Recent advances in crosslinking chemistry of biomimetic poly(ethylene glycol) hydrogels

The design and application of biomimetic hydrogels have become an important and integral part of modern tissue engineering and regenerative medicine. Many of these hydrogels are prepared from synthetic macromers (e.g., poly(ethylene glycol) or PEG) as they provide high degrees of tunability for matrix crosslinking, degradation, and modification. For a hydrogel to be considered biomimetic, it has to recapitulate key features that are found in the native extracellular matrix, such as the appropriate matrix mechanics and permeability, the ability to sequester and deliver drugs, proteins, and or nucleic acids, as well as the ability to provide receptor-mediated cell-matrix interactions and protease-mediated matrix cleavage. A variety of chemistries have been employed to impart these biomimetic features into hydrogel crosslinking. These chemistries, such as radical-mediated polymerizations, enzyme-mediated crosslinking, bio-orthogonal click reactions, and supramolecular assembly, may be different in their crosslinking mechanisms but are required to be efficient for gel crosslinking and ligand bioconjugation under aqueous reaction conditions. The prepared biomimetic hydrogels should display a diverse array of functionalities and should also be cytocompatible for in vitro cell culture and/or in situ cell encapsulation. The focus of this article is to review recent progress in the crosslinking chemistries of biomimetic hydrogels with a special emphasis on hydrogels crosslinked from poly(ethylene glycol)-based macromers

Photo-click hydrogels prepared from functionalized cyclodextrin and poly(ethylene glycol) for drug delivery and in situ cell encapsulation

Polymers or hydrogels containing modified cyclodextrin (CD) are highly useful in drug delivery applications, as CD is a cytocompatible amphiphilic molecule that can complex with a variety of hydrophobic drugs. Here, we designed modular photoclick thiol-ene hydrogels from derivatives of βCD and poly(ethylene glycol) (PEG), including βCD-allylether (βCD-AE), βCD-thiol (βCD-SH), PEG-thiol (PEGSH), and PEG-norbornene (PEGNB). Two types of CD-PEG hybrid hydrogels were prepared using radical-mediated thiol-ene photoclick reactions. Specifically, thiol-allylether hydrogels were formed by reacting multiarm PEGSH and βCD-AE, and thiol-norbornene hydrogels were formed by cross-linking βCD-SH and multiarm PEGNB. We characterized the properties of these two types of thiol-ene hydrogels, including gelation kinetics, gel fractions, hydrolytic stability, and cytocompatibility. Compared with thiol-allylether hydrogels, thiol-norbornene photoclick reaction formed hydrogels with faster gelation kinetics at equivalent macromer contents. Using curcumin, an anti-inflammatory and anticancer hydrophobic molecule, we demonstrated that CD-cross-linked PEG-based hydrogels, when compared with pure PEG-based hydrogels, afforded higher drug loading efficiency and prolonged delivery in vitro. Cytocompatibility of these CD-cross-linked hydrogels were evaluated by in situ encapsulation of radical sensitive pancreatic MIN6 β-cells. All formulations and cross-linking conditions tested were cytocompatible for cell encapsulation. Furthermore, hydrogels cross-linked by βCD-SH showed enhanced cell proliferation and insulin secretion as compared to gels cross-linked by either dithiothreitol (DTT) or βCD-AE, suggesting the profound impact of both macromer compositions and gelation chemistry on cell fate in chemically cross-linked hydrogels

Application of Salubrinal for Bone Fracture Healing

abstractThe long-term objective of this project is to commercialize a novel synthetic chemical agent, salubrinal, for treatment of bone growth and fracture healing. Bone morphogenetic proteins (BMPs) are clinically administered as growth stimulators for bone fracture healing. However, BMPs are not only expensive, but also stimulate ectopic bone formation and potentially induce cancer. A synthetic chemical agent that permits facile storage and administration could reduce costs, and provide longer shelf-life, and better bone healing outcomes. Currently, no synthetic chemical agents as a stimulator of fracture healing are clinically available. The research team recently identified “salubrinal,” a synthetic chemical agent, as a potential therapeutic stimulator of bone growth and fracture healing. An invention disclosure and a U.S. patent were filed. In this FORCES project, we are examining efficacy of salubrinal using a mouse model of closed tibia fracture. The results strongly indicate that salubrinal can accelerate bone fracture healing

Designer hydrogels: Shedding light on the physical chemistry of the pancreatic cancer microenvironment

Pancreatic ductal adenocarcinoma (PDAC) is currently the third leading cause of cancer mortality in the United States, with a 5-year survival of ∼8%. PDAC is characterized by a dense and hypo-vascularized stroma consisting of proliferating cancer cells, cancer-associated fibroblasts, macrophages and immune cells, as well as excess matrices including collagens, fibronectin, and hyaluronic acid. In addition, PDAC has increased interstitial pressures and a hypoxic/acidic tumor microenvironment (TME) that impedes drug delivery and blocks cancer-directed immune mechanisms. In spite of increasing options in targeted therapy, PDAC has mostly remained treatment recalcitrant. Owing to its critical roles on governing PDAC progression and treatment outcome, TME and its interplay with the cancer cells are increasingly studied. In particular, three-dimensional (3D) hydrogels derived from or inspired by components in the TME are progressively developed. When properly designed, these hydrogels (e.g., Matrigel, collagen gel, hyaluronic acid-based, and semi-synthetic hydrogels) can provide pathophysiologically relevant compositions, conditions, and contexts for supporting PDAC cell fate processes. This review summarizes recent efforts in using 3D hydrogels for fundamental studies on cell-matrix or cell-cell interactions in PDAC

