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Emergence of lyme arthritis after autologous chondrocyte transplantation
Stefan Marlovits *, Gelas Khanakah, Gabriele Striessnig, Vilmos Vécsei, Gerold Stanek Emergence of lyme arthritis after autologous chondrocyte transplantation University of Vienna, Vienna, Austria.
email: Stefan Marlovits (stefan.marlovits@akh-wien.ac.at)
*Correspondence to Stefan Marlovits, Department of Traumatology, University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria
Funded by:
Lorenz-Boehler-Society; Grant Number: 7/2001
Major of Vienna; Grant Number: 1774
Abstract
We report herein the first known incidence of the emergence of borrelial arthritis following autologous chondrocyte transplantation for repair of a cartilage defect. The patient had no recent manifestation of Lyme borreliosis, but 15 years earlier had had an expanding erythematous lesion after a tick bite. The current infection resulted in massive joint swelling, elevated body temperature, dissemination of the graft, and transplant failure. Results of routine bacteriologic studies were negative. A diagnosis of Lyme arthritis was first considered following the detection of Borrelia-specific serum antibodies. Additional evidence was provided when borrelial DNA sequences were detected in the synovial fluid through polymerase chain reaction. The diagnosis was confirmed by culture of Borrelia burgdorferi from the synovial fluid. The possibility of a dormant borrelial infection should be considered in patients who undergo repair of cartilage defects with autologous chondrocyte transplantation. We recommend that synovial fluid and joint tissue be screened for the presence of viable Borrelia before transplantation of an autologous graft.
Article Text
The implantation of autologous chondrocytes has stimulated new interest in the treatment of articular cartilage injuries, which are seen routinely in orthopedic clinical practice ([1]). Autologous chondrocytes can be harvested from a healthy part of the donor tissue, isolated, expanded in vitro, and finally implanted in high densities into the area of the defect. The implant is then covered with a periosteal membrane ([2]). Although the in vitro expansion of the cells and the surgery are performed under sterile conditions, the technique is subject to contamination hazards. It is not usually expected that the procedure itself would activate dormant infectious agents from the synovial cells or other joint tissues. Herein we present a case in which acute arthritis due to Borrelia burgdorferi infection developed after autologous chondrocyte transplantation.
Lyme borreliosis is the most frequently diagnosed tick-borne disease in the northern hemisphere and is highly endemic in several countries of central Europe ([3]). It is transmitted by ticks of the Ixodes ricinus complex. The basic characteristics of the disease are similar worldwide but there are regional variations, primarily between the illnesses found in America, Europe, and Asia ([3]). An asymptomatic infection may occur in 10% of persons bitten by ticks ([4]).
Lyme arthritis appears to occur more frequently in North America than in Europe and Asia, and monarticular or oligoarticular arthritis is the hallmark of untreated late Lyme borreliosis in North America ([3][5]). Attacks typically are intermittent and last from 3 days to 12 months. The knees are affected most often, but migratory arthritis is common and other large and small joints may be involved. Only very few Borrelia strains have been cultured from joint specimens worldwide ([6]). Laboratory diagnosis of Lyme arthritis requires detection of specific IgG antibodies, is supported by the demonstration of borrelial DNA in synovial fluid or tissue, and is confirmed by the isolation and cultivation of the agent ([7]).
CASE REPORT
The patient, a 38-year-old woman who was a frequent jogger, presented with a sports injury of her left knee. She had a ruptured anterior cruciate ligament and a traumatic cartilage lesion of the medial femoral condyle. The cartilage defect was verified radiologically with magnetic resonance imaging (MRI) using cartilage-specific sequences (dual T2-weighted turbo .
During arthroscopic repair of the anterior cruciate ligament with a patellar bone-tendon-bone autograft, a biopsy sample of hyaline cartilage (160 mg) was obtained from the intercondylar notch for cell processing and cultivation. The chondrocytes were enzymatically isolated and grown in monolayer culture. After reaching sufficient numbers, the cells were transferred onto a hyaluronic acid matrix (Hyalograft C; Fidia Advanced Polymers, Abano Terme, Italy) and cultivated in a 3-dimensional culture system ([8]). During the second surgery, the cartilage defect was debrided and cleaned. Chondrocyte transplantation was performed using a modification of the originally described technique ([2]), with a cell-carrier matrix and without a periosteal flap. The graft, a hyaluronan matrix sheet with chondrocytes, was trimmed to fit into the defect and was fixed with fibrin glue (Baxter International, Vienna, Austria). The cartilage defect was completely filled with the chondrocyte-matrix sheets .
