Kevin R. Stone

Regeneration of Meniscal Cartilage with Use of a Collagen Scaffold. Analysis of Preliminary Data

A collagen scaffold was designed for use as a template for the regeneration of meniscal cartilage and was tested in ten patients in an initial, Food and Drug Administration-approved, clinical feasibility trial. The goal of the study was to evaluate the implantability and safety of the scaffold as well as its ability to support tissue ingrowth. The study was based on the findings of in vitro and in vivo investigations in dogs that had demonstrated cellular ingrowth and tissue regeneration through the scaffold. Nine patients remained in the study for at least thirty-six months, and one patient voluntarily withdrew after three months for personal reasons. The collagen scaffold was found to be implantable and to be safe over the three-year period. Histologically, it supported regeneration of tissue in meniscal defects of various sizes. No adverse immunological reactions were noted on sequential serological testing. On second-look arthroscopy, performed either three or six months after implantation, gross and histological evaluation revealed newly formed tissue replacing the implant as it was resorbed. At thirty-six months, the nine patients reported a decrease in the symptoms. According to a scale that assigned 1 point for strenuous activity and 5 points for an inability to perform sports activity, the average score was 1.5 points before the injury, 3.0 points after the injury and before the operation, and 2.4 points at six months postoperatively, 2.2 points at twelve months, 2.0 points at twenty-four months, and 1.9 points at thirty-six months. According to a scale that assigned 0 points for no pain and 3 points for severe pain, the average pain score was 2.2 points preoperatively and 0.6 point thirty-six months postoperatively. One patient, who had had a repair of a bucket-handle tear of the medial meniscus and augmentation with the collagen scaffold, had retearing of the cartilage nineteen months after implantation. Another patient had débridement because of an irregular area of regeneration at the scaffold-meniscus interface twenty-one months after implantation. Magnetic resonance imaging scans demonstrated progressive maturation of the signal within the regenerated meniscus at three, six, twelve, and thirty-six months. These findings suggest that regeneration of meniscal cartilage through a collagen scaffold is possible. Additional studies are needed to determine long-term efficacy.

Porcine and bovine cartilage transplants in cynomolgus monkey: II. Changes in anti-Gal response during chronic rejection.

The recent advances in avoiding hyperacute rejection by producing transgenic pigs with complement regulatory proteins call for the analysis of posttransplantation changes in anti-Gal activity in the absence of hyperacute rejection. Transplantation of cynomolgus monkeys with porcine or bovine meniscus and articular cartilage enabled the study of anti-Gal IgG response to xenografts that are not subjected to hyperacute rejection. The cartilage implants were kept in suprapatellar pouches of the recipients for 1 or 2 months and anti-Gal activity was measured in the serum at various time intervals after transplantation. Within 2 weeks after transplantation, titer of anti-Gal IgG, in all transplanted monkeys, increased by 20- to 100-fold, as measured in ELISA with synthetic alpha-galactosyl epitopes linked to bovine serum albumin or with mouse laminin. Furthermore, binding of serum anti-Gal to porcine endothelial cells increased by 10-fold or more after transplantation. Complement-mediated cytotoxicity also increased by two- to eightfold after transplantation. The elevated activity of anti-Gal was maintained for the 2-month period during which the grafts were kept in the monkeys, and returned to the pretransplantation level 6 months after graft removal. All these data suggest that the primate immune system responds vigorously to alpha-galactosyl epitopes on xenografts by activating many B lymphocytes that produce increased amounts of anti-Gal IgG, which may also be of high affinity. These antibodies are likely to bind to the xenograft cells, even if these cells express low numbers of alpha-galactosyl epitopes. Such antibody binding may play an important role in chronic rejection of xenografts.

Porcine cartilage transplants in the cynomolgus monkey. III. Transplantation of alpha-galactosidase-treated porcine cartilage.

