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A Cell Electrofusion Chip for Somatic Cells Reprogramming.

Wu W, Qu Y, Hu N, Zeng Y, Yang J, Xu H, Yin ZQ - PLoS ONE (2015)

Bottom Line: Using this chip, we could efficiently fuse NIH3T3 cells and mouse embryonic stem cells (mESCs) to induce somatic cells reprogramming.We also found that fused cells demethylated gradually and 5-hydroxymethylcytosine (5hmC) was involved in the demethylation during the reprogramming.Thus, the cell electrofusion chip would facilitate reprogramming mechanisms research by improving efficiency of cell fusion and reducing workloads.

View Article: PubMed Central - PubMed

Affiliation: Southwest Eye Hospital, Southwest Hospital, Third Military Medical University, Chongqing, P.R. China; Key Lab of Visual Damage and Regeneration & Restoration of Chongqing, Chongqing, P.R. China.

ABSTRACT
Cell fusion is a potent approach to explore the mechanisms of somatic cells reprogramming. However, previous fusion methods, such as polyethylene glycol (PEG) mediated cell fusion, are often limited by poor fusion yields. In this study, we developed a simplified cell electrofusion chip, which was based on a micro-cavity/ discrete microelectrode structure to improve the fusion efficiency and to reduce multi-cell electrofusion. Using this chip, we could efficiently fuse NIH3T3 cells and mouse embryonic stem cells (mESCs) to induce somatic cells reprogramming. We also found that fused cells demethylated gradually and 5-hydroxymethylcytosine (5hmC) was involved in the demethylation during the reprogramming. Thus, the cell electrofusion chip would facilitate reprogramming mechanisms research by improving efficiency of cell fusion and reducing workloads.

No MeSH data available.


Related in: MedlinePlus

The structure and electric field distribution of cell electrofusion chip.(A) Cell trapping and pairing under positive-DEP force. (B) The electric field distribution in the cell electrofusion chip. (C) A 3D schematic of the cell electrofusion chip based on micro-cavity/ discrete microelectrode structure.
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pone.0131966.g001: The structure and electric field distribution of cell electrofusion chip.(A) Cell trapping and pairing under positive-DEP force. (B) The electric field distribution in the cell electrofusion chip. (C) A 3D schematic of the cell electrofusion chip based on micro-cavity/ discrete microelectrode structure.

Mentions: As shown in Fig 1C, this microfluidic chip consisted of two chiasm-shaped microelectrode arrays, which was fabricated on a SOI wafer. To provide good mechanical support for this microfluidic chip, we chose a SOI wafer with 430 μm thickness base silicon layer. And the buried SiO2 layer ensured desired electrical insulation. The two chiasm microelectrode arrays and serpentine-shaped microfluidic channel were fabricated by etching 35 μm thick top low-resistance silicon layers. On each microelectrode arrays, approximate 1.9×104 micro-cavity/ discrete microelectrode structures were integrated. In each micro-cavity structure, the exposure low-resistance silicon sidewall that was parallel to the microchannel served as a microelectrode, whereas the other two sidewalls, which were perpendicular to the microchannel, were fabricated by a SiO2 insulator. Since the separation distance between two adjacent micro-insulators was 20 μm, the width of microelectrode (exposure silicon between two adjacent micro-insulators) was also 20 μm. Each SiO2-Polysilicon-SiO2 micro-insulator was 60 μm in length and 20 μm in width. And it was composed of 600 nm thick SiO2 insulator wall, a 1.8 μm thick enclosing ploysilicon wall, which provided mechanical support [24]. In addition, the floating silicon, which was enclosed by the SiO2-Polysilicon-SiO2 micro-insulator, provided the sidewall of serpentine-shaped microfluidic channel.


A Cell Electrofusion Chip for Somatic Cells Reprogramming.

Wu W, Qu Y, Hu N, Zeng Y, Yang J, Xu H, Yin ZQ - PLoS ONE (2015)

The structure and electric field distribution of cell electrofusion chip.(A) Cell trapping and pairing under positive-DEP force. (B) The electric field distribution in the cell electrofusion chip. (C) A 3D schematic of the cell electrofusion chip based on micro-cavity/ discrete microelectrode structure.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4503441&req=5

pone.0131966.g001: The structure and electric field distribution of cell electrofusion chip.(A) Cell trapping and pairing under positive-DEP force. (B) The electric field distribution in the cell electrofusion chip. (C) A 3D schematic of the cell electrofusion chip based on micro-cavity/ discrete microelectrode structure.
Mentions: As shown in Fig 1C, this microfluidic chip consisted of two chiasm-shaped microelectrode arrays, which was fabricated on a SOI wafer. To provide good mechanical support for this microfluidic chip, we chose a SOI wafer with 430 μm thickness base silicon layer. And the buried SiO2 layer ensured desired electrical insulation. The two chiasm microelectrode arrays and serpentine-shaped microfluidic channel were fabricated by etching 35 μm thick top low-resistance silicon layers. On each microelectrode arrays, approximate 1.9×104 micro-cavity/ discrete microelectrode structures were integrated. In each micro-cavity structure, the exposure low-resistance silicon sidewall that was parallel to the microchannel served as a microelectrode, whereas the other two sidewalls, which were perpendicular to the microchannel, were fabricated by a SiO2 insulator. Since the separation distance between two adjacent micro-insulators was 20 μm, the width of microelectrode (exposure silicon between two adjacent micro-insulators) was also 20 μm. Each SiO2-Polysilicon-SiO2 micro-insulator was 60 μm in length and 20 μm in width. And it was composed of 600 nm thick SiO2 insulator wall, a 1.8 μm thick enclosing ploysilicon wall, which provided mechanical support [24]. In addition, the floating silicon, which was enclosed by the SiO2-Polysilicon-SiO2 micro-insulator, provided the sidewall of serpentine-shaped microfluidic channel.

Bottom Line: Using this chip, we could efficiently fuse NIH3T3 cells and mouse embryonic stem cells (mESCs) to induce somatic cells reprogramming.We also found that fused cells demethylated gradually and 5-hydroxymethylcytosine (5hmC) was involved in the demethylation during the reprogramming.Thus, the cell electrofusion chip would facilitate reprogramming mechanisms research by improving efficiency of cell fusion and reducing workloads.

View Article: PubMed Central - PubMed

Affiliation: Southwest Eye Hospital, Southwest Hospital, Third Military Medical University, Chongqing, P.R. China; Key Lab of Visual Damage and Regeneration & Restoration of Chongqing, Chongqing, P.R. China.

ABSTRACT
Cell fusion is a potent approach to explore the mechanisms of somatic cells reprogramming. However, previous fusion methods, such as polyethylene glycol (PEG) mediated cell fusion, are often limited by poor fusion yields. In this study, we developed a simplified cell electrofusion chip, which was based on a micro-cavity/ discrete microelectrode structure to improve the fusion efficiency and to reduce multi-cell electrofusion. Using this chip, we could efficiently fuse NIH3T3 cells and mouse embryonic stem cells (mESCs) to induce somatic cells reprogramming. We also found that fused cells demethylated gradually and 5-hydroxymethylcytosine (5hmC) was involved in the demethylation during the reprogramming. Thus, the cell electrofusion chip would facilitate reprogramming mechanisms research by improving efficiency of cell fusion and reducing workloads.

No MeSH data available.


Related in: MedlinePlus