacial fracture healing. In 24 adult male Sprague-Dawley rats, a standardized defect was created within the midportion of each zygomatic arch. One-half were treated with insulin growth factor type I administered with an osmotic infusion pump and the other half served as control subjects. At 2, 4, 8, and 12 weeks, animals from each group were killed and specimens of the defect obtained. Data were collected from radiographs and histological studies to compare the extent of bony repair. From this study, it appears that insulin growth factor type I could exert a potentiating effect on the repair of midfacial bone defects.     

The effect of recombinant human bone morphogenetic protein-2 (rhBMP-2) on the healing of full-thickness defects of articular cartilage

Articular cartilage has a limited capacity for repair. We investigated the effect of rhBMP-2 (recombinant human bone morphogenetic protein-2) on the healing of full-thickness osteochondral defects in adult New Zealand White rabbits. A single defect, three millimeters wide by three millimeters deep, was created in the trochlear groove of the right femur in eighty-nine rabbits. The defect was either left empty, filled with a plain collagen sponge, or filled with a collagen sponge impregnated with five micrograms of rhBMP-2. The animals were killed at four, eight, or twenty-four weeks, and the repair tissue was examined histologically and evaluated with use of a grading scale. The defects also were examined immunohistochemically for the presence of type-II collagen at four and eight weeks. The rate of bone repair was evaluated with fluorescent labeling of bone at two and four weeks and with use of fluorescence microscopy at eight weeks. Treatment with rhBMP-2 greatly accelerated the formation of new subchondral bone and improved the histological appearance of the overlying articular surface. At twenty-four weeks, the thickness of the repair cartilage was 70 per cent that of the normal adjacent cartilage and a new tidemark usually had formed between the repair cartilage and the underlying subchondral bone. The average total scores on the histological grading scale were significantly better (p < 0.01) for the defects treated with rhBMP-2 than for the untreated defects (those left empty or filled with a plain collagen sponge) at all time-points. Immunostaining with an antibody against type-II collagen showed the diffuse presence of this cartilage-specific collagen throughout the repair cartilage in the treated defects. The untreated defects demonstrated minimum staining with this antibody.     

The performance of subcutaneously injected Fertinex when used as the sole gonadotropin for in vitro fertilization stimulation

Adhesion formation constitutes a major problem following flexor tendon surgery in zone II. Expanded polytetrafluoroethylene (e-PTFE), a diffusible and biocompatible synthetic membrane, was investigated in rabbits with regard to the prevention of adhesion formation following tendon repair in zone II. An experimental model for biomechanical assessment of maximum tensile load to flex the distal interphalangeal joint 50 degrees from its resting position (MTL50) was used to estimate adhesion formation at intervals through 12 weeks. Tensile strength of the tendon repair was measured as well. On gross inspection, no apparent adverse tissue reactions or disruptions were observed. The e-PTFE group showed significantly lower MTL50 during the first 6 weeks after surgery, indicating less formation of restrictive adhesions compared with the control group. Tensile strength of tendon repair was similar in the 2 groups. Our results indicate that the e-PTFE membrane may be used as an adjunct to achieve more consistently reproducible results during the first 6 weeks following flexor tendon repair in zone II.     

Retinoic acid stimulates matrix calcification and initiates type I collagen synthesis in primary cultures of avian weight-bearing growth plate chondrocytes

