Lecture 12 - Scaffold Microfabrication

Question 1

Intro to Scaffold Microfabrication

Answer 1

• Mimicking natural tissue structures could be crucial to achieving engineered tissues
• Functionality of specific tissues is related to their complex architectures —> most tissues composed of more than one cell typed, usually involving microscale organization of cells and ECM components needed to perform specific function
• Incorporation of similar microscale structures in tissue engineering scaffolds may yield remedy for problems ranging from vascularization to cell function control/determination
• Key advantage of incorporating 3D microfeatures within tissue engineering scaffolds is recreation of spatial organization of different types of cells in same way as that found in target tissue

Question 2

Current Challenges of Scaffold Microfabrication

Answer 2

• Controlling cell-cell and cell-material interactions for multicellular engineered tissues
• Vascularization of engineered tissues for proper oxygen/nutrient delivery/metabolite removal
• Directing stem cell fate using microengineered platforms
• Difficult to mimic 3D microstructure of real tissues

Question 3

Approaches to Engineering 3D Microfabricated Tissues

Answer 3

• Photomask-based microfabrication
• Micromold-based microfabrication
• Photolabile rapid prototyping - stereolithography
• 3D printing

Question 4

Photomask-based Microfabrication

Answer 4

• This technique uses direct polymerization through ‘photomasks’
• Use of photomask in conjunction with polymers that crosslink due to exposure to light restricts the crosslinking of the polymer to the regions where the light is permitted to pass through the photomask
• As only the portions of the prepolymer (monomer, oligomer) residing below the transparent regions of the mask receive the necessary stimulus to initiate polymerization, only these areas are crosslinked
• Patterned hydrogels can be fabricated using this method —> acrylated poly(ethylene glycol), methacrylated hyaluronic acid, methacrylated gelatin, other polymer mixes containing these polymers

Question 5

Photomask

Answer 5

2D pattern printed onto a transparent sheet, designed such that light only passes through the mask following specific predetermined patterns

Question 6

Fabrication of Hepatic Tissues (Single Layer)

Answer 6

• Photopatterning of hepatocytes and PEG hydrogels

- Improves diffusive nutrient transport and viability

Question 7

Fabrication of Hepatic Tissues (Multiple Layers)

Answer 7

• Additive photopatterning of cellular hydrogels

- Patterning imporves nutrient transport and cell survival in 3D

Question 8

Micromold-based Microfabrication

Answer 8

• Robust and inexpensive fabrication approach that enables patterning of both planar and non-planar substrates
• Shape and size of patterns can be precisely controlled
• Involves use of elastomeric polymers to form molds and crosslinking of polymer solution between flat surface and patterned elastomer
• Crosslinking carried out by UV or temperature-assisted mechanisms

Question 9

Photolabile Rapid Prototype - Stereolithography

Answer 9

• 3D constructs built layer-by-layer from liquid photolabile polymers that can be cured when exposed to UV light
• Fabrication occurs on platform that is immersed in a container of liquid resin. Highly focused UV laser traces out each layer, solidifying the construct’s cross-sections
• After each layer is fabricated, platform is incrementally lowered and new layer of liquid resin is coated prior to laser tracing of next slice. Process repeated until construct is complete

Question 10

Photolabile Rapid Prototype - Stereolithography: Top-Down Approach

Answer 10

• Elevator where construct is fabricated moves inside a vat that is filled with a prepolymer solution
• When cells added to prepolymer solution, they sit in vat until all layers formed
• Can decrease cell viability since cells are continuously in contact with photoinitiator

Question 11

Photolabile Rapid Prototype - Stereolithography: Bottom-Up Approach

Answer 11

• Fresh cell-containing prepolymer solution is added onto each layer
• Allows uniform cell distribution and viability for at least 15 days
• Enables fabrication of multicellular constructs with varying cell types in each layer

Question 12

3D Printing

Answer 12

• 3D dispensing cell/hydrogel mix in liquid medium
• Creates scaffolds and other 3D architectures using both synthetic and natural materials
• One or two or more components can be applied in 3D printing based upon dispensing technology, typical for one- and two- component adhesives and sealants