engleski

Exploring seeding methods to evenly distribute cells in a 3D electrospun scaffold

The development of a novel flexible tissue engineering bioreactor requires optimal design and operating parameters. Initial uniform cell distribution is an important prerequisite as underpopulated areas can be a result of poor cell suspension but also nutrient deprivation and unsuccessful bioreactor design. Furthermore, cell distribution throughout the scaffold is often a design requirement of tissue-engineering products. To achieve even cell distribution, three seeding methods were tested for seeding efficiency. Several repeats of 3cm long electrospun PCL scaffolds were propped into three custom built stages and subsequently seeded with human hamstring cells, one with the scaffold filaments aligned along gravity, one aligned perpendicular to gravity, and one moved onto a rocking plate immediately after seeding. Cells were left to attach for 1 hr and subsequently fixed in 2.5% glutaraldehyde. Cell distribution was assessed using Osmium Tetraoxide stain and micro-CT analysis. Images were analysed using the Fiji 3D- Object counter to visualize cells above the threshold. Quantitative results were binned and plotted in a frequency distribution and heat map. SEM and Elemental x-ray analysis were used to confirm the findings. Results showed that cells readily adhere to the scaffold material and that seeding cells in a dynamic fashion greatly improved even cell distribution. The method of assessing this is a needed alternative for large tissue constructs as cryo-sectioning on large scale is time consuming and scaffold materials do not always withstand processing.

medical textiles ; electrospinning ; electroactive polymers ;

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