Biofunctionalized electrospun silk mats as a topical bioactive dressing for accelerated wound healing

Abstract

Materials able to deliver topically bioactive molecules represent a new generation of biomaterials. In this article, we describe the use of silk mats, made of electrospun nanoscale silk fibers containing epidermal growth factor (EGF), for the promotion of wound healing processes. In our experiments, we demonstrated that EGF is incorporated into the silk mats and slowly released in a time-dependent manner (25% EGF release in 170h). We tested these materials using a new model of wounded human skin-equivalents displaying the same structure as human skin and able to heal using the same molecular and cellular mechanisms found in vivo. This human three-dimensional model allows us to demonstrate that the biofunctionalized silk mats, when placed on the wounds as a dressing, aid the healing by increasing the time of wound closure by the epidermal tongue by 90%. The preservation of the structure of the mats during the healing period as demonstrated by electronic microscopy, the biological action of the dressing, as well as the biocompatibility of the silk demonstrate that this biomaterial is a new and very promising material for medical applications, especially for patients suffering from chronic wounds.

Fig. 1

(A) Electrospinning apparatus for silk mat preparation. The fibroin solution is mixed with the growth factor in the syringe before electrospinning the silk. (B) Methanol treated EGF-incorporated silk mat. The mat appears like a small piece of tissue that can be cut to fit the wound.

Fig. 2

SEM pictures of the electrospun silk mats. The mats consist of randomly distributed silk fibers of about 1 μm in diameter

Fig. 3

Construction of composite organotypic co-culture wound healing model. A wound is formed by punching a hole through the epithelium and collagen matrix. (A) Picture of the wound after the transfer of the punched human skin-equivalent onto a second collagen matrix that has undergone contraction (layer I), the resultant composite co-culture consists of two layers of contracted matrix and one layer of epithelium. (B and C) Schematic and picture of the wound with the silk mat, reepithelialization occurs as wounded keratinocytes migrate under the silk mats onto the collagen in layer I. The percentages of wound closure are a measure of the distance between the edges and between the epithelial tongues to obtain the percentage of wound closure. Scale bar (A) and (C): 10 mm.

Fig. 4

In vitro release profile of EGF from the electrospun silk mat at 37 ± 0.5 °C (n = 6) over 7 days.

Fig. 5

SEM pictures of the electrospun silk mats before contact on the wound (A and B), after 72 h soaked in PBS at 37 °C (C and D) and after contact on the wound for 72 h (E and F). Note the difference observed on the diameter of the fibers and the irregularities in the diameter of the fibers that contacted the wound. Bars: (A, C, and E): 20 μm; (b, d, and f): 5 μm

Fig. 6

Hematoxylin and Eosin staining of the wound after 24 (left column) and 48 h (right column). (A) and (B) are controls, without the silk dressing on top of the wound. (C) and (D) are the samples on which we applied a silk mat without EGF. (E) and (F) are the samples on which a silk mat containing the EGF has been applied. Lower magnification: scale bar equals 100 μm, higher magnification: scale bar equals 300 μm.

Fig. 7

Percentage of wound closure after 24 (gray) and 48 h (black). Data points are the average of n = 7 and error bars represent ± SD.