3D-printed ABCB5-positive stem cells for treating bilateral limbal stem cell deficiency

Abstract

Purpose : Limbal stem cells (LSC) residing in the limbus continually repopulate the corneal epithelium. Limbal stem cell deficiency (LSCD) occurs when these LSC are damaged (due to trauma such as burns) or missing (due to genetic conditions). Patients with LSCD are unable to regenerate the corneal epithelium, resulting in blindness due to invasion of the conjunctiva and neovascularization. For patients with unilateral LSCD, transplantation of autologous limbal tissue or ex vivo expanded limbal cells from the unaffected eye can be used to treat LSCD. However, patients with inflammation as well as those with severe pathologies resulting in total, bilateral LSCD have no source of autologous LSC and must rely on allogeneic transplants, associated with poor outcomes and requiring lifelong immunosuppression. Such patients would greatly benefit from an alternative autologous source of stem cells and reconstructed LSC niche to sustain donor stem cells. We previously demonstrated that human ABCB5+ LSC were capable of restoration of the corneal epithelium in an NSG mouse model of LSCD. We found that ABCB5 is also expressed by skin stem cells and hypothesized that these dermal cells could provide an alternative source of stem cells for corneal epithelial regeneration.

Methods : Human ABCB5+ dermal stem cells (DSC) expanded in vitro and purified by cell sorting were cultured in corneal differentiation media to determine their ability to transform into corneal epithelial cells. ABCB5+ DSC were also transplanted onto NSG mice with mechanically induced LSCD. In vitro, a 3D bioprinter was used to layer collagen and precisely deliver ABCB5+ DSC to the peripheral rim of human central corneas, their normal anatomical location, which we predict will enhance reconstruction of the LSC niche.

Results : Human ABCB5+ DSC were induced in vitro to express significant levels of PAX6 and KRT12 and mice transplanted with human purified ABCB5+ DSC had clearer corneas compared to mice transplanted with carrier only or ABCB5- cells (Fig 1). Bioprinted ABCB5+ DSC surrounding a human central cornea (Fig 2).

Conclusions : Our results support the use of 3D-printed ABCB5+ DSC as an alternative autologous source of stem cells to regenerate the corneal epithelium and reconstruct the LSC niche in patients with bilateral LSCD.

This is a 2021 ARVO Annual Meeting abstract.

 

ABCB5+ DSC transplanted onto NSG mice with mechanically induced LSCD.

 

ABCB5+ DSC bioprinted on the periphery of human central corneas.