Current microfluidic platforms for reverse engineering of cornea

Abstract

According to the World Health Organization, corneal blindness constitutes 5.1% of global blindness population. Surgical outcomes have been improved significantly in the treatment of corneal blindness. However, corneal transplantation is limited by global shortage of donor tissue, prompting researchers to explore alternative therapies such as novel ocular pharmaceutics to delay corneal disease progression. Animal models are commonly adopted for investigating pharmacokinetics of ocular drugs. However, this approach is limited by physiological differences in the eye between animals and human, ethical issues and poor bench-to-bedside translatability. Cornea-on-a-chip (CoC) microfluidic platforms have gained great attention as one of the advanced in vitro strategies for constructing physiologically representative corneal models. With significant improvements in tissue engineering technology, CoC integrates corneal cells with microfluidics to recapitulate human corneal microenvironment for the study of corneal pathophysiological changes and evaluation of ocular drugs. Such model, in complement to animal studies, can potentially accelerate translational research, in particular the pre-clinical screening of ophthalmic medication, driving clinical treatment advancement for corneal diseases. This review provides an overview of engineered CoC platforms with respect to their merits, applications, and technical challenges. Emerging directions in CoC technology are also proposed for further investigations, to accentuate preclinical obstacles in corneal research.

Keywords: Cornea-on-a-chip; Corneal microenvironment; Microfluidic platform; Pharmacokinetics; Tissue engineering.

Graphical abstract

Fig. 1

(A) Schematic anatomy of the human eyeball structure, with the layered structure of the cornea highlighted. (B) Detailed illustration of the five tissue layers within the cornea.

Fig. 2

Representative designs of CoC platforms for corneal epithelial barrier studies. (A) (i) Schematic overview of the hybrid microfluidic device and (ii) Confocal fluorescence images of corneal epithelial cells (green) and keratocytes (red) (Reproduced Ref. [56] with permission of Royal Society of Chemistry, Copyright 2009 Royal Society of Chemistry). (B) The photo and structural illustration of CEpOC, and its applicability using untargeted LC-MS equipment (Reproduced Ref. [57] with permission of Elsevier, Copyright 2021 Elsevier). (C) (i) Exploded assembly diagram of the DynaMiTES unit (Reproduced Ref. [59] with permission of Elsevier, Copyright 2017 Elsevier), and (ii) detailed components of the systematic ocular DynaMiTES (Reproduced Ref. [60] with permission of Elsevier, Copyright 2017 Elsevier). (D) Schematic diagram of (i) the dynamic CoC simulation of anterior ocular architecture, (ii) device fabrication, (iii) breakdown structure and (iv) principle of operation. (v) Photograph of demonstration with two dyes in the individual channels (Reproduced Ref. [34] with permission of Royal Society of Chemistry, Copyright 2018 Royal Society of Chemistry). (E) (i) Tear flow dynamics during eye blinking, (ii) illustration of the multi-layered flow perfusion CoC, and (iii) immunofluorescence photographs of ZO-1 (green) in HCEpCs (Reproduced Ref. [58] with permission of Royal Society of Chemistry, Copyright 2020 Royal Society of Chemistry). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 3

Corneal stroma chips with different applications. (A) (i) Schematic procedure of chip fabrication, (ii) the immunocytochemistry results, and (iii) effect of LBP pre-treatment on secretion of IL-6 and IL-8 proteins from keratocytes (Reproduced Ref. [61] with permission of Elsevier, Copyright 2021 Elsevier). (B) Fabrication process of the PDMS microfluidic device and aligned collagen fibril formation [62].

Fig. 4

Corneal endothelium-related CoC and representative eye blinking model. (A) (i) Schematic diagrams of cross-sectional culture zone, exploded view and the top view of the device with double-channel structure; (ii) photograph of the cornea chip; (iii) ZO-1 (green) and DAPI (blue) staining results in different conditions (‘Chip’ means the coculture of CEpCs and CEdCs; MFI: mean fluorescent intensity); (iv) CEpC migration results during wound healing under two different culture conditions (Reproduced Ref. [63] with permission of Elsevier, Copyright 2022 Elsevier). (B) (i) The diagram of the microfluidic device with multichannel; (ii) Microscopy images of different layers at 24 h after seeding (Reproduced Ref. [9] with permission of Creative Commons Attribution License, Copyright 2020 Creative Commons Attribution License). (C) Schematic of the (i) representative CoC setup to simulate eye blinking with an electromechanical actuator, and (ii) eyelid movement and changes of the tear film during spontaneous eye blinking [10]. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 5

Different corneal cell cultures using specific ECM environment. (A) (i) Schematic diagram of overall CEdC culture setup with biomimetic topography and electric fields (EF), and (ii) analysis of the protein with and without EFs (Reproduced Ref. [103] with permission of Elsevier, Copyright 2014 Elsevier). (B) (i) Silk film functionalization and (ii) preparation of 3D functional corneal stroma on the silk film [102].

Fig. 6

Representative CEdC-related research with various topographical patterns and in vitro corneal nerve models. (A) (i) Scanning electron microscope characterization and (ii) effect of topographical features on endothelial cell morphology (Reproduced Ref. [105] with permission of Elsevier, Copyright 2012 Elsevier) (B) Preparation process of PDMS substrate and (C) (i) patterned mold of natural white rose and (ii) substrate for cell culture (Reproduced Ref. [108] with permission of Elsevier, Copyright 2021 Elsevier). (D) Schematic diagram of 3D engineered corneal nerve model (Reproduced Ref. [109] with permission of Elsevier, Copyright 2016 Elsevier). (E) (i) Sketch map and picture of the corneal nerve-stroma model and (ii) fluorescence images at day 7 (Green: β tubulin-III labeled nerve bundles; Red: vimentin labeled CSCs; Blue: DAPI labeled nuclei) (Reproduced Ref. [110] with permission of Elsevier, Copyright 2022 Elsevier). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)