Influence of Polymer Chemistry on Cytokine Secretion from Polarized Macrophages

Of central importance to tissue engineering and drug delivery is identifying polymer parameters that increase or decrease specific cytokines in response to biomaterials. In this study, we have interrogated the effects of material descriptors and material characteristics on pro-inflammatory, pro-angiogenic, and naïve macrophages using polymeric particles (∼600 nm), functionalized with 13 different moieties. We characterized tumor necrosis factor-α (TNF-α) and interleukin-10 (IL-10) secretion for the three macrophage populations and used the quantitative structure–activity relationship method (QSAR) to accurately predict cytokine secretion for the different macrophage phenotypes. The findings presented here demonstrate that altering cellular responses to polymers can be achieved through exploiting material parameters. For pro-inflammatory macrophages, polarity and the ability to hydrogen bond appear to significantly impact TNF-α secretion while charge impacted pro-angiogenic macrophages. Naïve cells were impacted by charge in a similar manner as the pro-angiogenic cells; however, hydrophilicity also increased TNF-α secretion in these cells. For IL-10 secretion, hydrogen bonding was very negatively correlated with pro-inflammatory cells, whereas it was positively correlated with pro-angiogenic cells

How crosslinking Mechanisms of Methacrylated Gellan Gum Hydrogels Alter Macrophage Phenotype

In tissue engineering scaffolds, macrophages play a critical role in determining the host response to implanted biomaterials. Macrophage phenotype is dynamic throughout the host response, and a balance of phenotypes is essential for timely progression from injury to proper wound healing. Therefore, it is important to predict how materials will modulate the response of macrophages. In this study, we investigated the effect of methacrylated gellan gum hydrogels on macrophage phenotype and proliferation with the ultimate goal of improving rational design of biomedical implants. Naïve, along with classically and alternatively activated RAW 264.7 macrophages were seeded on methacrylated gellan gum hydrogels that were fabricated with different thiol-ene ratios and crosslinking mechanisms. Live/dead assays showed that all hydrogels supported cell attachment and proliferation. Stiffer substrates enhanced anti-inflammatory production of nitrites from both naïve and classically activated macrophages compared to the softer substrates. Moreover, arginine and CD206 expression – markers for alternatively activated macrophages – were inhibited by higher thiol content. Introducing ionic crosslinks using calcium did not influence the proliferation or polarization for any of the three macrophage phenotypes. Our results suggest that the macrophage phenotype shift from M1 to M2 is controlled by the different crosslinking mechanisms, physical properties, and the chemistry of methacrylated gellan gum hydrogels

Investigating the Synergistic Effects of Combined Modified Alginates on Macrophage Phenotype

Understanding macrophage responses to biomaterials is crucial to the success of implanted medical devices, tissue engineering scaffolds, and drug delivery vehicles. Cellular responses to materials may depend synergistically on multiple surface chemistries, due to the polyvalent nature of cell–ligand interactions. Previous work in our lab found that different surface functionalities of chemically modified alginate could sway macrophage phenotype toward either the pro-inflammatory or pro-angiogenic phenotype. Using these findings, this research aims to understand the relationship between combined material surface chemistries and macrophage phenotype. Tumor necrosis factor-α (TNF-α) secretion, nitrite production, and arginase activity were measured and used to determine the ability of the materials to alter macrophage phenotype. Cooperative relationships between pairwise modifications of alginate were determined by calculating synergy values for the aforementioned molecules. Several materials appeared to improve M1 to M2 macrophage reprogramming capabilities, giving valuable insight into the complexity of surface chemistries needed for optimal incorporation and survival of implanted biomaterials

Effects of arginine-based surface modifications of liposomes for drug delivery in Caco-2 colon carcinoma cells

