Meniscus matrix morphological composition: age-dependent evaluation in a swine model

Menisci are fibro-cartilaginous structures interposed between femoral condyle and tibial plateau, which have multiple functions in the stifle joint: act as shock absorbers, bear loaders and allow joint stability, congruity and lubrication (Sweigart et al., 2004; Proffen et al., 2012). It is well known that meniscal injuries lead to osteoarthritis and for these reasons, menisci are considered important target of investigation. Their important role in the knee wellness is only equalled by their deficiency in proper self-repairing.Nowadays, the gold standard technique is not just to remove the damaged meniscus, but to rebuild it or to replace it. For these reasons, studies are necessary to increase the knowledge about these small but essential structures (Streuli, 1999; Deponti et al., 2013). Composition and morphology are basic fundamental information for the development of engineered meniscal substitutes (Di Giancamillo et al., 2014). The analysis of the morphological, structural and biochemical changes, which occur during growth of the normal menisci, represent the goal of the present study. For this purpose, menisci from adult (7-month old), young (1-month old), and neonates (stillbirths) pigs were collected. Cellularity and glycosamiglycans (GAGs) deposition were evaluated by ELISA, while Collagen-1 and Collagen-2 were investigated by immunohistochemistry and Western blot analyses. Cellularity (P<0.01, all comparisons) and Collagen-1 (P<0.05, neonatal-young vs adult) decreased from neonatal to adult stage while GAGs (P<0.01 neonatal vs young-adult) and Collagen-2 (P<0.01 neonatal-young vs adult) showed the opposite trend. Immunohistochemistry revealed similar changes occurring during animal growth thus revealing that cellular phenotype, cellularity and protein expression, as well as fibers aggregation in the matrix, are dissimilar in the three ages analysed categories. These changes reflect the progressive menisci maturation and hyper-specialisation. We observed the correlation between biochemical and phenotype properties of swine menisci follow age-dependent changes during growth: starting with an immature cellular and fiber pattern to the mature organised and differentiated adult menisci.Acknowledgments: This work was funded by the “Finanziamento Piano Sviluppo Ateneo - Linea 2A

Meniscal femoral and tibial surfaces characterization in the swine model.

Menisci are wedge-like structures interposed, in the knee joint, between the femoral and the tibial articular heads (Kohn et al. 1995; Greis et al. 2002). Improving the articular surface, the cellular nutrition and the articular lubrication, they are essential structures for the prevention of gonarthrosis (Proctor et al.1989; Makris et al. 2011). This study is focused upon the relationship between the contact forces at the femoral and tibial surfaces and the corresponding structure of these meniscal surfaces. For this purpose, 20 adult (~9 months old) female pigs (Landrace x Large white, average weight 75–90 kg; n=80 meniscal samples) were obtained from a local slaughterhouse and dissected to isolate the menisci. Swine meniscal samples were evaluated from morphological (Safranin-O, Sirius Red and collagen type I and II) (Di Giancamillo et al. 2014), biochemical (DNA and glycosaminoglycans, or GAGs, contents) and biomechanical (compression and traction tests) points of view at the level of femoral and tibial meniscal surfaces. Results revealed a characterization of the meniscus which is biomechanical-dependent.  The femoral surface, morphologically characterized by the interposition of radial and oblique fibers and biomechanically by the femoral condyles compression, sliding and rolling forces, shows a higher compressive modulus (p<0.05) and a greater amount of cells and GAGs deposition (p<0.01 for each analysis). On the other hand, results from traction test revealed a higher tensile modulus (p<0.05) in the tibial surface, characterized by a circumferential arrangement of the fibers and a poorer GAGs deposition and cellular distribution (p<0.01). Results (summarized in the figure 1) from this work suggest that a biphasic “femoral-to-tibial” scaffold that mimic the different behavior and composition of the two meniscal surfaces could be useful in the light of meniscal replacement.

