Electromechanical delay of the knee extensor muscles is not altered after harvesting the patellar tendon as a graft for ACL reconstruction: implications for sports performance.

Although the scar tissue, which heals the donor site defect, has different elasticity from the neighbouring patellar tissue, it remains unclear if this scar tissue can lead to the changes of the electromechanical delay (EMD) of the knee extensor muscles. If such changes do exist, they can possibly affect both the utilization of the stored energy in the series elastic component, as well as the optimal performance of the knee joint movement. The purpose of this study was to investigate the influence of harvesting the patellar tendon during anterior cruciate ligament (ACL) reconstruction and the associated patellar tendon scar tissue development on the EMD of the rectus femoris (RF) and vastus medialis (VM) muscles. Seventeen patients who underwent an ACL reconstruction using the medial third of the patellar tendon were divided in two groups based upon their post-operative time interval. Maximal voluntary contraction from the knee extensors, surface EMG activity, and ultrasonographic measurements of the patellar tendon cross-section area were obtained from both knees. Our results revealed that no significant changes for the maximal voluntary contraction of the knee extensors and for the EMD of the RF and the VM muscles due to patellar scar tissue development after harvesting the tendon for ACL reconstruction. The EMD, as a component of the stretch reflex, is important for the utilization of the stored energy in the series elastic component and thus, optimal sports performance. However, from our results, it can be implied that the ACL reconstruction using a patellar tendon graft would not impair sports performance as far as EMD is concerned.

The presence of proprioceptive mechanoreceptors in the remnants of the ruptured ACL as a possible source of re-innervation of the ACL autograft.

This study investigated the presence of neural mechanoreceptors in the remnants of the ruptured ACL as a possible source of reinnervation of the ACL autologous graft. The remainder of the torn ACL was selected for further histological investigation from 17 patients during ACL reconstruction 3 months to 3.5 years after injury. Perioperatively two types of ACL remnant were identified. Fifteen patients had portions of ACL adapted at the PCL. In all of these patients we found mechanoreceptors (I and II). In five patients we found mushroomlike remnants which included either none or small numbers of mechanoreceptors. Free neural ends were found in both patient groups. There was a significant difference between the groups in regard to the mean number of mechanoreceptors I and II per slice. In conclusion, in patients with an ACL remnant adapted to the PCL, mechanoreceptors exist even 3 years after injury. If we accept that restoration of proprioception is the result of reinnervation of the ACL, leaving the ACL remnants as a source, if this is surgically possible without risk of Cyclop's lesion, may be of potential benefit to the patient.

Alterations of the extensor apparatus after anterior cruciate ligament reconstruction using the medial third of the patellar tendon.

PURPOSE: The objective of this study was the ultrasound evaluation of the donor defect of the patellar tendon (PT) and the radiologic evaluation of the patella after harvesting of the medial third as a bone-patella tendon-bone (BPTB) graft for anterior cruciate ligament (ACL) reconstruction. TYPE OF STUDY: This was a cohort study. METHODS: In 45 patients who had ACL reconstruction, the extensor apparatus of the donor side was studied using ultrasound cross-sections and radiographs (anteroposterior, lateral, and a tangential view of the patella) 3 to 70 months postoperatively. Patients were divided into two groups. The early postoperative group (3 to 30 months postoperative) consisted of 27 patients (group A) and the late postoperative group (31 to 70 months postoperative) consisted of 18 patients (group B). The healthy contralateral extensor apparatus was used as control. RESULTS: In group A, the standard ultrasound cross-section area of the PT increased by 20.48%, whereas in group B, it decreased by 4.88%. In group A, the patellar height was decreased by 9.21% in the donor side compared with the control. In group B, the patellar height was decreased by 7.02%. In group A, the Merchant's congruence angle increased by 11.59 degrees, and for group B, this angle increased by 3.82 degrees. This finding indicated that, after the 30th postoperative month, lateral displacement of the patella was not statistically significant (P =.38). In addition, no significant differences were found in the lateral patellofemoral angle in either group. CONCLUSIONS: Our study indicates that the tendon defect is always healed and the final tendon cross-section area is 95% of the contralateral after the 30th postoperative month. In addition, there was a nonsignificant slight lateral displacement of the patella. In contrast, other studies found shown that there is a slight medial displacement of the PT after using the central third as a BPTB graft.

Patellar fractures associated with medial-third bone-patellar tendon-bone autograft ACL reconstruction.

