Augmentation and repair tissue formation of the nucleus pulposus after partial nucleotomy in a rabbit model.

Disc degeneration alters disc height and mechanics of the spinal column and is associated with lower back pain. In preclinical studies gel-like materials or resorbable polymer-based implants are frequently used to rebuild the nucleus pulposus, aiming at tissue regeneration and restoration of tissue function. To compare the outcome of tissue repair, freeze-dried resorbable polyglycolic acid-hyaluronan (PGA/HA) implants without any bioactive components or bioactivated fibrin (fibrin-serum) was used in a degenerated disc disease model in New Zealand white rabbits. Animals with partial nucleotomy only served as controls. The T2-weighted/fat suppression sequence signal intensity in the nuclear region of operated discs as assessed by magnet resonance imaging was reduced in operated compared to healthy discs, indicating loss of water and did not change from week 1 to month 6 after surgery. Quantification of histological and immunohistochemical staining indicated that the implantation of PGA/HA leads to significantly more repair tissue compared to nucleotomy only. Type II collagen content of the repair tissue formed after PGA/HA or fibrin-serum treatment is significantly increased compared to controls with nucleotomy only. The data indicate that intervertebral disc augmentation after nucleotomy has a positive effect on repair tissue formation and type II collagen deposition as shown in the rabbit model.

Toward scaffold-based meniscus repair: effect of human serum, hyaluronic acid and TGF-ß3 on cell recruitment and re-differentiation.

OBJECTIVE: Repair approaches for the non-vascular meniscus are rarely developed. Recent strategies use scaffold-based techniques and inducing factors. The aim of the study was the investigation of cell recruitment and re-differentiation inducing factors for a scaffold-based meniscus repair approach. METHOD: 3D cultivation of in vitro expanded human meniscus-derived cells was performed in high-density cultures supplemented with 25% hyaluronic acid (HA), 10% human serum (HS) or 10 ng/ml transforming growth factor (TGF-ß3) compared to untreated controls. The in vitro cell recruitment potential of different HS concentrations was tested by chemotaxis assay. Analysis of chondrocytic markers (type I, II, IX collagen and proteoglycans) was performed on protein and gene expression level. RESULTS: Cells were attracted by 1-20% HS. 3D cultures supplemented with 10% HS and 25% HA showed meniscus-like gene expression profiles at day 7 with significantly increased cartilage oligomeric matrix protein (COMP) and aggrecan expression levels in the HS group and a slightly increased profile in the HA group compared to control. The TGF-ß3 group showed an additional induction of gene expression levels for type II and type IX collagen. Histological findings confirmed these results by proteoglycan and type I collagen staining in all groups and type II collagen staining only in the TGF-ß3 group. CONCLUSION: This study demonstrates that human meniscus cells are attracted by HS and allow for meniscal matrix formation in 3D culture in the presence of HA and HS, whereas TGF-ß3 additive does not initiate meniscal tissue. Regarding non-vascular meniscus repair, results of this study encourage scaffold-based repair approaches.

Expanded human meniscus-derived cells in 3-D polymer-hyaluronan scaffolds for meniscus repair.

Treatment options for lesions of the avascular region of the meniscus using regenerative medicine approaches based on resorbable scaffolds are rare. Recent approaches using scaffold-based techniques for tissue regeneration known from cartilage repair may be a promising treatment option for meniscal tears. The aim of the study was the investigation of meniscus matrix formation of in vitro expanded human meniscus-derived cells in a three-dimensional (3-D) bioresorbable polymer graft for meniscal repair approaches. Cultivation of the human meniscus cells was performed in a resorbable scaffold material made of polyglycolic acid (PGA) and hyaluronic acid, stabilized with fibrin glue. Cell viability and distribution of human meniscus cells in PGA-hyaluronan scaffolds were evaluated by fluorescein diacetate and propidium iodide staining. Verification of typical meniscal extracellular matrix molecules like type I and type III collagen was performed histologically, immunohistochemically and by gene expression analysis. In results, 3-D scaffold-based meniscus cultures showed high cell viability over an observational period of 21 days in PGA-hyaluronan scaffolds. On the protein level, type I collagen and proteoglycans were evident. Gene expression analysis confirmed the re-expression of meniscus-specific markers in PGA-hyaluronan scaffolds. This study demonstrated that in vitro expanded human meniscus cells allow for formation of meniscal matrix components when cultured in 3-D PGA-hyaluronan scaffolds stabilized with fibrin. These results encourage scaffold-based approaches for the treatment of meniscal lesions.

