Recombinant spider silk protein matrices facilitate multi-analysis of calcium-signaling in neural stem cell-derived AMPA-responsive neurons

Neural progenitors or stem cells (NSCs) show great promise in drug discovery and clinical application. Yet few efforts have been made to optimize biocompatible materials for such cells to be expanded and used in clinical conditions. We have previously demonstrated that NSCs are readily cultured on substrates of certain recombinant spider silk protein without addition of animal- or human-derived components. The question remains however whether this material allows differentiation into functional neurons and glia, and whether such differentiation can take place also when the NSCs are cultured within the material in a pseudo-3D context. Here we demonstrate that “foam”-like structures generated from recombinant spider silk protein (4RepCT) provided excellent matrices for the generation and multicellular analysis of functional excitatory neurons from NSCs without addition of animal- or human-derived components. NSCs isolated from the cerebral cortices of rat embryos were cultured on either 4RepCT matrices shaped as foam-like structures without coating, or on conventional polystyrene plates coated with poly-L-ornithine and fibronectin. Upon treatment with recombinant proteins including the growth factor BMP4 or a combination of BMP4 and the signaling factor Wnt3a, the cortical NSCs cultured in 4RepCT foam-like structures differentiated efficiently into neurons that responded to glutamate receptor agonists, such as AMPA, to at least the same extent as control cultures. Matrices derived from recombinant spider silk proteins thus provide a functional microenvironment for neural stem cells without any animal- or human-derived components, and can be employed in the development of new strategies in stem cell research and tissue engineering.

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Recombinant Spider Silk Protein Matrices Facilitate Differentiation of Neural Stem Cells Into Mature and Functional Neurons

Frontiers in Materials ◽

10.3389/fmats.2020.560372 ◽

2021 ◽

Vol 7 ◽

Author(s):

Michalina Lewicka ◽

Paola Rebellato ◽

Jakub Lewicki ◽

Per Uhlén ◽

Anna Rising ◽

...

Keyword(s):

Stem Cells ◽

Neural Stem Cells ◽

Spider Silk ◽

Silk Protein ◽

Great Promise ◽

Extracellular Signaling ◽

Spider Silk Protein ◽

Glutamate Receptor Agonists ◽

Protein Matrices ◽

Clinical Conditions

Neural stem cells (NSCs) show great promise in drug discovery and clinical application. Yet few efforts have been made to optimize biocompatible materials for such cells to be expanded and used in clinical conditions. We have previously demonstrated that NSCs are readily cultured on substrates of certain recombinant spider silk protein without addition of animal- or human-derived components. The question remains however whether this material allows differentiation into functional neurons, and whether such differentiation can take place also when the NSCs are cultured not only upon but also within the biodegradable material. Here we demonstrate that “foam”-like structures generated from recombinant spider silk protein (4RepCT) provided excellent matrices for the generation and multicellular analysis of functional excitatory neurons from NSCs without addition of animal- or human-derived components. NSCs isolated from the cerebral cortices of rat embryos were cultured at either 4RepCT matrices shaped as foam-like structures without coating, or on conventional polystyrene plates coated with poly-L-ornithine and fibronectin. Upon treatment with recombinant proteins including the extracellular signaling factor BMP4 or a combination of BMP4 and the signaling factor Wnt3a, the cortical NSCs cultured in 4RepCT foam-like structures differentiated efficiently into neurons that responded to glutamate receptor agonists, such as AMPA, to the same extent as control cultures. Matrices derived from recombinant spider silk proteins thus provide a functional microenvironment for neural stem cells with little or no animal- or human-derived components and can be employed in the development of new strategies in stem cell research and tissue engineering.

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Nonionic and zwitterionic forms of glycylglycylarginine as a part of spider silk protein: Spectroscopic and theoretical study

Spectrochimica Acta Part A Molecular and Biomolecular Spectroscopy ◽

10.1016/j.saa.2014.12.115 ◽

2016 ◽

Vol 152 ◽

pp. 557-571 ◽

Cited By ~ 4

Author(s):

Hatice Arı ◽

Talat Özpozan

Keyword(s):

Spider Silk ◽

Theoretical Study ◽

Silk Protein ◽

Spider Silk Protein

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Native-sized recombinant spider silk protein produced in metabolically engineered Escherichia coli results in a strong fiber

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1003366107 ◽

2010 ◽

Vol 107 (32) ◽

pp. 14059-14063 ◽

Cited By ~ 334

Author(s):

X.-X. Xia ◽

Z.-G. Qian ◽

C. S. Ki ◽

Y. H. Park ◽

D. L. Kaplan ◽

...

