Imaging of estrogen receptor transcriptional activity

Molecular imaging, i.e. the non-invasive imaging of target macromolecules and biological processes in living organisms, is providing a novel and powerful technology for the progress of biomedical research by facilitating the study of physio-pathological events in living organisms and by implementing the path towards novel drugs discovery. The ERE-Luc transgenic mouse line generated in 2001 in our laboratory is the first prototype of reporter animals, in which the firefly luciferase reporter gene is expressed in all tissues and regulated by the estrogen receptors (ERs) through an estrogen-responsive promoter. The luciferase activity is quantitatively detectable by evaluating photon emission through a charge-coupled device (CCD) camera and converting optical into digital imaging. The use of in vivo imaging of the ERE-Luc mice provided us with the opportunity to study the physiological fluctuations of ERs action in mice during the estrous cycle and to demonstrate that the state of ER transcriptional activity is regulated by circulating estradiol only in reproductive organs and liver, while in organs that are not linked with reproductive functions ERs are activated by signals other than estradiol through a mechanism known as “unliganded activation”. It is known that aging is associated with changes in physiological parameters, such as oxidative stress or immune senescence, that might influence ER transcriptional activity. Similarly, menopausal transition is linked with physiopathological alterations that are thought to be mediated by loss of estrogen signalling. Therefore, the principle objective of the present work is to study ER transcriptional activity at the menopause transition and during aging. We assayed bioluminescence imaging in non reproductive organs of ERE-Luc mice at different ages (3, 6, 12, 18 and 22 months-old) and after ovary removal, the most reliable model of menopause. Briefly, after systemic injection of the luciferase substrate, luciferine, we quantified luciferase activity in vivo by bioluminescence imaging and in vitro by an enzymatic assay on tissue extracts. Data are validated by measuring the expression of estrogen target genes, such as progesterone receptor. The results of these experiments will be discussed. Altogether, our study shows the extent to which ER transcriptional activity changes from 6 up to 22 months of age and after ovariectomy, providing insight in the relevance of unliganded activation of ERs activity in the onset of disorders associated with aging and menopause.