Growth factor effects on costal chondrocytes for tissue engineering fibrocartilage

Tissue-engineered fibrocartilage could become a feasible option for replacing tissues such as the knee meniscus or temporomandibular joint disc. This study employed five growth factors (insulin-like growth factor-I, transforming growth factor-beta1, epidermal growth factor, platelet-derived growth factor-BB, and basic fibroblast growth factor) in a scaffoldless approach with costal chondrocytes, attempting to improve biochemical and mechanical properties of engineered constructs. Samples were quantitatively assessed for total collagen, glycosaminoglycans, collagen type I, collagen type II, cells, compressive properties, and tensile properties at two time points. Most treated constructs had lower biomechanical and biochemical properties than the controls with no growth factors, suggesting a detrimental effect, but the treatment with insulin-like growth factor-I tended to improve the constructs. Additionally, the 6-week time point was consistently better than that at 3 weeks, with total collagen, glycosaminoglycans, and aggregate modulus doubling during this time. Further optimization of the time in culture and exogenous stimuli will be important in making a more functional replacement tissue.     

Eye ontogeny in Pleurodema bufoninum: A comparison with Pleurodema somuncurense (Anura,Leptodactylidae)

Vision is one of the main sensory systems in amphibians, and the eye structure is highly associated with habitat conditions. The ontogeny, as well as the adult structure, of the eye has been studied in only a few species. The life change after metamorphosis is accompanied by changes in the visual environment. The aim of this work is to describe the eye ontogeny of Pleurodema bufoninum and to compare it with that of Pleurodema somuncurense . Specimens of both Pleurodema species were processed for histology analysis at different stages of development, including the tadpole, postmetamorphic, and adult forms. Eyes in both Pleurodema species are composed of the 3 tunics, tunica fibrosa , tunica vasculosa , and tunica interna , and the lens. Additionally, in both, the iris presents a projection on its dorsal and ventral free ends that screens the cornea. This structure has been reported in the eye of several anuran species and is called the umbraculum, meniscus or pupillary nodule. Our results show that the structures related to light capture (retina and lens) appear early in larval life, while the components of the terrestrial‐life eye (scleral cartilage, specialized cornea, eyelids, nictitating membrane, and Harderian's gland) do not develop until the metamorphic climax, when the tadpole leaves the water. The adult eyes of P. bufoninum and P. somuncurense are very similar in structure and development.     

Effects of co‐cultures of meniscus cells and articular chondrocytes on PLLA scaffolds

The knee meniscus, a fibrocartilaginous tissue located in the knee joint, is characterized by heterogeneity in extracellular matrix and biomechanical properties. To recreate these properties using a tissue engineering approach, co‐cultures of meniscus cells (MCs) and articular chondrocytes (ACs) were seeded in varying ratios (100:0, 75:25, 50:50, 25:75, and 0:100) on poly‐L ‐lactic acid (PLLA) scaffolds and cultured in serum‐free medium for 4 weeks. Histological, biochemical, and biomechanical tests were used to assess constructs at the end time point. Strong staining for collagen and glycosaminoglycan (GAG) was observed in all groups. Constructs with 100% MCs were positive for collagen I and constructs cultured with 100% ACs were positive for collagen II, while a mixture of collagen I and II was observed in other co‐culture groups. Total collagen and GAG per construct increased as the percentage of ACs increased (27 ± 8 µg, 0% AC to 45 ± 8 µg, 100% ACs for collagen and 12 ± 4 µg, 0% ACs to 40 ± 5 µg, 100% ACs for GAG). Compressive modulus (instantaneous and relaxation modulus) of the constructs was significantly higher in the 100% ACs group (63 ± 12 and 22 ± 9 kPa, respectively) when compared to groups with higher percentage of MCs. No differences in tensile properties were noted among groups. Specific co‐culture ratios were identified mimicking the GAG/DW of the inner (0:100, 25:75, and 50:50) and outer regions (100:0) of the meniscus. Overall, it was demonstrated that co‐culturing MCs and ACs on PLLA scaffolds results in functional tissue engineered meniscus constructs with a spectrum of biochemical and biomechanical properties. Biotechnol. Bioeng. 2009;103: 808–816. © 2009 Wiley Periodicals, Inc.     

