Mechanical Loading Promoted Discogenic Differentiation of Human Mesenchymal Stem Cells Incorporated in 3D-PEG Scaffolds with RhGDF5 and RGD

Hydrogels have been described as ideal scaffolds for cells of 3D tissue constructs and hold strong promises with respect to in vitro 3D-cell-culture, where cells are isolated from native extracellular matrix (ECM). Synthesized polyethyleneglycol (PEG) hydrogels are appealing with regard to potential for cell therapy or as vehicles for drug delivery or even to regenerate tissue with similar hydrogel-like properties such as the nucleus pulposus of the intervertebral disc (IVD). Here, we tested whether incorporation of RGD motive would hinder discogenic differentiation of primary bone marrow-derived human mesenchymal stem cells (hMSCs) but favor proliferation of undifferentiated hMSCs. HMSCs were embedded in +RGD containing or without RGD PEG hydrogel and pre-conditioned with or without growth and differentiation factor-5 (rhGDF-5) for 13 days. Afterwards, all hMSCs-PEG gels were subsequently cyclically loaded (15% strain, 1Hz) for 5 consecutive days in a bioreactor to generate an IVD-like phenotype. Higher metabolic activity (resazurin assay) was found in groups with rhGDF5 in both gel types with and without RGD. Cell viability and morphology measured by confocal laser microscopy and DNA content showed decreased values (~60%) after 18 days of culture. Real-time RT-PCR of an array of 15 key genes suspected to be distinctive for IVD cells revealed moderate response to rhGDF5 and mechanical loading as also shown by histology staining. Preconditioning and mechanical loading showed relatively moderate responses revealed from both RT-PCR and histology although hMSCs were demonstrated to be potent to differentiate into chondrocyte-progenitor cells in micro- mass and 3D alginate bead culture.

Experimental determination of barium isotope fractionation during diffusion and adsorption processes at low temperatures

Variations in barium (Ba) stable isotope abundances measured in low and high temperature environments have recently received increasing attention. The actual processes controlling Ba isotope fractionation, however, remain mostly elusive. In this study, we present the first experimental approach to quantify the contribution of diffusion and adsorption on mass- dependent Ba isotope fractionation during transport of aqueous Ba2+ ions through a porous medium. Experiments have been carried out in which a BaCl2 solution of known isotopic composition diffused through u-shaped glass tubes filled with silica hydrogel at 10 C and 25 C for up to 201 days. The diffused Ba was highly fractionated by up to -2.15‰ in d137/134Ba, despite the low relative difference in atomic mass. The time-dependent isotope fractionation can be successfully reproduced by a diffusive transport model accounting for mass-dependent differences in the effective diffusivities of the Ba isotope species (D137Ba/D134Ba = (m134/m137)^b). Values of b extracted from the transport model were in the range of 0.010–0.011. Independently conducted batch experiments revealed that adsorption of Ba onto the surface of silica hydrogel favoured the heavier Ba isotopes (a = 1.00015 ± 0.00008). The contribution of adsorption on the overall isotope fractionation in the diffusion experiments, however, was found to be small. Our results contribute to the understanding of Ba isotope fractionation pro- cesses, which is crucial for interpreting natural isotope variations and the assessment of Ba isotope ratios as geochemical proxies.