Biodegradable and Biocompatible Graphene-based Scaffolds for Functional Neural Tissue Engineering: A Strategy Approach Using Dental Pulp Stem Cells and Biomaterials

Neural tissue engineering aims to restore function of nervous system tissues using biocompatible cell-seeded scaffolds. Graphene-based scaffolds combined with stem cells deserve special attention to enhance tissue regeneration in a controlled manner. However, it is believed that minor changes in scaffold biomaterial composition, internal porous structure, and physicochemical properties can impact cellular growth and adhesion. The current work aims to investigate in vitro biological effects of 3D graphene oxide (GO)/sodium alginate (GOSA) and reduced GOSA (RGOSA) scaffolds on dental pulp stem cells (DPSCs) in terms of cell viability and cytotoxicity. Herein, the effects of the 3D scaffolds, coating conditions, and serum supplementation on DPSCs functions are explored extensively. Biodegradation analysis revealed that addition of GO enhanced the degradation rate of composite scaffolds. Compared to the 2D surface, the cell viability of 3D scaffolds was higher (p <0.0001), highlighting the optimal initial cell adhesion to the scaffold surface and cell migration through pores. Moreover, the cytotoxicity study indicated that the incorporation of graphene supported higher DPSCs viability. It is also shown that when the mean pore size of scaffold increases, DPSCs activity decreases. In terms of coating conditions, poly-l-lysine (PLL) was the most robust coating reagent that improved cell-scaffold adherence and DPSCs metabolism activity. The cytotoxicity of GO-based scaffolds showed that DPSCs can be seeded in serum-free media without cytotoxic effects. This is critical for human translation as cellular transplants are typically serum-free. These findings suggest that proposed 3D GO-based scaffolds have favourable effects on the biological responses of DPSCs.

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Biodegradable and Biocompatible Graphene-based Scaffolds for Functional Neural Tissue Engineering: A Strategy Approach Using Dental Pulp Stem Cells and Biomaterials

10.1101/2021.01.12.426431 ◽

2021 ◽

Author(s):

Negar Mansouri ◽

Said Al-Sarawi ◽

Dusan Losic ◽

Jagan Mazumdar ◽

Jillian Clark ◽

...

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Cell Viability ◽

Dental Pulp ◽

Biological Effects ◽

Neural Tissue ◽

Dental Pulp Stem Cells ◽

Neural Tissue Engineering ◽

3D Scaffolds ◽

Serum Free

AbstractNeural tissue engineering aims to restore function of nervous system tissues using biocompatible cell-seeded scaffolds. Graphene-based scaffolds combined with stem cells deserve special attention to enhance tissue regeneration in a controlled manner. However, it is believed that minor changes in scaffold biomaterial com-position, internal porous structure, and physicochemical properties can impact cellular growth and adhesion. The current work aims to investigate in vitro biological effects of 3D graphene oxide (GO)/sodium alginate (GOSA) and reduced GOSA (RGOSA) scaffolds on dental pulp stem cells (DPSCs) in terms of cell viability and cytotoxicity. Herein, the effects of the 3D scaffolds, coating conditions, and serum supplementation on DPSCs functions are explored extensively. Biodegradation analysis revealed that addition of GO enhanced the degradation rate of composite scaffolds. Compared to the 2D surface, the cell viability of 3D scaffolds was higher (p <0.0001), highlighting the optimal initial cell adhesion to the scaffold surface and cell migration through pores. Moreover, the cytotoxicity study indicated that the incorporation of graphene supported higher DPSCs viability. It is also shown that when the mean pore size of scaffold increases, DPSCs activity decreases. In terms of coating conditions, poly-l-lysine (PLL) was the most robust coating reagent that improved cell-scaffold adherence and DPSCs metabolism activity. The cytotoxicity of GO-based scaffolds showed that DPSCs can be seeded in serum-free media without cytotoxic effects. This is critical for human translation as cellular transplants are typically serum-free. These findings suggest that proposed 3D GO-based scaffolds have favourable effects on the biological responses of DPSCs.

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Biodegradable and Biocompatible Graphene‐based Scaffolds for Functional Neural Tissue Engineering: A Strategy Approach Using Dental Pulp Stem Cells and Biomaterials

Biotechnology and Bioengineering ◽

10.1002/bit.27891 ◽

2021 ◽

Author(s):

Negar Mansouri ◽

Said Al‐Sarawi ◽

Dusan Losic ◽

Jagan Mazumdar ◽

Jillian Clark ◽

...