The effect of retinoic acid (RA) on primary cultures of growth plate chondrocytes obtained from weight-bearing joints was examined, Chondrocytes were isolated from the tibial epiphysis of 6- to 8-week-old broiler-strain chickens and cultured in either serum-containing or serum-free media. RA was administered at low levels either transiently or continuously after the cells had become established in culture. Effects of RA on cellular protein levels, alkaline phosphatase (AP) activity, synthesis of proteoglycan (PG), matrix calcification, cellular morphology, synthesis of tissue-specific types of collagen, and level of matrix metalloproteinase (MMP) activity were explored. RA treatment generally increased AP activity and stimulated mineral deposition, especially if present continuously. RA also caused a shift in cell morphology from spherical/polygonal to spindle-like. This occurred in conjunction with a change in the type of collagen synthesized: type X and II collagens were decreased, while synthesis of type I collagen was increased. There was also a marked increase in the activity of MMP. Contrasting effects of continuous RA treatment on cellular protein levels were seen: they were enhanced in serum-containing media, but decreased in serum-free HL-1 media. Levels of RA as low as 10 nM significantly inhibited PG synthesis and caused depletion in the levels of PG in the medium and cell-matrix layer. Thus, in these appendicular chondrocytes, RA suppressed chondrocytic (PG, cartilage-specific collagens) and enhanced osteoblastic phenotype (cell morphology, type I collagen, alkaline phosphatase, and mineralization).     

Nitric oxide modulates basal and endothelin-induced coronary artery vascular smooth muscle cell proliferation and collagen levels

The mechanisms governing the pathological accumulation of collagen in the extracellular matrix following angioplasty are complex, but may involve interactions between endothelium-derived paracrine agents and vascular cellular components. We tested the hypothesis that nitric oxide (NO) directly decreases collagen levels and decreases endothelin (ET-1)-stimulated increases in levels of specific collagen subtypes in coronary vascular smooth muscle cells (VSMC). Cultured VSMC were incubated for 48 h with the NO donor CAS 754 (10(-4) M), ET-1 (10(-8) M), or ET-1 plus CAS 754. In some experiments, angiotensin II (Ang II; 10(-8) M) was utilized in place of ET-1. Soluble collagen types I and III were quantitated with an ELISA method, and cell counts were performed. CAS 754 significantly inhibited cell proliferation (-17+/-2% v control), basal total protein synthesis (-65+/-7% v control), and basal collagen type I levels (-39+/-6% v control), but not collagen type III levels. ET-1 and Ang II both significantly stimulated cell proliferation (26+/-5% v control), total protein synthesis (169+/-6% v control), and collagen type I levels (200+/-11% v control). Ang II, but not ET-1, significantly increased collagen type III levels. Co-incubations of ET-1 and CAS 754 resulted in a significant decrease in cell proliferation, protein synthesis, and collagen levels (-23+/-2% v control, 90+/-5% v control, and 63+/-3% v control, respectively) compared to ET-1 alone. In contrast, co-incubation of Ang II and CAS 754 had no significant effect on cell proliferation, protein synthesis, and collagen levels seen with Ang II alone. These results demonstrate that NO inhibits basal collagen levels and cell division. Additionally, NO alters ET-1 stimulation of VSMC proliferation, protein synthesis, and production of extracellular matrix components. Thus, an imbalance in key endothelium-derived compounds could significantly impact upon extracellular matrix deposition following mechanical revascularization.     

Effect of alignment of the transplanted graft extracellular matrix on cellular repopulation and newly synthesized collagen

This study examines the effect of alignment of the transplanted graft extracellular matrix on cellular repopulation and new collagen synthesis. The lateral half of the patellar tendon was harvested as a tendon graft from Lewis rats and frozen at -80 degrees C. In order to maintain the original alignment of the graft extracellular matrix, the graft was transplanted to a same size defect in the patellar tendon of other Lewis rats (group I). For controls, the graft was transplanted in a lax condition after excision of only the distal half of the lateral side of a patellar tendon (group II). After transplantation, six animals in each group were killed at 3, 7, 14, and 28 days. Cellular repopulation was assessed by using fibrillar-actin (F-actin) labeling with rhodamine-phalloidin, and new collagen synthesis was detected by means of a polyclonal antibody against type III collagen aminopropeptide (pN collagen III). Collagen fibril profiles were observed under the transmission electron microscope. On the 3rd day after transplantation, no specific fluorescence was detected in either group. Specific labeling for F-actin and pN collagen III, however, was observed at both ends of the graft in both groups at 1 week and throughout the graft at 2 weeks after transplantation. Consistent with the actin bundles' orientation, pN collagen III was aligned parallel to the longitudinal axis of the graft in group I. Collagen fibrils with a smaller diameter, mixed evenly and everywhere with a larger diameter, increased gradually in group I. However, smaller collagen fibrils in group II increased more slowly and were distributed unevenly. In summary, the dense collagen arrangement in the native patellar tendon determined the alignment of the repopulating cells, and the distribution of newly synthesized collagen might be affected by the actin cytoskeleton within the repopulating cells.     