BACKGROUND Studies on transplantation of porcine meniscus and articular cartilage into monkeys are important for evaluating the possible use of such tissues in humans. In addition, such studies shed light on the chronic xenograft rejection process in primates. Transplantation of porcine cartilage into cynomolgus monkeys for 2 months results in a many-fold increase in anti-Gal activity and in a strong cellular inflammatory response of T lymphocytes and macrophages within the implants. The objective of this study was to determine whether elimination of Galalpha1-3Galbeta1-4GlcNAc-R (alpha-gal epitopes) from the xenograft may alter the immune response and the inflammatory reaction. METHODS Porcine meniscus and articular cartilage specimens were treated with recombinant alpha-galactosidase (100 U/ml), and the absence of alpha-gal epitopes was assessed by the binding of the monoclonal anti-Gal antibody M86. The treated cartilage specimens were transplanted into the suprapatellar pouch of cynomolgus monkeys. The immune response to cartilage was monitored in the serum and the inflammatory reaction was assessed in the xenografts, which were explanted after 2 months. RESULTS Incubation with alpha-galactosidase resulted in complete removal of alpha-gal epitopes from the cartilage. The increase in anti-Gal activity in the transplanted monkeys was marginal. However, most monkeys produced antibodies to antigens specific to porcine cartilage. The inflammatory response within the alpha-galactosidase-treated xenografts was much lower than in nontreated cartilage and the proportion of T lymphocytes within the cellular infiltrates was greatly reduced. CONCLUSIONS Treatment of cartilage xenografts with alpha-galactosidase successfully removes alpha-gal epitopes from porcine cartilage. Transplantation of the treated cartilage results in the production of only anti-porcine cartilage-specific antibodies and a reduced inflammatory response consisting primarily of macrophages infiltrating into the cartilage.

Replacement of human anterior cruciate ligaments with pig ligaments: a model for anti-non-gal antibody response in long-term xenotransplantation.

Background. Understanding anti-non-gal antibody response is of significance for success in xenotransplantation. Long-term anti-non-gal response in humans was studied in patients transplanted with porcine patellar tendon (PT) lacking &agr;-gal epitopes, for replacing ruptured anterior cruciate ligament (ACL). Methods. Porcine PTs were treated with recombinant &agr;-galactosidase to eliminate &agr;-gal epitopes and with glutaraldehyde for moderate cross-linking of collagen fibers. The processed pig PTs were implanted to replace ruptured ACL in patients. Results. In five of six evaluable subjects, the xenografts have continued to function for over two years and passed all functional stability assessments. Thus, processed porcine PT seems to be appropriate for replacing ruptured human ACL. Enzyme-linked immunosorbent assay and Western blot studies indicated that all subjects produced anti-non-gal antibodies against multiple pig xenoproteins, but not against human ligament proteins. Production of anti-non-gal antibodies peaked two to six months posttransplantation and disappeared after two years. Conclusions. These antibodies contribute to a low-level inflammatory process that aids in gradual xenograft replacement by infiltrating host fibroblasts that align with the pig collagen “scaffold” and secrete collagen matrix. The assays monitoring anti-non-gal antibodies will help to determine whether long-term survival of live organ xenografts requires complete suppression of this antibody response.

Long-term survival of concurrent meniscus allograft transplantation and repair of the articular cartilage: A PROSPECTIVE TWO- TO 12-YEAR FOLLOW-UP REPORT

We describe 119 meniscal allograft transplantations performed concurrently with articular cartilage repair in 115 patients with severe articular cartilage damage. In all, 53 (46.1%) of the patients were over the age of 50 at the time of surgery. The mean follow-up was for 5.8 years (2 months to 12.3 years), with 25 procedures (20.1%) failing at a mean of 4.6 years (2 months to 10.4 years). Of these, 18 progressed to knee replacement at a mean of 5.1 years (1.3 to 10.4). The Kaplan-Meier estimated mean survival time for the whole series was 9.9 years (sd 0.4). Coxs proportional hazards model was used to assess the effect of covariates on survival, with age at the time of surgery (p = 0.026) and number of previous operations (p = 0.006) found to be significant. The survival of the transplant was not affected by gender, the severity of cartilage damage, axial alignment, the degree of narrowing of the joint space or medial versus lateral allograft transplantation. Patients experienced significant improvements at all periods of follow-up in subjective outcome measures of pain, activity and function (all p-values < 0.05), with the exception of the seven-year Tegner index score (p = 0.076).