The patient was discharged from the hospital 4 days after the surgery, with no signs of infection or other symptoms. Postoperative rehabilitation included continuous passive motion and no weight bearing for 8 weeks. The postoperative MRI 4 weeks after transplantation showed filling of the defect, indicated by different signal intensities of the graft compared with the surrounding hyaline cartilage . Five weeks after transplantation, the patient presented with diffuse swelling of the operated knee, moderate pain, and a body temperature of 37.2°C (mainly at night). No other joints were affected, and no other signs or symptoms were present.
For further analysis, peripheral blood was drawn and the effusion in the affected knee was aspirated. Specimens were submitted for routine bacteriology studies, but results were all negative. Results of all of the following standard blood tests were normal: red blood cell count, white blood cell count, electrolytes, total protein, bilirubin, creatinine, blood urea nitrogen, amylase, lipase, alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, gamma glutamyl transferase, lactate dehydrogenase, creatine kinase, blood fats, and blood coagulation. Only the C-reactive protein level was elevated (9.6 mg/ml; normal 1.0). Urinalysis and chest radiography revealed no abnormalities. Serum samples were tested for antibodies to B burgdorferi by enzyme immunoassay (EIA) and showed high concentrations of IgG antibody to Borrelia (6.2 EIA units; positive result 1) (unit calculation: optical density [OD] of sample/OD cutoff where cutoff = 0.250 OD). A reevaluation of the medical history revealed that the patient had had an expanding erythematous lesion after a tick bite 15 years before the transplantation; the rash was not confirmed by a physician nor was it treated. No additional history of any recent tick bites or manifestations of Lyme borreliosis was discovered.
Samples of cultured cells and joint fluid were divided into several fractions for direct microscopy, cytospins, histologic and immunohistologic staining, and polymerase chain reaction (PCR) analysis. Histologic evaluation of the unstained synovial specimens revealed mainly polymorphonuclear cells and some chondrocytes attached to loose hyaluronic acid fibers. For the PCR assay, a 2-step procedure was used. First, the presence of Borrelia was investigated by using part of the flagellin gene of Borrelia ([9]). The primer and the TaqMan probe for fluorescent online detection were designed to react with all Borrelia species. Second, for the detection of specific sequences of B burgdorferi subsp sensu stricto, Borrelia afzelii, and Borrelia garinii, sequences of the 5S and 23S intergenic spacer regions were used ([10]). Samples were typed using the ABI/Prism 7700 Sequence Detector (Applied BioSystems, Foster City, CA) with 3 different probes for the spacer regions and different kinds of dye. The reaction cycles were as follows: 2 minutes at 50°C and 15 minutes at 95°C, followed by 60 cycles of 15 seconds at 95°C and 1 minute at 60°C.
For the cultivation of B burgdorferi, 1-ml samples of each specimen (synovial fluid and cultivated chondrocytes) were placed in vials containing 7 ml BSK II medium (Sigma-Aldrich, St. Louis, MO) and incubated at 33°C. The presence of spirochetes was checked by darkfield microscopy every week until detected. Spirochete cultures were tested by PCR, and Borrelia strains were identified using the PCR systems described above.
PCR study of the synovial fluid specimen yielded positive results for Borrelia, and growth of spirochetes was observed after 21 days of incubation in BSK II medium. The isolate was identified as the genospecies B burgdorferi subsp sensu stricto by the PCR techniques described above. As a result of these findings, samples of the biomaterial used in the surgical intervention (hyaluronic acid membrane) were tested for the presence of Borrelia and were negative by both methods (culture and PCR).