Mesenchymal stem cells as trophic mediators

Adult marrow-derived Mesenchymal Stem Cells (MSCs) are capable of dividing and their progeny are further capable of differentiating into one of several mesenchymal phenotypes such as osteoblasts, chondrocytes, myocytes, marrow stromal cells, tendon-ligament fibroblasts, and adipocytes. In addition, these MSCs secrete a variety of cytokines and growth factors that have both paracrine and autocrine activities. These secreted bioactive factors suppress the local immune system, inhibit fibrosis (scar formation) and apoptosis, enhance angiogenesis, and stimulate mitosis and differentiation of tissue-intrinsic reparative or stem cells. These effects, which are referred to as trophic effects, are distinct from the direct differentiation of MSCs into repair tissue. Several studies which tested the use of MSCs in models of infarct (injured heart), stroke (brain), or meniscus regeneration models are reviewed within the context of MSC-mediated trophic effects in tissue repair.     

Sex Difference in Contusion of the Medial Femoral Condyle Rim-Association with MRI Occult Meniscal Tears in the ACL Deficient Knee

BackgroundMeniscal tears, specifically lateral meniscal tears, have a larger than expected un-derdiagnosis rate in the presence of an ACL tear. The purpose of our study was to search for an MRI bone contusion pattern associated with MRI occult meniscal tears in patients with an ACL tear, specifically a contusion of the rim of the medial femoral condyle (RMFC). Our hypothesis was that there would be a significant association between RMFC contusions and MRI occult meniscal tears in patients with an ACL tear. We also searched for a difference between sexes with respect to the presence of the RMFC contusion in the setting of an occult meniscal tear. We also categorized the type, size, and location of these occult meniscal tears in the setting of an ACL tear.Methods This was a retrospective study that examined characteristics of occult meniscal tears and their association with a RMFC bone contusion. IRB approval was obtained. The date range of the study was June 2009 through December 2015. 6392 consecutive knee MRI reports in patients with an ACL deficient knee were reviewed. The study group included 22 patients with MRI occult meniscal tears, the control group included 110 patients. Relevant statistical values were calculated.ResultsThe most common type of occult meniscal tears were small radial and small longitudinal tears of the lateral meniscus. Occult meniscal tears were associated with an RMFC contusion in the study group (p=0.0457), particularly in males (p = 0.0003). In males with a torn ACL, the sensitivity of an RMFC contusion for an occult meniscal tear was 80%.ConclusionIn males with an ACL tear, there was a significant association between a contusion of the RMFC and an occult meniscal tear (commonly small radial or small peripheral partial-thickness longitudinal tears). RMFC contusions were reliably identified by radiologists in this study.Level of Evidence: II     

Structured three-dimensional co-culture of mesenchymal stem cells with meniscus cells promotes meniscal phenotype without hypertrophy

Menisci play a crucial role in weight distribution, load bearing, shock absorption, lubrication, and nutrition of articular cartilage within the knee joint. Damage to the meniscus typically does not heal spontaneously due to its partial avascular nature. Partial or complete meniscectomy is a common clinical treatment of the defective meniscus. However, this procedure ultimately leads to osteoarthritis due to increased mechanical stress to the articular cartilage. Meniscus tissue engineering offers a promising solution for partial or complete meniscus deficiency. Mesenchymal stem cells (MSC) have the potential to differentiate into meniscal fibrochondrocyte as well as deliver trophic effects to the differentiated cells. This study tested the feasibility of using MSC co-cultured with mature meniscal cells (MC) for meniscus tissue engineering. Structured cell pellets were created using MC and MSC at varying ratios (100:0, 75:25, 50:50, 25:75, and 0:100) and cultured with or without transforming growth factor-beta 3 supplemented chondrogenic media for 21 days. The meniscal and hypertrophic gene expression, gross appearance and structure of the pellets, meniscus extracellular matrix (ECM), histology and immunohistochemistry of proteoglycan and collagen were evaluated. Co-culture of MC with MSC at 75:25 demonstrated highest levels of collagen type I and glycosaminoglycans (GAG) production, as well as the lowest levels of hypertrophic genes, such as COL10A1 and MMP13. All co-culture conditions showed better meniscus ECM production and hypertrophic inhibition as compared to MSC culture alone. The collagen fiber bundles observed in the co-cultures are important to produce heterogenic ECM structure of meniscus. In conclusion, co-culturing MC and MSC is a feasible and efficient approach to engineer meniscus tissue with enhanced ECM production without hypertrophy.     

