Browsing by Author "Nguyen, Han"

Now showing 1 - 5 of 5
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    Dissolvable microgel-templated macroporous hydrogels for controlled cell assembly
    (Elsevier, 2022) Jiang, Zhongliang; Lin, Fang-Yi; Jiang, Kun; Nguyen, Han; Chang, Chun-Yi; Lin, Chien-Chi; Biomedical Engineering, School of Engineering and Technology
    Mesenchymal stem cells (MSCs)-based therapies have been widely used to promote tissue regeneration and to modulate immune/inflammatory response. The therapeutic potential of MSCs can be further improved by forming multi-cellular spheroids. Meanwhile, hydrogels with macroporous structures are advantageous for improving mass transport properties for the cell-laden matrices. Herein, we report the fabrication of MSC-laden macroporous hydrogel scaffolds through incorporating rapidly dissolvable spherical cell-laden microgels. Dissolvable microgels were fabricated by tandem droplet-microfluidics and thiol-norbornene photopolymerization using a novel fast-degrading macromer poly(ethylene glycol)-norbornene-dopamine (PEGNB-Dopa). The cell-laden microgels were subsequently encapsulated within another bulk hydrogel matrix, whose porous structure was generated efficiently by the rapid degradation of the PEGNB-Dopa microgels. The cytocompatibility of this in situ pore-forming approach was demonstrated with multiple cell types. Furthermore, adjusting the stiffness and cell adhesiveness of the bulk hydrogels afforded the formation of solid cell spheroids or hollow spheres. The assembly of solid or hollow MSC spheroids led to differential activation of AKT pathway. Finally, MSCs solid spheroids formed in situ within the macroporous hydrogels exhibited robust secretion of HGF, VEGF-A, IL-6, IL-8, and TIMP-2. In summary, this platform provides an innovative method for forming cell-laden macroporous hydrogels for a variety of future biomedical applications.
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    Dual Functionalization of Gelatin for Orthogonal and Dynamic Hydrogel Cross-Linking
    (American Chemical Society, 2021) Kim, Min Hee; Nguyen, Han; Chang, Chun-Yi; Lin, Chien-Chi; Biomedical Engineering, School of Engineering and Technology
    Gelatin based hydrogels are widely used in biomedical fields owing to its abundance of bioactive motifs that support cell adhesion and matrix remodeling. While inherently bioactive, unmodified gelatin exhibits temperature-dependent rheology and solubilizes at body temperature, making it unstable for three-dimensional (3D) cell culture. Therefore, the addition of chemically reactive motifs is required to render gelatin-based hydrogels with highly controllable crosslinking kinetics and tunable mechanical properties that are critical for 3D cell culture. This article provides a series of methods toward establishing orthogonally crosslinked gelatin-based hydrogels for dynamic 3D cell culture. In particular, we prepared dually functionalized gelatin macromers amenable for sequential, orthogonal covalent crosslinking. Central to this material platform is the synthesis of norbornene-functionalized gelatin (GelNB), which forms covalently crosslinked hydrogels via orthogonal thiol-norbornene click crosslinking. Using GelNB as the starting material, we further detail the methods for synthesizing gelatin macromers susceptible to hydroxyphenylacetic acid (HPA) dimerization (i.e., GelNB-HPA) and hydrazone bonding (i.e., GelNB-CH) for on-demand matrix stiffening. Finally, we outline the protocol for synthesizing a gelatin macromer capable of adjusting hydrogel stress-relaxation via boronate ester bonding (i.e., GelNB-BA). The combinations of these orthogonal chemistries affords a wide range of gelatin based hydrogels as biomimetic matrices in tissue engineering and regenerative medicine applications.
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    Triple click chemistry for crosslinking, stiffening, and annealing of gelatin-based microgels
    (Royal Society of Chemistry, 2024-03-28) Chang, Chun-Yi; Nguyen, Han; Frahm, Ellen; Kolaczyk, Keith; Lin, Chien-Chi; Biomedical Engineering, Purdue School of Engineering and Technology
    Microgels are spherical hydrogels with physicochemical properties ideal for many biomedical applications. For example, microgels can be used as individual carriers for suspension cell culture or jammed/annealed into granular hydrogels with micron-scale pores highly permissive to molecular transport and cell proliferation/migration. Conventionally, laborious optimization processes are often needed to create microgels with different moduli, sizes, and compositions. This work presents a new microgel and granular hydrogel preparation workflow using gelatin-norbornene-carbohydrazide (GelNB-CH). As a gelatin-derived macromer, GelNB-CH presents cell adhesive and degradable motifs while being amenable to three orthogonal click chemistries, namely the thiol-norbornene photo-click reaction, hydrazone bonding, and the inverse electron demand Diels-Alder (iEDDA) click reaction. The thiol-norbornene photo-click reaction (with thiol-bearing crosslinkers) and hydrazone bonding (with aldehyde-bearing crosslinkers) were used to crosslink the microgels and to realize on-demand microgel stiffening, respectively. The tetrazine-norbornene iEDDA click reaction (with tetrazine-bearing crosslinkers) was used to anneal microgels into granular hydrogels. In addition to materials development, we demonstrated the value of the triple-click chemistry granular hydrogels via culturing human mesenchymal stem cells and pancreatic cancer cells.
