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Open Access Research article

Glycosylated VCAM-1 isoforms revealed in 2D western blots of HUVECs treated with tumoral soluble factors of breast cancer cells

Delina Montes-Sánchez123, Jose Luis Ventura12, Irma Mitre12, Susana Frías12, Layla Michán12, Aurora Espejel-Nuñez4, Felipe Vadillo-Ortega5 and Alejandro Zentella12*

Author Affiliations

1 Departamento de Medicina Genómica y Toxicología Ambiental, Instituto de Investigaciones Biomédicas, UNAM. Ciudad Universitaria, Circuito Interior apartado postal 70228, CP04510, México DF

2 Departamento de Bioquímica, Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", SSA Vasco de Quiroga no 15 Colonia Sección XVI, Delegación Tlalpan. CP.4080, México DF

3 Dirección de Investigación Básica, Instituto Nacional de Cancerología, SSA Av San Fernando no 22 Col. Sección XVI, Tlalpan, CP 14080, México DF

4 Dirección de Investigación, Instituto Nacional de Perinatología, SSA Montes Urales 800 Torre de Investigación 5° piso, Colonia Lomas Virreyes, Delegación Miguel Hidalgo, CP11000, México DF

5 Edificio de Investigación, Facultad de Medicina, UNAM. Ciudad Universitaria, Circuito Interior apartado postal 70228, CP04510, México DF

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BMC Chemical Biology 2009, 9:7  doi:10.1186/1472-6769-9-7

Published: 22 November 2009

Abstract

Background

Several common aspects of endothelial phenotype, such as the expression of cell adhesion molecules, are shared between metastasis and inflammation. Here, we analyzed VCAM-1 variants as biological markers of these two types of endothelial cell activation. With the combination of 2-DE and western blot techniques and the aid of tunicamycin, we analyzed N-glycosylation variants of VCAM-1 in primary human endothelial cells stimulated with either TNF or tumoral soluble factors (TSF's) derived from the human breast cancer cell line ZR75.30.

Results

Treatments induced a pro-adhesive endothelial phenotype. 2D western blots analysis of cells subjected to both treatments revealed the expression of the two known VCAM-1 isoforms and of previously unknown isoforms. In particular TSFZR75.30 induced an isoform with a relative molecular mass (Mr) and isoelectric point (pI) of 75-77 kDa and 5.0, respectively.

Conclusion

The unknown isoforms of VCAM-1 that were found to be overexpressed after treatment with TSF's compared with TNF, could serve as biomarkers to discriminate between inflammation and metastasis. 2D western blots revealed three new VCAM-1 isoforms expressed in primary human endothelial cells in response to TSF stimulation. Each of these isoforms varies in Mr and pI and could be the result of differential glycosylation states.