Photodynamic therapy (PDT) is a clinically effective cancer treatment which is based on cytotoxicity of reactive oxygen species generated by an interaction between photosensitizer and light. In recent years, many detailed mechanisms for PDT-induced cell death have been reported to improve the efficacy of PDT, and applications to other diseases have been sought. Imaging technique will provide additional information on the mechanism that has not been obtained by conventional methods. This article reviews recent mechanistic studies on PDT-induced cell death and introduces applicable imaging techniques for these studies.

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Fan-shaped keratocytes from fish epidermis are an excellent model system for studying mechanisms of amoeboid cell locomotion. Their overall shape and size remain roughly constant during locomotion, and the detailed morphology of their cell margin can be described mathematically by the GRE model. Previously, we developed a method for quantitative shape analysis during locomotion of the keratocytes, using shape prediction calculated according to the GRE model. In this paper, we present an improved version of our difference-from-prediction method for high resolution analysis of cell shape changes during locomotion. By superimposing the difference-from-prediction plot of cell outline onto the original microscope image, subtle but significant changes on local cell margin have been visually detected and quantitatively evaluated. By using this analysis, the way cells move on surfaces can be described in detail.

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The effects of progesterone on intracellular Ca²⁺ levels have been examined in immortalized hypothalamic neurons (GT1-7) with fluorescence microscopy. The effects observed were classified into two types: 1) the rapid effects and 2) the slow effects. To observe the rapid effects of progesterone, we added these reagents to cells at 10 μM. Upon stimulation with progesterone, the increase in intracellular Ca²⁺ level was observed at around 40% of cells within two minutes. The progesterone-induced Ca²⁺ signals were also observed with membrane-impermeable progesterone conjugated to BSA. These results suggest that progesterone stimulates Ca²⁺ signals in a nongenomic way by acting on a target in the plasma membrane. Next, to observe the slow effects, cells were incubated with progesterone at 100 nM for 72 hours. The incubation for 72 hours significantly increased the intracellular Ca²⁺ level and stimulated the spontaneous fluctuations. Further, the 72h incubation significantly suppressed GABA-induced Ca²⁺ signals. We have also measured effects of tributyltin, a possible analogue of steroid hormones, on intracellular Ca²⁺ levels in GT1-7 cells. The effects were similar to those of progesterone, implying that tributyltin might mimic the progesterone action on intracellular Ca²⁺ in GT1-7 cells.

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To explore the proteolysis of extracellular matrix by the invasion of growing tumor cells, a metastatic pharyngeal carcinoma cell line (Detroit 562) was cultured in Type-I collagen gel, and enzymatic and confocal laser-scanning microscopic (CLSM) analyses were carried out. Cells grew exponentially and degraded the collagen matrix. Concomitant with this growth were increased expressions of both urokinase-type plasminogen activator and matrix metalloproteinases (MMPs), which were the major proteolytic enzymes involved in the hydrolysis of the matrix. MMPs were identified as the 72-kDa and the 92-kDa species. CLSM demonstrated that colonies formed holes (micro-voids) and connecting tunnels around their peripheries. These findings and direct visualization suggest extensive proteolysis of pericellular matrix by the activated MMPs, providing an in vitro model for the invasive spread of tumor cells.

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