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. 2020 Dec 23;9(13):40.
doi: 10.1167/tvst.9.13.40. eCollection 2020 Dec.

Development of a Platelet Lysate-Based Printable, Transparent Biomaterial With Regenerative Potential for Epithelial Corneal Injuries

Hannah Frazer  1 , Jingjing You  1 , Zhi Chen  2 , Sepidar Sayyar  2   3 , Xiao Liu  2 , Adam Taylor  2 , Chris Hodge  1   2   4 , Gordon Wallace  2   3 , Gerard Sutton  1   2   4
Affiliations

Development of a Platelet Lysate-Based Printable, Transparent Biomaterial With Regenerative Potential for Epithelial Corneal Injuries

Hannah Frazer et al. Transl Vis Sci Technol. .

Abstract

Purpose: To develop a human platelet lysate (hPL)-based bioink that is transparent and also encompasses the regenerative properties of hPL to facilitate wound healing.

Methods: The effect of different batches of hPLand fetal bovine serum (FBS) on corneal epithelial cell growth and scratch assay was first examined using Incucyte Zoom. Various combinations of human fibrinogen (concentration range from 0.2 to 5 mg/mL) and thrombin (concentration from 1 to 10 U/mL) were combined with hPL to generate nine types of potential bioink. Rheology, transparency, and cell compatibility of bioinks were assessed and compared. The final selected bioink was used in an ex vivo corneal model to examine its ability in re-epithelization.

Results: No significant difference was detected in cell proliferation and wound healing tests between different hPL batches at the same concentration. Scratch assay data showed that hPL had significantly higher effect on wound healing than FBS. Comparing across the nine bioinks, bioink 5 consisting of 10% hPL, 2 mg/mL fibrinogen, and 5 U/mL thrombin demonstrated all required mechanical and cellular properties and was able to regenerate the full-thickness epithelium ex vivo.

Conclusions: The results showed that a transparent and adhesive bioink can be generated by combining hPL, fibrinogen, and thrombin together. The bioink can be directly applied to a human cornea to promote corneal re-epithelization with huge potential applications in corneal injuries.

Translational relevance: The developed transparent hPL-based ink with its adhesive and healing ability showed that it could be used as a new treatment option for corneal injuries.

Keywords: bioink; corneal wound healing; fibrin; human platelet lysate.

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Conflict of interest statement

Disclosure: H. Frazer (P); J. You, iFix Medical (P, I, F); Z. Chen, None; S. Sayyar, None; X. Liu, None; A. Taylor, None; C. Hodge, None; G. Wallace, None; G. Sutton, iFix Medical (P, I)

Figures

Figure 1.
Figure 1.
Comparison of effect of various hPL batch and FBS on corneal epithelial cell growth. (A) Confluence curve generated for each test condition showing cells can reach 100% confluence in all conditions. (B) Mean growth rate analysis showed that cells treated with 5% CCP5 had the lowest mean growth rate, which is significantly lower than 10% CCA4 and 10% FBS.
Figure 2.
Figure 2.
Significantly higher wound closure rate was found in hPL treated compared to FBS group (P < 0.05), and DMEM/F12 only group showed no closure of the wound as a negative control. Treatment conditions: (A) hPL, (B) FBS, and (C) DMEM only. (D) Comparison of the mean healing rate among the conditions.
Figure 3.
Figure 3.
Bioinks tested showed high transparency. (A) Mean percentage transmittance across wavelengths of the visible spectrum (400–700 nm) ± SEM for various bioinks showed that most bioinks have great than 90% transmittance except bioinks 8 and 9. (B) A representative image of the gel formed by the formulation.
Figure 4.
Figure 4.
Storage (G′), loss (G″) moduli and tan (δ) of the various bioinks ± SEM over a time-sweep oscillation of 1Hz at 34°C for 10 minutes for each bioink.
Figure 5.
Figure 5.
Storage (G′) of bioink 5 is higher than loss (G″) moduli across 10°C to 40°C.
Figure 6.
Figure 6.
Corneal epithelial cell confluence can be reached in majority of bioinks within 1 week (except bioinks 6, 8, and 9). (A) percentage of confluence for corneal epithelial cells cultured at a density of 6,000 cells per bioink (n = 6 per condition). (B) The mean rate of confluence as a percentage per day for all nine bioinks.
Figure 7.
Figure 7.
Re-epithelialization of an ex vivo, ulcerated, human cornea that had been previously rejected for transplant use. iFixInk was applied at days 1 and 4 and fixed, cryosectioned, and stained with H&E at day 7. (A) The initial ulcer, with the ulcerated regions emphasized in red. (B) The size of the epithelial-free surface after wound-debriding. (C) H&E staining of the central part of the cornea shows complete re-epithelialization with a thickness and morphology comparable to that of a healthy human cornea. (D) H&E staining of the central part of a de-epithelialized cornea for comparison.
See this image and copyright information in PMC

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