Keyword(s):

Escherichia Coli ◽

Spider Silk ◽

Silk Protein ◽

Spider Silk Protein ◽

Engineered Escherichia Coli ◽

Metabolically Engineered Escherichia Coli

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The Winding Road of Cardiac Regeneration—Stem Cell Omics in the Spotlight

Cells ◽

10.3390/cells7120255 ◽

2018 ◽

Vol 7 (12) ◽

pp. 255 ◽

Cited By ~ 4

Author(s):

Miruna Mihaela Micheu ◽

Alina Ioana Scarlatescu ◽

Alexandru Scafa-Udriste ◽

Maria Dorobantu

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Fate ◽

Cell Biology ◽

Molecular Mechanisms ◽

Unmet Need ◽

Cardiac Regeneration ◽

Cell Types ◽

Great Promise ◽

Omics Data

Despite significant progress in treating ischemic cardiac disease and succeeding heart failure, there is still an unmet need to develop effective therapeutic strategies given the persistent high-mortality rate. Advances in stem cell biology hold great promise for regenerative medicine, particularly for cardiac regeneration. Various cell types have been used both in preclinical and clinical studies to repair the injured heart, either directly or indirectly. Transplanted cells may act in an autocrine and/or paracrine manner to improve the myocyte survival and migration of remote and/or resident stem cells to the site of injury. Still, the molecular mechanisms regulating cardiac protection and repair are poorly understood. Stem cell fate is directed by multifaceted interactions between genetic, epigenetic, transcriptional, and post-transcriptional mechanisms. Decoding stem cells’ “panomic” data would provide a comprehensive picture of the underlying mechanisms, resulting in patient-tailored therapy. This review offers a critical analysis of omics data in relation to stem cell survival and differentiation. Additionally, the emerging role of stem cell-derived exosomes as “cell-free” therapy is debated. Last but not least, we discuss the challenges to retrieve and analyze the huge amount of publicly available omics data.

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Presence of β-Turn Structure in Recombinant Spider Silk Dissolved in Formic Acid Revealed with NMR

Molecules ◽

10.3390/molecules27020511 ◽

2022 ◽

Vol 27 (2) ◽

pp. 511

Author(s):

Yu Suzuki ◽

Takanori Higashi ◽

Takahiro Yamamoto ◽

Hideyasu Okamura ◽

Takehiro K. Sato ◽

...

Keyword(s):

Mechanical Properties ◽

Formic Acid ◽

Spider Silk ◽

Chemical Shifts ◽

Repetitive Sequences ◽

Silk Protein ◽

Random Coil ◽

Oh Groups ◽

Spider Silk Protein ◽

Turn Structure

Spider dragline silk is a biopolymer with excellent mechanical properties. The development of recombinant spider silk protein (RSP)-based materials with these properties is desirable. Formic acid (FA) is a spinning solvent for regenerated Bombyx mori silk fiber with excellent mechanical properties. To use FA as a spinning solvent for RSP with the sequence of major ampullate spider silk protein from Araneus diadematus, we determined the conformation of RSP in FA using solution NMR to determine the role of FA as a spinning solvent. We assigned 1H, 13C, and 15N chemical shifts to 32-residue repetitive sequences, including polyAla and Gly-rich regions of RSP. Chemical shift evaluation revealed that RSP is in mainly random coil conformation with partially type II β-turn structure in the Gly-Pro-Gly-X motifs of the Gly-rich region in FA, which was confirmed by the 15N NOE data. In addition, formylation at the Ser OH groups occurred in FA. Furthermore, we evaluated the conformation of the as-cast film of RSP dissolved in FA using solid-state NMR and found that β-sheet structure was predominantly formed.

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Stem cell application in neurorehabilitation

Oxford Textbook of Neurorehabilitation ◽

10.1093/med/9780198824954.003.0014 ◽

2020 ◽

pp. 169-184

Author(s):

Sebastian Jessberger ◽

Armin Curt ◽

Roger A. Barker

Keyword(s):

