Nanofiber Enabled Layer-by-Layer Approach Toward Three-Dimensional Tissue Formation

Abstract

Taking rapid and efficient formation of functional tissues as our long-term goal, we discuss in this study a new and generic approach toward formation of multilayered three-dimensional (3D) tissues using nanofibers. 3:1 poly (epsilon-caprolactone) (PCL) (8% w/v)/collagen (8.0% w/v) solution was electrospun into nanofibers with an average diameter of 454.5 +/- 84.9 nm. The culture of human dermal fibroblasts (NHDF) on PCL/collagen nanofibers showed a high initial cell adhesion (88.1 +/- 1.5%), and rapid cell spreading with spindle morphology. Three-dimensional multilayered cell-nanofiber constructs were built with alternating NHDF seeding (1 x 10(5)cells/layer) and PCL/collagen nanofiber collection on site of electrospinning, where almost all the seeded cells retained in the constructs. The formed construct showed layered structure with uniform cell distribution in between layers of PCL/collagen nanofibers. In the 3D constructs, cells continuously proliferated and deposited new extracellular matrix. By culturing either fibroblast/fiber layered constructs or keratinocyte/fibroblast/fiber layered constructs, dermal-like tissues or bilayer skin tissues (containing both epidermal and dermal layers) were consequently produced within 1 week. Taken together, the present study reports a novel approach to 3D multilayered tissue formation using a bottom-up, on-site layer-by-layer cell assembly while electrospinning. This approach has marked potentials to form functional tissues composed of multiple types of cells, heterogeneous scaffold composition, and customized specific microenvironment for cells.

Electrospinning jets and nanofibrous structures

“…These scaffolds were found conducive to fibroblast growth and proliferation. Yang et al 152 created a layered structure of polymers and cells by electrospinning PCL and collagen together, seeding human dermal fibroblasts, and then repeating the polymer and cell layers. These cellularized constructs were done in sheet form and not with tubes.…”

Patterned Electrospun Nanofiber Matrices Via Localized Dissolution: Potential for Guided Tissue Formation

“…[ 1 ] Due to their dimensional similarity to the native tissue extracellular matrix (ECM), electrospun matrices have been explored for versatile applications in regenerative medicine aiming to promote cell-fi ber interactions for regulation of cell phenotype, [ 2 ] direct tissue regeneration via contact guidance, [ 3 ] support stem-cell proliferation and differentiation, [ 4 ] and create three-dimensional (3D) tissues with cells encapsulated inside the fi bers [ 5,6 ] or embedded among the fi bers. [ 7 ] …”

Nanotechnology meets regenerative medicine: a new frontier?

“…Yang et al explored a 3-D multilayered cell-nanofiber scaffold with alternating layers of human dermal FBs or keratinocytes seeded on PCL/collagen nanofibrous sheets. The results indicated that both kinds of cells seeded could effectively proliferate and contribute to the formation of new ECM, which would indicate the potential to be a substitute of the skin [51] .…”