In English

Development of a Microfluidic Platform for Cell Migration Studies along Gradients

Elin Bernson
Göteborg : Chalmers tekniska högskola, 2012. 72 s.
[Examensarbete på avancerad nivå]

Cell migration is a cellular fate process essential for embryogenesis, tissue regeneration and wound healing. In vivo cell migration is promoted by soluble chemoattrac-tants which stimulate cell movement, a process called chemotaxis. Migration is also controlled by cell surface interactions where focal adhesions are formed between cell integrins and attachment peptides, such as RGD (Arginine-‐Glycine-‐Aspartic acid), in the extra cellular matrix. It has been shown that by varying the spacing of these attachment peptides on a surface, cell attachment, spreading and focal adhesion formation can be controlled. In addition, varying the spacing of attachment peptides is known to influence cell fate processes, such as apoptosis, proliferation and differentiation. The hypothesis that we wish to investigate is that cell migration is affected by the spacing of attachment peptides. The aim of this project was to design and evaluate an experimental system that could monitor cell migration as a function of attachment peptide spacing by simultaneously exposing cells to a chemotactic gradient in the culture media. Therefore, Au nano‐patterned surfaces, with Au nano-‐dots placed in hexagonal pattern with an inter-particle spacing increasing gradient-‐wise from 65 to 85 nm over 6 mm (Spatz group, MPI Intelligent Systems, Stuttgart) were used to control the spacing of a cyclic RGD attachment peptide, which preferentially binds to the Au nano-‐dots. These surfaces were combined with a microfluidic network capable of generating stable concentration gradients of chemoattractants. The microfluidic chip was designed and evaluated with help of a finite element method (Comsol, Stockholm, Sweden) where simulations were performed and concentration gradient formation, velocity, and flow and shear stress profiles were investigated. Several designs were tested and parameters such as placing of inlets and outlets, channel dimensions and connections, were evaluated. Based on the simulation data, the final microfluidic chip was designed, produced and experimen-tally characterized. The design is a diffusion based gradient generator capable of generating a concentration gradient inside a flow-‐free cell culture chamber, ensuring no shear stress induced migration in the main cell culture channel. The microfluidic chip was further combined with the cRGD functionalized surfaces and it was shown that Human Umbilical Vein Endothelial Cells (HUVECs) attach and spread and that they can be cultured in the system for at least five days during static conditions (longest time evaluated). The developed system is able to generate relevant concentration gradients of a chemoattractant factor, the microfluidic channel can be combined with an Au nano-‐dot patterned substrate, and cells can be cultured and monitored in situ, both in live and fixed/stained states, using bright field, phase contrast and fluorescent microscopy. In conclusion, a microfluidic platform was developed in which it is possible to study the effect of attachment peptide spacing during directed cell migration.

Nyckelord: Cell surface interactions, cRGD attachment peptide spacing, Microfluidics, Cell migration

Publikationen registrerades 2013-06-13. Den ändrades senast 2013-06-17

CPL ID: 178392

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