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Poly-?-lysine based hydrogels as synthetic substrates for the expansion of corneal endothelial cells for transplantation.

ABSTRACT: Dysfunction of the corneal endothelium (CE) resulting from progressive cell loss leads to corneal oedema and significant visual impairment. Current treatments rely upon donor allogeneic tissue to replace the damaged CE. A donor cornea shortage necessitates the development of biomaterials, enabling in vitro expansion of corneal endothelial cells (CECs). This study investigated the use of a synthetic peptide hydrogel using poly-?-lysine (p?K), cross-linked with octanedioic-acid as a potential substrate for CECs expansion and CE grafts. P?K hydrogel properties were optimised to produce a substrate which was thin, transparent, porous and robust. A human corneal endothelial cell line (HCEC-12) attached and grew on p?K hydrogels as confluent monolayers after 7 days, whereas primary porcine CECs (pCECs) detached from the p?K hydrogel. Pre-adsorption of collagen I, collagen IV and fibronectin to the p?K hydrogel increased pCEC adhesion at 24?h and confluent monolayers formed at 7 days. Minimal cell adhesion was observed with pre-adsorbed laminin, chondroitin sulphate or commercial FNC coating mix (fibronectin, collagen and albumin). Functionalisation of the p?K hydrogel with synthetic cell binding peptide H-Gly-Gly-Arg-Gly-Asp-Gly-Gly-OH (RGD) or ?2?1 integrin recognition sequence H-Asp-Gly-Glu-Ala-OH (DGEA) resulted in enhanced pCEC adhesion with the RGD peptide only. pCECs grown in culture at 5 weeks on RGD p?K hydrogels showed zonula occludins 1 staining for tight junctions and expression of sodium-potassium adenosine triphosphase, suggesting a functional CE. These results demonstrate the p?K hydrogel can be tailored through covalent binding of RGD to provide a surface for CEC attachment and growth. Thus, providing a synthetic substrate with a therapeutic application for the expansion of allogenic CECs and replacement of damaged CE.

SUBMITTER: Kennedy S

PROVIDER: S-EPMC6726667 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Poly-ε-lysine based hydrogels as synthetic substrates for the expansion of corneal endothelial cells for transplantation.

Kennedy Stephnie S Lace Rebecca R Carserides Constandinos C Gallagher Andrew G AG Wellings Donald A DA Williams Rachel L RL Levis Hannah J HJ

Journal of materials science. Materials in medicine 20190904 9

Dysfunction of the corneal endothelium (CE) resulting from progressive cell loss leads to corneal oedema and significant visual impairment. Current treatments rely upon donor allogeneic tissue to replace the damaged CE. A donor cornea shortage necessitates the development of biomaterials, enabling in vitro expansion of corneal endothelial cells (CECs). This study investigated the use of a synthetic peptide hydrogel using poly-ε-lysine (pεK), cross-linked with octanedioic-acid as a potential subs ...[more]

PMID: 31485761

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Project description:PurposeCorneal endothelial cell density (ECD) gradually decreases after corneal transplantation by unknown biologic, biophysical, or immunologic mechanism. Our purpose was to assess the association between donor corneal endothelial cell (CEC) maturity in culture and postoperative endothelial cell loss (ECL) after successful corneal transplantation.DesignProspective cohort study.ParticipantsThis cohort study was conducted at Baptist Eye Institute, Kyoto, Japan, between October 2014 and October 2016. It included 68 patients with a 36-month follow-up period who had undergone successful Descemet stripping automated endothelial keratoplasty (DSAEK) or penetrating keratoplasty.MethodsHuman CECs (HCECs) from remaining peripheral donor corneas were cultured and evaluated for maturity by surface markers (CD166+, CD44-/dull, CD24-, and CD105-) using fluorescence-activated cell sorting. Postoperative ECD was assessed according to the mature-differentiated HCEC contents: high-maturity group: > 70%, middle-maturity group: 10% to 70%, low-maturity group: < 10%. The successful rate of ECD maintained at 1500 cells/mm2 at 36 months postoperative was analyzed using the log-rank test.Main outcome measuresEndothelial cell density and ECL at 36 months postoperative.ResultsThe 68 included patients (mean [standard deviation] age 68.1 [13.6] years, 47.1% women, 52.9% DSAEK). The high, middle, and low-maturity groups included 17, 32, and 19 eyes, respectively. At 36 months postoperative, the mean (standard deviation) ECD significantly decreased to 911 (388) cells/mm2 by 66% in the low-maturity group, compared with 1604 (436) by 40% and 1424 (613) cells/mm2 by 50% in the high and middle-maturity groups (P < 0.001 and P = 0.007, respectively) and the low-maturity group significantly failed to maintain ECD at 1500 cells/mm2 at 36 months postoperative (P < 0.001). Additional ECD analysis for patients who underwent DSAEK alone displayed a significant failure to maintain ECD at 1500 cells/mm2 at 36 months postoperative (P < 0.001).ConclusionsThe high content of mature-differentiated HCECs expressed in culture by the donor peripheral cornea was coincident with low ECL, suggesting that a high-maturity CEC content predicts long-term graft survival. Understanding the molecular mechanism for maintaining HCEC maturity could elucidate the mechanism of ECL after corneal transplantation and aid in developing effective interventions.Financial disclosuresProprietary or commercial disclosure may be found after the references.

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Poly-?-lysine based hydrogels as synthetic substrates for the expansion of corneal endothelial cells for transplantation.

Publications

Poly-ε-lysine based hydrogels as synthetic substrates for the expansion of corneal endothelial cells for transplantation.

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