After confirmation of the diagnosis of Lyme arthritis by serology, PCR, and culture, antibiotic treatment was instituted (amoxicillin 500 mg 3 times per day for 3 weeks). The arthritis improved during clinical followup. Serologic screening by EIA showed high concentrations of IgG antibody to Borrelia in the acute phase of the illness, at the initiation of therapy, and at 2 weeks, 4 weeks, 2 months, 6 months, and 12 months thereafter, with a sharp increase at the initiation of therapy and a decrease thereafter. The concentration of IgG antibodies did not subsequently change significantly during a period of 11 months . IgM antibody levels were not elevated.
Three months after transplantation of the graft, MRI of the knee showed that the graft had dissolved, leaving the cartilage defect completely unfilled, with exposure of the subchondral bone (Figure 1).
DISCUSSION
The intrinsic capacity of cartilage to repair chondral injuries is poor, and symptomatic full-thickness chondral lesions in the knee pose a difficult management issue. The first published study about autologous chondrocyte transplantation indicated good-to-excellent clinical results in 14 of 16 patients treated in the weight-bearing areas of the femoral condyles ([2]). Since then, there have been more than 5,000 such procedures worldwide. The long-term durability of the treatment in 1 series of 94 patients with 2-9 years of followup was reported as clinically good to excellent in 92% of cases in which the procedure was used to treat isolated femoral condyles and 89% of those in which it was used to treat osteochondritis dissecans ([11]). Further modification of the technique introduced 3-dimensional culture systems and the use of biologic matrices (scaffolds) as cell-carrier systems. Currently, 2 scaffolds are in clinical use: a collagen sponge and a hyaluronic acid matrix. The use of such 3-dimensional, matrix-based cell-carrier systems provides a template for the growth of the chondrocytes and contributes to their phenotypic stability. The scaffolds make it easier to deliver the cells to the injured site, and no periosteal membrane is necessary ([8]).
In this report, we provide information about the persistence for many years of B burgdorferi in the tissue of the knee joint, which caused borrelial arthritis with graft failure and dissemination of the transplanted chondrocytes. Although the patient was previously asymptomatic, the spirochete may have been present in the joint for years, and immune surveillance was perhaps altered by the surgical procedure. Innate immune responses to B burgdorferi lipoproteins are found during episodes of arthritis ([12]). In addition, there are marked adaptive immune responses to many spirochetal proteins, and humoral immune response is the host's most effective defense against B burgdorferi ([13]). Patients with Lyme arthritis usually have higher titers of Borrelia-specific antibody than do patients with any other manifestations of the illness ([12]). Our patient had a high concentration of Borrelia antibodies at the time of surgery. The antibody concentration, however, does not allow retrospective determination of the time of infection. The patient reported a history only of an expanding skin lesion 15 years before the surgery, which was not confirmed by a physician or treated with antibiotics. Antibiotic treatment in the intervening years or any recent tick bite was also not reported, which supported the hypothesis that B burgdorferi was present in the joint asymptomatically for 15 years.
In animal models of Lyme borreliosis, the development of humoral immunity correlates with a significant reduction in the number of organisms ([4]). However, the immune system is often unable to completely resolve the infection, and organisms remain in lower numbers at localized sites ([14]). In humans, these sites are the skin, the joints, and the nervous system, and the organism can often be recovered by culture or PCR years after infection ([4]). The local persistence of B burgdorferi in the joint over a long period of time might be related to the exacerbations of symptoms after chondrocyte cell transplantation. B burgdorferi is difficult to detect in synovial fluid, and cultures are positive only rarely ([6]). However, a high percentage of patients with Lyme arthritis (85%) have evidence of B burgdorferi DNA, detected by PCR, in the synovial fluid ([15]). In the patient described herein, Borrelia apparently persisted for years without causing symptoms. After the surgical intervention, the spirochetes were detectable by PCR; however, their presence was assessed not only by PCR, but also by culture. This is clearly the best confirmation of present infection.
In conclusion, we recommend that patients who undergo biologic repair of cartilage defects with autologous chondrocyte transplantation should be carefully interviewed about their outside activities in tick-infested areas. Laboratory tests should include screening for specific antibodies, and culture and PCR should be performed with specimens of synovial fluid, harvested cartilage, and adjacent tissue. Our findings indicate the need for controlled prospective clinical studies to assess the frequency of dormant Borrelia in synovial cells or adjacent tissue.
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