Repair of a post-traumatic cartilage defect with a cell-free polymer-based cartilage implant: a follow-up at two years by MRI and histological review.

Microfracture is frequently used as the first line of treatment for the repair of traumatic cartilage defects. We present the clinical and histological results 18 months to two-years after treatment in a 26-year-old male with a post-traumatic chondral defect of the medial femoral condyle managed by microfracture covered with chondrotissue, a cell-free cartilage implant made of a resorbable polyglycolic acid felt and hyaluronic acid.

Chemokine profile of synovial fluid from normal, osteoarthritis and rheumatoid arthritis patients: CCL25, CXCL10 and XCL1 recruit human subchondral mesenchymal progenitor cells.

OBJECTIVE: The microfracture technique activates mesenchymal progenitors that enter the cartilage defect and form cartilage repair tissue. Synovial fluid (SF) has been shown to stimulate the migration of subchondral progenitors. The aim of our study was to determine the chemokine profile of SF from normal, rheumatoid arthritis (RA) and osteoarthritis (OA) donors and evaluate the chemotactic effect of selected chemokines on human subchondral progenitor cells. METHOD: Chemokine levels of SF were analyzed using human chemokine antibody membrane arrays. The chemotactic potential of selected chemokines on human mesenchymal progenitors derived from subchondral cortico-spongious bone was tested using 96-well chemotaxis assays. Chemokine receptor expression of subchondral progenitors was assessed by real-time gene expression analysis and immuno-histochemistry. RESULTS: Chemokine antibody array analysis showed that SF contains a broad range of chemokines. Ten chemokines that showed significantly reduced levels in RA or OA compared to normal SF or robustly high levels in all SF tested were used for further chemotactic analysis. Chemotaxis assays showed that the chemokines MDC/CCL22, CTACK/CCL27, ENA78/CXCL5 and SDF1α/CXCL12 significantly inhibited migration of progenitors, while TECK/CCL25, IP10/CXCL10 and Lymphotactin/XCL1 effectively stimulated cell migration. MCP1/CCL2, Eotaxin2/CCL24 and NAP2/CXCL7 showed no chemotactic effect on subchondral progenitors. Gene expression and immuno-histochemical analysis of corresponding chemokine receptors document presence of low levels of chemokine receptors in subchondral progenitors, with the CXCL10 receptor CXCR3 showing the highest expression level. CONCLUSION: These results suggest that SF contains chemokines that may contribute to the recruitment of human mesenchymal progenitors from the subchondral bone in microfracture.

[Cryopreservation of bovine embryos]. / Beitrag zur Kryokonservierung von Rinderembryonen.

The method of cryopreservation of embryos aged seven days, proposed for embryo transfer with cattle by Niemann (1985), was tested under production conditions on three cattle breeding farms and three experimental animal units. The number of donors was 128, and 467 intact embryos were obtained from them and were cryopreserved in semen straw. Following thawing, 455 were recovered, and 439 (96.5 percent) of these were suitable for transfer. A pregnancy rate of 49.0 percent was recorded from 412 transfers. This rate was differentiated by oestric cycle conditions of heifer recipients, which gave percentages of 46.0 among recipients of seven-day old embryos, 45.7 for eight-day recipients, and 65.8 for six-day recipients. Related to pregnancy results recorded on the same units from transfer of freshly collected seven-day embryos, the efficiency coefficient was 0.69 (550 fresh transfers = 65.4 percent and 222 cryopreserved transfers = 48.2 percent). The method is recommended for general field practice.