Three-dimensional culture of human meniscal cells: Extracellular matrix and proteoglycan production

<p>Abstract</p> <p>Background</p> <p>The meniscus is a complex tissue whose cell biology has only recently begun to be explored. Published models rely upon initial culture in the presence of added growth factors. The aim of this study was to test a three-dimensional (3D) collagen sponge microenvironment (without added growth factors) for its ability to provide a microenvironment supportive for meniscal cell extracellular matrix (ECM) production, and to test the responsiveness of cells cultured in this manner to transforming growth factor-β (TGF-β).</p> <p>Methods</p> <p>Experimental studies were approved prospectively by the authors' Human Subjects Institutional Review Board. Human meniscal cells were isolated from surgical specimens, established in monolayer culture, seeded into a 3D scaffold, and cell morphology and extracellular matrix components (ECM) evaluated either under control condition or with addition of TGF-β. Outcome variables were evaluation of cultured cell morphology, quantitative measurement of total sulfated proteoglycan production, and immunohistochemical study of the ECM components chondroitin sulfate, keratan sulfate, and types I and II collagen.</p> <p>Result and Conclusion</p> <p>Meniscal cells attached well within the 3D microenvironment and expanded with culture time. The 3D microenvironment was permissive for production of chondroitin sulfate, types I and II collagen, and to a lesser degree keratan sulfate. This microenvironment was also permissive for growth factor responsiveness, as indicated by a significant increase in proteoglycan production when cells were exposed to TGF-β (2.48 μg/ml ± 1.00, mean ± S.D., vs control levels of 1.58 ± 0.79, p < 0.0001). Knowledge of how culture microenvironments influence meniscal cell ECM production is important; the collagen sponge culture methodology provides a useful in vitro tool for study of meniscal cell biology.</p

ACL reconstruction combined with lateral monoloop tenodesis can restore intact knee laxity

Purpose An anterior cruciate ligament (ACL) injury is often combined with injury to the lateral extra-articular structures, which may cause a combined anterior and rotational laxity. It was hypothesised that addition of a ‘monoloop’ lateral extra-articular tenodesis (mLET) to an ACL reconstruction would restore anteroposterior, internal rotation and pivot-shift laxities better than isolated ACL reconstruction in combined injuries. Method Twelve cadaveric knees were tested, using an optical tracking system to record the kinematics through 0°–100° of knee flexion with no load, anterior and posterior translational forces (90 N), internal and external rotational torques (5 Nm), and a combination of an anterior translational (90 N) plus internal rotational load (5 Nm). They were tested intact, after sectioning the ACL, sectioning anterolateral ligament (ALL), iliotibial band (ITB) graft harvest, releasing deep ITB fibres, hamstrings tendon ACL reconstruction, mLET combined with ACL reconstruction, and isolated mLET. Two-way repeated-measures ANOVA compared laxity data across knee states and flexion angles. When differences were found, paired t tests with Bonferroni correction were performed. Results In the ACL-deficient knee, cutting the ALL significantly increased anterior laxity only at 20°–30°, and only significantly increased internal rotation at 50°. Additional deep ITB release significantly increased anterior laxity at 40°–90° and caused a large increase of internal rotation at 20°–100°. Isolated ACL reconstruction restored anterior drawer, but significant differences remained in internal rotation at 30°–100°. After adding an mLET there were no remaining differences with anterior translation or internal rotation compared to the intact knee. With the combined injury, isolated mLET allowed abnormal anterior translation and rotation to persist. Conclusions Cutting the deep fibres of the ITB caused large increases in tibial internal rotation laxity across the range of knee flexion, while cutting the ALL alone did not. With ACL deficiency combined with anterolateral deficiency, ACL reconstruction alone was insufficient to restore native knee rotational laxity. However, combining a ‘monoloop’ lateral extra-articular tenodesis with ACL reconstruction did restore native knee laxity