Patella fractures following anterior cruciate ligament (ACL) reconstruction are a recognized but rarely reported complication. To our knowledge, 24 reports of patella fractures after ACL reconstruction using the central-third patella-tendon autograft have been reported in the literature. Patellar fractures associated with the use of the medial-third bone-patellar tendon-bone autograft have not been reported. This article describes four cases of patellar fractures in 478 ACL reconstructions between 1992 and 1999, using the medial third of the patellar tendon graft. All of them were transverse fractures of the patella but only one was displaced. All patients suffered local injury to the donor knee between 2 and 4 months postoperatively. No significant differences in the final outcome were noticed between the cases complicated with patellar fracture and those with uncomplicated ACL reconstructions.

Simultaneous ligation of CD5 and CD28 with monoclonal antibodies restores impaired immunostimulatory function in human renal cell carcinoma.

Tumor cells, including renal cell carcinoma (RCC) cells, do not effectively stimulate T lymphocyte responses against specific antigens presented on their surface. Reasons for this low immunogenicity may include low or absent expression of MHC class I and/or class II molecules, as well as accessory and costimulatory molecules. We used tumor cell pretreatment with cytokines, together with monoclonal antibodies (mAbs) directed at receptors for costimulatory molecules, to render RCC cells immunostimulatory. Interferon-gamma or tumor necrosis factor-alpha pretreatment enhanced expression of MHC class I and class II molecules, as well as CD54, but had only minimal effects on T cell activation. A CD28 mAb, or an even more effective combination of CD28 and CD5 mAb, induced strong primary proliferative responses of allogeneic resting T lymphocytes. Cytokine pretreatment further augmented this T cell response in vitro and allowed T cell expansion and establishment of T cell lines. Stimulation of T cells with autologous RCC cells resulted in a similar T cell activation but with the expansion of cytolytic T cells directed at autologous MHC class II molecules. These experiments demonstrate that cytokines combined with costimulatory mAbs are useful for increasing the immunogenicity of tumor cells. They also indicate. however, that autologous MHC class II expression on tumor cells, together with strong costimulation, may lead to the activation of autoreactive T cells.

Posterior cruciate ligament architecture: evaluation under microsurgical dissection.

PURPOSE: Our objective was to verify the fiber anatomy of the posterior cruciate ligament (PCL) and to measure the main dimensions and the femoral and tibial attachment site distances of the ligament after microsurgical dissection. We hypothesized that PCL anatomy is more complex than the 2 traditionally characterized bands. TYPE OF STUDY: This is a purely anatomic description of microdissections of the PCL, focused on the fine anatomy of the ligament. MATERIALS AND METHODS: Twenty-four fresh-frozen cadaveric knees were dissected using magnifying loupes and an operative microscope, being careful to avoid creating artificially separated bundles. The main dimensions of the PCL were measured using a micrometer. RESULTS: The anterior, central, posterior-longitudinal, and posterior-oblique were the 4 fiber regions identified based on their orientation and the osseous sites of their insertions. These were partially separable anatomically but were functionally distinct. The anterior and central fiber regions made up the bulk of the ligament, while the remaining 15% consisted of the posterior fiber regions. During manual joint motion, the behavior of these fiber regions was observed. The anterior fiber region appeared to be the most nonisometric and remained in tension mainly between 30 degrees and 90 degrees of flexion. The posterior fiber regions appeared to be the most isometric (especially the posterior-oblique) and remained in tension mainly in extension and partially in deep flexion. The central fiber region appeared to have an intermediate behavior and remained in tension mainly between 30 degrees and 120 degrees of flexion. Additionally, it appeared to be the widest of all fiber regions. CONCLUSIONS: These findings should be of interest and help in interpreting some of the anatomy encountered during arthroscopic examination of the PCL, both from the anterior and posterior lateral portals. Furthermore, this information should prove useful in selecting treatment for the PCL.

Interleukin-12 requires initial CD80-mediated T-cell activation to support immune responses toward human breast and ovarian carcinoma.