Spinal Cord ◽

Stem Cells ◽

Stem Cell ◽

Adult Brain ◽

Proper Function ◽

Great Promise ◽

Neuropsychiatric Diseases ◽

Brain And Spinal Cord ◽

The Brain

A number of diseases of the brain and spinal cord are associated with substantial neural cell death and/or disruption of correct and functional neural networks. In the past, a variety of therapeutic strategies to rescue these systems have been proposed along with agents to induce functional plasticity within the remaining central nervous system (CNS) structures. In the case of injury or neurodegenerative disease these approaches have only met with limited success, indicating the need for novel approaches to treat diseases of the adult CNS. Recently, the idea of recruiting endogenous or transplanting stem cells to replace lost structures within the adult brain or spinal cord has gained significant attention, along with in situ reprogramming, and opened up novel therapeutic avenues in the context of regenerative medicine. Here we review recent advances in our understanding of how endogenous stem cells may be a part of pathological processes in certain neuropsychiatric diseases and summarize recent clinical and preclinical data suggesting that stem cell-based therapies hold great promise as a future treatment option in a number of diseases disrupting the proper function of the adult CNS.

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Stem cell-based treatment of kidney diseases

Experimental Biology and Medicine ◽

10.1177/1535370220915901 ◽

2020 ◽

Vol 245 (10) ◽

pp. 902-910

Author(s):

Binbin Pan ◽

Guoping Fan

Keyword(s):

Chronic Kidney Disease ◽

Stem Cells ◽

Acute Kidney Injury ◽

Stem Cell ◽

Kidney Disease ◽

Cell Therapy ◽

Kidney Diseases ◽

Kidney Injury ◽

Great Promise ◽

Kidney Organoids

Kidney dysfunction, including chronic kidney disease and acute kidney injury, is a globally prevalent health problem. However, treatment regimens are still lacking, especially for conditions involving kidney fibrosis. Stem cells hold great promise in the treatment of chronic kidney disease and acute kidney injury, but success has been hampered by insufficient incorporation of the stem cells in the injured kidney. Thus, new approaches for the restoration of kidney function after acute or chronic injury have been explored. Recently, kidney organoids have emerged as a useful tool in the treatment of kidney diseases. In this review, we discuss the mechanisms and approaches of cell therapy in acute kidney injury and chronic kidney disease, including diabetic kidney disease and lupus nephritis. We also summarize the potential applications of kidney organoids in the treatment of kidney diseases. Impact statement Stem cells hold great promise in regenerative medicine. Pluripotent stem cells have been differentiated into kidney organoids to understand human kidney development and to dissect renal disease mechanisms. Meanwhile, recent studies have explored the treatment of kidney diseases using a variety of cells, including mesenchymal stem cells and renal derivatives. This mini-review discusses the diverse mechanisms underlying current renal disease treatment via stem cell therapy. We postulate that clinical applications of stem cell therapy for kidney diseases can be readily achieved in the near future.

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Interfacial Behavior of Recombinant Spider Silk Protein Parts Reveals Cues on the Silk Assembly Mechanism

Langmuir ◽

10.1021/acs.langmuir.8b02381 ◽

2018 ◽

Vol 34 (39) ◽

pp. 11795-11805 ◽

Cited By ~ 7

Author(s):

Linnea Nilebäck ◽

Suvi Arola ◽

Mathias Kvick ◽

Arja Paananen ◽

Markus B. Linder ◽

...

Keyword(s):

Spider Silk ◽

Silk Protein ◽

Interfacial Behavior ◽

Assembly Mechanism ◽

Spider Silk Protein

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Designing stem cell niches for differentiation and self-renewal

Journal of The Royal Society Interface ◽

10.1098/rsif.2018.0388 ◽

2018 ◽

Vol 15 (145) ◽

pp. 20180388 ◽

Cited By ~ 35

Author(s):

Hannah Donnelly ◽

Manuel Salmeron-Sanchez ◽

Matthew J. Dalby

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Regenerative Medicine ◽

Regulatory Networks ◽

Regulatory Mechanisms ◽

Great Promise ◽

Regenerative Capacity ◽

Cell Niche ◽

Stem Cell Niches

Mesenchymal stem cells, characterized by their ability to differentiate into skeletal tissues and self-renew, hold great promise for both regenerative medicine and novel therapeutic discovery. However, their regenerative capacity is retained only when in contact with their specialized microenvironment, termed the stem cell niche . Niches provide structural and functional cues that are both biochemical and biophysical, stem cells integrate this complex array of signals with intrinsic regulatory networks to meet physiological demands. Although, some of these regulatory mechanisms remain poorly understood or difficult to harness with traditional culture systems. Biomaterial strategies are being developed that aim to recapitulate stem cell niches, by engineering microenvironments with physiological-like niche properties that aim to elucidate stem cell-regulatory mechanisms, and to harness their regenerative capacity in vitro . In the future, engineered niches will prove important tools for both regenerative medicine and therapeutic discoveries.

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