Neuronal Axon Mimetics For In Vitro Analysis Of Neurological Diseases, Myelination, And Drug Screening

Abstract

Aspects of the present invention provide improved methods and apparatus for use in in vitro modeling of the interaction of cells with cellular constructs/parts/axons, including axon mimetics and use of three-dimensional fibers.

1 . A cell-mimetic device comprising:
 a three dimensional structure comprising a plurality of fibers, said plurality of fibers having an average stiffness post-curing of between about 0.1 and about 300 kPa, and an average diameter of between about 0.1 and about 50 micrometers; and   a support structure connected to the three dimensional structure; wherein the stiffness is calculated as any of Young's modulus, bulk modulus, shear modulus, and dynamic modulus; and post-curing stiffness is measured after equilibration in an aqueous solution buffered at pH 7.0-7.4.   
     
     
         2 . (canceled) 
     
     
         3 . The device of  claim 1 , wherein the post-curing stiffness is between about 0.1 and 100 kPa. 
     
     
         4 . The device of  claim 1 , wherein the post-curing stiffness is between about 0.1 and 10 kPa. 
     
     
         5 . The device of  claim 1 , wherein the post-curing stiffness is between about 0.1 and 1 kPa. 
     
     
         6 . The device of  claim 4 , wherein the average diameter is between about 0.1 and about 10 micrometers. 
     
     
         7 . The device of  claim 4 , wherein the average diameter is between about 0.1 and about 1 micrometers. 
     
     
         8 . (canceled) 
     
     
         9 . (canceled) 
     
     
         10 . The device of  claim 6 , wherein the fibers are formed of compliant polymers or hydrogels. 
     
     
         11 . (canceled) 
     
     
         12 . The device of f  claim 10 , wherein the fibers are PEG, pHEMA, PDMS, polyacrylamide, hyaluronic methacrylate, or any viscoelastic polymer, or a derivative of the foregoing. 
     
     
         13 . The device of  claim 12 , wherein the fibers are modified by a surface ligand. 
     
     
         14 . The device of  claim 6 , wherein the average stiffness is constant over the three dimensional structure. 
     
     
         15 . The device of  claim 6 , wherein the average diameter is constant over the three dimensional structure. 
     
     
         16 . The device of  claim 6 , wherein the surface ligand density or type is constant over the three dimensional structure. 
     
     
         17 . The device of  claim 6 , wherein the plurality of fibers are arranged as one or more piles in the three dimensional structure, and at least one of fiber diameter, fiber stiffness, surface ligand density, and surface ligand type varies along at least one dimension of the three dimensional structure. 
     
     
         18 . The device of  claim 6 , wherein the three dimensional structure represents at least one of a model of a tissue, and a model of neuronal axons. 
     
     
         19 . The device of  claim 6 , wherein the fibers in the three dimensional structure are formed of a stretchable fiber material. 
     
     
         20 . The device of  claim 19 , wherein the stretchable fiber material is PDMS, pHEMA, hyaluronic methacrylate, or any viscoelastic polymer. 
     
     
         21 . The device of  claim 20 , wherein the three dimensional structure can be elastically deformed in at least one dimension by at least 5%. 
     
     
         22 . The device of  claim 21 , further comprising a substrate material attached to, and at least as elastically deformable in the at least one dimension as, the three dimensional structure. 
     
     
         23 . A method of studying cells in vitro, comprising:
 providing a three dimensional structure comprising a plurality of fibers, said plurality of fibers having an average stiffness post-curing of between about 0.1 and about 300 kPa, and an average diameter of between about 0.1 and about 50 micrometers; and a support structure connected to the three dimensional structure;   providing a cell-mimetic device comprising a three dimensional structure comprising a plurality of fibers,   contacting the cell-mimetic device with a population of cells;   studying at least one feature of an interaction of the population of cells with the cell mimetic device; and   studying at least one feature of an interaction between cells of the same cell type or of different cell types within the cell-mimetic device.   
     
     
         24 . The method of  claim 23 , wherein the cells are neural cells or oligodendrocytes. 
     
     
         25 . (canceled) 
     
     
         26 . (canceled) 
     
     
         27 . (canceled) 
     
     
         28 . (canceled) 
     
     
         29 . (canceled) 
     
     
         30 . An assay device comprising:
 a substrate;   a fiber support attached to the substrate; and   a plurality of fibers, each of the plurality of fibers having a length and spanning from the substrate to the fiber support such that each fiber is suspended in air or fluid along at least part of the fiber length, and the plurality of fibers having:   an average stiffness of between about 0.1 and about 300 kPa; and   an average diameter of between about 0.1 and about 50 micrometers; wherein the stiffness is calculated as any of Young's modulus, bulk modulus, shear modulus, and dynamic modulus;   wherein the post-curing stiffness is between about 0.1 and 10 kPa;   wherein the average diameter is between about 0.1 and about 10 micrometers; and   wherein the post-curing stiffness is measured after equilibration in an aqueous solution buffered at pH 7.0-7.4.   
     
     
         31 . (canceled) 
     
     
         32 . (canceled) 
     
     
         33 . (canceled) 
     
     
         34 . (canceled) 
     
     
         35 . (canceled) 
     
     
         36 . (canceled) 
     
     
         37 . (canceled) 
     
     
         38 . (canceled) 
     
     
         39 . The device of  claim 30 , wherein the fibers are formed of compliant polymers or hydrogels. 
     
     
         40 . (canceled) 
     
     
         41 . The device of  claim 39 , wherein the fibers are PEG, pHEMA, PDMS, polyacrylamide, hyaluronic methacrylate, or any viscoelastic polymer or derivatives thereof. 
     
     
         42 . The device of  claim 30 , wherein the fibers are modified by a surface ligand. 
     
     
         43 . The device of  claim 30 , wherein the average stiffness is constant over the three dimensional structure. 
     
     
         44 . The device of  claim 30 , wherein the average diameter is constant over the three dimensional structure. 
     
     
         45 . The device of  claim 30 , wherein the surface ligand density or type is constant over the three dimensional structure. 
     
     
         46 . The device of  claim 41 , wherein the plurality of fibers are arranged as one or more piles in the three dimensional structure and at least one of fiber diameter, fiber stiffness, and surface ligand density and type varies along at least one dimension of the three dimensional structure. 
     
     
         47 . The device of  claim 46 , wherein the three dimensional structure represents at least one of a model of a tissue, and a model of neuronal axons. 
     
     
         48 . The device of  claim 30 , wherein the fibers in the three dimensional structure are formed of a stretchable fiber material. 
     
     
         49 . An assay method comprising:
 given the device of  claim 30 :   contacting the assay device with a population of cells and   studying at least one feature of an interaction of the population of cells with the device.   
     
     
         50 . The assay method of  claim 49 , wherein the cells are oligodendrocytes and the at least one feature of the interaction is myelination of the plurality of fibers. 
     
     
         51 . The assay method of  claim 50 , wherein the studying comprises determining, for at least one of the plurality of fibers, both an extent of myelination along a longitudinal axis of the fiber and a thickness of myelin. 
     
     
         52 . The assay method of  claim 51 , wherein the longitudinal extent and thickness of myelin are determined from a microscopy images. 
     
     
         53 . (canceled) 
     
     
         54 . The device of  claim 5 , wherein the average diameter is between about 0.1 and about 1 micrometers. 
     
     
         55 . The device of  claim 17 , wherein the three dimensional structure represents at least one of a model of a tissue, and a model of neuronal axons.