Neural cell adhesion molecule (NCAM) is a type of cell surface glycoprotein and a member of the immunoglobulin superfamily. It has been reported that NCAM may be associated with perineural invasion by malignant salivary gland tumors such as adenoid cystic carcinoma. We have previously demonstrated that NCAM is constitutively expressed in the human salivary gland tumor cell line HSG, in vitro. In the present study, we have aimed to clarify the hypothesis that NCAM-mediated inhibition of salivary gland tumor proliferation is caused by homophilic binding and involves the prevention of signal transduction for perineural invasion using HSG cells. NCAM mRNA and protein expression was found to decrease in a dose-dependent manner upon treatment with the anti-NCAM antibody (MAb NCAM) for 24 h. The MTT assay showed a significant reduction in the number of viable HSG cells. Confocal laser microscopy showed that HSG cells underwent apoptosis after treatment with MAb NCAM. The activation of caspases 3, 7 and 9 was observed in HSG cells after treatment with MAb NCAM, thus confirming that apoptosis was induced by the activated caspases. Apaf-1 activity was also detected in HSG cells in a dose-dependent manner after treatment with MAb NCAM. The up-regulation of TGF-β1-mediated NCAM expression appeared to lead to the activation of homophilic NCAM binding, further accelerating HSG cell proliferation. In addition, the localization of NCAM in adenoid cystic carcinomas (ACCs) was examined using an immunohistochemical method. NCAM was slightly to moderately positive in 9 of 13 cases (69.2%) of ACC. These findings suggest that NCAM is associated not only with a cell-to-cell adhesion mechanism, but also with tumorigenesis, including growth, development and perineural invasion in human salivary gland tumors.

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