Collagen scaffold arrays for combinatorial screening of biophysical and biochemical regulators of cell behavior.
Ontology highlight
ABSTRACT:
SUBMITTER: Caliari SR
PROVIDER: S-EPMC4282613 | biostudies-literature | 2015 Jan
REPOSITORIES: biostudies-literature
Publications
Collagen scaffold arrays for combinatorial screening of biophysical and biochemical regulators of cell behavior.
Caliari Steven R SR Gonnerman Emily A EA Grier William K WK Weisgerber Daniel W DW Banks Jessica M JM Alsop Aurora J AJ Lee Jae-Sung JS Bailey Ryan C RC Harley Brendan A C BA
Advanced healthcare materials 20140702 1
Arrays of 3D macroporous collagen scaffolds with orthogonal gradations of structural and biomolecular cues are described. Gradient maker technology is applied to create linear biomolecular gradients within microstructurally distinct sections of a single CG scaffold array. The array set up is used to explore cell behaviors including proliferation and regulation of stem cell fate. ...[more]
Similar Datasets
Project description:Here, we have developed a novel method for forming hydrogel arrays using surfaces patterned with differential wettability. Our method for benchtop array formation is suitable for enhanced-throughput, combinatorial screening of biochemical and biophysical cues from chemically defined cell culture substrates. We demonstrated the ability to generate these arrays without the need for liquid handling systems and screened the combinatorial effects of substrate stiffness and immobilized cell adhesion peptide concentration on human mesenchymal stem cell (hMSC) behavior during short-term 2-dimensional cell culture. Regardless of substrate stiffness, hMSC initial cell attachment, spreading, and proliferation were linearly correlated with immobilized CRGDS peptide concentration. Increasing substrate stiffness also resulted in increased hMSC initial cell attachment, spreading, and proliferation; however, examination of the combinatorial effects of CRGDS peptide concentration and substrate stiffness revealed potential interplay between these distinct substrate signals. Maximal hMSC proliferation seen on substrates with either high stiffness or high CRGDS peptide concentration suggests that some baseline level of cytoskeletal tension was required for hMSC proliferation on hydrogel substrates and that multiple substrate signals could be engineered to work in synergy to promote mechanosensing and regulate cell behavior.Statement of significanceOur novel array formation method using surfaces patterned with differential wettability offers the advantages of benchtop array formation for 2-dimensional cell cultures and enhanced-throughput screening without the need for liquid handling systems. Hydrogel arrays formed via our method are suitable for screening the influence of chemical (e.g. cell adhesive ligands) and physical (stiffness, size, shape, and thickness) substrate properties on stem cell behavior. The arrays are also fully compatible with commercially available micro-array add-on systems, which allows for simultaneous control of the insoluble and soluble cell culture environment. This study used hydrogel arrays to demonstrate that synergy between cell adhesion and mechanosensing can be used to regulate hMSC behavior.
| S-EPMC4794413 | biostudies-literature
Project description:Three-dimensional (3D) culture, which can simulate in vivo microenvironments, has been increasingly used to study tumor cell biology. Since most preclinical anti-glioma drug tests still rely on conventional 2D cell culture, we established a collagen scaffold for 3D glioma cell culture. Glioma cells cultured on these 3D scaffolds showed greater degree of dedifferentiation and quiescence than cells in 2D culture. 3D-cultured cells also exhibited enhanced resistance to chemotherapeutic alkylating agents, with a much higher proportion of glioma stem cells and upregulation of O6-methylguanine DNA methyltransferase (MGMT). Importantly, tumor cells in 3D culture showed chemotherapy resistance patterns similar to those observed in glioma patients. Our results suggest that 3D collagen scaffolds are promising in vitro research platforms for screening new anti-glioma therapeutics.
| S-EPMC5302961 | biostudies-literature
Project description:Glaucoma is a degenerative eye disease in which damage to the optic nerve leads to a characteristic loss of vision. The primary risk factor for glaucoma is an increased intraocular pressure that is caused by an imbalance of aqueous humor generation and subsequent drainage through the trabecular meshwork (TM) drainage system. The small size, donor tissue limitations, and high complexity of the TM make it difficult to research the relationship between the TM cells and their immediate environment. Thus, a biomaterial-based approach may be more appropriate for research manipulations and in vitro drug development platforms. In this work, human TM (hTM) cells were cultured on various collagen scaffolds containing different glycosaminoglycans (GAGs) and different pore architectures to better understand how hTM cells respond to changes in their extracellular environment. Cellular response was measured by quantifying cellular proliferation and expression of an important extracellular matrix protein, fibronectin. The pore architecture of the scaffolds was altered using freeze-casting technique to make both large and small pores that were aligned or with a non-aligned random structure. The composition of the scaffolds was altered with the addition of chondroitin sulfate and/or hyaluronic acid. It was found that the hTM cells grown on large pore scaffolds proliferate more than those grown on small pores. There was an increase in the fibronectin expression with the incorporation of GAGs, and its morphology was changed by the underlying pore architecture. This work will help provide an insight into the behavior of hTM cells when introducing changes in their microenvironment.