In vitro characterization of stem/progenitor cells from semitendinosus and gracilis tendons as a possible new tool for cell-based therapy for tendon disorders

Purpose: this study was conducted to characterize tendon stem/progenitor cells (TSPCs) isolated from human semitendinosus and gracilis tendons in terms of stemness properties and multi-differentiation potential. Methods: TSPCs were isolated from waste portions of semitendinosus and gracilis tendons from three donors who underwent anterior cruciate ligament reconstruction. TSPCs were plated in culture until passage 4, when experiments to assess cell proliferation, viability and clonogenic ability were performed. The immunophenotype of TSPCs was evaluated by cytofluorimetric analysis. The in vitro osteogenic, chondrogenic, adipogenic and tenogenic potential was evaluated using biochemical, histological and gene expression analysis to detect specific differentiation markers. Statistical analysis was performed using Student’s t-test. Results: after a few passages in culture the cell populations showed a homogeneous fibroblast-like morphology typical of mesenchymal stem cells. The average doubling time of TSPCs increased from 52.4±4.8 at passage 2 to 100.8±23.4 hours at passage 4. The highest percentage of colonies was also found at passage 4 (4.7±2.3%). TSPCs showed the typical mesenchymal phenotype, with high expression of CD73, CD90 and CD105 and no expression of CD34 and CD45. Cells induced to differentiate toward osteogenic lineage showed significant upregulations of ALP activity (+189%, p<0.05) and calcified matrix deposition (+49%, p<0.05) compared with undifferentiated cells; culture in chondrogenic medium also provoked a significant increase in glycosaminoglycan levels (+108%, p<0.05). On the other hand, TSPCs were not able to respond to adipogenic stimuli. Scleraxis gene expression and decorin gene expression, considered tenogenic markers, were already very high in control cells, and culture in tenogenic medium further increased these values although not significantly. Conclusions: our data show that it is possible to isolate TSPCs from very small fragments of tissue and that they show the typical features of MSCs and multi-differentiation potential, above all toward osteogenic and chondrogenic lineages. Clinical relevance: this study can be considered one of the first attempts to clarify the biology of tendon cell populations, focusing in particular on the potential applicability of this cell source for future regenerative medicine purposes

Tactile and proprioceptive temporal discrimination are impaired in functional tremor.

In order to obtain further information on the pathophysiology of functional tremor, we assessed tactile discrimination threshold and proprioceptive temporal discrimination motor threshold values in 11 patients with functional tremor, 11 age- and sex-matched patients with essential tremor and 13 healthy controls. Tactile discrimination threshold in both the right and left side was significantly higher in patients with functional tremor than in the other groups. Proprioceptive temporal discrimination threshold for both right and left side was significantly higher in patients with functional and essential tremor than in healthy controls. No significant correlation between discrimination thresholds and duration or severity of tremor was found. Temporal processing of tactile and proprioceptive stimuli is impaired in patients with functional tremor. The mechanisms underlying this impaired somatosensory processing and possible ways to apply these findings clinically merit further research

The Role of Peritumoral Depapillation and Its Impact on Narrow-Band Imaging in Oral Tongue Squamous Cell Carcinoma

A recent study reported that the occurrence of depapillated mucosa surrounding oral tongue squamous cell carcinomas (OTSCC) is associated with perineural invasion (PNI). The present study evaluates the reliability of depapillation as a PNI predictor and how it could affect narrow-band imaging (NBI) performance. This is thus a retrospective study on patients affected by OTSCC submitted to radical surgery. The preoperative endoscopy was evaluated to identify the presence of depapillation. Differences in distribution between depapillation and clinicopathological variables were analyzed. NBI vascular patterns were reported, and the impact of depapillation on those was studied. We enrolled seventy-six patients. After evaluation of the preoperative endoscopies, 40 (53%) patients had peritumoral depapillation, while 59 (78%) had a positive NBI pattern. Depapillation was strongly correlated to PNI, 54% vs. 28% (p = 0.022). Regarding the NBI pattern, there was no particular association with depapillation-associated tumors. The presence of depapillation did not affect the intralesional pattern detected by the NBI, while no NBI-positive pattern was found in the depapillation area. Finally, the NBI-guided resection margins were not affected by depapillation. Peritumoral depapillation is a reliable feature for PNI in OTSCC. NBI margin detection is not impaired by depapillation

Perceptual tactile and proprioceptive thresholds, TDT and TDMT in cerebellar ataxia and healthy subjects.

<p>PTT = perceptual tactile threshold; PPT = perceptual proprioceptive threshold; TDT = tactile temporal discrimination threshold; TDMT = proprioceptive temporal discrimination movement thresholds; FDI = first dorsal interosseous; FCR = flexor carpi radialis.</p>*<p>Significant with Bonferroni’s correction.</p><p>All neurophysiological data are given as mean ±SE.</p