Testosterone and 17ß-estradiol have opposite effects on podocyte apoptosis that precedes glomerulosclerosis in female estrogen receptor knockout mice.

Podocyte damage and apoptosis are thought to be important if not essential in the development of glomerulosclerosis. Female estrogen receptor knockout mice develop glomerulosclerosis at 9 months of age due to excessive ovarian testosterone production and secretion. Here, we studied the pathogenesis of glomerulosclerosis in this mouse model to determine whether testosterone and/or 17ß-estradiol directly affect the function and survival of podocytes. Glomerulosclerosis in these mice was associated with the expression of desmin and the loss of nephrin, markers of podocyte damage and apoptosis. Ovariectomy preserved the function and survival of podocytes by eliminating the source of endogenous testosterone production. In contrast, testosterone supplementation induced podocyte apoptosis in ovariectomized wild-type mice. Importantly, podocytes express functional androgen and estrogen receptors, which, upon stimulation by their respective ligands, have opposing effects. Testosterone induced podocyte apoptosis in vitro by androgen receptor activation, but independent of the TGF-ß1 signaling pathway. Pretreatment with 17ß-estradiol prevented testosterone-induced podocyte apoptosis, an estrogen receptor-dependent effect mediated by activation of the ERK signaling pathway, and protected podocytes from TGF-ß1- or TNF-α-induced apoptosis. Thus, podocytes are target cells for testosterone and 17ß-estradiol. These hormones modulate podocyte damage and apoptosis.

Oestrogen receptors pathways to oestrogen responsive elements: the transactivation function-1 acts as the keystone of oestrogen receptor (ER)beta-mediated transcriptional repression of ERalpha.

Oestrogen receptors (ER)alpha and beta modify the expression of genes involved in cell growth, proliferation and differentiation through binding to oestrogen response elements (EREs) located in a number of gene promoters. Transient transfection of different luciferase reporter vectors 3xEREs-Vit, 2xEREs-tk and ERE-C3 showed that the transactivation capacity of both ER subtypes was influenced by 1) the nature of the inducer (oestradiol (E2), phyto- and anti-oestrogen (AE)), 2) the structure of the promoter (nucleotidic sequence, number of ERE, length of the promoter sequence) and 3) the cell line (containing endogenous ER (MCF-7) or in which ER was stably expressed (MDA-MB-231-HE-5 (ERalpha+) or MDA-MB-231-HERB (ERbeta+)). ER subtype did not display the same efficacy on the different constructions in the presence of E2 and of AE according to the cell (e.g. in MCF-7 cells: tk>>Vit>>C3 approximately 0 while in MDA-MB-231 cells: Vit>>tk approximately C3). E2 response was higher in MCF-7 cells, probably due to higher ER expression level (maximal at 10(-10)M instead of 10(-8)M for E2 in HE-5 cells). Finally, the same ligand could exert opposite activities on the same promoter according to the ER isoform expressed: in the MDA-MB-231 cells, AE acted as inducers of the C3 promoter via ERbeta whereas ERalpha/AE complexes down-regulated this promoter. Approximately 70% of breast tumours express ER and most tumour cells coexpress both ER isotypes. Thus, different types of ER dimers can be formed in such tumours (ERbeta or ERalpha homodimers or ERalpha/ERbeta heterodimers). We therefore studied the influence of the coexistence of the two ERs on the ligand-induced transcriptional process following transient transfection of ERalpha in ERbeta+ cells, and inversely ERbeta in ERalpha+ cells. ERbeta-transfection inhibited the E2- and genistein-induced ERalpha-dependent transcription on all promoters in all cell lines except C3 in MCF-7; this inhibitory effect was lost following transfection of ERbeta deleted of its AF-1 (ERbeta-AF-2). These results suggest that the dominant negative properties of ERbeta are mainly due to its AF-1 function. Interestingly, transfection of an ERbeta-AF-2 construct into MCF-7 cells potentiated the transcription inhibitory capacity of 4-OH-tamoxifen (OHT) on the Vit and tk promoters. Thus, (1) OHT exerts an agonistic activity through the AF-1 function of ER and (2) expression of ERbeta in breast cancer cells seems to favour the AE treatment. Contrary to ERbeta, ERalpha-transfection had little effect on ERbeta transactivation capacity in HERB cells. Finally, the ratio ERalpha/ERbeta constitutes one decisive parameters to orientate the transcriptional mechanism of a target gene in the presence of agonist as well as of antagonist ligands.