Biopsy samples. Phase-contrast photomicrograph of tumor explants isola…

Biopsy samples. Phase-contrast photomicrograph of tumor explants isolated from HNSCC biopsies in culture after 2-4 weeks with cells growing directly from the explants in a monolayer (A ?00, B ?00 original magnification).Liebertz et al. Head Neck Oncology 2010, 2:5 http://www.headandneckoncology.org/content/2/1/Page 7 ofFigure 4 Establishment of the USC-HN1 cell line. (A) Phase contrast microphotograph of growing USC-HN1 cells showing numerous mitotic cells (rounded, luminescent cells) with a tightly, adherent squamous cell morphology. (B) Cytology of the USC-HN1 cell line shows malignant cells with large nuclei and nucleoli and an abundance of cytoplasm typical of squamous cells (Cytospin, Wright-Giemsa stain, ?00 original magnification).prominent nucleoli. Mitotic activity is abundant including atypical forms such as ring and tripolar configurations. Intercellular bridges are focally present but faint.Immunophenotype of USC-HN1 in cell culture and in situImmunophenotypic characterization of USC-HN1 cells in culture and from tumors grown in Nude mice demonstrated similarity to the original tumor and confirmed akeratinizing squamous cell carcinoma (Figure 6). USCHN1 cells demonstrate strong, uniform nuclear p53 and Rb expression, as well as positive expression of keratin, Ecadherin, EGFr, and CD44 in situ consistent with HNSCC as well as the original tumor biopsy [4,7,13,19]. There is decreased staining of E-cadherin and CD44 in situ compared with cultured cells. Conversely, EGFr staining is stronger in those cells grown as heterotransplants. CDFigure 5 Heterotransplantation of USC-HN1 cell line into Nude mice. (A) Appearance of subcutaneous USC-HN1 tumor in Nude mouse. (B and C) Low and high magnification of USC-HN1 Nude mouse heterotransplant showing a poorly differentiated squamous cell carcinoma arranged as a sheet with areas of tumor necrosis and bluntly infiltrative borders. The tumor cells are tightly cohesive featuring from moderate to abundant eosinophilic cytoplasm. The nuclear to cytoplasmic ratio is markedly increased with nuclei showing frequent, prominent nucleoli. Mitotic activity is abundant including atypical forms such as ring and tripolar configurations. Intercellular bridges are focally present but faint (H E ?00 and ?00 original magnification).Liebertz et al. Head Neck Oncology 2010, 2:5 http://www.headandneckoncology.org/content/2/1/Page 8 ofFigure 6 Immunoperoxidase staining of USC-HN1 cells in Nude mouse heterotransplant and in cytospin preparations for HNSCC classification markers. Photomicrograph of immunoperoxidase staining of original tumor biopsy (top), PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/16989806 IHC stained formalin-fixed paraffinembedded tissue sections of USC-HN1 Nude mouse heterotransplant (middle), and USC-HN1 cells from culture in a cytospin preparation (bottom) for keratin, E-cadherin, EGFr, CD44, p53 and Rb (?00 original magnification).staining is decreased in the heterotransplant and cytospin in comparison with the original tumor biopsy.USC-HN1 characterization by flow cytometryTable 2 Analysis of USC-HN1 cytokines and surface markers by FACS.Positive Target FABP5 CD24 EGFR c-Kit CD74 IL-13Ra CD44v6 CD133 MFI 97.49 49.74 7.87 10.71 0.26 0.15 2.63 0.04 3.02 31.80 6.87 Capecitabine 6.87 37.70 6.55 6.87 8.89 13.27 38.84 1310.47 *** 203.69 *** 90.52 *** 198.78 ** 14.01 ** 9.16 ** 7.89 * 9.38 * 20.67 * Isotype Control Antibody Isotype Control Antibody 0.01 0.01 4.50 0.01 0.01 5.47 0.06 7.The phenotype of USC-HN1 cell line was further characterized by surfac.