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Review
. 2015 Apr 15;34(8):987-1008.
doi: 10.15252/embj.201490756. Epub 2015 Mar 12.

Application of Biomaterials to Advance Induced Pluripotent Stem Cell Research and Therapy

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Free PMC article
Review

Application of Biomaterials to Advance Induced Pluripotent Stem Cell Research and Therapy

Zhixiang Tong et al. EMBO J. .
Free PMC article

Abstract

Derived from any somatic cell type and possessing unlimited self-renewal and differentiation potential, induced pluripotent stem cells (iPSCs) are poised to revolutionize stem cell biology and regenerative medicine research, bringing unprecedented opportunities for treating debilitating human diseases. To overcome the limitations associated with safety, efficiency, and scalability of traditional iPSC derivation, expansion, and differentiation protocols, biomaterials have recently been considered. Beyond addressing these limitations, the integration of biomaterials with existing iPSC culture platforms could offer additional opportunities to better probe the biology and control the behavior of iPSCs or their progeny in vitro and in vivo. Herein, we discuss the impact of biomaterials on the iPSC field, from derivation to tissue regeneration and modeling. Although still exploratory, we envision the emerging combination of biomaterials and iPSCs will be critical in the successful application of iPSCs and their progeny for research and clinical translation.

Keywords: biomaterials; disease modeling; expansion; induced pluripotent stem cells; reprogramming.

