Smooth muscle tissue engineering in crosslinked electrospun gelatin scaffolds

Abstract

Crosslinked, multi-layer electrospun gelatin fiber scaffolds with generally ±45 degree fiber orientation have been used to grow human umbilical vein smooth muscle cells (HUVSMCs) to create a vascular tunica media graft. Scaffolds of different fiber diameter (2-5 μm in wet state), pore size, and porosity (16-21% in wet state) were assessed in terms of cell adherence and viability, cell proliferation, and migration in both in-plane and transverse directions through the scaffold as a function of time under static cell culture conditions. HUVSMC cell viability reached between 80 and 92% for all scaffolds after 9 days in culture. HUVSMCs adhered, elongated, and orientated in the fiber direction, and migrated through a scaffold thickness of 200-235 μm 9 days post-seeding under static conditions. The best scaffold was then used to assess the tissue engineering of HUVSMCs under dynamic conditions for a rotating, cell seeded, tubular scaffold in the bioreactor containing the culture medium. Dynamic conditions almost doubled the rate of cell proliferation through the scaffold, forming full tissue throughout a scaffold of 250-300 μm thickness 6 days post-seeding.

Modeling, simulations, and optimization of smooth muscle cell tissue engineering for the production of vascular grafts

“…The predicted cell front propagation through the depth of the scaffold was based on the definition of the cell front at a normalized cell density (cell area fraction) of 0.5 for the cell layer attached to the fiber surface. Figure a,b, demonstrate good agreement between the predictions of this study and the experimental data by Elsayed et al (b), with the line of best fit through the predicted values after 3, 6, and 9 days of cell culture passing through the origin and crossing the experimental data for both scaffolds. Using these sets of experimental data, the cell migration coefficient in the transverse ( x ‐) and in‐plane ( y ‐) directions was fitted to the values presented in Table for a static bioreactor, and as can be seen, it was found that the migration coefficient of the SMCs is 10 times higher in the transverse direction through the fibrous scaffold than in the in‐plane direction; these values were used for all scaffolds in the validation and parametric studies in a static bioreactor in section 4.1.…”

“…Computational simulations of smooth muscle arterial tissue engineering were conducted for both a static and a dynamic bioreactor. In each case‐study, a first set of simulations were conducted with the aim at validating the theoretical model and fitting the values for the cell migration coefficients, D x , D y , by comparing the predictions against experimental data (Elsayed et al, a, b). The second stage of computational simulations aimed at the optimization of the microstructural parameters of the scaffold, by conducting a series of parametric studies with different values of the scaffold porosity and fiber diameter and constructing maps from which the best parameters for a scaffold would be selected on the basis of maximum cell proliferation within the shortest time, whereas the predicted oxygen concentration is above the threshold that maintains cell survival.…”

“…Table presents the value of the input parameters used for the computational simulations to simulate experiments presented by Elsayed et al (a, b). In the experiments, gelatin fiber scaffolds were electrospun as stacks of alternating +45 and −45° unidirectional fiber layers, forming a bidirectional fiber assembly to optimize the mechanical properties of the scaffold (Elsayed et al, ).…”

“…In the SMC tissue engineering experiments described by Elsayed et al (a, b), samples of the cell‐seeded scaffolds were removed after a certain number of days in the culture medium under static or dynamic conditions and subjected to a number of tests. For scanning electron microscopy (SEM), the samples were washed in phosphate buffered saline solution (phosphate buffered saline solution), fixed in 10 wt% glutaraldehyde overnight, dehydrated in a graded series of ethanol/water solution and dried in a vacuum desiccator overnight.…”

“…Cell migration has been the focus of research within our group and we have identified directional differences in the 3D migration of cells, depending on the size and shape of cells in relation with the pores of the scaffold. In particular, such differences have been revealed in experimental tissue engineering studies between smooth muscle cells (SMCs; Elsayed, Lekakou, Labeed, & Tomlins, b) and osteoblasts (Salifu, Lekakou, & Labeed, ) which points to the need for different phenomenological models of cell migration and proliferation for each type of cells with respect to the scaffold pore size and geometry. Therefore, a 3D cell migration model was adopted in our analysis for the SMC tissue engineering in fibrous scaffolds, with transverse and in‐plane migration where different migration coefficients may be assigned in each direction.…”