Diffusion Dynamics, Moments, and Distribution of First Passage Time on the Protein-Folding Energy Landscape, with Applications to Single Molecules

We study the dynamics of protein folding via statistical energy-landscape theory. In particular, we concentrate on the local-connectivity case with the folding progress described by the fraction of native conformations. We obtain information for the first passage-time (FPT) distribution and its moments. The results show a dynamic transition temperature below which the FPT distribution develops a power-law tail, a signature of the intermittency phenomena of the folding dynamics. We also discuss the possible application of the results to single-molecule dynamics experiments

Improving Cross-linking of Degradable Thiol-acrylate Hydrogels via Peptide Design

poster abstractHydrogels fabricated from poly (ethylene glycol) (PEG) based macromers are ideal for drug delivery and tissue engineering applications. Recently, a new visible light-mediated photopolymerization scheme was developed to fabricate cytocompatible and degradable poly (ethylene glycol)-diacrylate (PEGDA) hydrogels. Co-polymerization of mono-cysteine peptides (e.g. CRGDS) with PEGDA offers the gels with cell adhesion property. However, this approach causes significant reduction in network crosslinking density, in part due to chain transfer of thiols to acrylates. The goal of the project is to improve the network cross-linking efficiency of this peptide-immobilized PEGDA hydrogel for cell culture. We hypothesized that the incorporation of bi-functional bis-cysteine peptides or silk fibroin will produce hydrogels with enhanced stiffness. The shear moduli of the gels were characterized via oscillatory rheometry in strain-sweep (0.1-5%) mode. Hydrolytic degradation of the gels as a function of time was also evaluated by rheometry. Cytocompatibility of the hydrogel system will be assessed by in situ encapsulation of 3T3 fibroblasts. Cell metabolic activity was determined by Alamar-Blue assay. We found that the bis-cysteine peptide enhanced gel crosslinking, as compared with mono-cysteine peptide. Incorporation of silk fibroin protein also exhibited enhancement in gel stiffness. However, the optimum concentration of incorporated silk fibroin presented an increased shear modulus compared to gels containing only the mono-cysteine peptide. Ongoing work is focused on fine-tuning gel formulations and degradation, as well as on evaluating the cytocompatibility of these visible-light cured thiol-acrylate hydrogels

Orthogonal enzymatic reactions for rapid crosslinking and dynamic tuning of PEG–peptide hydrogels

Stiffening of the extracellular matrix is a hallmark in cancer progression, embryonic development, and wound healing. To mimic this dynamic process, our work explored orthogonal enzymatic reactions capable of modulating the properties of poly(ethylene glycol) (PEG)–peptide hydrogels. A hepta-mutant bacterial transpeptidase sortase A (SrtA7M) was used to ligate two PEG–peptide macromers (i.e., PEG-YLPRTG and NH2-GGGG-PEG) into a primary hydrogel network. The hydrogels were dynamically stiffened using mushroom tyrosinase (MT), which oxidized tyrosine residues into di-tyrosine and led to increased matrix stiffness. After confirming the expression and enhanced catalytic activity of SrtA7M, we investigated the cytocompatibility of the enzymatic reaction with a mouse insulinoma cell line, MIN6. In addition, we altered peptide substrate concentrations and evaluated their influence on primary hydrogel network properties and MT-triggered stiffening. Using a pancreatic cancer cell line, COLO-357, the effect of MT-triggered stiffening on spheroid formation was investigated. We found that cell spheroids formed in hydrogels that were exposed to MT were significantly smaller than spheroids formed without MT incubation, suggesting that matrix stiffening played a crucial role in the sizes of cancer cell spheroids. Through utilizing highly specific and orthogonal enzymatic reactions, this hydrogel platform permits rapid and mild in situ cell encapsulation, as well as dynamic control of matrix stiffness for investigating the role of matrix stiffening on cell fate processes

Chemotaxonomic Analysis of the Venom Composition within the Ant Genus Strumigenys (Hymenoptera, Formicidae) in Taiwan

In Taiwan, the ant genus Strumigenys is represented by 13 species, nine of which being endemic to this island. Classic morphological taxonomy can be complex and may lead to equivoque identification within this group. To clarify subtle species assignments, we investigated the venom composition of five Strumigenys species, using SPME extraction and GC/MS analyses, and searched for a suitable chemical marker. Our results indicate that three out of the five species tested showed enough specificity in their chemical profiles to allow clear differentiation. However, the two remaining species could not be distinguished from each other on the basis of their venom composition. We further assessed the phylogenetic relationships between the five species, analyzing both morphological and chemical characters. Our clusters revealed congruency between some species associations and suggested that the analysis of venom composition may apply, at least partially, to Strumigenys chemosystematics. However, important discrepancies also appeared, signifying that selective pressures for chemical diversification have operated differentially during the speciation and dispersal processes within this genus inTaiwan