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Tissue and cellular characterisation of nucleolin in a murine model of corneal angiogenesis

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Abstract

Purpose

Corneal neovascularisation (CNV), with consequent loss of transparency, is due to an imbalance of proangiogenic factors. Cell-surface nucleolin (NCL) has been associated with neo-angiogenesis. There are studies identifying NCL translocation from nucleus to the cell surface, which is essential for endothelial cell proliferation. To find the possible role of NCL in the generation of corneal neovessels, the aim of this study is to characterise the NCL presence and cell-localisation in non-injured corneas, as well as to describe the changes in NCL cell and tissue localisation in CNV, and to analyse the effect of bevacizumab on NCL cellular and tissular distribution.

Methods

Suture-induced CNV was performed in mice. The corneal tissues were obtained and the histological and co-immunofluorescence assays were performed using different proteins, such as CD31, cadherin and isolectin B4. To determine the possible role of VEGF in NCL presence and localisation in our CNV model, bevacizumab was concomitantly used.

Results

Nucleolin was principally observed in the nucleus of the basal epithelial cells of normal corneas. Interestingly, angiogenesis-induced changes were observed in the localisation of NCL, not only in tissue but also at the cellular level where NCL was extranuclear in epithelial cells, stromal cells and neovessels. In contrast, these changes were reverted when bevacizumab was used. Besides, NCL was able to stain only aberrant corneal neovessels in comparison with retinal vessels.

Conclusions

NCL mobilisation outside the nucleus during angiogenesis could have a possible role as a proangiogenic molecule in the corneal tissue.

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Acknowledgments

The authors wish to thank Juan E. Jiménez and Mohamed Ali Pereyra Morales for their technical support. Joaquín Quiroz-Mercado received a scholarship from CONACYT-CVU: 315182. This article is part of the requirements for the Doctoral Degree of Joaquín Quiroz-Mercado in the “Programa de Doctorado en Producción Animal y Ciencias de la Salud, Facultad de Medicina Veterinaria y Zootecnia, UNAM”.

The authors have full control over all primary data and they agree to allow Graefes Archive for Clinical and Experimental Ophthalmology to review their data upon request.

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Authors and Affiliations

  1. Research Unit, Institute of Ophthalmology Conde de Valenciana Foundation, Chimalpopoca 14, 06800, Mexico City, Mexico

    Joaquín Quiroz-Mercado, Norma Ramírez-Velázquez, Graciela Partido, Maria Carmen Jiménez-Martínez & Yonathan Garfias

  2. Faculty of Veterinary Medicine and Animal Husbandry, Universidad Nacional Autónoma de México, Avenida Universidad 3000, 04510, Mexico City, Mexico

    Joaquín Quiroz-Mercado

  3. Department of Biochemistry, Faculty of Medicine, Universidad Nacional Autónoma de México, Avenida Universidad 3000, 04510, Mexico City, Mexico

    Edgar Zenteno, Raúl Chávez, Concepción Agundis-Mata, Maria Carmen Jiménez-Martínez & Yonathan Garfias

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  1. Joaquín Quiroz-Mercado
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Correspondence to Yonathan Garfias.

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Funding

CONACYT SALUD-160286; 71291, CONACYT-SEP-CIENCIA-BASICA 167438, DGAPA UNAM-PAPIIT IA203514 and Conde de Valenciana Foundation provided financial support in the form of grants. The sponsor had no role in the design or conduct of this research.

Conflict of interest

All authors certify that they have no affiliations with or involvement in any organisation or entity with any financial interest (such as honoraria; educational grants; participation in speakers’ bureaus; membership, employment, consultancy, stock ownership, or other equity interest; or expert testimony or patent-licencing arrangements) or non-financial interest (such as personal or professional relationships, affiliations, knowledge or beliefs) in the subject matter or materials discussed in this manuscript.

Ethical approval

All applicable international, national and institutional guidelines for the care and use of animals were followed. All procedures were in accordance with the ethical standards of the Instituto de Oftalmología Conde de Valenciana.

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Quiroz-Mercado, J., Ramírez-Velázquez, N., Partido, G. et al. Tissue and cellular characterisation of nucleolin in a murine model of corneal angiogenesis. Graefes Arch Clin Exp Ophthalmol 254, 1753–1763 (2016). https://doi.org/10.1007/s00417-016-3409-8

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