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Creators/Authors contains: "Harley, Brendan A. C."

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  1. Abstract

    Current treatments for craniomaxillofacial (CMF) defects motivate the design of instructive biomaterials that can promote osteogenic healing of complex bone defects. We report methods to promote in vitro osteogenesis of human mesenchymal stem cells (hMSCs) within a model mineralized collagen scaffold via the incorporation of ascorbic acid (vitamin C), a key factor in collagen biosynthesis and bone mineralization. An addition of 5 w/v% ascorbic acid into the base mineralized collagen scaffold significantly changes key morphology characteristics including porosity, macrostructure, and microstructure. This modification promotes hMSC metabolic activity, ALP activity, and hMSC‐mediated deposition of calcium and phosphorous. Additionally, the incorporation of ascorbic acid influences osteogenic gene expression (BMP‐2,RUNX2,COL1A2) and delays the expression of genes associated with osteoclast activity and bone resorption (OPN,CTSK), though it reduces the secretion of OPG. Together, these findings highlight ascorbic acid as a relevant component for mineralized collagen scaffold design to promote osteogenic differentiation and new bone formation for improved CMF outcomes.

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  2. Abstract

    Biomaterials that replicate patterns of microenvironmental signals from the stem cell niche offer the potential to refine platforms to regulate stem cell behavior. While significant emphasis has been placed on understanding the effects of biophysical and biochemical cues on stem cell fate, vascular‐derived or angiocrine cues offer an important alternative signaling axis for biomaterial‐based stem cell platforms. Elucidating dose‐dependent relationships between angiocrine cues and stem cell fate are largely intractable in animal models and 2D cell cultures. In this study, microfluidic mixing devices are leveraged to generate 3D hydrogels containing lateral gradients in vascular density alongside murine hematopoietic stem cells (HSCs). Regional differences in vascular density can be generated via embossed gradients in cell, matrix, or growth factor density. HSCs co‐cultured alongside vascular gradients reveal spatial patterns of HSC phenotype in response to angiocrine signals. Notably, decreased Akt signaling in high vessel density regions led to increased expansion of lineage‐positive hematopoietic cells. This approach offers a combinatorial tool to rapidly screen a continuum of microenvironments with varying vascular, biophysical, and biochemical cues to reveal the influence of local angiocrine signals on HSC fate.

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  3. Abstract

    Hematopoietic stem cells (HSCs) reside in the bone marrow within niches that provide microenvironmental signals in the form of biophysical cues, bound and diffusible biomolecules, and heterotypic cell–cell interactions that influence HSC fate decisions. This study seeks to inform the development of a synthetic culture platform that promotes ex vivo HSC expansion without exhaustion. A library of methacrylamide‐functionalized gelatin (GelMA) hydrogels is used to explore remodeling and crosstalk from mesenchymal stromal cells (MSCs) on the expansion and quiescence of murine HSCs. The use of a degradable GelMA hydrogel enables MSC‐mediated remodeling, yielding dynamic shifts in the matrix environment over time. An initially low‐diffusivity hydrogel for co‐culture of hematopoietic stem and progenitor cells to MSCs facilitates maintenance of an early progenitor cell population over 7 days. Excitingly, this platform promotes retention of a quiescent HSC population compared to HSC monocultures. These studies reveal MSC‐density–dependent upregulation of MMP‐9 and changes in hydrogel mechanical properties (ΔE= 2.61 ± 0.72) suggesting MSC‐mediated matrix remodeling may contribute to a dynamic culture environment. Herein, a 3D hydrogel is reported for ex vivo HSC culture, in which HSC expansion and quiescence is sensitive to hydrogel properties, MSC co‐culture, and MSC‐mediated hydrogel remodeling.

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  4. Abstract

    The ability of the extracellular matrix (ECM) to instruct progenitor cell differentiation has generated excitement for the development of materials‐based regenerative solutions. Described a nanoparticulate mineralized collagen glycosaminoglycan (MC‐GAG) material capable of inducing in vivo skull regeneration without exogenous growth factors or ex vivo progenitor cell‐priming is described previously. Here, the contribution of titrating stiffness to osteogenicity is evaluated by comparing noncrosslinked (NX‐MC) and crosslinked (MC) forms of MC‐GAG. While both materials are osteogenic, MC demonstrates an increased expression of osteogenic markers and mineralization compared to NX‐MC. Both materials are capable of autogenously activating the canonical BMPR signaling pathway with phosphorylation of Smad1/5. However, unlike NX‐MC, human mesenchymal stem cells cultured on MC demonstrate significant elevations in the major mechanotransduction mediators YAP and TAZ expression, coincident with β‐catenin activation in the canonical Wnt signaling pathway. Inhibition of YAP/TAZ activation reduces osteogenic expression, mineralization, and β‐catenin activation in MC, with less of an effect on NX‐MC. YAP/TAZ inhibition also results in a reciprocal increase in Smad1/5 phosphorylation and BMP2 expression. The results indicate that increasing MC‐GAG stiffness induces osteogenic differentiation via the mechanotransduction mediators YAP/TAZ and the canonical Wnt signaling pathway, whereas the canonical BMPR signaling pathway is activated independent of stiffness.

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