Figures

Figure 1
Figure 1
Overview of biomaterial-based strategies for enhancing the safety and efficiency of existing iPSC technologies and to better probe iPSC biology and control cell fate in vitro and in vivo Biomaterials may be employed to facilitate all steps of iPSC production and consequently may help address pressing limitations of current derivation, expansion, and differentiation protocols. Traditional viral iPSC reprogramming methods, though efficient, are concerning due to insertional mutagenesis. Conversely, non-integrating iPSC reprogramming methods have lower efficiencies and require elaborate experimental procedures. To overcome the concerns associated with traditional iPSC reprogramming methods, biomaterial-based nano-/microparticles can be used to control the release kinetics of reprogramming factors to potentially avoid viral insertion, increase efficiency, and introduce simpler and less labor-intensive approaches for reprogramming. Furthermore, these nano-/microparticles can also be used to deliver soluble factors and small molecules for the expansion and differentiation of iPSCs (Corradetti et al, 2012). In parallel, biocompatible synthetic substrates can be engineered with patterned physicochemical cues and functionalized with surface-tethered factors to emulate native components of stem cell niches (Watt & Huck, 2013). Making use of biomaterial-based nano-/microparticles and biocompatible synthetic substrates can improve the scalability of traditional expansion and differentiation protocols because they can be reproducibly synthesized on large scales (as they are chemically defined) and at relatively low costs. The reduction in cost and labor will be key for the large-scale production of iPSCs and their progeny. Given the costs associated with the initial development and manufacturing of biomaterials, the cost of biomaterial-based iPSC production could be higher than that of traditional protocols at the early stage. However, in the long run, the application of biomaterials could render the iPSC production and differentiation processes more efficient (e.g. reducing the required concentrations of reprogramming factors via controlling their spatiotemporal presentation). We envision that the overall cost of biomaterial-based protocols will be significantly lower than that of traditional protocols.
Figure 2
Figure 2
Biomaterial-based approaches for improved iPSC reprogramming (A) Well-defined, biomaterial-based micro-/nanoparticles can be formulated and engineered to load multiple reprogramming factors (e.g. Sox2, Oct4, and Klf4). The controlled distribution of different factors—on the surface of the particle (factor a) and entrapped in the particle (factor b)—can be readily achieved during the particle formulation process, and an additional factor (c), if required, can be loaded onto the particle via a stimuli-responsive linker. Given the biodegradability of chosen biomaterials, the varied distribution of multiple reprogramming factors (surface adsorption versus homogeneous encapsulation) and the degradation characteristics of the carrier particle dictate the spatiotemporally controlled release profiles of different factors such as sustained, zero-order release (curve b) and initial burst release followed by slower release or maintenance dose (curve a). Meanwhile, the surface-immobilized stimuli-responsive linker can be cleaved in response to environmental triggers, for example, pH and temperature, to release factor c on demand (e.g. pulsed release every other day, curve c). (B) Nanoparticle-based, smart, artificial transcription factor that is surface-functionalized with nuclear-targeting sequence, DNA-binding domains, and activators for the relevant transcription factors, enabling efficient nuclear localization and effective gene regulation. (C) The biomaterial substrate can be engineered with specific surface anisotropy or microgroove features, which in turn controls cell morphology and, as a result, mediates the epigenetic regulation and cellular reprogramming. (D) Reprogramming factor-laden biomaterials can be delivered into different intracellular loci via multiple engineering approaches, therefore efficiently modulating cell phenotype from inside-out. For example, high-throughput microfluidic technology can be employed to rapidly generate transient membrane disruption on the cells, enabling efficient intracellular localization of phenotype-altering agents without significantly impairing the target cells. Alternatively, this can be achieved via engineering methods such as microinjection or electroporation. Figure partially adapted with permission from Xu et al (2013b) and Patel et al (2014).
Figure 3
Figure 3
Emerging applications of biomaterial-based targeted modulation of cell phenotype and gene regulation with potential application for reprogramming somatic cells (A) Confocal microscopy shows the intracellular localization of phenotype-altering agent-doped PLGA MPs in mesenchymal stem cells that can release agents for several weeks following internalization (A1) a mesenchymal stem cell, (A2) a MIN6 beta cell, and (A3) a RAW 264.7 macrophage. This robust particle platform could potentially serve as an intracellular depot for sustained presentation of reprogramming factors to achieve efficient iPSC derivation from multiple somatic cell types. Scale bars, 10 μm. Green (DiO stain), membrane; red (rhodamine 6 g), particles; blue (Hoechst), nuclei; Adapted with permission from Ankrum et al (2014b). (B) One potential biomaterial strategy for controlled regulation of gene expression is nanoparticle-based artificial transcription factors (NanoScript). This platform could be potentially adopted for the activation or expression of pluripotency-associated genes for improved iPSC derivation. B1: NanoScript is devised to emulate the structure and function of TFs by assembling the principle components, DBD, AD, and NLS, onto a single 10-nm gold nanoparticle via molecular linkers. This design enables the penetration through plasma membrane and entrance into the nuclear membrane through NLS–nuclear receptor coupling. NanoScript interacts with DNA and triggers transcriptional activity leading to desired gene regulation. B2: transmission electron microscopy (TEM) micrograph demonstrates the localization of NanoScript clusters within the nucleus (scale bar = 200 nm), with the inset showing individual nanoparticles (scale bar, 100 nm). Adapted with permission from Patel et al (2014).
Figure 4
Figure 4
The development of PSC/iPSC expansion substrates: advancing from a complex, chemically undefined, feeder layer-based system to simple, synthetic polymeric substrates with improved efficiency, scalability, and reproducibility (A) MEF feeder layers support PSC adhesion and self-renewal via their specific secretome contents. (B) ECM-coated substrates composed of an undefined mixture of ECM proteins such as Matrigel™. (C) Surface-tethered functional epitopes derived from ECM components such as E-cadherin and vitronectin (VN)-derived peptides [e.g. heparin-binding peptide, GKKQRFRHRNRKG (Klim et al, 2010)]. (D) Synthetic, polymeric substrates support the attachment and self-renewal of iPSCs via interface-mediated adsorption of essential adhesive ECM components from the culture medium. Examples for such substrates include the following: (1) ultraviolet-/ozone-modified TCPS; (2) poly(methyl vinyl ether-alt-maleic anhydride); (3) poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl)ammonium hydroxide] (PMEDSAH). Figure adapted and modified with permission from Celiz et al (2014).
Figure 5
Figure 5
High-throughput biomaterial arrays enable simultaneous evaluation of multiple cell–matrix and cell–cell interactions to identify conditions for efficient expansion and controlled differentiation of iPSCs or their progeny (A) Multiple cellular microenvironmental cues and their combinations can be engineered and their interactions with iPSCs-/iPSC-derived progeny can be evaluated in a combinatorial manner on a single arrayed substrate. Microenvironmental cues can be presented to cells via (a) a coculture of multiple cellular components mimicking native cell–cell interactions; (b) substrate stiffness; (c) substrate-induced topographic cues generated by micro-/nanopatterning; (d) spatiotemporally controlled presentation of soluble factors; (e) tethered bioactive factors via substrate surface functionalization. (B) In a relevant example, single Oct4-GFP-positive human ESCs were plated onto the polymer arrays with 496 polymer combinations. Individual cells were initially seeded and attached at a near-clonal density within each spot (white arrowheads in the day 1 panel). Cell response was varied from days 1 to 7 on spots coated with three different polymers (two repeats of each), as shown by the staining of cell nuclei (Hoechst blue) and the level of GFP expression. The 16E-30% polymer was not supportive of cell attachment or survival, whereas the 6F-30% polymer exhibited moderate support for cell growth but did not maintain pluripotency. A hit polymer, the “9” homopolymer, supported robust proliferation of ESCs without differentiation, which was further confirmed by the immunostaining of pluripotency markers Oct4 (green) and SSEA4 (red). (C) In another relevant example, murine ESCs were placed on a combinatorial ECM array and stained with X-gal (blue area indicating the activity of a β-galactosidase reporter fused into a fetal liver-specific gene Ankrd17) after 3 days of culture. The combination of collagen I (C1), collagen III (C3), laminin (L), and fibronectin (Fn) (top red box) induced elevated reporter activity compared to cells on C3 + L (bottom gray box). Scale bar, 250 μm; inset scale bar, 50 μm. The right bar graph depicts the quantification of “blue” image area on each spot of the ECM mixtures, with specific compositions illustrated by the color legend (C4: collagen IV). The cells cultured on matrix mixture C1 + C3 + L + FN elicited ∼27-fold higher Ankrd17 reporter activity than that of the cultures on C3 + L. Error bars, s.e.m. (n = duplicate spots). (B) and (C) were adapted with permission from Flaim et al (2005) and Mei et al (2010), respectively.

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