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Abstract The biochemical and physical properties of a scaffold can be tailored to elicit specific cellular responses. However, it is challenging to decouple their individual effects on cell‐material interactions. Here, we solvent‐cast 3D printed different ratios of high and low molecular weight (MW) poly(caprolactone) (PCL) to fabricate scaffolds with significantly different stiffnesses without affecting other properties. Ink viscosity was used to match processing conditions between inks and generate scaffolds with the same surface chemistry, crystallinity, filament diameter, and architecture. Increasing the ratio of low MW PCL resulted in a significant decrease in modulus. Scaffold modulus did not affect human mesenchymal stromal cell (hMSC) differentiation under osteogenic conditions. However, hMSC response was significantly affected by scaffold stiffness in chondrogenic media. Low stiffness promoted more stable chondrogenesis whereas high stiffness drove hMSC progression toward hypertrophy. These data illustrate how this versatile platform can be used to independently modify biochemical and physical cues in a single scaffold to synergistically enhance desired cellular response. 

Abstract 3D printing is a popular fabrication technique because of its ability to produce complex architectures. Melt‐based 3D printing is widely used for thermoplastic polymers like poly(caprolactone) (PCL), poly(lactic acid) (PLA), and poly(lactic‐co‐glycolic acid) (PLGA) because of their low processing temperatures. However, traditional melt‐based techniques require processing temperatures and pressures high enough to achieve continuous flow, limiting the type of polymer that can be printed. Solvent‐cast printing (SCP) offers an alternative approach to print a wider range of polymers. Polymers are dissolved in a volatile solvent that evaporates during deposition to produce a solid polymer filament. SCP, therefore, requires optimizing polymer concentration in the ink, print pressure, and print speed to achieve desired print fidelity. Here, capillary flow analysis shows how print pressure affects the process‐apparent viscosity of PCL, PLA, and PLGA inks. Ink viscosity is also measured using rheology, which is used to link a specific ink viscosity to a predicted set of print pressure and print speed for all three polymers. These results demonstrate how this approach can be used to accelerate optimization by significantly reducing the number of parameter combinations. This strategy can be applied to other polymers to expand the library of polymers printable with SCP. 

Osteoarthritis (OA) is the most prevalent musculoskeletal disorder and a leading cause of disability globally. Although many efforts have been made to treat this condition, current tissue engineering (TE) and regenerative medicine strategies fail to address the inflammatory tissue environment that leads to the rapid progression of the disease and prevents cartilage tissue formation. First, this review addresses in detail the current antiinflammatory therapies for OA with a special emphasis on pharmacological approaches, gene therapy, and mesenchymal stromal cell (MSC) intra-articular administration, and discusses the reasons behind the limited clinical success of these approaches at enabling cartilage regeneration. Then, we analyze the state-of-the-art TE strategies and how they can be improved by incorporating immunomodulatory capabilities such as the optimization of biomaterial composition, porosity and geometry, and the loading of anti-inflammatory molecules within an engineered structure. Finally, the review discusses the future directions for the new generation of TE strategies for OA treatment, specifically focusing on the spatiotemporal modulation of anti-inflammatory agent presentation to allow for tailored patient-specific therapies. 

Functional repair of osteochondral (OC) tissue remains challenging because the transition from bone to cartilage presents gradients in biochemical and physical properties necessary for joint function. Osteochondral regeneration requires strategies that restore the spatial composition and organization found in the native tissue. Several biomaterial approaches have been developed to guide chondrogenic and osteogenic differentiation of human mesenchymal stem cells (hMSCs). These strategies can be combined with 3D printing, which has emerged as a useful tool to produce tunable, continuous scaffolds functionalized with bioactive cues. However, functionalization often includes one or more post-fabrication processing steps, which can lead to unwanted side effects and often produce biomaterials with homogeneously distributed chemistries. To address these challenges, surface functionalization can be achieved in a single step by solvent-cast 3D printing peptide-functionalized polymers. Peptide-poly(caprolactone) (PCL) conjugates were synthesized bearing hyaluronic acid (HA)-binding (HAbind–PCL) or mineralizing (E3–PCL) peptides, which have been shown to promote hMSC chondrogenesis or osteogenesis, respectively. This 3D printing strategy enables unprecedented control of surface peptide presentation and spatial organization within a continuous construct. Scaffolds presenting both cartilage-promoting and bone-promoting peptides had a synergistic effect that enhanced hMSC chondrogenic and osteogenic differentiation in the absence of differentiation factors compared to scaffolds without peptides or only one peptide. Furthermore, multi-peptide organization significantly influenced hMSC response. Scaffolds presenting HAbind and E3 peptides in discrete opposing zones promoted hMSC osteogenic behavior. In contrast, presenting both peptides homogeneously throughout the scaffolds drove hMSC differentiation towards a mixed population of articular and hypertrophic chondrocytes. These significant results indicated that hMSC behavior was driven by dual-peptide presentation and organization. The downstream potential of this platform is the ability to fabricate biomaterials with spatially controlled biochemical cues to guide functional tissue regeneration without the need for differentiation factors. 

Three-dimensional (3D) printing of biodegradable polymers has rapidly become a popular approach to create scaffolds for tissue engineering. This technique enables fabrication of complex architectures and layer-by-layer spatial control of multiple components with high resolution. The resulting scaffolds can also present distinct chemical groups or bioactive cues on the surface to guide cell behavior. However, surface functionalization often includes one or more post-fabrication processing steps, which typically produce biomaterials with homogeneously distributed chemistries that fail to mimic the biochemical organization found in native tissues. As an alternative, our laboratory developed a novel method that combines solvent-cast 3D printing with peptide-polymer conjugates to spatially present multiple biochemical cues in a single scaffold without requiring post-fabrication modification. Here, we describe a detailed, stepwise protocol to fabricate peptide-functionalized scaffolds and characterize their physical architecture and biochemical spatial organization. We used these 3D-printed scaffolds to direct human mesenchymal stem cell differentiation and osteochondral tissue formation by controlling the spatial presentation of cartilage-promoting and bone-promoting peptides. This protocol also describes how to seed scaffolds and evaluate matrix deposition driven by peptide organization. 

Three-dimensional (3D) printing of biodegradable polymers has rapidly become a popular approach to create scaffolds for tissue engineering. This technique enables fabrication of complex architectures and layer-by-layer spatial control of multiple components with high resolution. The resulting scaffolds can also present distinct chemical groups or bioactive cues on the surface to guide cell behavior. However, surface functionalization often includes one or more post-fabrication processing steps, which typically produce biomaterials with homogeneously distributed chemistries that fail to mimic the biochemical organization found in native tissues. As an alternative, our laboratory developed a novel method that combines solvent-cast 3D printing with peptide-polymer conjugates to spatially present multiple biochemical cues in a single scaffold without requiring post-fabrication modification. Here, we describe a detailed, stepwise protocol to fabricate peptide-functionalized scaffolds and characterize their physical architecture and biochemical spatial organization. We used these 3D-printed scaffolds to direct human mesenchymal stem cell differentiation and osteochondral tissue formation by controlling the spatial presentation of cartilage-promoting and bone-promoting peptides. This protocol also describes how to seed scaffolds and evaluate matrix deposition driven by peptide organization.