For the first time, we demonstrated that functional neural-like cells can be derived from neural crestderived limbal cells. The aim of this study is now to investigate whether mouse and human limbal neurosphere cells can differentiate into retinal like cells both in vivo and in vitro after exposure to a developing retinal microenvironment. We have previously demonstrated that adult mouse LNS are neural crest-derived limbal stromal stem/progenitor cells. They can generate functional neural-like cells in vitro. We now report that these cells have the potential for differentiation towards a retinal lineage. The expression of photoreceptor specific markers at a transcript level indicates endogenous expression of these photoreceptor specific genes. Moreover, the major retinal synapse component syntaxin3 and sensory cilia were also observed, as would be expected in differentiated retinal cells. After transplantation into the SRS, expression of rhodopsin, recoverin and syntaxin3 were also detected in grafted LNS cells. For the first time, this work demonstrates that neural crest-originated limbal stromal stem/ progenitor cells have the potential to generate retinal-like cells both in vitro and in vivo. We VE-821 further investigated whether LNS can be generated from aged human eyes, and whether they have a similar potential to generate retinal cells. We generated LNS from aged human limbal tissue from donors up to 97 years of age. The upregulation of retinal progenitor markers such as Lhx2, Pax6 and Rx was noted following culture in permissive conditions in vitro. To the best of our knowledge, this is the first evidence showing that human LNS have the plasticity to express retinal progenitor markers. However, mature photoreceptor markers were not observed. The lack of further cell maturation suggests that more comprehensive intrinsic and extrinsic regulation is needed cf. mouse cells. Extrinsic factors released by PN1–3 mouse retinal cells may be insufficient in promoting further differentiation of human cells towards a mature retinal cell lineage. We also co-cultured human LNS with fetal week 7–8 human retinal cells. LNS did not appear to differentiate towards a retinal lineage in this circumstance. This may be due to the fact that retinal tissue/cells at the gestational stage of Fwk 7–8, have not started rod genesis. Our observation is consistent with a previous report that rod promoting activity is only observed in retinal cells at the peak of rod genesis, but not at an early developmental stage or in adult retinal cells. Due to ethical concerns, we were unable to access later stage human fetal retinal tissues. However, it is encouraging that human LNS expressed retinal progenitor markers when exposed to several defined factors including Shh, Taurine and RA. Shh has been shown to be involved in the formation of the ventral optic cup, specification of dorso-ventral polarity in the optic vesicle, and governing of ocular morphogenesis. Besides specification of the eye field during embryonic development, Shh also has been implicated in the control of retinal development in vertebrates and is required for the maintenance of retinal progenitor cell proliferation.