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Dental Pulp ◽

Neural Tissue ◽

Dental Pulp Stem Cells ◽

Neural Tissue Engineering

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Average cell viability levels of human dental pulp stem cells: an accurate combinatorial index for quality control in tissue engineering

Cytotherapy ◽

10.1016/j.jcyt.2012.11.017 ◽

2013 ◽

Vol 15 (4) ◽

pp. 507-518 ◽

Cited By ~ 9

Author(s):

Miguel Angel Martin-Piedra ◽

Ingrid Garzon ◽

Ana Celeste Oliveira ◽

Camilo Andres Alfonso-Rodriguez ◽

Maria Carmen Sanchez-Quevedo ◽

...

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Quality Control ◽

Cell Viability ◽

Dental Pulp ◽

Dental Pulp Stem Cells ◽

Average Cell ◽

Human Dental Pulp

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Electrical Stimulation Promotes Stem Cell Neural Differentiation in Tissue Engineering

Stem Cells International ◽

10.1155/2021/6697574 ◽

2021 ◽

Vol 2021 ◽

pp. 1-14

Author(s):

Hong Cheng ◽

Yan Huang ◽

Hangqi Yue ◽

Yubo Fan

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Stem Cell ◽

Electrical Stimulation ◽

Neural Differentiation ◽

Biological Effects ◽

Neural Tissue ◽

Neural Regeneration ◽

Neural Tissue Engineering ◽

Conductive Materials

Nerve injuries and neurodegenerative disorders remain serious challenges, owing to the poor treatment outcomes of in situ neural stem cell regeneration. The most promising treatment for such injuries and disorders is stem cell-based therapies, but there remain obstacles in controlling the differentiation of stem cells into fully functional neuronal cells. Various biochemical and physical approaches have been explored to improve stem cell-based neural tissue engineering, among which electrical stimulation has been validated as a promising one both in vitro and in vivo. Here, we summarize the most basic waveforms of electrical stimulation and the conductive materials used for the fabrication of electroactive substrates or scaffolds in neural tissue engineering. Various intensities and patterns of electrical current result in different biological effects, such as enhancing the proliferation, migration, and differentiation of stem cells into neural cells. Moreover, conductive materials can be used in delivering electrical stimulation to manipulate the migration and differentiation of stem cells and the outgrowth of neurites on two- and three-dimensional scaffolds. Finally, we also discuss the possible mechanisms in enhancing stem cell neural differentiation using electrical stimulation. We believe that stem cell-based therapies using biocompatible conductive scaffolds under electrical stimulation and biochemical induction are promising for neural regeneration.

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Vasculogenesis from Human Dental Pulp Stem Cells Grown in Matrigel with Fully Defined Serum-Free Culture Media

Biomedicines ◽

10.3390/biomedicines8110483 ◽

2020 ◽

Vol 8 (11) ◽

pp. 483

Author(s):

Jon Luzuriaga ◽

Jon Irurzun ◽

Igor Irastorza ◽

Fernando Unda ◽

Gaskon Ibarretxe ◽

...

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Cell Therapy ◽

Dental Pulp ◽

Culture Media ◽

Dental Pulp Stem Cells ◽

Adult Brain ◽

Serum Free ◽

Human Dental Pulp

The generation of vasculature is one of the most important challenges in tissue engineering and regeneration. Human dental pulp stem cells (hDPSCs) are some of the most promising stem cell types to induce vasculogenesis and angiogenesis as they not only secrete vascular endothelial growth factor (VEGF) but can also differentiate in vitro into both endotheliocytes and pericytes in serum-free culture media. Moreover, hDPSCs can generate complete blood vessels containing both endothelial and mural layers in vivo, upon transplantation into the adult brain. However, many of the serum free media employed for the growth of hDPSCs contain supplements of an undisclosed composition. This generates uncertainty as to which of its precise components are necessary and which are dispensable for the vascular differentiation of hDPSCs, and also hinders the transfer of basic research findings to clinical cell therapy. In this work, we designed and tested new endothelial differentiation media with a fully defined composition using standard basal culture media supplemented with a mixture of B27, heparin and growth factors, including VEGF-A165 at different concentrations. We also optimized an in vitro Matrigel assay to characterize both the ability of hDPSCs to differentiate to vascular cells and their capacity to generate vascular tubules in 3D cultures. The description of a fully defined serum-free culture medium for the induction of vasculogenesis using human adult stem cells highlights its potential as a relevant innovation for tissue engineering applications. In conclusion, we achieved efficient vasculogenesis starting from hDPSCs using serum-free culture media with a fully defined composition, which is applicable for human cell therapy purposes.

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METTL3-mediated M6a Modification Regulates Cell Cycle Progression of Dental Pulp Stem Cells

10.21203/rs.3.rs-136528/v1 ◽

2021 ◽

Author(s):

Haiyun Luo ◽

Wenjing Liu ◽

Yanli Zhang ◽

Xiao Jiang ◽

Shiqing Wu ◽

...

