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Early differentiation patterning of mouse embryonic stem cells in response to variations in alginate substrate stiffness.

Candiello J, Singh SS, Task K, Kumta PN, Banerjee I - J Biol Eng (2013)

Bottom Line: Upon generation of functionally mature ESC derived islet-like cells, they need to be implanted into diabetic patients to restore the loss of islet activity.Encapsulation in alginate microcapsules is a promising route of implantation, which can protect the cells from the recipient's immune system.The insight into these biophysical phenomena found in this study can be used along with other cues to enhance the differentiation of embryonic stem cells toward a specific lineage fate.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA. pkumta@pitt.edu.

ABSTRACT

Background: Embryonic stem cells (ESCs) have been implicated to have tremendous impact in regenerative therapeutics of various diseases, including Type 1 Diabetes. Upon generation of functionally mature ESC derived islet-like cells, they need to be implanted into diabetic patients to restore the loss of islet activity. Encapsulation in alginate microcapsules is a promising route of implantation, which can protect the cells from the recipient's immune system. While there has been a significant investigation into islet encapsulation over the past decade, the feasibility of encapsulation and differentiation of ESCs has been less explored. Research over the past few years has identified the cellular mechanical microenvironment to play a central role in phenotype commitment of stem cells. Therefore it will be important to design the encapsulation material to be supportive to cellular functionality and maturation.

Results: This work investigated the effect of stiffness of alginate substrate on initial differentiation and phenotype commitment of murine ESCs. ESCs grown on alginate substrates tuned to similar biomechanical properties of native pancreatic tissue elicited both an enhanced and incrementally responsive differentiation towards endodermal lineage traits.

Conclusions: The insight into these biophysical phenomena found in this study can be used along with other cues to enhance the differentiation of embryonic stem cells toward a specific lineage fate.

No MeSH data available.


Related in: MedlinePlus

Representative image of cell morphology of ES-D3 cells during spontaneous differentiation on alginate gels of varying Young’s modulus. Cells on softer alginate gels (516 Pa, Figure 3A) were generally more numerous, but smaller after 3 days of spontaneous differentiation, while cells on stiffer substrates (1337 Pa, Figure 3B) formed larger clumps in a shorter time frame.
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Figure 2: Representative image of cell morphology of ES-D3 cells during spontaneous differentiation on alginate gels of varying Young’s modulus. Cells on softer alginate gels (516 Pa, Figure 3A) were generally more numerous, but smaller after 3 days of spontaneous differentiation, while cells on stiffer substrates (1337 Pa, Figure 3B) formed larger clumps in a shorter time frame.

Mentions: It was consistently observed in repeated experiments that some of the softer alginate gels were not robust enough to support the ES-D3 cells. A large portion of the cells sunk to the bottom of the well and attached to the underlying plastic of the 24-well plate. Hence these gel conditions: 6 mg/ml alginate concentration at 1× and 2× crosslinking concentrations along with 8 mg/ml alginate concentration at 1× crosslinking concentration were excluded from further analysis. For the gels that were able to support cell culture, phase contrast imaging of ES-D3 cells showed a clumped morphology (Figure 2). These clumps were not stagnant on the gel surface, but rather migrated on the gel surface, forming larger clumps as the cells proliferated and combined. Also, it was noted that the ESD3 cells generally formed larger clumps more immediately on stiffer gels (Figure 2B) in contrast to those on softer gel substrates (Figure 2A).


Early differentiation patterning of mouse embryonic stem cells in response to variations in alginate substrate stiffness.

Candiello J, Singh SS, Task K, Kumta PN, Banerjee I - J Biol Eng (2013)

Representative image of cell morphology of ES-D3 cells during spontaneous differentiation on alginate gels of varying Young’s modulus. Cells on softer alginate gels (516 Pa, Figure 3A) were generally more numerous, but smaller after 3 days of spontaneous differentiation, while cells on stiffer substrates (1337 Pa, Figure 3B) formed larger clumps in a shorter time frame.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3643844&req=5

Figure 2: Representative image of cell morphology of ES-D3 cells during spontaneous differentiation on alginate gels of varying Young’s modulus. Cells on softer alginate gels (516 Pa, Figure 3A) were generally more numerous, but smaller after 3 days of spontaneous differentiation, while cells on stiffer substrates (1337 Pa, Figure 3B) formed larger clumps in a shorter time frame.
Mentions: It was consistently observed in repeated experiments that some of the softer alginate gels were not robust enough to support the ES-D3 cells. A large portion of the cells sunk to the bottom of the well and attached to the underlying plastic of the 24-well plate. Hence these gel conditions: 6 mg/ml alginate concentration at 1× and 2× crosslinking concentrations along with 8 mg/ml alginate concentration at 1× crosslinking concentration were excluded from further analysis. For the gels that were able to support cell culture, phase contrast imaging of ES-D3 cells showed a clumped morphology (Figure 2). These clumps were not stagnant on the gel surface, but rather migrated on the gel surface, forming larger clumps as the cells proliferated and combined. Also, it was noted that the ESD3 cells generally formed larger clumps more immediately on stiffer gels (Figure 2B) in contrast to those on softer gel substrates (Figure 2A).

Bottom Line: Upon generation of functionally mature ESC derived islet-like cells, they need to be implanted into diabetic patients to restore the loss of islet activity.Encapsulation in alginate microcapsules is a promising route of implantation, which can protect the cells from the recipient's immune system.The insight into these biophysical phenomena found in this study can be used along with other cues to enhance the differentiation of embryonic stem cells toward a specific lineage fate.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA. pkumta@pitt.edu.

ABSTRACT

Background: Embryonic stem cells (ESCs) have been implicated to have tremendous impact in regenerative therapeutics of various diseases, including Type 1 Diabetes. Upon generation of functionally mature ESC derived islet-like cells, they need to be implanted into diabetic patients to restore the loss of islet activity. Encapsulation in alginate microcapsules is a promising route of implantation, which can protect the cells from the recipient's immune system. While there has been a significant investigation into islet encapsulation over the past decade, the feasibility of encapsulation and differentiation of ESCs has been less explored. Research over the past few years has identified the cellular mechanical microenvironment to play a central role in phenotype commitment of stem cells. Therefore it will be important to design the encapsulation material to be supportive to cellular functionality and maturation.

Results: This work investigated the effect of stiffness of alginate substrate on initial differentiation and phenotype commitment of murine ESCs. ESCs grown on alginate substrates tuned to similar biomechanical properties of native pancreatic tissue elicited both an enhanced and incrementally responsive differentiation towards endodermal lineage traits.

Conclusions: The insight into these biophysical phenomena found in this study can be used along with other cues to enhance the differentiation of embryonic stem cells toward a specific lineage fate.

No MeSH data available.


Related in: MedlinePlus