Effects of the type of meniscal lesion on knee function

The relationship between the type of meniscal lesion (bucket-handle, flap or degenerative tears) and preoperative knee function (5.1±6.2 days before arthroscopic meniscectomy) was studied in 35 patients. Patients with bucket-handle tears (group B, n=12) had large knee extension work deficits during maximal voluntary contractions at 30° and 180°/s (Kin-Com dynamometer). These work deficits were accompanied by proportional decreases in the prime mover or agonist-EMG levels (VM: vastus medialis and/or VL: vastus lateralis). These patients had smaller deficits during flexion movements at 30°/s. Moreover, a larger number of negative clinical signs and symptoms (pain at rest and during tests, locking, thigh atrophy, extension and flexion movement deficits larger than 10°) were found with an equal or a higher prevalence in group B than in the other groups. Patients with flap tears (group F, n=15) had deficits in work and agonist-EMG activity (VM and VL) only during the extension tests. In contrast, patients with degenerative tears (group D, n=8) had a work deficit and a concomitant decrease in the EMG level of the medial gastrocnemius (MG), only during the flexion test at 30°/s. Comparable mean knee function scores, as measured by the Lysholm and Gillquist questionnaire [35], were obtained for the three groups of patients, suggesting that this measure was not sensitive enough to discern functional differences related to types of meniscal tears. The results of this study have demonstrated a link between the type of meniscal lesion and the consequent preoperative knee joint disability profile as defined by comparison with the sound leg. These results emphasize the need to consider meniscal lesion type and an individual's preoperative strength deficit when group comparisons of patients are made or the effects of therapy (arthroscopic surgery and rehabilitation) are evaluated.     

Automated hexahedral meshing of knee cartilage structures – application to data from the osteoarthritis initiative

We propose a fully automated methodology for hexahedral meshing of patient-specific structures of the human knee obtained from magnetic resonance images, i.e. femoral/tibial cartilages and menisci. We select eight patients from the Osteoarthritis Initiative and validate our methodology using MATLAB on a laptop computer. We obtain the patient-specific meshes in an average of three minutes, while faithfully representing the geometries with well-shaped elements. We hope to provide a fundamentally different means to test hypotheses on the mechanisms of disease progression by integrating our patient-specific FE meshes with data from individual patients. Download both our meshes and software at http://im.engr.uconn.edu/downloads.php.     

Inflammatory molecules produced by meniscus and synovium in early and end-stage osteoarthritis: a coculture study

The aim of this study was to identify the molecules and pathways involved in the cross-talk between meniscus and synovium that may play a critical role in osteoarthritis (OA) pathophysiology. Samples of synovium and meniscus were collected from patients with early and end-stage OA and cultured alone or cocultured. Cytokines, chemokines, metalloproteases, and their inhibitors were evaluated at the gene and protein levels. The extracellular matrix (ECM) changes were also investigated. In early OA cultures, higher levels of interleukin-6 (IL-6) and IL-8 messenger RNA were expressed by synovium and meniscus in coculture compared with meniscus cultured alone. RANTES release was significantly increased when the two tissues were cocultured compared with meniscus cultured alone. Increased levels of matrix metalloproteinase-3 (MMP-3) and MMP-10 proteins, as well as increased release of glycosaminoglycans and aggrecan CS846 epitope, were observed when synovium was cocultured with meniscus. In end-stage OA cultures, increased levels of IL-8 and monocyte chemoattractant protein-1 (MCP-1) proteins were released in cocultures compared with cultures of meniscus alone. Chemokine (C-C motif) ligand 21 (CCL21) protein release was higher in meniscus cultured alone and in coculture compared with synovium cultured alone. Increased levels of MMP-3 and 10 proteins were observed when tissues were cocultured compared with meniscus cultured alone. Aggrecan CS846 epitope release was increased in cocultures compared with cultures of either tissue cultured alone. Our study showed the production of inflammatory molecules by synovium and meniscus which could trigger inflammatory signals in early OA patients, and induce ECM loss in the progressive and final stages of OA pathology.