Keyword(s):

Cell Cycle ◽

Tissue Engineering ◽

Stem Cells ◽

Flow Cytometry ◽

Differentially Expressed Genes ◽

Dental Pulp ◽

Cell Cycle Control ◽

Dental Pulp Stem Cells ◽

Differentially Expressed ◽

Expression Level

Abstract Background: Dental pulp stem cells (DPSCs) exhibited self-renewal, pluripotency capacity and served as promising cells source in endodontic regeneration and tissue engineering. Meanwhile, the regenerative capacity of DPSCs is limited and reduced in long lifespan. N6-methyladenosine (m6A) is the most prevalent, reversible internal modification in RNAs. The methyltransferases complex and demethylases mediated m6A methylation and cooperated to impact various biological processes associated with stem cell fate determination. However, the biological effect of m6A methylation in DPSCs remained unclear. Methods: Cell surface markers and differentiation potential of primary DPSCs were identified and m6A immunoprecipitation with deep sequencing (m6A RIP-seq) was used to uncover characteristics of m6A modifications in DPSCs transcriptome. Expression level of m6A-related genes were evaluated in immature/mature pulp tissues and cells. Lentiviral vectors were constructed to knockdown or overexpress methyltransferase like 3 (METTL3). Cell morphology, viability, senescence and apoptosis were further analyzed by β-galactosidase, TUNEL staining and flow cytometry. Bioinformatic analysis combing m6A RIP and shMETTL3 RNA-seq was used to functionally enrich overlapped genes and screen target of METTL3. Cell cycle distributions were assayed by flow cytometry and m6A RIP-qPCR was used to confirm METTL3 mediated m6A methylation in DPSCs. Results: Here, m6A peaks distribution, binding area and motif in DPSCs were first revealed by m6A RIP-seq. We also found a relative high expression level of METTL3 in immature DPSCs with superior regenerative potential and METTL3 knockdown induced cell apoptosis and senescence. Furthermore, Conjoint analysis of m6A RIP and RNA-sequencing showed differentially expressed genes affected by METTL3 depletion was mainly enriched in cell cycle, mitosis and alteration of METTL3 expression resulted in cell cycle arrest which indicated METTL3 make essential effect in cell cycle control. To further investigate underlying mechanisms, we explored proteins interaction network of differentially expressed genes and Polo-like Kinase 1 (PLK1), a critical cycle modulator was identified as target of METTL3-mediated m6A methylation in DPSCs. Conclusions: These results revealed m6A methylated hallmarks in DPSCs and a regulatory role of METTL3 in cell cycle control. Our study shed light on therapeutic approaches in vital pulp therapy and serve new insight in stem cells based tissue engineering.

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Evaluation of Resin-Based Material Containing Copaiba Oleoresin (Copaifera Reticulata Ducke): Biological Effects on the Human Dental Pulp Stem Cells

Biomolecules ◽

10.3390/biom10070972 ◽

2020 ◽

Vol 10 (7) ◽

pp. 972

Author(s):

Roberta Souza D’Almeida Couto ◽

Maria Fernanda Setubal Destro Rodrigues ◽

Leila Soares Ferreira ◽

Ivana Márcia Alves Diniz ◽

Fernando de Sá Silva ◽

...

Keyword(s):

Stem Cells ◽

Dental Pulp ◽

Biological Effects ◽

Dental Pulp Stem Cells ◽

Nodule Formation ◽

Alizarin Red ◽

Pulp Capping ◽

Hsp 27 ◽

Dental Pulp Capping ◽

Human Dental Pulp

The purpose of this study was to analyze in vitro the biological effects on human dental pulp stem cells triggered in response to substances leached or dissolved from two experimental cements for dental pulp capping. The experimental materials, based on extracts from Copaifera reticulata Ducke (COP), were compared to calcium hydroxide [Ca(OH)2] and mineral trioxide aggregate (MTA), materials commonly used for direct dental pulp capping in restorative dentistry. For this, human dental pulp stem cells were exposed to COP associated or not with Ca(OH)2 or MTA. Cell cytocompatibility, migration, and differentiation (mineralized nodule formation (Alizarin red assay) and gene expression (RT-qPCR) of OCN, DSPP, and HSP-27 (genes regulated in biomineralization events)) were evaluated. The results showed that the association of COP reduced the cytotoxicity of Ca(OH)2. Upregulations of the OCN, DSPP, and HSP-27 genes were observed in response to the association of COP to MTA, and the DSPP and HSP-27 genes were upregulated in the Ca(OH)2 + COP group. In up to 24 h, cell migration was significantly enhanced in the MTA + COP and Ca(OH)2 + COP groups. In conclusion, the combination of COP with the currently used materials for dental pulp capping [Ca(OH)2 and MTA] improved the cell activities related to pulp repair (i.e., cytocompatibility, differentiation, mineralization, and migration) including a protective effect against the cytotoxicity of Ca(OH)2.

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