Allen M H, Robinson M K, Stephens P E, MacDonald D M, Barker J N
Dunhill Dermatology Laboratory, St. John's Institute of Dermatology, London, U.K.
J Invest Dermatol. 1996 Apr;106(4):611-5. doi: 10.1111/1523-1747.ep12345385.
E-selectin is an endothelial adhesion molecule that binds carbohydrate epitopes on leukocytes and has been implicated in a potential pathway of tumor metastasis. Keratinocyte cell lines express similar carbohydrate epitopes, one of which, sialyl Lewis X (SL-X) is a ligand for E-selectin and is also expressed by squamous cell carcinomas (SCC) in situ. The functional role of keratinocyte selectin ligands was investigated using a soluble E-selectin chimaeric protein (pE-sel-Ig) containing pig lectin-like and epidermal growth factor-like domains fused to human IgG. After incubation of keratinocyte cell lines (A431 and SVK14) and normal keratinocytes with pE-sel Ig, binding was quantified by flow cytometry. Frozen sections of SCC were overlaid with pE-sel Ig and binding was visualized immunoenzymatically. Immunolabeling was undertaken using monoclonal antibodies (CSLEX-1 and HECA-452), which label E-selectin ligands including sialyl Lewis X. E-selectin bound strongly to A431 and SVK14 cells; the degree of binding paralleled staining intensity with CSLEX-1 antibody. HECA-452 antibody stained A431 cells strongly but SVK14 cells only weakly. Normal keratinocytes and normal epidermis did not express CSLEX-1 or HECA-452 antigens or bind E-selectin. Serial sections of SCC revealed close correlation between fusion protein binding and antibody staining. Antibody pretreatment of tumor sections with CSLEX-1 blocked fusion protein binding, whereas HECA-452 antibody only slightly reduced fusion protein binding. pE-sel Ig pretreated with YT11.1 antibody failed to bind to A431 or SVK14 cells or to SCC. These studies provide functional evidence that SL-X/E-selectin pathways may be important in SCC metastasis and that A431 and SVK14 cells provide a good model to investigate these mechanisms.
E选择素是一种内皮黏附分子，它能与白细胞上的碳水化合物表位结合，并与肿瘤转移的潜在途径有关。角质形成细胞系表达相似的碳水化合物表位，其中之一唾液酸化路易斯X（SL-X）是E选择素的配体，原位鳞状细胞癌（SCC）也表达该配体。利用一种可溶性E选择素嵌合蛋白（pE-sel-Ig）研究角质形成细胞选择素配体的功能作用，该蛋白含有与人类IgG融合的猪凝集素样结构域和表皮生长因子样结构域。用pE-sel Ig孵育角质形成细胞系（A431和SVK14）及正常角质形成细胞后，通过流式细胞术对结合情况进行定量分析。将pE-sel Ig覆盖在SCC的冰冻切片上，通过免疫酶法观察结合情况。使用单克隆抗体（CSLEX-1和HECA-452）进行免疫标记，这些抗体可标记包括唾液酸化路易斯X在内的E选择素配体。E选择素与A431和SVK14细胞强烈结合；结合程度与CSLEX-1抗体的染色强度平行。HECA-452抗体对A431细胞染色强烈，但对SVK14细胞染色较弱。正常角质形成细胞和正常表皮不表达CSLEX-1或HECA-452抗原，也不结合E选择素。SCC的连续切片显示融合蛋白结合与抗体染色之间密切相关。用CSLEX-1对肿瘤切片进行抗体预处理可阻断融合蛋白结合，而HECA-452抗体仅略微降低融合蛋白结合。用YT11.1抗体预处理的pE-sel Ig无法与A431或SVK14细胞或SCC结合。这些研究提供了功能证据，表明SL-X/E选择素途径可能在SCC转移中起重要作用，且A431和SVK14细胞为研究这些机制提供了一个良好的模型。