Decreased type II/type I TGF-beta receptor ratio in cells derived from human atherosclerotic lesions. Conversion from an antiproliferative to profibrotic response to TGF-beta1

Atherosclerosis and postangioplasty restenosis may result from abnormal wound healing. The present studies report that normal human smooth muscle cells are growth inhibited by TGF-beta1, a potent wound healing agent, and show little induction of collagen synthesis to TGF-beta1, yet cells grown from human vascular lesions are growth stimulated by TGF-beta1 and markedly increase collagen synthesis. Both cell types increase plasminogen activator inhibitor-1 production, switch actin phenotypes in response to TGF-beta1, and produce similar levels of TGF-beta activity. Membrane cross-linking of 125I-TGF-beta1 indicates that normal human smooth muscle cells express type I, II, and III receptors. The type II receptor is strikingly decreased in lesion cells, with little change in the type I or III receptors. RT-PCR confirmed that the type II TGF-beta1 receptor mRNA is reduced in lesion cells. Transfection of the type II receptor into lesion cells restores the growth inhibitory response to TGF-beta1, implying that signaling remains responsive. Because TGF-beta1 is overexpressed in fibroproliferative vascular lesions, receptor-variant cells would be allowed to grow in a slow, but uncontrolled fashion, while overproducing extracellular matrix components. This TGF-beta1 receptor dysfunction may be relevant for atherosclerosis, restenosis and related fibroproliferative diseases.     

Human chondrocytes proliferate and produce matrix components in microcarrier suspension culture

Chondrocytes propagated in monolayer culture proliferate and change into 'fibroblastoid'-like cells. This change is characterized by a shift in production of collagen type II to I and from high- to low-molecular-weight proteoglycans. When propagated in three-dimensional culture, chondrocytes have limited ability to divide but re-express their original characteristics. The goal of the present study was to determine whether a microcarrier suspension culture system would support chondrocyte proliferation and phenotype expression. Our experiments indicate that a collagen type I microcarrier (cellagen) best supported chondrocyte proliferation and phenotype expression. Cells in cellagen microcarriers multiplied at least twentyfold within 2 weeks and had doubling times of 2 to 3 d. Viable and metabolically active cells were retrieved with ease. The harvested chondrocytes had no detectable staining for collagen type I and stained intensely for collagen type II. Our studies demonstrate that the microcarrier suspension culture system supports growth and enhances expression of the 'chondrocytic' phenotype. Attachment to a constrained surface and the fluid shear forces on the microcarriers during suspension culture may have helped chondrocytes to reacquire their rounded shape and produce cartilage matrix components.     

Interactions between anemone toxin II and veratridine on single neuronal sodium channels