The Frequency of Baker's Cysts Associated with Meniscal Tears

We reviewed the films of 1760 patients who had mag netic resonance image scanning of the knee joint to assess the most common pathologic changes associ ated with an incidental finding of a Bakers cyst. Of the 1760 knees scanned, Bakers cysts were noted in 238. The cysts were classified as small (55%), medium (30%), or large (15%) and were primarily found on the medial side of the knee (94%). The menisci were eval uated and changes were classified as complete tears, where signal contacts the surface, or degenerative intrasubstance tears. One hundred eleven (47%) com plete meniscal tears were found, and 88 (37%) degen erative tears. The majority of tears were found in the posterior horn of the medial meniscus (65 complete tears and 45 degenerative tears). Thus, 199 tears were found in 170 knees, and 106 of the 170 knees (62%) had tears of the posterior horn of the medial meniscus. Bakers cysts are frequent findings on physical exam inations and on magnetic resonance imaging scans of the knee. They are thought to be due to intraarticular pathologic changes, usually posterior meniscal tears. This study documents the association between Bakers cysts and meniscal tears and notes that a complete tear is not necessary for the cyst to be present.

3D MRI volume sizing of knee meniscus cartilage

Meniscal replacement by allograft and meniscal regeneration through collagen meniscal scaffolds have been recently reported. To evaluate the effectiveness of a replaced or regrown meniscal cartilage, a method for measuring the size and function of the regenerated tissue in vivo is required. To solve this problem, we developed and evaluated a magnetic resonance imaging (MRI) technique to measure the volume of meniscal tissues. Twenty-one intact fresh cadaver knees were evaluated and scanned with MRI for meniscal volume sizing. The sizing sequence was repeated six times for each of 21 lateral and 12 medial menisci. The menisci were then excised and measured by water volume displacement. Each volume displacement measurement was repeated six times. The MRI technique employed to measure the volume of the menisci was shown to correspond to that of the standard measure of volume and was just as precise. However, the MRI technique consistently underestimated the actual volume. The average of the coefficient of variation for lateral volumes was 0.04 and 0.05 for the water and the MRI measurements, respectively. For medial measurements it was 0.04 and 0.06. The correlation for the lateral menisci was r = 0.45 (p = 0.04) and for the medial menisci it was r = 0.57 (p = 0.05). We conclude that 3D MRI is precise and repeatable but not accurate when used to measure meniscal volume in vivo and therefore may only be useful for evaluating changes in meniscal allografts and meniscal regeneration templates over time.

Surgical technique of meniscal replacement

Meniscal replacement by allograft and prosthesis is under laboratory and investigational clinical practice. In order to succeed, a replacement must duplicate the exact mechanical function of the original meniscal cartilage. The technique of replacement described in this article permits minimal disruption of the joint tissues, accurate placement of the meniscal horns, and secure fixation of the meniscal synovial junction.

Autogenous replacement of the meniscus cartilage: Analysis of results and mechanisms of failure

Autogenous replantation of meniscal cartilage (resection of 80% of the meniscus cartilage followed by immediate replantation) was performed in 14 dogs as a control arm of a meniscal replacement study. The purpose was to assess the ability of the excised tissue to heal to the intact rim and function as a meniscus cartilage. This procedure is an idealized model of allografting meniscus cartilage in that the tissue is fresh, autogenous, and perfectly sized. If this procedure did not succeed, it seemed likely to the authors that allografting meniscal cartilage would have diminished chances for success. Evaluation of these replant failures led us to speculate that the causes and mechanisms might include slow or incomplete revascularization, inadequate mechanical fixation or stabilization, and, perhaps, some type of rejection phenomenon not examined or confirmed in the present study. We believe these mechanisms will be particularly deleterious for allografted meniscal cartilages and recommend further extensive evaluation of meniscal allografts before wide clinical use.

Surgical technique for articular cartilage transplantation to full-thickness cartilage defects in the knee joint

Abstract Focal arthritic defects in the knee lead to pain, swelling, and dysfunction. Treatment of the defects has includeddrilling, abrasion, and grafting. This report describes our surgical technique of autogenous articular cartilage grafting of arthritic and traumatic articular cartilage lesions. Articular cartilage grafting can be performed as a single arthroscopic outpatient procedure. The mixture of articular cartilage and cancellous bone appears to provide a supportive matrix for cartilage formation. Pain relief is excellent if careful surgical technique and a defined rehabilitation program is followed. Further collagen typing data and additional biopsies will reveal more about the durability of the newly formed cartilage.