The chondrogenic differentiation of bone marrow-derived human mesenchymal stem cells (MSCs) in a collagen type I hydrogel, which is in clinical use for matrix-based autologous chondrocyte transplantation (ACT), was investigated. Collagen hydrogels with 2.5 x 10(5) MSCs/mL were fabricated and cultured for 3 weeks in a serum-free, defined, chondrogenic differentiation medium containing 10 ng/mL TGF-beta1 or 100 ng/mL BMP-2. Histochemistry revealed morphologically distinct, chondrocyte-like cells, surrounded by a sulfated proteoglycan-rich extracellular matrix in the TGF-beta1 and BMP-2 treated group, with more elongated cells seen in the BMP-2 treated group. Immunohistochemistry detected collagen type II (Col II) in the TGF-beta1 and BMP-2 treated group. Collagen type X (Col X) staining was positive in the TGF-beta1 but only very weak in the BMP-2 treated group. RT-PCR analyses revealed a specific chondrogenic differentiation with the expression of the cartilage specific marker genes Col II, Col X, and aggrecan (AGN) in the TGF-beta1 and the BMP-2 treated group, with earlier expression of these marker genes in the TGF-beta1 treated group. Interestingly, MSC-gels cultured in DMEM with 10% FBS (control) indicated few isolated chondrocyte-like cells but no expression of Col II or Col X could be detected. The results show, that MSCs cultured in a collagen type I hydrogel are able to undergo a distinct chondrogenic differentiation pathway, similar to that described for MSCs cultured in high-density pellet cultures. These findings are valuable in terms of ex vivo predifferentiation or in situ differentiation of MSCs in collagen hydrogels for articular cartilage repair.

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This study reports the T 1 and T 2 relaxation rates of rhodamine-labeled anionic magnetic nanoparticles determined at 7, 11.7, and 17.6 T both in solution and after cellular internalization. Therefore cells were incubated with rhodamine-labeled anionic magnetic nanoparticles and were prepared at decreasing concentrations. Additionally, rhodamine-labeled anionic magnetic nanoparticles in solution were used for extracellular measurements. T 1 and T 2 were determined at 7, 11.7, and 17.6 T. T 1 times were determined with an inversionrecovery snapshot-flash sequence. T 2 times were obtained from a multispin-echo sequence. Inductively coupled plasmamass spectrometry was used to determine the iron content in all samples, and r 1 and r 2 were subsequently calculated. The results were then compared with cells labeled with AMI-25 and VSOP C-200. In solution, the r 1 and r 2 of rhodamine-labeled anionic magnetic nanoparticles were 4.78/379 (7 T), 3.28/389 (11.7 T), and 2.00/354 (17.6 T). In cells, the r 1 and r 2 were 0.21/56 (7 T), 0.19/37 (11.7 T), and 0.1/23 (17.6 T). This corresponded to an 11-to 23-fold decrease in r 1 and an 8-to 15-fold decrease in r 2 . A decrease in r 1 was observed for AMI-25 and VSOP C-200. AMI-25 and VSOP exhibited a 2-to 8-fold decrease in r 2 . In conclusion, cellular internalization of iron oxide nanoparticles strongly decreased their T 1 and T 2 potency. Magn Reson Med 64:1607-1615, 2010. V C 2010 Wiley-Liss, Inc.Key words: iron oxide nanoparticles; relaxivities; cellular internalization; high fieldThe unique soft-tissue contrast achieved by magnetic resonance imaging (MRI) originates from the variations of longitudinal (T 1 ) and transversal (T 2 /T 2 *) proton relaxation rates between different tissues. T 1 -and T 2 -shortening contrast agents are commonly used to enhance contrast. T 1 -shortening results in enhanced signal and/or faster imaging due to a higher value of available longitudinal magnetization for repetitive excitations. Because of faster spin relaxation, T 2 -shortening results in less signal but even higher contrast. As all contrast agents lower T 1 and T 2 , the ratio of the molar relaxivities (r 1 /r 2 ) is commonly used to describe their features.Superparamagnetic iron oxides particles (SPIOs) mainly shorten T 2 /T 2 * (r 1 /r 2 at 0.47 T: 0.2-0.6) (1-3); therefore, they are considered as ''negative contrast'' agents. For high field strengths above 2 T, r 1 decreases to zero and r 2 approaches a constant nonzero value. This results in a significant shift of the r 1 /r 2 ratio to lower values (r 1 /r 2 at 7 T: 0.01-0.1) (3-6). Therefore, at field strengths above 7 T, T 2 /T 2 *-weighted imaging sequences are commonly used for SPIO-enhanced MRI. In contrast to ''positive'' extracellular Gadolinium (Gd) chelates (r 1 /r 2 of Gd-DTPA at 0.47 T: >0.6), SPIOs remain in the healthy vasculature and are internalized by reticuloendothelial system cells after intravenous injection. Depending on their physicochemical properties, they are applied to MR-angiography, liver imaging (Ku...

A promising approach for the repair of osteochondral defects is the use of a scaffold with a well-defined cartilage-bone interface. In this study, we used a multiphasic composite scaffold with an upper collagen I fibre layer for articular cartilage repair, separated by a hydrophobic interface from a lower polylactic acid (PLA) part for bone repair. Focusing initially on the engineering of cartilage, the upper layer was seeded with human mesenchymal stem cells (hMSCs) suspended in a collagen I hydrogel for homogeneous cell distribution. The constructs were cultured in a defined chondrogenic differentiation medium supplemented with 10 ng/ml transforming growth factor-beta1 (TGFbeta1) or in DMEM with 10% fetal bovine serum as a control. After 3 weeks a slight contraction of the collagen I fibre layer was seen in the TGFbeta1-treated group. Furthermore, a homogeneous cell distribution and chondrogenic differentiation was achieved in the upper third of the collagen I fibre layer. In the TGFbeta1-treated group cells showed a chondrocyte-like appearance and were surrounded by a proteoglycan and collagen type II-rich extracellular matrix. Also, a high deposition of glycosaminoglycans could be measured in this group and RT-PCR analyses confirmed the induction of chondrogenesis, with the expression of cartilage-specific marker genes, such as aggrecan and collagen types II and X. This multiphasic composite scaffold with the cartilage layer on top might be a promising construct for the repair of osteochondral defects.