Modifiable hydrogels have revealed tremendous insight into how physical characteristics of cells’ 3D environment drive stem cell lineage specification. However, in native tissues, cells do not passively receive signals from their niche. Instead they actively probe and modify their pericellular space to suit their needs, yet the dynamics of cells’ reciprocal interactions with their pericellular environment when encapsulated within hydrogels remains relatively unexplored. Here, we show that human bone marrow stromal cells (hMSC) encapsulated within hyaluronic acid-based hydrogels modify their surroundings by synthesizing, secreting and arranging proteins pericellularly or by degrading the hydrogel. hMSC’s interactions with this local environment have a role in regulating hMSC fate, with a secreted proteinaceous pericellular matrix associated with adipogenesis, and degradation with osteogenesis. Our observations suggest that hMSC participate in a bi-directional interplay between the properties of their 3D milieu and their own secreted pericellular matrix, and that this combination of interactions drives fate.

Osteoarthritis (OA) is a common cause of pain and disability and is often associated with the degeneration of articular cartilage. Lesions to the articular surface, which are thought to progress to OA, have the potential to be repaired using tissue engineering strategies; however, it remains challenging to instruct cell differentiation within a scaffold to produce tissue with appropriate structural, chemical and mechanical properties. We aimed to address this by driving progenitor cells to adopt a chondrogenic phenotype through the tailoring of scaffold composition and physical properties. Monomeric type-I and type-II collagen scaffolds, which avoid potential immunogenicity associated with fibrillar collagens, were fabricated with and without chondroitin sulfate (CS) and their ability to stimulate the chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells was assessed. Immunohistochemical analyses showed that cells produced abundant collagen type-II on type-II scaffolds and collagen type-I on type-I scaffolds. Gene expression analyses indicated that the addition of CS – which was released from scaffolds quickly – significantly upregulated expression of type II collagen, compared to type-I and pure type-II scaffolds. We conclude that collagen type-II and CS can be used to promote a more chondrogenic phenotype in the absence of growth factors, potentially providing an eventual therapy to prevent OA.

A thorough understanding of how proteins induce nanoparticle (NP) aggregation is crucial when designing in vitro and in vivo assays and interpreting experimental results. This knowledge is also crucial when developing nano-applications and formulation for drug delivery systems. In this study, we found that extraction of immunoglobulin G (IgG) from cow serum results in lower polystyrene NPs aggregation. Moreover, addition of isolated IgG or fibrinogen to fetal cow serum enhanced this aggregation, thus demonstrating that these factors are major drivers of NP aggregation in serum. Counter-intuitively, NP aggregation was inversely dependent on protein concentration; i.e., low protein concentrations induced large aggregates, whereas high protein concentrations induced small aggregates. Protein-induced NP aggregation and aggregate size were monitored by absorbance at 400 nm and dynamic light scattering, respectively. Here, we propose a mechanism behind the protein concentration dependent aggregation; this mechanism involves the effects of multiple protein interactions on the NP surface, surface area limitations, aggregation kinetics, and the influence of other serum proteins.

The transcriptional profile induced by hypoxia plays important roles in the chondrogenic differentiation of marrow stromal/stem cells (MSC) and is mediated by the Hypoxia Inducible Factor complex. However, various compounds can also stabilise HIF's oxygen‐responsive element, HIF‐1α, at normoxia and mimic many hypoxia‐induced cellular responses. Such compounds may prove efficacious in cartilage tissue engineering, where microenvironmental cues may mediate functional tissue formation. Here, we investigated three HIF stabilising compounds, which each have distinct mechanisms of action, to understand how they differentially influenced the chondrogenesis of human bone marrow‐derived MSC (hBM‐MSC) in vitro. hBM‐MSCs were chondrogenically‐induced in TGF‐β₃‐containing media in the presence of HIF‐stabilising compounds. HIF‐1α stabilisation was assessed by HIF‐1α immunofluorescence staining, expression of HIF target and articular chondrocyte specific genes by qPCR, and cartilage‐like extracellular matrix (ECM) production by immunofluorescence and histochemical staining. We demonstrate that all three compounds induced similar levels of HIF‐1α nuclear localisation. However, whilst the 2‐oxoglutarate analogue Dimethyloxalylglycine (DMOG) promoted upregulation of a selection of HIF target genes, Desferrioxamine (DFX) and Cobalt Chloride (CoCl₂), compounds that chelate or compete with Fe²⁺, respectively, did not. Moreover, DMOG induced a more chondrogenic transcriptional profile, which was abolished by Acriflavine, an inhibitor of HIF‐1α‐HIF‐β binding, whilst the chondrogenic effects of DFX and CoCl₂ were more limited. Together, these data suggest that HIF‐1α function during hBM‐MSC chondrogenesis may be regulated by mechanisms with a greater dependence on 2‐oxoglutarate than Fe²⁺ availability. These results may have important implications for understanding cartilage disease and developing targeted therapies for cartilage repair.