Promotion of hMDSC differentiation by combined action of scaffold material and TGF-ß superfamily growth factors.

Objective: Herein we propose a combined action of collagen type I (CA) or synthetic collagen-like-peptide functionalized with the cell adhesive RGD motif (PEG-CLP-RGD) hydrogels and selected growth factors to promote chondrogenic differentiation of human muscle-derived stem cells (hMDSCs) under normal and reduced oxygen conditions. Methods: hMDSCs were set for differentiation towards chondrogenic lineage using BMP-7 and TGF-ß3. Cells were seeded onto hydrogels loaded with growth factors (75ng/scaffold) and cultured for 28 days under normal (21%) and severe hypoxic (1%) conditions. Chondrogenesis was evaluated by monitoring collagen type II and GAG deposition, and quantification of ACAN expression by RT-PCR. Results: Sustained release of TGFß3 from the hydrogels was observed, 8.7 ± 0.5% of the initially loaded amount diffused out after 24 h from both substrates. For the BMP-7 growth factor, 14.8 ± 0.3% and 18.2 ± 0.6% of the initially loaded amount diffused out after 24 h from CA and CLP-RGD, respectively. The key findings of this study are: i) the self-supporting hydrogels themselves can stimulate hMDSC chondrogenesis by inducing gene expression of cartilage-specific proteoglycan aggrecan and ECM production; ii) the effect of dual BMP-7 and TGF-ß3 loading was more pronounced on CA hydrogel under normal oxygen conditions; iii) dual loading on PEG-CLP-RGD hydrogels did not have the synergistic effect, TGF-ß3 was more effective under both oxygen conditions; iv) BMP-7 can improve chondrogenic effect of TGF-ß3 on CA scaffolds, and hydrogels loaded with both growth factors can induce cartilage formation in hMDSC cultures. Conclusion: Our results support the potential strategy of combining implantable hydrogels functionalized with differentiation factors toward improving cartilaginous repair.

Investigation of Cancer Cell Migration and Proliferation on Synthetic Extracellular Matrix Peptide Hydrogels.

Chemical and mechanical properties of a tumor microenvironment are essential players in cancer progression, and it is important to precisely control the extracellular conditions while designing cancer in vitro models. The study investigates synthetic hydrogel matrices from multi-arm polyethylene glycol (PEG) functionalized with collagen-like peptide (CLP) CG(PKG)4(POG)4(DOG)4 alone and conjugated with either cell adhesion peptide RGD (mimicking fibronectin) or IKVAV (mimicking laminin). Human glioblastoma HROG36, rat C6 glioma cells, and A375 human melanoma cells were grown on the hydrogels and monitored for migration, proliferation, projected cell area, cell shape index, size and number, distribution of focal contacts in individual cells, and focal adhesion number. PEG-CLP-RGD induced migration of both glioma cell lines and also stimulated proliferation (assessed as metabolic activity) of HROG36 cells. Migration of C6 cells were also stimulated by PEG-CLP-IKVAV. These responses strongly correlated with the changes in adhesion and morphology parameters of individual cells - projected cell area, cell shape index, and focal contact number. Melanoma A375 cell proliferation was increased by PEG-CLP-RGD, and this was accompanied by a decrease in cell shape index. However, neither RGD nor IKVAV conjugated to PEG-CLP stimulated migratory capacity of A375 cells. Taken together, the study presents synthetic scaffolds with extracellular matrix (ECM)-mimicking peptides that allow for the exploration of the effect of ECM signaling to cancer cells.

Photografting and Patterning of Poly(ethylene glycol) Methacrylate Hydrogel on Glass for Biochip Applications.

An efficient procedure for chemical initiator-free, in situ synthesis of a functional polyethylene glycol methacrylate (PEG MA) hydrogel on regular glass substrates is reported. It is demonstrated that self-initiated photografting and photopolymerization driven by UV irradiation can yield tens of nanometer-thick coatings of carboxy-functionalized PEG MA on the aldehyde-terminated borosilicate glass surface. The most efficient formulation for hydrogel synthesis contained methyl methacrylic acid (MAA), 2-hydroxyethyl methacrylate (HEMA), and PEG methacrylate (PEG10MA) monomers (1:1:1). The resulting HEMA/PEG10MA/MAA (HPMAA) coatings had a defined thickness in the range from 11 to 50 nm. The physicochemical properties of the synthesized HPMAA coatings were analyzed by combining water contact angle measurements, stylus profilometry, imaging null ellipsometry, and atomic force microscopy (AFM). The latter technique was employed in the quantitative imaging mode not only for direct probing of the surface topography but also for swelling behavior characterization in the pH range from 4.5 to 8.0. The estimated high swelling ratios of the HPMAA hydrogel (up to 3.2) together with its good stability and resistance to nonspecific protein binding were advantageous in extracellular matrix mimetics via patterning of fibronectin (FN) at a resolution close to 200 nm. It was shown that the fabricated FN micropatterns on HPMAA were equally suitable for single-cell arraying, as well as controlled cell culture lasting at least for 96 h.