Imaging Cell-Matrix Interactions in 3D Collagen Hydrogel Culture Systems.

3D hydrogels better replicate in vivo conditions, and yield different results from 2D substrates. However, imaging interactions between cells and the hydrogel microenvironment is challenging because of light diffraction and poor focal depth. Here, cryosectioning and vibrating microtomy methods and fixation protocols are compared. Collagen I/III hydrogel sections (20-100 µm) are fixed with paraformaldehyde (2%-4%) and structurally evaluated. Cryosectioning damaged hydrogels, and vibrating microtomy (100 µm, 2%) yielded the best preservation of microstructure and cell integrity. These results demonstrate a potential processing method that preserves hydrogel and cell integrity, permitting imaging of cell interactions with the microenvironment.

Hydrogels That Allow and Facilitate Bone Repair, Remodeling, and Regeneration.

Bone defects can originate from a variety of causes, including trauma, cancer, congenital deformity, and surgical reconstruction. Success of the current "gold standard" treatment (i.e., autologous bone grafts) is greatly influenced by insufficient or inappropriate bone stock. There is thus a critical need for the development of new, engineered materials for bone repair. This review describes the use of natural and synthetic hydrogels as scaffolds for bone tissue engineering. We discuss many of the advantages that hydrogels offer as bone repair materials, including their potential for osteoconductivity, biodegradability, controlled growth factor release, and cell encapsulation. We also discuss the use of hydrogels in composite devices with metals, ceramics, or polymers. These composites are useful because of the low mechanical moduli of hydrogels. Finally, the potential for thermosetting and photo-cross-linked hydrogels as three-dimensionally (3D) printed, patient-specific devices is highlighted. Three-dimensional printing enables controlled spatial distribution of scaffold materials, cells, and growth factors. Hydrogels, especially natural hydrogels present in bone matrix, have great potential to augment existing bone tissue engineering devices for the treatment of critical size bone defects.

Glioma-astrocyte interactions on white matter tract-mimetic aligned electrospun nanofibers.

Gliomas are highly invasive forms of brain cancer comprising more than 50% of brain tumor cases in adults, and astrocytomas account for ∼60-70% of all gliomas. As a result of multiple factors, including enhanced migratory properties and extracellular matrix remodeling, even with current standards of care, mean survival time for patients is only ∼12 months. Because glioblastoma multiforme (GBM) cells arise from astrocytes, there is great interest in elucidating the interactions of these two cell types in vivo. Previous work performed on two-dimensional assays (i.e., tissue culture plastic and Boyden chamber assays) utilizes substrates that lack the complexities of the natural microenvironment. Here, we employed a three-dimensional, electrospun poly-(caprolactone) (PCL) nanofiber system (NFS) to mimic some features of topographical properties evidenced in vivo. Co-cultures of human GBM cells and rat astrocytes, as performed on the NFS, showed a significant increase in astrocyte GFAP expression, particularly in the presence of extracellular matrix (ECM) deposited by GBM cells. In addition, GBM migration increased in the presence of astrocytes or soluble factors (i.e., conditioned media). However, the presence of fixed astrocytes acted as an antagonist, lowering GBM migration rates. This data suggests that astrocytes and GBM cells interact through a multitude of pathways, including soluble factors and direct contact. This work demonstrates the potential of the NFS to duplicate some topographical features of the GBM tumor microenvironment, permitting analysis of topographical effects in GBM migration.

Glioblastoma behaviors in three-dimensional collagen-hyaluronan composite hydrogels.

Glioblastoma multiforme (GBM) tumors, which arise from glia in the central nervous system (CNS), are one of the most deadly forms of human cancer with a median survival time of ∼1 year. Their high infiltrative capacity makes them extremely difficult to treat, and even with aggressive multimodal clinical therapies, outcomes are dismal. To improve understanding of cell migration in these tumors, three-dimensional (3D) multicomponent composite hydrogels consisting of collagen and hyaluronic acid, or hyaluronan (HA), were developed. Collagen is a component of blood vessels known to be associated with GBM migration; whereas, HA is one of the major components of the native brain extracellular matrix (ECM). We characterized hydrogel microstructural features and utilized these materials to investigate patient tumor-derived, single cell morphology, spreading, and migration in 3D culture. GBM morphology was influenced by collagen type with cells adopting a rounded morphology in collagen-IV versus a spindle-shaped morphology in collagen-I/III. GBM spreading and migration were inversely dependent on HA concentration; with higher concentrations promoting little or no migration. Further, noncancerous astrocytes primarily displayed rounded morphologies at lower concentrations of HA; in contrast to the spindle-shaped (spread) morphologies of GBMs. These results suggest that GBM behaviors are sensitive to ECM mimetic materials in 3D and that these composite hydrogels could be used to develop 3D brain mimetic models for studying migration processes.