Restenosis is the single most important factor limiting a favorable long-term outcome following mechanical revascularization. The vascular endothelium, through the release of key regulatory compounds, may regulate vascular structure by exerting fundamental control over collagen synthesis following injury to the vessel wall. We tested the hypothesis that endothelin (ET-1), an endothelium-derived peptide previously shown to be increased in pathological states, differentially stimulates porcine coronary vascular smooth muscle cell collagen types I and III synthesis. Monocultures of porcine coronary vascular smooth muscle were exposed to varying concentrations of endothelin over a 24-96-h time period. The medium was assayed for soluble collagen types I and III using a sensitive and specific ELISA method. Experiments were also done with the ET-1 antagonists PD 145065 and BQ123. Cell counts and viability were serially monitored. Experiments were also conducted with angiotensin II (A-II). A-II and ET-1 stimulated cell proliferation. ET-1 maximally stimulated collagen type I synthesis at 48 h at an optimal concentration of 10(-8) M, with no significant stimulation of collagen type III synthesis. The ETA specific antagonist BQ123 significantly inhibited the stimulatory effects of ET-1. A-II also stimulated collagen type I synthesis above basal levels, but was less efficacious than endothelin (95 +/- 5%, A-II, v 189 +/- 14% ET-1). In contrast to ET-1, A-II stimulated collagen type III synthesis (31 +/- 6% above basal, compared to -4 +/- 5% for ET-1). Results are also reported using smooth muscle cells from porcine aorta. The data demonstrate that ET-1 and A-II stimulate collagen synthesis by coronary artery vascular smooth muscle, and that they exert a differential effect over the two types of collagen that are present in the intima following balloon injury. Thus, the over expression of key regulatory compounds by endothelium following balloon injury could enhance collagen deposition and, consequently, play an integral role in intimal hyperplasia and restenosis.     

Chondrocyte-seeded collagen matrices implanted in a chondral defect in a canine model

The objective of our study was to evaluate reparative tissues formed in chondral defects in an adult canine model implanted with cultured autologous articular chondrocytes seeded in type I and II collagen GAG matrices. Two defects were produced in the trochlea grooves of the knees of 21 dogs, with cartilage removed down to the tidemark. This study includes the evaluation of 36 defects distributed among five treatment groups: Group A, type II collagen matrix seeded with autologous chondrocytes under a sutured type II collagen flap; Group B, type I collagen matrices seeded with chondrocytes under a sutured fascia flap; Group C, unseeded type I collagen matrix implanted under a sutured fascia flap; Group D, fascia lata flap alone; and Group E, untreated defects. All animals were killed 15 weeks after implantation. Six other defects were created at the time of death and evaluated immediately after production as 'acute defect controls'. In three additional defects, unseeded matrices were sutured to the defect and the knee closed and reopened after 30 min to determine if early displacement of the graft was occurring; these defects served as 'acute implant controls'. The areal percentages of four tissue types in the chondral zone of the original defect were determined histomorphometrically: fibrous tissue (FT); hyaline cartilage (HC); transitional tissue (TT, including fibrocartilage); and articular cartilage (AC). New tissue formed in the remodeling subchondral bone underlying certain defects was also assessed. Bonding of the repair tissue to the subchondral plate and adjacent cartilage, and degradation of the adjacent tissues were evaluated. There were no significant differences in the tissues filling the original defect area of the sites treated with chondrocyte-seeded type I and type II matrices. Most of the tissue in the area of the original defect in all of the groups was FT and TT. The areal percentage of HC plus AC was highest in group E, with little such tissue in the cell-seeded groups, and none in groups C and D. The greatest total amount of reparative tissue, however, was found in the cell-seeded type II matrix group. Moreover, examination of the reparative tissue formed in the subchondral region of defects treated with the chondrocyte-seeded collagen matrices (Groups A and B) demonstrated that the majority of the tissue was positive for type II collagen and stained with safranin O. These results indicate an influence of the exogenous chondrocytes on the process of chondrogenesis in this site. In all groups with implants (A-D), 30(50% of the FT and TT was bonded to the adjacent cartilage. Little of this tissue (6-22%) was attached to the subchondral plate, which was only about 50% intact. Remarkable suture damage was found in sections from each group in which sutures were used. Harvest sites showed no regeneration of normal articular cartilage, 18 weeks after the biopsy procedure. Future studies need to investigate other matrix characteristics, and the effects of cell density and incubation of the seeded sponges prior to implantation on the regenerative response.