Photofrin mediated PDT in normal rat brain: assessment on apoptosis as a quantitative biological endpoint

摘要

The extent of the apoptotic response of normal rat brain tissue to Photofrin-mediated photodynamic therapy was determined following sub-necrotic light exposures. Wistar rats were injected with 12.5 mg kg$+$MIN@1$/ Photofrin and the brain surface was exposed to 1-17 J cm$+$MIN@2$/ of 630 nm light 24 hours later. In order to identify apoptotic cells on tissue sections 24 hour post treatment, the TUNEL assay was used to fluorescently label DNA cleavage, with propidium iodide as a control counter stain. Confocal laser scanning microscopy was used to quantify the local spatial density of apoptotic bodies and to determine their depth distribution in the tissue from the irradiated surface. Radiant exposures of 1-3 J cm$+$MIN@2$/ produced the highest apoptotic response, which occurred in well-circumscribed volumes, without evidence of gross tissue necrosis. Up to 40 percent of all cells in the treatment field were positively stained, with more apoptotic bodies observed at the edge of the lesion. The number of TUNEL-positive cells was lower at 5 and 17 J cm$+$MIN@2$/. The apoptotic depth distribution was correlated with the light fluence distribution in the tissue over a limited depth range. Beyond this range there is evidence of a threshold for apoptosis, while at small depth and height radiant exposures there is a significant reduction in observed apoptosis. Hence, there is evidence that apoptosis is 'inhibited' by the presence of PDT-induced necrosis.