Biology subjects

Henry, J. J.

Publications and source records attributed to Henry, J. J..

3 recordsLinked to original sources

bioRxiv2020

Single-cell analyses of the corneal epithelium: Unique cell types and gene expression profiles

Corneal Epithelial Stem Cells (CESCs) and their proliferative progeny, the Transit Amplifying Cells (TACs), are responsible for homeostasis and maintaining corneal transparency. Owing to our limited knowledge of cell fates and gene activity within the cornea, the search for unique markers to identify and isolate these cells remains crucial for ocular surface reconstruction. We performed single-cell RNA sequencing of corneal epithelial cells from stage 49-51 Xenopus larvae. We identified five main clusters with distinct molecular signatures, which represent apical, basal and keratocyte cell types as well as two discrete proliferative cell types in the bi-layered epithelium. Our data reveal several novel genes expressed in corneal cells and spatiotemporal changes in gene expression during corneal differentiation. Through gene regulatory network analysis, we identified key developmental gene regulons, which guide these different cell states. Our study offers a detailed atlas of single-cell transcriptomes in the frog corneal epithelium. In future, this work will be useful to elucidate the function of novel genes in corneal homeostasis, wound healing and cornea regeneration, which includes lens regeneration in Xenopus. SUMMARY STATEMENTThis study identifies cell types and transcriptional heterogeneity in the corneal epithelium that regulate its differentiation, and facilitates the search for corneal stem cell markers.

developmental biology↗

bioRxiv2019

Understanding the basis of CYP26 mediated regulation of lens regeneration using ex vivo eye cultures and 4-oxo-RA

PURPOSEXenopus has the remarkable ability to regenerate a lens from the basal cornea epithelial cells in response to signals from the retina. Previous work demonstrated that the Retinoic Acid (RA) metabolizing enzyme CYP26 is expressed in the cornea, and that its activity is required for lens regeneration. Gaps remain in our knowledge as to whether CYP26 is needed only to attenuate RA signaling via RA elimination, or whether it also acts to generate retinoid metabolites, such as 4-oxo-RA, to act as signaling ligands. Other key questions are why CYP26 antagonism, but not exogenous retinoids, can reduce cell division in the cornea, and when during regeneration CYP26 is important.\n\nMATERIALS AND METHODSEx vivo cultures supplemented with RA, 4-oxo-RA, or the CYP26 inhibitor Liarozole were used to assay the effects of these compounds on lens regeneration. Similarly, corneas were explanted, cultured in the presence of these compounds, and assayed for mitotic changes by counting anti-Histone H3 positive nuclei. qPCRs validated responsiveness to these compounds.\n\nRESULTSEx vivo cultures showed that when the media was supplemented with the RA metabolite 4-oxo-RA in addition to Liarozole, lens regeneration was still inhibited. 4-oxo-RA also does not rescue the loss of cell division in the cornea that is observed upon CYP26 antagonism. Liarozole inhibited regeneration when added 12 hours after lentectomy, but not when added 48 hours after.\n\nCONCLUSIONSThese data show that the necessity of CYP26 is not explained as a generator of 4-oxo-RA for regeneration. Moreover, Liarozole-induced mitotic reduction is not explained by 4-oxo-RA deficiency. These results support a model of RA-independent mitotic regulation by CYP26, though other retinoid metabolites may be active. Finally, CYP26 activity is only needed between 12 and 48 hours post-surgery, showing that its action is required only during the earliest stages of lens regeneration.\n\nFinancial interestsThe authors declare no competing financial interests.

developmental biology↗

bioRxiv2018

FGF1 Promotes Xenopus laevis Lens Regeneration

BackgroundThe frog Xenopus laevis has notable regenerative capabilities, including that of the lens. The neural retina provides the factors that trigger lens regeneration from the cornea, but the identity of these factors is largely unknown. In contrast to the cornea, fibroblast growth factors FGF1, 8, and 9 are highly expressed within the retina, and are potential candidates for those factors. The purpose of this study is to determine whether specific FGF proteins can induce lens formation, and if perturbation of FGFR signaling inhibits lens regeneration.\n\nMethodsA novel cornea epithelial culture method was developed to investigate the sufficiency of FGFs in lens regeneration. Additionally, transgenic larvae expressing dominant negative FGFR1 were used to investigate the necessity of FGFR signaling in lens regeneration.\n\nResultsTreatment of cultured corneas with FGF1 induced lens regeneration in a dose-dependent manner, whereas treatment with FGF2, FGF8, or FGF9 did not result in significant lens regeneration. Inhibition of FGFR signaling decreased the lens regeneration rate for in vitro eye cultures.\n\nConclusionThe culture techniques developed here, and elsewhere, have provided reliable methods for examining the necessity of various factors that may be involved in lens regeneration. Based on the results demonstrated in this study, we found that FGF1 signaling and FGFR activation are key factors for lens regeneration in Xenopus.

developmental biology↗