Assembly of FN-silk with laminin-521 to integrate hPSCs into a three-dimensional culture for neural differentiation

Åstrand, Carolina; Chotteau, Véronique; Falk, Anna; Hedhammar, My

doi:10.1039/c9bm01624d

Cited by 10 publications

(13 citation statements)

References 38 publications

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“…Generation and characterization of silk-bioengineered VM organoids. In an attempt to create more homogeneous VM organoids and further reduce organoid-to-organoid variability, we evaluated a biomaterial made of recombinant spider-silk protein 43,44 that can self-assemble into a biocompatible cell scaffold. Silk scaffolds in the form of a network of microfiber solution were obtained by placing a 20 μl droplet at the bottom center of a hydrophobic well and then introducing air bubbles by repeatedly pipetting air into the droplet (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids

et al. 2021

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Three-dimensional brain organoids have emerged as a valuable model system for studies of human brain development and pathology. Here we establish a midbrain organoid culture system to study the developmental trajectory from pluripotent stem cells to mature dopamine neurons. Using single cell RNA sequencing, we identify the presence of three molecularly distinct subtypes of human dopamine neurons with high similarity to those in developing and adult human midbrain. However, despite significant advancements in the field, the use of brain organoids can be limited by issues of reproducibility and incomplete maturation which was also observed in this study. We therefore designed bioengineered ventral midbrain organoids supported by recombinant spider-silk microfibers functionalized with full-length human laminin. We show that silk organoids reproduce key molecular aspects of dopamine neurogenesis and reduce inter-organoid variability in terms of cell type composition and dopamine neuron formation.

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Section: Resultsmentioning

confidence: 99%

“…The FN-silk/LN-111 mixture was incubated 10 min at RT before usage. About 10 µM Y-27632 dihydrochloride was added 44 . Air bubbles were introduced into the droplet by pipetting up and down (10-15 strokes), creating a dense foam.…”

Section: Methodsmentioning

confidence: 99%

Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids

et al. 2021

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“…[29,[37][38][39] Because of the critical role of LNs in neural tissue development, [40] several different strategies have been developed to incorporate full-size LN in engineered ECM mimicking hydrogels to mimic the native 3D microenvironment better. Whereas affinity-based interactions [41] or physical trapping of LN in the hydrogel network [42] reduce the risk of interfering with LN structure and function, the LN gradually dissociates from the hydrogels over time. Covalent conjugation can result in more efficient retention of LN in the hydrogels [43,44] but can compromise LN function if not carefully optimized.…”

Section: Introductionmentioning

confidence: 99%

Bioorthogonally Cross-Linked Hyaluronan-Laminin Hydrogels for 3D Neuronal Cell Culture and Biofabrication

Jury

Matthiesen

Boroojeni

et al. 2021

Preprint

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Laminins (LNs) are key components in the extracellular matrix of neuronal tissues in the developing brain and neural stem cell niches. LN-presenting hydrogels can provide a biologically relevant matrix for the 3D culture of neurons towards development of advanced tissue models and cell-based therapies for the treatment of neurological disorders. Biologically derived hydrogels are rich in fragmented LN and are poorly defined concerning composition, which hampers clinical translation. Engineered hydrogels require elaborate and often cytotoxic chemistries for cross-linking and LN conjugation and provide limited possibilities to tailor the properties of the materials. Here we show a modular hydrogel system for neural 3D cell culture, based on hyaluronan (HA) and poly(ethylene glycol) (PEG), that is cross-linked and functionalized with human recombinant LN 521 using bioorthogonal copper-free click chemistry. Encapsulated human neuroblastoma cells demonstrate high viability and grow into spheroids. Neuroepithelial stem cells (lt-NES) cultured in the hydrogels can undergo spontaneous differentiation to neural fate and demonstrate significantly higher viability than cells cultured without LN. The hydrogels further support the structural integrity of 3D bioprinted structures and maintain high viability of syringe extruded lt-NES, which can facilitate the development of advanced neuronal tissue and disease models and translation of stem cell-based therapies.

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Section: Spidroin-based Biomaterials For Regenerative Therapiesmentioning

confidence: 99%

“…In this 3D structure, human pluripotent stem cells (hPSC) developed into 3D cell constructs supported by a microfibrillar network. After the initial expansion, neurons will be induced to differentiate into a continuous layer of neuroectodermal tissue and further differentiate into neuronal subtypes [116]. Another ideal source of stem cells proposed for neural regeneration is DPSCs because of their close relationship with the embryonic origin of neurons and easily available properties.…”

Section: Vascularizationmentioning

confidence: 99%

Spidroin-Based Biomaterials in Tissue Engineering: General Approaches and Potential Stem Cell Therapies

Zhang

Wang

et al. 2021

Stem Cells International

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Spider silks are increasingly gaining interest for potential use as biomaterials in tissue engineering and biomedical applications. Owing to their facile and versatile processability in native and regenerated forms, they can be easily tuned via chemical synthesis or recombinant technologies to address specific issues required for applications. In the past few decades, native spider silk and recombinant silk materials have been explored for a wide range of applications due to their superior strength, toughness, and elasticity as well as biocompatibility, biodegradation, and nonimmunogenicity. Herein, we present an overview of the recent advances in spider silk protein that fabricate biomaterials for tissue engineering and regenerative medicine. Beginning with a brief description of biological and mechanical properties of spidroin-based materials and the cellular regulatory mechanism, this review summarizes various types of spidroin-based biomaterials from genetically engineered spider silks and their prospects for specific biomedical applications (e.g., lung tissue engineering, vascularization, bone and cartilage regeneration, and peripheral nerve repair), and finally, we prospected the development direction and manufacturing technology of building more refined and customized spidroin-based protein scaffolds.

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Assembly of FN-silk with laminin-521 to integrate hPSCs into a three-dimensional culture for neural differentiation

Abstract: The functionalized recombinant spider silk protein FN-silk can self-assemble into a 3D microfiber network. When combined with recombinant laminin521 it provides a 3D culture system suitable for expansion of hPSCs and following neural differentiation.

Cited by 10 publications

References 38 publications

Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids

Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids

Bioorthogonally Cross-Linked Hyaluronan-Laminin Hydrogels for 3D Neuronal Cell Culture and Biofabrication

Spidroin-Based Biomaterials in Tissue Engineering: General Approaches and Potential Stem Cell Therapies

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