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    Viscoelastic hydrogels for interrogating pancreatic cancer-stromal cell interactions
    (Elsevier, 2023-02-04) Lin, Fang-Yi; Chang, Chun-Yi; Nguyen, Han; Li, Hudie; Fishel, Melissa L.; Lin, Chien-Chi; Pediatrics, School of Medicine
    The tumor microenvironment (TME) is known to direct cancer cell growth, migration, invasion into the matrix and distant tissues, and to confer drug resistance in cancer cells. While multiple aspects of TME have been studied using in vitro, ex vivo, and in vivo tumor models and engineering tools, the influence of matrix viscoelasticity on pancreatic cancer cells and its associated TME remained largely unexplored. In this contribution, we synthesized a new biomimetic hydrogel with tunable matrix stiffness and stress-relaxation for evaluating the effect of matrix viscoelasticity on pancreatic cancer cell (PCC) behaviors in vitro. Using three simple monomers and Reverse-Addition Fragmentation Chain-Transfer (RAFT) polymerization, we synthesized a new class of phenylboronic acid containing polymers (e.g., poly (OEGA-s-HEAA-s-APBA) or PEHA). Norbornene group was conjugated to HEAA on PEHA via carbic anhydride, affording a new NB and BA dually modified polymer - PEHNBA amenable for orthogonal thiol-norbornene photopolymerization and boronate ester diol complexation. The former provided tunable matrix elasticity, while the latter gave rise to matrix stress-relaxation (or viscoelasticity). The new PEHNBA polymers were shown to be highly cytocompatible for in situ encapsulation of PCCs and cancer-associated fibroblasts (CAFs). Furthermore, we demonstrated that hydrogels with high stress-relaxation promoted spreading of CAFs, which in turns promoted PCC proliferation and spreading in the viscoelastic matrix. Compared with elastic matrix, viscoelastic gels upregulated the secretion of soluble proteins known to promote epithelial-mesenchymal transition (EMT). This study demonstrated the crucial influence of matrix viscoelasticity on pancreatic cancer cell fate and provided an engineered viscoelastic matrix for future studies and applications related to TME.
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    Viscoelastic stiffening of gelatin hydrogels for dynamic culture of pancreatic cancer spheroids
    (Elsevier, 2024) Nguyen, Han; Lin, Chien-Chi; Biomedical Engineering, Purdue School of Engineering and Technology
    The tumor microenvironment (TME) in pancreatic adenocarcinoma (PDAC) is a complex milieu of cellular and non-cellular components. Pancreatic cancer cells (PCC) and cancer-associated fibroblasts (CAF) are two major cell types in PDAC TME, whereas the non-cellular components are enriched with extracellular matrices (ECM) that contribute to high stiffness and fast stress-relaxation. Previous studies have suggested that higher matrix rigidity promoted aggressive phenotypes of tumors, including PDAC. However, the effects of dynamic viscoelastic matrix properties on cancer cell fate remain largely unexplored. The focus of this work was to understand the effects of such dynamic matrix properties on PDAC cell behaviors, particularly in the context of PCC/CAF co-culture. To this end, we engineered gelatin-norbornene (GelNB) based hydrogels with a built-in mechanism for simultaneously increasing matrix elastic modulus and viscoelasticity. Two GelNB-based macromers, namely GelNB-hydroxyphenylacetic acid (GelNB-HPA) and GelNB-boronic acid (GelNB-BA), were modularly mixed and crosslinked with 4-arm poly(ethylene glycol)-thiol (PEG4SH) to form elastic hydrogels. Treating the hybrid hydrogels with tyrosinase not only increased the elastic moduli of the gels (due to HPA dimerization) but also concurrently produced 1,2-diols that formed reversible boronic acid-diol bonding with the BA groups on GelNB-BA. We employed patient-derived CAF and a PCC cell line COLO-357 to demonstrate the effect of increasing matrix stiffness and viscoelasticity on CAF and PCC cell fate. Our results indicated that in the stiffened environment, PCC underwent epithelial-mesenchymal transition. In the co-culture PCC and CAF spheroid, CAF enhanced PCC spreading and stimulated collagen 1 production. Through mRNA-sequencing, we further showed that stiffened matrices, regardless of the degree of stress-relaxation, heightened the malignant phenotype of PDAC cells. STATEMENT OF SIGNIFICANCE: The pancreatic cancer microenvironment is a complex milieu composed of various cell types and extracellular matrices. It has been suggested that stiffer matrices could promote aggressive behavior in pancreatic cancer, but the effect of dynamic stiffening and matrix stress-relaxation on cancer cell fate remains largely undefined. This study aimed to explore the impact of dynamic changes in matrix viscoelasticity on pancreatic ductal adenocarcinoma (PDAC) cell behavior by developing a hydrogel system capable of simultaneously increasing stiffness and stress-relaxation on demand. This is achieved by crosslinking two gelatin-based macromers through orthogonal thiol-norbornene photochemistry and post-gelation stiffening with mushroom tyrosinase. The results revealed that higher matrix stiffness, regardless of the degree of stress relaxation, exacerbated the malignant characteristics of PDAC cells.