| S-EPMC8360228 | biostudies-literature
Project description:3D microenvironmental parameters control cell behavior, but can be challenging to investigate over a wide range of conditions. Here, a combinatorial hydrogel platform is developed that uses light-mediated thiol-norbornene chemistry to encapsulate cells within hydrogels with biochemical gradients made by spatially varied light exposure. Specifically, mesenchymal stem cells are photoencapsulated in norbornene-modified hyaluronic acid hydrogels functionalized with gradients (0-5?mM) of peptides that mimic cell-cell or cell-matrix interactions, either as single or orthogonal gradients. Chondrogenesis varied spatially in these hydrogels based on the local biochemical formulation, as indicated by Sox9 and aggrecan expression levels. From 100 combinations investigated, discrete hydrogels are formulated and early gene expression and long-term cartilage-specific matrix production are assayed and found to be consistent with screening predictions. This platform is a scalable, high-throughput technique that enables the screening of the effects of multiple biochemical signals on 3D cell behavior.
| S-EPMC5807520 | biostudies-literature
Project description:The peptide spanning residues 35 to 55 of the protein myelin oligodendrocyte glycoprotein (MOG) has been studied extensively in its role as a key autoantigen in the neuroinflammatory autoimmune disease multiple sclerosis. Rodents and nonhuman primate species immunized with this peptide develop a neuroinflammatory condition called experimental autoimmune encephalomyelitis, often used as a model for multiple sclerosis. Over the last decade, the role of citrullination of this antigen in the disease onset and progression has come under increased scrutiny. We recently reported on the ability of these citrullinated MOG35–55 peptides to aggregate in an amyloid-like fashion, suggesting a new potential pathogenic mechanism underlying this disease. The immunodominant region of MOG is highly conserved between species, with the only difference between the murine and human protein, a polymorphism on position 42, which is serine in mice and proline for humans. Here, we show that the biophysical and biochemical behavior we previously observed for citrullinated murine MOG35–55 is fundamentally different for human and mouse MOG35–55. The citrullinated human peptides do not show amyloid-like behavior under the conditions where the murine peptides do. Moreover, we tested the ability of these peptides to stimulate lymphocytes derived from MOG immunized marmoset monkeys. While the citrullinated murine peptides did not produce a proliferative response, one of the citrullinated human peptides did. We postulate that this unexpected difference is caused by disparate antigen processing. Taken together, our results suggest that further study on the role of citrullination in MOG-induced experimental autoimmune encephalomyelitis is necessary.
| S-EPMC10060747 | biostudies-literature
Project description:Biomaterial strategies for regenerating multitissue structures require unique approaches. One strategy is to design scaffolds so that their local biophysical properties can enhance site-specific effects of an otherwise heterogeneous biomolecular environment. This investigation examined the role of biomaterial physical properties (relative density, mineral content) on the human mesenchymal stem cell phenotype in the presence of mixed soluble signals to drive osteogenesis or chondrogenesis. We tested a series of three-dimensional collagen-glycosaminoglycan scaffolds with properties inspired by extracellular matrix characteristics across the osteotendinous interface (tendon, cartilage, and bone). We found that selective scaffold mineralization induced a depressed chondrogenic response compared with nonmineralized groups as demonstrated by gene expression and histological analyses. Interestingly, the greatest chondrogenic response was found in a higher density, nonmineralized scaffold variant despite increased contraction and cellular condensation in lower density nonmineralized scaffolds. In fact, the lower density scaffolds demonstrated a significantly higher expression of osteogenic transcripts as well as ample mineralization after 21 days of culture. This effect may be due to local stiffening of the scaffold microenvironment as the scaffold contracts, leading to increased cell density, accelerated differentiation, and possible endochondral ossification as evidenced by a transition from a glycosaminoglycan (GAG)-rich milieu to higher mineralization at later culture times. These findings will help shape the design rules for graded biomaterials to regenerate distinct fibrillar, fibrocartilagenous, and mineralized regions of orthopedic interfaces.
