Flickinger K S, Judware R, Lechner R, Carter W G, Culp L A
Department of Molecular Biology and Microbiology, Case Western Reserve University, School of Medicine, Cleveland, Ohio 44106.
Exp Cell Res. 1994 Jul;213(1):156-63. doi: 10.1006/excr.1994.1185.
Three human neuroblastoma cell lines, with or without N-myc amplification, were evaluated for their integrin expression patterns as cultured cells, as well as their nude mouse-borne tumors obtained after subcutaneous (ectopic) or adrenal gland (orthotopic) injection. IMR-32 and LaN1 cells (with amplified N-myc) do not express any of the common integrin subunits that recognize fibronectin or collagens, as determined by immunoprecipitation of cell extracts with specific monoclonal antibodies; the same was true for all subcutaneous or adrenal tumors from IMR-32 or LaN1, indicating that they are not essential during primary tumor formation at either site. SK-N-SH cells (with diploid N-myc) express beta 1, alpha 2, and alpha 3 subunits of expected sizes (with alpha 2 uncleaved at 145 kDa) but do not express alpha 1, alpha 4, alpha 5, alpha V, or beta 3. This expression pattern was conserved in all first-round subcutaneous and adrenal tumor cell populations, as well as in second-round subcutaneous tumors derived from a first-round subcutaneous tumor (no tumors expressed beta 3). One significant difference was noted between subcutaneous and adrenal tumor populations: all first- and second-round subcutaneous tumors expressed high levels of alpha V subunit, while adrenal tumors did not express any alpha V. This result suggests some essential function for alpha V beta 1 during subcutaneous primary tumor formation. Integrin patterns were also evaluated by fluorescence-activated cell sorting. SK-N-SH and its derivative tumors expressed heterogeneous amounts of beta 1 and alpha 2 at the cell surface, while only subcutaneous tumor cells expressed alpha V. Parental SK-N-SH cells contained two subpopulations, half of which expresses alpha 3, while the other half does not; all subcutaneous tumor cells retained this two-subpopulation pattern, indicating that primary tumor formation does not lead to clonal dominance of alpha 3- or alpha 3+ cell types in larger primary tumors. While these results suggest a correlation between N-myc amplification and down-regulation of integrin expression in neuroblastoma, they demonstrate conservation of integrin expression during two rounds of primary tumor formation at ectopic or orthotopic sites in a mouse model system, induction and/or selection for alpha V beta 1 expression at the subcutaneous site, and clonal heterogeneity in alpha 3 beta 1 expression throughout primary tumor development.
对三种有无N-myc扩增的人神经母细胞瘤细胞系，评估了其作为培养细胞的整合素表达模式，以及皮下（异位）或肾上腺（原位）注射后获得的裸鼠体内肿瘤的整合素表达模式。通过用特异性单克隆抗体免疫沉淀细胞提取物测定，IMR-32和LaN1细胞（N-myc扩增）不表达识别纤连蛋白或胶原蛋白的任何常见整合素亚基；IMR-32或LaN1的所有皮下或肾上腺肿瘤也是如此，这表明它们在任一部位的原发性肿瘤形成过程中并非必需。SK-N-SH细胞（N-myc为二倍体）表达预期大小的β1、α2和α3亚基（α2在145 kDa处未裂解），但不表达α1、α4、α5、αV或β3。这种表达模式在所有第一轮皮下和肾上腺肿瘤细胞群体中以及在源自第一轮皮下肿瘤的第二轮皮下肿瘤中（无肿瘤表达β3）均保持不变。皮下和肾上腺肿瘤群体之间存在一个显著差异：所有第一轮和第二轮皮下肿瘤均高表达αV亚基，而肾上腺肿瘤不表达任何αV。这一结果表明αVβ1在皮下原发性肿瘤形成过程中具有某些重要功能。还通过荧光激活细胞分选评估了整合素模式。SK-N-SH及其衍生肿瘤在细胞表面表达不同量的β1和α2，而只有皮下肿瘤细胞表达αV。亲代SK-N-SH细胞包含两个亚群，其中一半表达α3，另一半不表达；所有皮下肿瘤细胞均保留这种双亚群模式，这表明原发性肿瘤形成不会导致较大原发性肿瘤中α3+或α3-细胞类型的克隆优势。虽然这些结果表明N-myc扩增与神经母细胞瘤中整合素表达下调之间存在相关性，但它们证明了在小鼠模型系统中异位或原位部位两轮原发性肿瘤形成过程中整合素表达的保守性、皮下部位αVβ1表达的诱导和/或选择以及整个原发性肿瘤发展过程中α3β1表达的克隆异质性。