One possible reason for the poor immunogenicity of tumors is the induction of peripheral tolerance by tumor cells that fail to deliver costimulatory signals. Furthermore, T cells stimulated with wild-type tumor cells often fail to secrete cytokines. The present study has been undertaken to identify cytokines that cooperate with CD80 in T-cell activation in vitro toward human breast and ovarian carcinoma cell lines. Tumor cell-mediated T-lymphocyte activation was analyzed directly in allogeneic mixed lymphocyte/tumor cell cultures as proliferation and effector functions were assessed in cytotoxic T-cell assays. Interleukin-7 (IL-7) amplified the proliferative response toward CD80-transfected breast and ovarian carcinomas and stimulated predominantly CD4+ T lymphocytes. IL-12 represses the proliferative response of naive T cells but cooperates with CD80-mediated activation during secondary stimulations. In long-term T-cell cultures, IL-12 synergizes with CD80 expression to stimulate cytolytic CD8+ T-cell lines, which recognize a breast carcinoma line in a human histocompatibility leukocyte antigen-restricted manner. These studies illustrate that costimulation is necessary for tumor cells to function as alloantigen-presenting cells. Furthermore, when added after the priming of T cells with CD80-transfected tumor cells, IL-12 could be helpful in propagating sufficient T-cell numbers to be used in adoptive transfers during cellular immunotherapy.

Potential of CD80-transfected human breast carcinoma cells to induce peptide-specific T lymphocytes in an allogeneic human histocompatibility leukocyte antigens (HLA)-A2.1+-matched situation.

Allogeneic human histocompatibility leukocyte antigen (HLA)-matched tumor cell lines that have been made immunogenic by the transfer of genes encoding for costimulatory molecules such as CD80 are considered to be potential vaccines for the induction of systemic immune reactions in cancer patients. We used a human HLA-A2.1+ CD80-transfected breast carcinoma cell line (KS-CD80) and investigated in vitro the efficiency at which antigen (Ag)-specific responses were induced following the stimulation of allogeneic HLA-A2.1-matched T lymphocytes. The influenza matrix protein M1 was used as a model Ag. It was either endogenously expressed or exogenously loaded as a peptide (matrix protein), and the frequency of the generated specific T cells was determined. The expression of CD80 in KS cells was required for an effective activation and expansion of Ag-specific T cells. This response was augmented following the pretreatment of KS-CD80 cells with interferon-gamma and tumor necrosis factor-alpha. Interleukin-4 (IL-4), IL-7, and IL-12 further increased T-cell expansion. IL-7 was best at supporting the generation of T cells with Ag specificity. This investigation demonstrates that allogeneic CD80+ tumor cells can induce Ag-specific, HLA-restricted T lymphocytes at a high frequency. Our study supports the use of allogeneic cell lines for the induction of specific T-cell responses in tumor patients.

T cell activation by monoclonal antibodies bound to tumor cells by a cell surface displayed single-chain antibody.

Tumor cells often lack the costimulatory molecules necessary for T cell activation. However, the transformation of cells with more than one stimulatory molecule is a difficult procedure. We therefore developed a retroviral vector for the expression of a cell membrane anchored single-chain antibody fragment (scFv) directed against the hapten 4-ethoxymethylene-2-phenyl-2-oxazoline-5-one (phOx). Proteins and peptides can be readily modified with this hapten, thus, enabling them to be bound to cells with the cell surface displayed anti-phOx scFv. To test combinations of surface-bound stimulatory molecules on T cell activation, SK-Mel63 human melanoma cells expressing the membrane anchored anti-phOx scFv were incubated with phOx-labeled mAbs against CD3, CD28 and CD5. Cells presenting a given mixture of modified anti-CD3 and anti-CD28 molecules stimulated T cell activation better than any single antibody and a given mixture of anti-CD3, anti-CD28 and anti-CD5 provided a stimulatory response higher than the best double combination. However, the relative concentrations are very important and must be carefully chosen. Concentrations of antibodies giving good T cell responses when used alone can block synergistic effects.

Nerve and vessel injuries during high tibial osteotomy combined with distal fibular osteotomy: a clinically relevant anatomic study.

