Expression pattern differences between osteoarthritic chondrocytes and mesenchymal stem cells during chondrogenic differentiation.

OBJECTIVE: The use of mesenchymal stem cells (MSCs) for cartilage regeneration is hampered by lack of knowledge about the underlying molecular differences between chondrogenically stimulated chondrocytes and MSCs. The aim of this study was to evaluate differences in phenotype and gene expression between primary human chondrocytes and MSCs during chondrogenic differentiation in three-dimensional (3D) pellet culture (PC). MATERIALS AND METHODS: Chondrocytes isolated from cartilage samples obtained during total knee alloarthroplastic procedure (N=8) and MSCs, purified from bone marrow aspirates of healthy donors (N=8), were cultivated in PC under chondrogenic conditions. Immunohistology and quantitative reverse transcribing PCR (RT-PCR) were performed for chondrogenic-specific markers (i.e., Sox9, Collagen II). Global gene expression of the so-cultivated chondrocytes and MSCs was assessed by a novel approach of microarray-based pathway analysis. Refinement of data was done by hypothesis-driven gene expression omnibus (GEO) dataset comparison. Validation was performed with separate samples in transforming growth factor (TGF)ß+ or TGFß- conditions by use of quantitative real-time RT-PCR. RESULTS/CONCLUSIONS: Chondrogenic commitment of both cell types was observed. Interestingly, chondrocytes demonstrated an upregulated fatty acid/cholesterol metabolism which may give hints for future optimization of culture conditions. The novel microarray-based pathway analysis applied in this study seems suitable for the evaluation of whole-genome based array datasets in case when hypotheses can be backed with already existing GEO datasets. Future experiments should further explore the different metabolic behaviour of chondrocytes and MSC.

Ovine bone marrow mesenchymal stem cells: isolation and characterization of the cells and their osteogenic differentiation potential on embroidered and surface-modified polycaprolactone-co-lactide scaffolds.

The current study was undertaken with the goal being isolation, cultivation, and characterization of ovine mesenchymal stem cells (oMSC). Furthermore, the objective was to determine whether biological active polycaprolactone-co-lactide (trade name PCL) scaffolds support the growth and differentiation of oMSC in vitro. The oMSC were isolated from the iliac crest of six merino sheep. Three factors were used to demonstrate the MSC properties of the isolated cells in detail. (1) Their ability to proliferate in culture with a spindle-shaped morphology, (2) presence of specific surface marker proteins, and (3) their capacity to differentiate into the three classical mesenchymal pathways, osteoblastic, adipogenic, and chondrogenic lineages. Furthermore, embroidered PCL scaffolds were coated with collagen I (coll I) and chondroitin sulfate (CS). The porous structure of the scaffolds and the coating with coll I/CS allowed the oMSC to adhere, proliferate, and to migrate into the scaffolds. The coll I/CS coating on the PCL scaffolds induced osteogenic differentiation of hMSC, without differentiation supplements, indicating that the scaffold also has an osteoinductive character. In conclusion, the isolated cells from the ovine bone marrow have similar morphologic, immunophenotypic, and functional characteristics as their human counterparts. These cells were also found to differentiate into multiple mesenchymal cell types. This study demonstrates that embroidered PCL scaffolds can act as a temporary matrix for cell migration, proliferation, and differentiation of oMSC. The data presented will provide a reliable model system to assess the translation of MSC-based therapy into a variety of valuable ovine experimental models under autologous settings.

Immunohistochemical in situ characterization of orthopedic implants on polymethyl metacrylate embedded cutting and grinding sections.

When investigating the tissue reaction on orthopedic implants, the cellular activity at the bone-implant interface is of special interest. Preparation of undecalcified bone sections with methylmetacrylate (MMA)-based resins allows evaluation of the host tissue reactions with the implant in situ. However, the technical workup is demanding and few reports exist on the immunohistochemical characterization of these sections. Rat (R), sheep (S), and human (H) samples were investigated. R specimens contained intramedullary rods in the rat tibia. S specimens were sheep tibiae with an external fixator. H specimens were obtained from deceased patients. Specimens were embedded in MMA-based Technovit 9100N using cold polymerization. Sections of 10-15 microm thickness were obtained and prepared for immunohistochemical staining. Good morphological detail was preserved in all specimens providing information about mineralization, recent bone formation, and bone-implant contact. The following antibodies could reproducibly be detected specifically: Osteopontin (R, S, H), Osteonectin, Cathepsin D (R, S), von Willebrand factor (R, H), Osteocalcin, ED 1 (R), CD 3, CD 68, Keratin (H). Control procedures without adding primary antibodies showed no unspecific staining. Reliable detection of immunohistochemical markers of bone resorption, bone formation, inflammation, and angiogenesis at undecalcified sections with the implant in situ appears promising in enhancing our understanding of the cellular activity and cell-matrix interactions at the bone-implant interface.

Using a self-control training procedure to increase appropriate behavior.

The present study evaluated a technique for teaching self-control and increasing desirable behaviors among adults with developmental disabilities. Results showed that when participants were initially given the choice between an immediate smaller reinforcer and a larger delayed reinforcer, all participants repeatedly chose the smaller reinforcer. Concurrent fixed-duration/progressive-duration reinforcement schedules then were introduced in which initially both the smaller and larger reinforcers were available immediately. Thereafter, progressively increasing delays were introduced for the schedule associated with the larger reinforcer only. When initial short-duration requirements for access to the larger reinforcer were gradually increased, participants repeatedly selected the larger reinforcer, thereby demonstrating increased self-control.