Figure 5.

Modulation of signaling kinases and transcription factors in tissue-isolated (ER+) MCF-7 human breast tumor xenografts from female nude rats exposed to control LD 12:12, dLEN or dLEN supplemented with nighttime melatonin and treated with 4OH-TAM. (A) Study I - Total and phospho- Western blot analysis of total tissue lysates from tissue-isolated MCF-7 breast tumor xenografts harvested rats in dLEN lighting schedule and treated with vehicle (dLEN) or 4OH-TAM (dLEN+TAM) or a LD 12:12 lighting schedule and treated with vehicle (12L:12D) or 4OH-TAM (12L:12D+TAM). (B) Study II- Total and phospho- Western blot analysis of total tissue lysates from breast tumor xenografts harvested rats in dLEN lighting schedule and treated with vehicle (dLEN) or 4OH-TAM (dLEN+TAM) or in dLEN but supplemented with nighttime melatonin and treated with vehicle (dLEN+MLT) or 4OH-TAM (dLEN+MLT+TAM). In both studies, tumors were harvested at 2400 hr (mid-dark phase) from 3 animals in each group. Total cell lysates (120 μg of protein per sample) from each tumor were analyzed by Western blot for expression of phosphorylated and total forms of ERK1/2, AKT, NF-kB, SRC, FAK, STAT3, and CREB. β-actin was used as a control for equal loading. (C) Study II - Total tissue lysates from tissue-isolated MCF-7 (ERα+) breast tumor xenografts harvested from nude rats housed under dLEN photoperiod (dLEN) or dLEN and supplemented with nighttime melatonin (dLEN+MLT). As above, tumors were harvested at 2400 h (mid-dark phase) from 3 animals in each group. Total cellular protein was analyzed by Western blotting for expression of total and phosphorylated ERα (p-ERα S118 and S167). β-actin was used as a control for equal loading.