| S-EPMC4161190 | biostudies-literature
Project description:Despite great progress in biomaterial design strategies for replacing damaged articular cartilage, prevention of stem cell-derived chondrocyte hypertrophy and resulting inferior tissue formation is still a critical challenge. Here, by using engineered biomaterials and a high-throughput system for screening of combinatorial cues in cartilage microenvironments, we demonstrate that biomaterial cross-linking density that regulates matrix degradation and stiffness-together with defined presentation of growth factors, mechanical stimulation, and arginine-glycine-aspartic acid (RGD) peptides-can guide human mesenchymal stem cell (hMSC) differentiation into articular or hypertrophic cartilage phenotypes. Faster-degrading, soft matrices promoted articular cartilage tissue formation of hMSCs by inducing their proliferation and maturation, while slower-degrading, stiff matrices promoted cells to differentiate into hypertrophic chondrocytes through Yes-associated protein (YAP)-dependent mechanotransduction. in vitro and in vivo chondrogenesis studies also suggest that down-regulation of the Wingless and INT-1 (WNT) signaling pathway is required for better quality articular cartilage-like tissue production.
| S-EPMC7244269 | biostudies-literature
Project description:The nematode Caenorhabditis elegans is an important model organism in genetic research and drug screening because of its relative simplicity, ease of maintenance, amenability to simple genetic manipulation, and relevance to human biology. However, their small size and mobility make nematodes difficult to physically manipulate, particularly with spatial and temporal precision. We have developed a microfluidic device to overcome these challenges and enable fast behavior-based chemical screening in C. elegans. The key components of this easy-to-use device allow rapid loading and housing of C. elegans in a chamber array for chemical screening. A simple two-step loading process enables simultaneous loading of a large number of animals within a few minutes without using any expensive/active off-chip components. In addition, chemicals can be precisely delivered to the worms and exchanged with high temporal precision. To demonstrate this feature and the ability to measure time dependent responses to chemicals, we characterize the transient response of worms exposed to different concentrations of anesthetics. We then use the device to study the effect of chemical signals from hermaphrodite worms on male behavior. The ability of the device to maintain a large number of free moving animals in one field of view over a long period of time permits us to demonstrate an increase in the incidence of a specific behavior in males subjected to worm-conditioned medium. Because our device allows monitoring of a large number of worms with single-animal resolution, we envision that this platform will greatly expedite chemical screening in C. elegans.
| S-EPMC3924777 | biostudies-literature
Project description:Disease models, including in vitro cell culture and animal models, have contributed significantly to developing diagnostics and treatments over the past several decades. The successes of traditional drug screening methods were generally hampered by not adequately mimicking critical in vivo features, such as a 3D microenvironment and dynamic drug diffusion through the extracellular matrix (ECM). To address these issues, we developed a 3D dynamic drug delivery system for cancer drug screening that mimicks drug dissemination through the tumor vasculature and the ECM by creating collagen-embedded microfluidic channels. Using this novel 3D ECM microsystem, we compared viability of tumor pieces with traditionally used 2D methods in response to three different drug combinations. Drug diffusion profiles were evaluated by simulation methods and tested in the 3D ECM microsystem and a 2D 96-well setup. Compared with the 2D control, the 3D ECM microsystem produced reliable data on viability, drug ratios, and combination indeces. This novel approach enables higher throughput and sets the stage for future applications utilizing drug sensitivity predicting algorithms based on dynamic diffusion profiles requiring only minimal patient tissue. Our findings moved drug sensitivity screening closer to clinical implications with a focus on testing combinatorial drug effects, an option often limited by the amount of available patient tissues.
| S-EPMC8172531 | biostudies-literature
Project description:The abnormal Wnt signaling pathway leads to a high expression of β-catenin, which causes several types of cancer, particularly colorectal cancer (CRC). The inhibition of tankyrase (TNKS) activity can reduce cancer cell growth, invasion, and resistance to treatment by blocking the Wnt signaling pathway. A pharmacophore search and pharmacophore docking were performed to identify potential TNKS inhibitors in the training databases. The weighted MM/PBSA binding free energy of the docking model was calculated to rank the databases. The reranked results indicated that 26.98% of TNKS inhibitors that were present in the top 5% of compounds in the database and near an ideal value ranked 28.57%. The National Cancer Institute database was selected for formal virtual screening, and 11 potential TNKS inhibitors were identified. An enzyme-based experiment was performed to demonstrate that of the 11 potential TNKS inhibitors, NSC295092 and NSC319963 had the most potential. Finally, Wnt pathway analysis was performed through a cell-based assay, which indicated that NSC319963 is the most likely TNKS inhibitor (pIC50 = 5.59). The antiproliferation assay demonstrated that NSC319963 can decrease colorectal cancer cell growth; therefore, the proposed method successfully identified a novel TNKS inhibitor that can alleviate CRC.
| S-EPMC8773749 | biostudies-literature