In situ collagen assembly for integrating microfabricated three-dimensional cell-seeded matrices

Gillette, Brian M.; Jensen, Jacob A.; Tang, Beixian; Yang, Genevieve; Bazargan-Lari, Ardalan; Zhong, Ming; Sia, Samuel K.

doi:10.1038/nmat2203

Cited by 207 publications

(173 citation statements)

References 31 publications

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“…Using CF NB ðGPOÞ 9 , we were also able to photo-pattern collagen and gelatin films demonstrating the potential for local immobilization of CMP conjugated bioactive components (10,22,23) to collagen-containing tissue engineering scaffolds (Fig. 2D) (24,25).…”

Section: Resultsmentioning

confidence: 96%

Targeting collagen strands by photo-triggered triple-helix hybridization

Foss

Summerfield

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

169

244

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Collagen remodeling is an integral part of tissue development, maintenance, and regeneration, but excessive remodeling is associated with various pathologic conditions. The ability to target collagens undergoing remodeling could lead to new diagnostics and therapeutics as well as applications in regenerative medicine; however, such collagens are often degraded and denatured, making them difficult to target with conventional approaches. Here, we present caged collagen mimetic peptides (CMPs) that can be phototriggered to fold into triple helix and bind to collagens denatured by heat or by matrix metalloproteinase (MMP) digestion. Peptidebinding assays indicate that the binding is primarily driven by stereo-selective triple-helical hybridization between monomeric CMPs of high triple-helical propensity and denatured collagen strands. Photo-triggered hybridization allows specific staining of collagen chains in protein gels as well as photo-patterning of collagen and gelatin substrates. In vivo experiments demonstrate that systemically delivered CMPs can bind to collagens in bones, as well as prominently in articular cartilages and tumors characterized by high MMP activity. We further show that CMP-based probes can detect abnormal bone growth activity in a mouse model of Marfan syndrome. This is an entirely new way to target the microenvironment of abnormal tissues and could lead to new opportunities for management of numerous pathologic conditions associated with collagen remodeling and high MMP activity.A s the most abundant protein in mammals, collagens play a crucial role in tissue development and regeneration, and their structural or metabolic abnormalities are associated with debilitating genetic diseases and various pathologic conditions. Although collagen remodeling occurs during development and normal tissue maintenance, particularly for renewing tissues (e.g., bones), excess remodeling activity is commonly seen in tumors, arthritis, and many other chronic wounds. During collagen remodeling, large portions of collagens are degraded and denatured by proteolytic enzymes, which can be explored for diagnostic and therapeutic purposes. Since unstructured proteins are not ideal targets for rational drug design, library approaches have been employed to develop monoclonal antibody (1, 2) and peptide probes (3) that specifically bind to cryptic sites in collagen strands that become exposed when denatured. However, these probes suffer from poor pharmacokinetics (4), and/or low specificity, and binding affinity (5).We envisioned that triple helix, the hallmark structural feature of collagen, could provide a unique targeting mechanism for the denatured collagens. The triple helix is nearly exclusively seen in collagens except as small subdomains in a few noncollagen proteins (6). Considering its striking structural similarity to the DNA double helix in terms of multiplex formation by periodic interchain hydrogen bonds along the polymer backbone (6), we thought that a small peptide sequence with strong triple-helix prope...

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Section: Resultsmentioning

confidence: 96%

Targeting collagen strands by photo-triggered triple-helix hybridization

Foss

Summerfield

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

169

244

View full text Add to dashboard Cite

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“…14 This smaller, rounded morphology phenomenon was been observed with other 3D environments, including Matrigel, 13,14 collagen, 35,36 and PEG-L-PA thermogel [a poly-(ethylene glycol)-b-poly (Lalanina) gel]. 37 Investigations by Kumar and others [38][39][40] used different features of the extracellular environment to observe changes in cell morphology and related cell behaviors.…”

Section: D Effects Of Tgf-b1mentioning

confidence: 82%

Response of Fibroblasts to Transforming Growth Factor-β1 on Two-Dimensional and in Three-Dimensional Hyaluronan Hydrogels

Chen

Thibeault

2012

Tissue Engineering Part A

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Transforming growth factor-b1 (TGF-b1), an important cytokine with multiple functions, is secreted during wound healing. Previous studies have utilized two-dimensional (2D) cell culture to elucidate the functions of TGF-b1; however, 2D culture does not represent the complex three-dimensional (3D) in vivo environment. Using a synthetic hyaluronan (HA) extracellular matrix (ECM) hydrogel, we investigated the effect of TGF-b1 on fibroblasts cultured in three conditions-on tissue culture polystyrene (TCP), on HA (2D), and in HA (3D). After TGF-b1 treatment (0.1 to 20 ng/mL), morphological features and ECM regulation were analyzed by immunocytochemistry, Western blot, quantitative polymerase chain reaction, and zymogram assays. On TCP, cells showed the typical spindle shape with strong alpha smooth muscle actin (a-SMA) staining of cytoplasmic myofilaments along the cell axes after TGF-b1 treatment; on HA (2D), spindle-shape cells showed little a-SMA staining; in HA (3D), cells were smaller and rounded with less a-SMA deposition. The a-SMA gene and protein expression on TCP were significantly upregulated by TGF-b1, but TGF-b1 did not induce a-SMA expression in the presence of HA (both 2D and 3D). 3D HA culture significantly downregulated collagen I, III, and fibronectin expression, increased matrix metalloproteinase 1 and 2 (MMP1/MMP2) activity, upregulated MMP1 mRNA and downregulated TIMP3 mRNA expression. This study suggested that exogenous HA, particularly in 3D culture, appears to suppress ECM production, enhances ECM degradation and remodeling, and inhibits myofibroblast differentiation without decreasing TGF-b receptor expression.

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“…11,12 Molecular self-assembly is a useful tool to fabricate supramolecular architectures, and many biomolecules (e.g., peptides and proteins) have been reported to selfassemble into hydrogels with a nanofibrous structure. [13][14][15] Although molecular self-assembly is a fairly new method for the formation of nano-structured scaffolds, the mechanical property of the self-assembled hydrogels is very low, which limits its applications to tissue regeneration. Using a thermally induced phase separation combined with particle leaching technique, we recently prepared 3D nanofibrous gelatin (NF-Gelatin) scaffolds with high surface areas, high porosities, well-interconnected macropores, and nanofibrous pore wall structures.…”

Section: Introductionmentioning

confidence: 99%