Molecular profile of organ culture-stored corneal epithelium: Lgr5 is a potential new phenotypic marker of residual human corneal limbal epithelial stem cells

Abstract

Abstract. Long-term preservation of corneal limbal epithelium may decrease its quality and change the molecular signature of the limbal epithelial stem cells. In this study we have investigated the molecular profile of isolated corneal epithelial cells that have been in storage for an extended time. Isolated cells were characterised by the expression profile of different cytokeratins and markers of squamous metaplasia (vimentin and α-actin). Furthermore, we examined global markers of adult stem cells including p63α and ABCG2 but also LGR5 as a novel stem cell marker. Immunocytochemical staining and PCR analysis of p63α, ABCG2 and LGR5 revealed the existence of side-population cells with a stem-cell phenotype and maintenance of corneal limbal stem cell properties. LGR5 expression can be related to cellular stemness and can be considered as a new phenotypic marker of residual human corneal limbal stem cells. However, the existence of CK10 together with co-expressed α-actin and vimentin suggests that the corneas investigated were under oxidative stress and showed evidence of squamous metaplasia.

Culturing Limbal Epithelial Cells of Long-term Stored Corneal Donors (Organ Culture) In Vitro – A Stepwise Linear Regression Algorithm

“…Ex vivo expansion of LECs involves the culture of LECs harvested either from the patient [11], a living relative [16], or a cadaver [17]. It has been suggested that limbal tissue that is taken from a cadaver donor should be placed onto the recipientʼs eye within 72 hours of the donorʼs death in order to maintain cell viability [18,19]. However, it is a common practice in the United Kingdom and Europe to store corneoscleral discs intended for corneal transplantation for up to 4 weeks in an organ culture medium at 28-37 °C [20].…”

Repairing the corneal epithelium using limbal stem cells or alternative cell-based therapies

“…This is evidenced by the rapidly increasing numbers of manuscripts published over the last two decades with the keyword “limbal stem cell” searchable in PubMed (Figure 1). The quest for a bona fide LSC marker has lead to the discovery of a number of molecules that can potentially be utilized as either positive selection markers such as p63 [15], Lgr5 [16], Tcf4[17], CD157 [18], CD71 low /Integrin α6 high [19], TrkA [20], N-Cadherin [21], ABCG2 [22, 23], Cytokeratin 15[24] and ABCB5[25], or as negative selection markers, e.g. ALDH dim [26], RHAMM bright [26] and Connexin-43 [27].…”

Leucine-rich G Protein-coupled Receptor-5 Is Significantly Increased in the Aqueous Humor of Human Eye with Proliferative Diabetic Retinopathy

“…LGR5 was primarily known as a marker of self-renewing stem cells in rapidly proliferating epithelia under physiological conditions or upon injury, including the stomach [ 3 ], small intestine, colon [ 4 ], and so on [ 5 6 7 8 9 10 ]. In the visual system, it has been reported that LGR5 was detected in only two ocular tissues: the cornea in human [ 11 12 13 ] and the retina in mouse. LGR5 expressing retinal amacrine cells of mouse retina were the first piece of evidence that LGR5 was also expressed in neuronal but not stem cell lineage [ 14 15 ], and were known to possess regenerative capacity and function as an endogenous regenerative source in adult mice [ 14 ].…”