Based on our clinical experience and an anatomical study, we examined the conditions under which injury to the popliteal artery, tibial nerve or peroneal nerve and its branches may occur during high tibial osteotomy. In 250 high tibial osteotomies performed in our department, we observed the following intraoperative complications. (1) The popliteal artery was severed in 1 patient and repaired by the same surgical team using a microsurgical technique. (2) A tibial nerve paresis also occurred in 1 patient. (3) In 3 patients, temporary palsy of the anterior tibialis muscle was documented. (4) In 4 other patients, palsy of the extensor hallucis longus occurred. To investigate the causes of these complications in the popliteal artery, tibial nerve and branches of the peroneal nerve, we dissected the neurovascular structures surrounding the area of the osteotomy in 10 cadaveric knees and performed a high tibial osteotomy in another 13 cadaveric knees. We concluded the following. (1) The popliteal artery and tibial nerve are protected, at the level of the osteotomy, behind the popliteus and tibialis posterior muscles. Damage can occur only by placing the Hohman retractor behind the muscles. The insertion of the muscles is very close to the periosteum and can be separated only with a scalpel. (2) The tibialis anterior muscle is innervated by a group of branches arising from the deep branch of the peroneal nerve. In two-thirds of the dissected knees, we found a main branch close to the periosteum, which can be damaged by dividing the muscle improperly or due to improper placement and pressure of the Hohman retractor. This may explain the partially reversible muscle palsy. (3) The extensor hallucis longus is also innervated by 2-3 thin branches, arising from the deep branch of the peroneal nerve, but in 25% of the specimens, only one large branch was found. This branch is placed under tension by manipulating the distal tibia forward. Thus, it may be damaged by the Hohman retractor during distal screw fixation, tensioned by hyperextension or directly injured during midshaft fibular osteotomy.

Difficulties and early results of the endoscopic carpal tunnel release using the modified Chow technique.

In reporting on the preliminary results of our series of 76 patients, this paper aims to identify potentially complicating aspects of endoscopic carpal tunnel release (ECTR) using the two-portal Chow technique, and to recommended solutions, based on our early experience, which enhance the ease and safety of this minimally invasive technique. Of the first 24 patients, 16 cases required conversion to an open procedure. Based on these initial cases, we developed certain modifications of the Chow technique which precluded any need for open conversion in the 60 remaining cases. During a follow-up interval ranging from 4 to 24 months, there was no recurrence of carpal tunnel symptoms, and the average time to resumption of work activity was 14 days. The complication rate was 5% and included one case of transient hypesthesia, one case of extended hematoma, and one hypersensitive scar. All complications resolved at subsequent follow-up. In our experience, correct positioning of the hand, proper injection of local anesthetic, use of magnifying loupes, and correct use of instruments are essential for a safe and successful procedure.

Gene transfer of costimulatory molecules into a human colorectal cancer cell line: requirement of CD54, CD80 and class II MHC expression for enhanced immunogenicity.

Colorectal cancer is considered a non-immunogenic malignany. One strategy to augment the immunogenicity of such tumours is represented by the expression of costimulatory molecules by gene transfer. Using transfected variants of the human colorectal cancer cell line SW480 we tested various costimulatory molecules (CD80, CD86, CD54) and a class II major histocompatibility complex (MHC) allele (HLA-DR3) alone or in combination on their ability to support primary T-lymphocyte activation in vitro. Expression of CD80 or CD86 similarly as the combination of both was not sufficient to induce proliferation of human allogeneic T cells. Expression of CD54 together with CD80 strongly augmented the costimulatory function of CD80, as observed in the presence of a CD3 monoclonal antibody (mAb), but did not lead directly to a T-cell response against modified tumour cells. Importantly, SW480 cells coexpressing CD54, CD80 and the HLA-DR3 allele effectively promoted T-lymphocyte proliferation. Moreover, the use of such CD54+/CD80+/HLA-DR3+ SW480 variants for repetitive stimulations resulted in the generation of T-cell lines predominantly composed of CD8+ T cells exhibiting class I MHC restricted cytolytic activity towards untransfected SW480 tumour cells. This demonstrates that the generation of immunogenic tumour cell variants, i.e. for the use as cellular vaccines, requires multiple genetic alterations in the case of non-immunogenic human tumours cells, such as colorectal cancer cells.

Strong immunogenic potential of a B7 retroviral expression vector: generation of HLA-B7-restricted CTL response against selectable marker genes.

The stimulation of a specific immune response is an attractive goal in cancer therapy. Gene transfer of co-stimulatory molecules and/or cytokine genes into tumor cells and the injection of these genetically modified cells leads to tumor rejection by syngeneic hosts and the induction of tumor immunity. However, the development of host immune response could be either due to the introduced immunomodulatory genes or due to vector components. In this study, human renal cell carcinoma cell lines were modified by a retrovirus to express the co-stimulatory molecule B7-1 together with the hygromycin/thymidine kinase fusion protein (HygTk) as positive and negative selection markers. These B7-1-transduced renal cell carcinoma cell lines were able significantly to activate allogeneic T cell proliferation. The cytolytic activity of these T cells was determined by employing several transduced and nontransduced renal cell carcinoma cell lines as targets. Evidence for a strong vector-specific T cell reactivity induced by the Hyg/Tk protein was obtained in autologous renal cell carcinoma systems. Antibody blocking experiments as well as peptide binding assays demonstrated an HLA-B7-restricted T cell response directed against both the Hyg and the Tk genes. Thus, the vector itself may mask the generation of immune reactivity against tumor antigens and may even detract from it. Vectors with immunogenic potential may be useful for tumor vaccination via cross priming in vivo, whereas antivector reactivities would be detrimental in situations where gene defects are being corrected and where long term expression of a therapeutic protein is required.

