Chitosan–gelatin biopolymers as carrier substrata for limbal epithelial stem cells

Abstract

The aim of this work was to evaluate semi-synthetic biopolymers based on chitosan (CH) and gelatin (G) as potential in vitro carrier substrata for human limbal epithelial cells (hLECs). To that end, human corneal epithelial cells (HCE) were cultured onto different CH-G membranes. None of the polymers were cytotoxic and cell proliferation was higher when CH was functionalized with G. Expression levels of corneal epithelial markers (K3, K12, E-caherin, desmoplakin, and zonula occludens (ZO)-1) were better maintained in HCE cells grown on CH-G 20:80 membranes than other proportions. Consequently, CH-G 20:80 was chosen for the subsequent expansion of hLECs. Cells derived from limbal explants were successfully expanded on CH-G 20:80 membranes using a culture medium lacking components of non-human animal origin. The expression levels found for corneal (K3 and K12) and limbal epithelial stem cells (K15) specific markers were similar to or higher than those found in limbal cells grown onto the control substratum. Our results demonstrate that CH-G 20:80 membranes are suitable for the expansion and maintenance of stem cells derived from the limbal niche. These results strongly support the use of polymers as alternative substrata for the transplantation of cultivated limbal cells onto the ocular surface.

Gelatin-Based Materials in Ocular Tissue Engineering

“…Gelatin and its derivatives have been used as potential scaffolds for corneal epithelium [13], corneal endothelium [20] and retinal pigment epithelium [21], as a bioartificial corneal stroma [19] and as a potential bioadhesive in treatment of retinal detachment [17]. The range of crosslinking options used to strengthen gelatin scaffolds in this field to date is extensive.…”

“…Research into gelatin-based biomaterials for cornea tissue engineering is still preclinical and has mainly focused on providing substrates for cultivation and delivery of different corneal cell types [13]. These include the growth and delivery of corneal endothelial cells [73,74,107], growth of epithelial or limbal cells for corneal surface delivery [13,108], and investigation into the use of gelatin as a stromal replacement [18,19,69,108].…”

“…De la Mata and colleagues recently reported the use of a LESC carrier composed of chitosan and gelatin covalently bound through crosslinking with glutaraldehyde and subsequently reduced with sodium borohydride [13]. Introduction of chitosan was an attempt to mimic the glycosaminoglycan composition of the native limbus, thought to be important in maintaining the “stemness” of seeded LESCs.…”

Biopolymers in Medical Implants

“…Most often the only way to get back normal vision in such cases is by LESC transplantation. Some of the recent research reports in this area using gelatin as a biopolymer of interest includes; work done by De la Mata et al, using LESC carrier composed of gelatin and chitosan covalently bound through crosslinking with glutaraldehyde and subsequently reduced with sodium borohydride (de la Mata et al 2013). Another study report revealed the use of cushioned gelatin films loaded with epidermal growth factor as a therapeutic bandage to enhance wound healing in epithelial scars in an in vivo rabbit model (Hori et al 2007).…”

Macroporous thin membranes for cell transplant in regenerative medicine

“…This is the case of the anchorage ring of cornea prosthesis, for which a thin biointegrable biostable material is required [1][2][3][4][5][6][7][8][9]. In other cases a patch of a biostable material for cell carrier can be pressed on the damaged tissue to induce a paracrinic effect by a continuous growth factor delivery, this could find application in wound healing, and cornea or skin regeneration [10][11][12][13][14][15][16][17]. Pore architecture of these scaffolds must be quite particular to fit simultaneously requirements of high porosity (as large as possible), pore interconnectivity, and 3 pore size adequate for cell invasion, small thickness and mechanical resistance.…”