Surface-directed assembly of cell-laden microgels

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Entry by Sandeep Koshy, AP 225, Fall 2010

Title: Surface Directed Assembly of Cell-Laden Microgels

Authors: Yanan Du,1,2 Majid Ghodousi,1,2 Edward Lo,1,2 Mahesh K. Vidula,1,2 Onur Emiroglu,1,2 Ali Khademhosseini1,2

Journal: Biotechnology and Bioengineering

Volume: 105

Issue: 3

Pages: 655-662

Summary

This work by Du et al. exploits the use of surface interactions to direct self-assembly of cell-laden microgels for tissue engineering. The goal of this strategy is to have well defined control over the geometric arrangement of hydrogel blocks containing different cell types in order to create a microtissue with defined cell architecture. The authors added an aqueous phase containing hydrophilic polymer hydrogel blocks with cells onto a glass slide containing hydrophilic patterns. They were able to demonstrate that the hydrogels would assemble on the glass surface in a defined pattern and that a secondary crosslinking step could be used to stabilize the pattern and allow removal of the pattern from the surface. Various parameters were optimized for the process including the concentration of the hydrogel polymer blocks, the amount of detergent present in the aqueous phase and secondary crosslinking parameters. This technique could potentially be used to assemble hydrogel blocks with different cell types in order to produce a tissue engineered microtissue of defined spatial organization, which may be useful for regenerative medicine applications.

Soft Matter Keywords: polymer, surface interactions, surface tension, hydrophobic, hydrophilic, hydrogel

Experimental Summary

Microcontact printing on glass surfaces

Glass slides were treated with 70% sulfuric acid, 30% hydrogen peroxide (Piranha solution) to clean their surface. Polidimethylsiloxane (PDMS) stamps bearing the desired pattern were spin-coated with 1% octadecyltrichlorosilane (OTS) in hecane, The PDMS patterns were then put into contact with the glass slide for 2 minutes in order to transfer the pattern.

Microgel fabrication

Photolithography was used to pattern microgels from polyethylene glycol dimethacrylate(PEG) dissolved in phosphate buffered saline (PBS). After the addition of a photoinitiator, the prepolymer mixture was placed between two cover slips beneath a photomask and exposed to ultraviolet (UV) light to crosslink the polymer into gels of a defined size. This procedure could also be performed with the presence of living cells within the prepolymer mixture to generate cell-laden microgels.

Surface assembly of microgels on patterned glass slides

The pattern glass slide was wetted with PBS and the microgels were transferred onto it using a needle. The microgels were allowed to establish interactions with the glass and assembled for 3 minutes, at which point the glass slide was tilted to remove excess liquid. Secondary crosslinking was achieved by using PEG premolymer as the aqueous phase instead of water. Exposure to a second UV step after pattern formation was used to establish crosslinks between the patterned microgels.

Fig 1. Schematic of microgel fabrication and surface-directed assembly.

Results

Surface assembly of microgels on patterned glass slides

Fig 2. Effect of PEG molecular weight, concentration and microgel size on surface-directed assembly.

Once placed on the bulk glass slides, the hydrophilic microgels clustered in the regions where the hydrophilic OTS pattern was created, driven by the hydrophilic surface force. The authors first tested the effect of the molecular weight of PEG macromer on eventual self-assembly. They noted that higher molecular weight PEG macromers were more hydrophilic due to an increased number of ester groups and saw performance increase with higher molecular weight macromers. The effect of macromer concentration showed that lower macromer concentration increased the ability to direct self-assembly presumably since the less dense polymer blocks were able to move more freely in solution to assemble properly. This hypothesis regarding the requirement of mobility of the hydrogels was also supported by the result that smaller hydrogel blocks showed improved assembly.


'Use of surfactant to improve pattern resolution

Fig 3. Effect of surfactant on microgel surface-directed assembly.


The authors noted poor pattern fidelity at the edges of the pattern due to a tendency of water to form rounded drops to minimize the air-water interface near the edge of the hydrophilic pattern. They achieved higher pattern resolution by including a surfactant (Tween 20) and showed that more precise pattern edges could be achieved.


Secondary crosslinking to create stable hydrogel cluster

Fig 4. Secondary crosslinking of microgels.

It was then shown that the patterns could be crosslinked and removed from the surface of the glass by using a second exposure to UV light. The hydrogels were mixed with prepolymer solution containing photoinitiator and this was used to chemically crosslink the gels. The secondary crosslinking polymer concentration was shown to be important for fidelity, with lower concentrations resulting in less excess crosslinked polymer inhibiting pattern resolution.


Creation of cell-laden microtissues

Fig 5. Assembly of cell-laden microgels.


Finally, it was shown that this technique could be used to generate microtissue constructs containing cells. Cell viability was shown to be acceptable using a live dead stain which revealed cells were still actively metabolizing after all of the processing steps. The authors suggest that this technique could be used to assemble microgel blocks containing different cell types into a controlled sheet that mimics the organization of native tissue for applications in regenerative medicine.