Detection of photodynamic therapy-induced early apoptosis in human salivary gland tumor cells in vitro and in a mouse tumor model

Abstract

We studied the detection of apoptosis of malignant human salivary gland tumor cells induced by photodynamic therapy (PDT) using the photosensitizer mono-L-aspartyl chlorin e6 (NPe6) in vitro and in vivo in mice receiving transplanted human salivary gland tumor (HSG) cells. An immunohistocytochemical method using a monoclonal antibody (MoAb), M30, which reacts with the product resulting from the cleavage of cytokeratin (CK) 18 by activated caspase, was applied to detect the apoptosis of HSG cells induced by PDT. Significant amounts of immunoreactive products were observed in the cytoplasm of HSG cells after PDT. In vitro, M30-positive cells increased from 2 h after PDT, increased rapidly from 8 h and reached a peak 24 h after PDT. In vivo, a peak of early apoptosis was confirmed two hours after PDT. In comparison with DNA fragmentation detected by the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method, the destroyed tumor cells were observed sporadically 24 h after PDT. These results suggest that immunohistocytochemical staining with the MoAb M30 may be useful for detecting early apoptosis induced by PDT. Futhermore, PDT using NPe6 is effective in inducing apoptosis of HSG cells at an early stage, which suggests the possibility of the therapy being ideal for treatment of human malignant neoplasms.