Anterior cruciate ligament reconstruction with the press-fit technique. 2-5 years followed-up of 42 patients.

42 patients underwent anterior cruciate ligament (ACL) reconstruction with the press-fit technique. The ACL was reconstructed with a bone-tendon-bone graft from the medial third of the patellar tendon. The graft was stabilized without screws in the femur and tibia by press-fit. To imitate the anatomical functioning of the ACL, the femoral bone block was placed with the tendon close to the over-the-top position. The tibial block was then placed in a trough on the tibia, so that the ligament fibres were parallel and tight during extension and slightly inverted during flexion. At evaluation mean 41 (25-61) months postoperatively, the mean Lysholm score was 93 (80-100) points, the mean activity level was 6 (3-10) points, and the mean translation of the tibia head, measured by the KT-1000 arthrometer (side-to-side difference), was 2 (0-7) mm. Only 3 of the patients suffered loss of extension (5 degrees). Patients who underwent reconstruction at least 4 months after the injury had better results than those who were operated earlier. The press-fit method allowed for anatomic substitution of the ACL with a stable graft without the disadvantages associated with screws. This method gave early postoperative functioning of the knee and good mid-term results.

Detection of cytosine deaminase in genetically modified tumor cells by specific antibodies.

Bacterial cytosine deaminase (CD) converts the non-toxic prodrug 5-fluorocytosine (5-FC) into 5-fluorouracil (5-FU), which is toxic for mammalian cells. Therefore, the CD gene is used in cancer gene therapy to achieve high local concentration of a toxic metabolite without significant systemic toxicity. To allow the detection of CD expression at the protein level, we raised both polyclonal rabbit antisera and a monoclonal antibody (mAb) against a histidine-tagged CD fusion protein. The specificity of the polyclonal antisera and the mAb was confirmed by immunohistochemistry, immunoblot analysis, and immunoprecipitation using CD-expressing tumor cell lines. Furthermore, the antibodies can be used for ELISA assays and flow cytometry. Finally, the CD protein could be demonstrated in frozen tissue sections of CD-modified tumors in a rat tumor model using the anti-CD serum. With these antibodies, CD expression can now be monitored throughout in vitro and in vivo gene transfer studies, including clinical protocols relying on the CD suicide gene strategy.

Differential function of CD80- and CD86-transfected human melanoma cells in the presence of IL-12 and IFN-gamma.

Introduction of co-stimulatory molecules like CD80 and CD86 represents a means to augment the immunogenicity of tumor cells and to induce immune responses directed at tumor antigens. Here we compared CD80- and CD86-transfected human melanoma cells to induce primary immune responses by their capacity to promote proliferation of human allogeneic resting T lymphocytes. CD80- and CD86-transfected SkMel63 melanoma cells induced T cell activation to a comparable degree, which was found to be independent of the cell surface density of these co-stimulatory molecules. Co-expression of CD80 and CD86 did not result in a synergistic increase in T cell proliferation. Both CD80 and CD86 transfectants induced the proliferation of isolated CD4+ or CD8+ T cells. Exogenous IL-2, IL-4 and tumor necrosis factor-alpha respectively enhanced primary T cell proliferation independent of CD80 or CD86 expression. Interestingly, differential activities of CD80 and CD86 were observed following stimulation of resting T cells in the presence of IL-12. Whereas IL-12 increased T cell proliferation in the presence of CD86-transfected melanoma cells, it exhibited an inhibitory function in the presence of CD80-expressing SkMel63 cells. Experimental evidence indicates that this inhibitory effect was mediated by IFN-gamma since (I) IFN-gamma secretion of stimulated T cells was augmented by IL-12, (II) exogenous IFN-gamma also inhibited T cell proliferation induced by CD80- but not CD86-transfected SkMel63 cells and (III) the inhibitory effect of IL-12 was blocked by an anti-IFN-gamma mAb.