Liposomal encapsulation of chemotherapeutics improves circulation time and decreases off-target effects through the enhanced permeability and retention (EPR) effect. Improving the efficacy of these drug carriers through surface modification could benefit patients. A library of arginine derivatives was conjugated to liposomes through carbodiimide chemistry. Both unmodified and modified liposomes were loaded with doxorubicin and exposed to Caco-2 colon carcinoma cells to measure the half maximal inhibitory concentration (IC50). Most of the modifications improved the toxicity of doxorubicin. Principal component analysis (PCA) was used to uncover correlations between physicochemical properties (lipophilicity (log P), partition coefficient (log D), number of hydrogen bond donors, number of hydrogen bond acceptors, freely rotating bonds, surface tension, polarization surface area, and isoelectric point) and the IC50 of encapsulated doxorubicin. Generalized rules for improved toxicity were also developed, which stated that improved drug carriers should have at least 4 hydrogen bond donors, between 4 and 6 freely rotating bonds, an isoelectric point above 5.5, and a log P between -2 and -1. Using these relationships along with previously obtained correlations for macrophages, selective targeting and the understanding of how to rationally design such drug carriers can be improved

Identifying Factors of Microparticles Modified with Arginine Derivatives That Induce Phenotypic Shifts in Macrophages

Macrophages are key players in the progression of many diseases, ranging from rheumatoid arthritis to cancer. Drug delivery systems have the potential not only to transport payloads to diseased tissue but also to influence cell behavior. Here, poly(N-isopropylacrylamide-co-acrylic acid) (pNIPAm-co-AAc) microparticles were modified with 14 different arginine derivatives. These particles were then incubated with interleukin-4 or lipopolysaccharide-stimulated macrophages or naïve macrophages (RAW 264.7). The phenotypic state of the macrophages was assessed by measuring arginase activity, tumor necrosis factor-α (TNF-α) secretion, and nitrite production. Partial least-squares analysis revealed material properties and descriptors that shifted the macrophage phenotype for the three cell conditions in this study. Material descriptors relating to secondary bonding were suggested to play a role in shifting phenotypes in all three macrophage culture conditions. These findings suggest that macrophage responses could be altered through drug delivery vehicles, and this method could be employed to assist in screening potential candidates

Core-shell Hydrogel Microcapsules for Improved Islets Encapsulation

Islets microencapsulation holds great promise to treat type 1 diabetes. Currently used alginate microcapsules often have islets protruding outside capsules, leading to inadequate immuno-protection. A novel design of microcapsules with core–shell structures using a two-fluid co-axial electro-jetting is reported. Improved encapsulation and diabetes correction is achieved in a single step by simply confining the islets in the core region of the capsules

Transfer of assembled collagen fibrils to flexible substrates for mechanically tunable contact guidance cues

Contact guidance or bidirectional migration along aligned fibers modulates many physiological and pathological processes such as wound healing and cancer invasion. Aligned 2D collagen fibrils epitaxially grown on mica substrates replicate many features of contact guidance seen in aligned 3D collagen fiber networks. However, these 2D collagen self-assembled substrates are difficult to image through, do not have known or tunable mechanical properties and cells degrade and mechanically detach collagen fibrils from the surface, leading to an inability to assess contact guidance over long times. Here, we describe the transfer of aligned collagen fibrils from mica substrates to three different functionalized target substrates: glass, polydimethylsiloxane (PDMS) and polyacrylamide (PA). Aligned collagen fibrils can be efficiently transferred to all three substrates. This transfer resulted in substrates that were to varying degrees resistant to cell-mediated collagen fibril deformation that resulted in detachment of the collagen fibril field, allowing for contact guidance to be observed over longer time periods. On these transferred substrates, cell speed is lowest on softer contact guidance cues for both MDA-MB-231 and MTLn3 cells. Intermediate stiffness resulted in the fastest migration. MTLn3 cell directionality was low on soft contact guidance cues, whereas MDA-MB-231 cell directionality marginally increased. It appears that the stiffness of the contact guidance cue regulates contact guidance differently between cell types. The development of this collagen fibril transfer method allows for the attachment of aligned collagen fibrils on substrates, particularly flexible substrates, that do not normally promote aligned collagen fibril growth, increasing the utility of this collagen self-assembly system for the fundamental examination of mechanical regulation of contact guidance