Recognition of Cervical Cancer Cells by the Amaranthus leucocarpus Lectin
Daniela Illescas-Barbosa
Faculty of Medicine and Surgery, Universidad Autónoma Benito Juárez de Oaxaca, Ex Hacienda de Aguilera S/N, Calzada San Felipe del Agua, C.P. 68120, Oaxaca de Juárez, Oaxaca, México.
Pedro Antonio Hernández-Cruz
Faculty of Medicine and Surgery, Universidad Autónoma Benito Juárez de Oaxaca, Ex Hacienda de Aguilera S/N, Calzada San Felipe del Agua, C.P. 68120, Oaxaca de Juárez, Oaxaca, México.
Itandehui Belem Gallegos-Velasco
Faculty of Medicine and Surgery, Universidad Autónoma Benito Juárez de Oaxaca, Ex Hacienda de Aguilera S/N, Calzada San Felipe del Agua, C.P. 68120, Oaxaca de Juárez, Oaxaca, México.
Saira Karina Ramírez-Thomé
Faculty of Dentistry, Universidad Autónoma Benito Juárez de Oaxaca, Avenida Universidad S/N, Ex-Hacienda de Cinco Señores, Ciudad Universitaria, C.P. 68120, Oaxaca de Juárez, Oaxaca, México.
Taurino Amilcar Sosa-Velasco
Faculty of Dentistry, Universidad Autónoma Benito Juárez de Oaxaca, Avenida Universidad S/N, Ex-Hacienda de Cinco Señores, Ciudad Universitaria, C.P. 68120, Oaxaca de Juárez, Oaxaca, México and Nursing Faculty, Universidad Autónoma Benito Juárez de Oaxaca, Avenida Universidad S/N, Ex-Hacienda de Cinco Señores, Ciudad Universitaria, C.P. 68120, Oaxaca de Juárez, Oaxaca, México.
Edgar Zenteno
Faculty of Medicine, Universidad Nacional Autónoma de México, Circuito Interior, Av. Universidad 3000, Ciudad Universitaria, Coyoacán, C.P. 04510, Mexico City, Mexico.
Carlos Josué Solórzano-Mata
*
Faculty of Medicine and Surgery, Universidad Autónoma Benito Juárez de Oaxaca, Ex Hacienda de Aguilera S/N, Calzada San Felipe del Agua, C.P. 68120, Oaxaca de Juárez, Oaxaca, México and Faculty of Dentistry, Universidad Autónoma Benito Juárez de Oaxaca, Avenida Universidad S/N, Ex-Hacienda de Cinco Señores, Ciudad Universitaria, C.P. 68120, Oaxaca de Juárez, Oaxaca, México.
*Author to whom correspondence should be addressed.
Abstract
Cervical cancer is strongly associated with human papillomavirus (HPV) infection and is accompanied by alterations in cellular glycosylation. This study evaluated glycoproteins recognized by Amaranthus leucocarpus lectin (ALL), which binds O-linked carbohydrate structures including T and Tn antigens, in HPV-positive and HPV-negative cervical cancer cells. HPV-positive CaLo and SiHa cells and HPV-negative ViBo cells were examined using lectin cytochemistry, flow cytometry, electrophoresis, Western blotting and lectin-based precipitation. Healthy cervical tissues (n = 5), cervical cancer tissues (n = 5) and cell lines were evaluated by fluorescence microscopy (n=3). The specificity of ALL lectin binding in the cell lines was also assessed by inhibition using specific carbohydrates and glycoproteins (GalNAc, lactose, fetuin and bovine submaxillary mucin). ALL showed greater recognition of HPV-positive cells than of HPV-negative ViBo cells. Inhibition assays using GalNAc and fetuin completely prevented lectin recognition in the cell lines. Flow cytometry (n=2) demonstrated ALL lectin-positive populations of 93.1% in CaLo, 97.2% in SiHa and 90.0% in ViBo cells, with mean fluorescence intensity values of 48,700, 44,300 and 29,800, respectively; the corresponding means reported in Table I were 48,700, 44,300 and 29,800. Western blot analysis (n=3) of total cell lysates revealed approximately 65- and 100-kDa lectin-reactive bands in all three cell lines. Analysis of precipitated glycoproteins demonstrated bands of approximately 100, 50–70 and 25 kDa in CaLo and SiHa cells, whereas a prominent approximately 25-kDa band was observed in ViBo cells. These preliminary findings indicate differences in ALL-reactive O-glycoprotein patterns among the cervical cancer cell lines examined. Further studies using additional cell models and direct molecular characterization are required to determine the relationship between HPV status and these glycosylation patterns.
Keywords: Cervical cancer, human papillomavirus, Amaranthus leucocarpus, lectin cytochemistry, O-glycosylation, T antigen, Tn antigen, glycoproteins, flow cytometry, Western blotting, lectin binding