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The bioluminescence signal, displayed as a pseudocolored scale, corresponds to the radiance (photons?sec?1?cm?2?sr?1) emitted from each tumor due to the reaction between luciferin substrate and luciferase enzyme. In the first group of mice which received no agents (Fig.?(Fig.3A),3A), radiance continued to increase from day 4 to day 8, consistent with continuous/unrestricted growth of luciferase-bearing cancer cells in the breast fat pad environment. In the second group of mice, the increase in radiance between day 7 and day 8 was reversed by application of ALA-PDT at day 7, resulting in a 45% drop in signal (Fig.?(Fig.3B).3B). However, radiance from these tumors at day 8 was still higher than at day 4, indicating room for improvement in the Temsirolimus ic50 treatment protocol. In contrast, all animals that received Vit D/ALA-PDT treatment showed a statistically significant reduction (82?��?6%, P?selleck kinase inhibitor actually corresponded to changes in the number of living cells after treatment, tumors were histologically examined after biopsy; see below). Figure 3 Noninvasive monitoring of treatment responses in breast cancer tumors. Photographs illustrate typical changes in individual mice; BLI signal superimposed upon white light image. (A) Control mouse that received neither Vit D treatment nor ALA-PDT; (B) ... Vit D stimulates differentiation and proliferation in MDA-MB-231-luc tumors To examine the effect of Vit D pretreatment on differentiation and proliferation status, tumor MRIP tissues were stained for E-cadherin and Ki-67, respectively. When compared with saline-treated control tumors, E-cadherin expression was significantly increased in Vit D pretreated tumors (Fig.?(Fig.4A).4A). E-cadherin was mainly expressed in cells within ductal structures in the tumors that received Vit D, whereas E-cadherin in saline-treated tumors was undetectable (Fig.?(Fig.4A).4A). Expression of nuclear Ki-67 was also induced, being observed in the majority of cells within tumors receiving Vit D preconditioning (Fig.?(Fig.4B).4B). Quantitatively, E-cadherin and Ki-67 were increased 9.6?��?1.7-fold and 10.7?��?2.8-fold, respectively, in Vit D-preconditioned tumors relative to nonconditioned controls (Fig.?(Fig.4C4C and D). Figure 4 Vit D preconditioning increases the status of proliferation and differentiation in MDA-MB-231-luc tumor cells. Images show paraffin sections of the tumors, immunostained with antisera to (A), E-Cadherin (red fluorescence), and (B), Ki-67 (green fluorescence). ... Vit D preconditioning increases ALA-induced phototoxicity To assess cell-death responses after treatment, a TUNEL assay was performed on formalin-fixed/paraffin-embedded tissues (Fig.?(Fig.5).5).