3D Tissue Scaffolds

Our goal is to develop advanced measurement tools and standards for measuring polymer scaffold properties and their impact on biological response.  

Use of 3D tissue scaffolds as a template for regeneration is the basis of tissue engineering. These tools will enable a better understanding cell-scaffold interactions, including identification of the relationships of cell response on 2D surfaces to that in 3D scaffolds, and will facilitate improved design of future scaffold-based medical products. These measurement tools and standards advance the ability of researchers to develop scaffolds that direct stem cell differentiation.

Drivers: 

• The U.S. is in a healthcare cost crisis. We spend more of our GDP (16%) on healthcare than any other nation; second on the list Switzerland at 11%.
• Bill "H.R. 1862 Regenerative Medicine Promotion Act of 2011" was introduced in May 2011 and calls for NIST to develop "consensus standards regarding scientific issues critical to regulatory approval of regenerative medicine products". 
• Bone grafting is the second most common transplant procedure (blood is first) with 500,000 bone grafts performed each year in the US; 2.2 million procedures worldwide
• Diabetes, a major tissue engineering target (pancreatic islets), affects 6 million patients and costs $125B/yr (6% of US healthcare spending).
• We spend $35 billion annually (3% of healthcare costs) to care for the 100,000 patients with end stage organ failure waiting for organ transplants.

Reference Material Scaffolds: Reference material scaffolds are being developed that can serve as a calibration point for comparing scaffold measurements between different labs. The first generation reference scaffolds have been deployed and focus on scaffold structure and porosity. A second generation reference scaffold is under development that will focus on measuring cell response (adhesion and proliferation) to 3D scaffolds.  
 

First Generation - Scaffold Structure: A reference scaffold has been developed with input from ASTM (F04.42.WK6507). The scaffolds were made by freeform fabrication since this approach offers the tightest control over scaffold structural morphology. Scaffold structure and porosity have been characterized using microscopy, gravimetrics and μCT imaging. These well-characterized reference scaffolds can serve as standards during development of scaffolds-based products where structure and scaffold porosity are measured.

Left: Three different reference material scaffolds in their packaging: RM8395, RM8396 and RM8397. Middle: Table of RM scaffold properties. Right: 3D X-ray tomograph of RM8397.

• RM8395 (200 micron strut spacing)
• RM8396 (300 micron strut spacing)
• RM8397 (450 micron strut spacing)

Second Generation - Cell Response: Second generation reference material scaffolds are being developed that are characterized for cell response. These are freeform fabricated scaffolds that fit into a 96-well plate for cell culture experiments. One unit will proved 24 scaffolds that have been structurally characterized and for which the cell responses of cell adhesion and proliferation have been measured. The reference scaffold plate can be run as a control in cell culture experiments to serve as a calibration point between different labs.  

Left: Reference material scaffolds RM8394 are under development. One unit is a 96-well plate with 24 scaffolds. RM8394 will have strut diameter 300 mm, strut spacing 500 mm and porosity 50%. Middle: 3D X-ray tomograph of RM8394. Right: Fluorescence micrograph of MC3T3-E1 osteoblasts culture 1 d on RM8394. Red staining is actin and green staining is the nucleus.

3D Scaffold Libraries: Cells respond differently to different materials. Chemistry, mechanics and structure of the materials have strong impact on whether cells adhere, proliferate, migrate or differentiate. We have pioneered the development of a suite of combinatorial screening methods for 3D scaffolds. Previous combinatorial approaches for screening cell-material interactions have focused on planar (2D) surfaces or films. However, biomaterials are commonly used in 3D scaffolds and cells behave differently when cultured in a 3D environment.  Thus, "combi" tests for many types of scaffolds and properties have been developed for a "scaffoldomics" approach.   

"Combi" approaches for screening a wide range of 3D scaffolds have been developed to comprise a "scaffoldomics" approach.  

Links to Scaffold Library Fabrication YouTube Videos:

• Salt-leached scaffold libraries
  
• Hydrogel scaffold libraries 

Cell-Material Interactions: Combi screens provide exciting "hits" that we can explore with more rigorously. Our goal is to enable design of improved scaffolding materials by determining how 3D scaffold properties influence stem cell differentiation. Much of our current knowledge of biomaterials is phenomenological.  In order for tissue engineering to advance, a mechanistic understanding of how material properties direct stem cell function must be developed.

Left: Primary human bone marrow stromal cells (hBMSCs) deposit mineralized matrix when encapsulated in stiff poly(ethylene glycol) tetramethacrylate (PEGTM) hydrogels (21 d). Middle: During culture in PEGTM gels, hBMSCs deposit a matrix containing fibronectin (7 d). Right: RGDS peptides, antibodies that block integrin a_v , and antibodies that block integrin b₃ , inhibit PEGTM-modulus-induced hBMSC mineralization (14 d, 5% PEGTM gels, 11 kPa compressive modulus). These results suggest that interactions between RGD peptides and a_vb₃ integrins (not b₁ integrins) are required for osteogenic differentiation of hBMSCs in stiff PEGTM gels.

• Deployed first reference material for the tissue engineering industry: reference material scaffolds developed in collaboration with ASTM that enable inter-lab comparison of scaffold measurements (RM 8395, RM8396, RM8397)
• Developed a suite of combi methods for screening 3D tissue scaffolds comprising a scaffoldomics approach.
• Developed the world's first combinatorial method for screening cell response to 3D tissue scaffold properties.
• First demonstration that the modulus of 3D scaffolds can direct osteoblast differentiation