Sterility and absence of histopathological defects in nonreproductive organs of a mouse ERbeta-null mutant

Abstract

Estrogen signaling is mediated by estrogen receptors alpha (ERalpha) and beta (ERbeta). Although a consensus has now been reached concerning many physiological functions of ERalpha, those of ERbeta are still controversial: When housed and examined in two distant laboratories, mice originating from the same initial ERbeta mutant exhibited widely different phenotypes, which were themselves different from the phenotype of another ERbeta mutant previously generated in our laboratory. Because, in addition to a knockout insertion in exon 3, all these mouse mutants displayed alternative splicing transcripts, we have now constructed a ERbeta mouse mutant (ERbeta(ST)(L-/L-)) in which exon 3 was cleanly deleted by Cre/LoxP-mediated excision and was devoid of any transcript downstream of exon 3. Both females and males were sterile. The histology of the ovary was mildly affected, and no histological defects were detected in other organs, neither in females nor in males. Our present results, which are in contrast with previously published data, suggest that, with the notable exception of male and female reproduction, ERbeta is not required in the mouse for the development and homeostasis of the major body systems.

Fig. 1.

Normal morphology and cell proliferation in the ventral prostate of 16-month-old ERβ_ST^L−/L− mice. (a–d) Hematoxylin and eosin-stained sections of ventral prostate. (c and d) Higher magnification of functionally hyperplastic alveoli. (e–h) Immunostaining for detection of the cell proliferation marker Ki67 in ventral prostate. Arrows point to Ki67-positive epithelial cells (red signals). (g and h) Enlargement of the black frames from e and f, respectively. E, prostatic epithelium; M, smooth muscle; VP1 and VP2, alveoli without and with functional hyperplasia, respectively. Four males of each genotype were used in each type of assay. (Scale bar: a and b, 500 μm; c–f, 50 μm.)

Fig. 2.

The ERβ_ST^L−/L− mutation does not alter architecture and neuronal density of the somatosensory cortex and does not elicit astrocytosis in 16-month-old mice. (a–d) Coronal sections at similar levels of the brains of WT and ERβ_ST^L−/L− males. (e–h) Immunodetection of GFAP (red signals) and DAPI counterstain (blue nuclei) in the paraventricular nucleus (e and f) and amygdala (g and h). C, cerebral cortex. Five males of each genotype were used for histology. (Scale bar: a and b, 500 μm; c and d, 250 μm; e–h, 30 μm.)

Fig. 3.

Absence of myeloproliferation in 16- to 19-month-old ERβ_ST^L−/L− mice. (a–d) Sections of hematopoietic compartment of the spleen (a and b) and bone marrow (c and d) of ERβ_ST^L−/L− females (hematoxylin and eosin). Myeloid precursors (MP) appear as large cells, containing (i) a large, pale-stained nucleus displaying prominent nucleoli and a finely granular chromatin and (ii) a relatively low abundance of lightly stained cytoplasm. In contrast, erythroid precursors (EP) are medium- to small-sized cells, with dark nuclei and low amount of darkly stained cytoplasm. MK, megakaryocyte. Nine mice of each sex and genotype were used for histology. (Scale bar: 8 μm.)

Fig. 4.

Normal morphology of the lung and heart of 16- to 19-month-old ERβ_ST^L−/L− mice. (a and b) Normal cellular composition of the lung of ERβ_ST^L−/L− females (hematoxylin and eosin). (c and d) Normal cardiomyocyte size and intercellular spaces in the heart of ERβ_ST^L−/L− males (modified Mallory's trichrome). (e and f) Normal pattern of f-actin distribution in the sarcomeres of ERβ_ST^L−/L−hearts; sections are stained with FITC-conjugated phalloidin. (g and h) Normal structure of intercalated discs in ERβ_ST^L−/L− hearts; transmission electron microscopy. (i) Normal mean alveolar surface in ERβ_ST^L−/L− female lungs (arbitrary units; pixels × 10³; means ± SEM). (j) Normal mean left ventricular wall surface in ERβ_ST^L−/L− male heart (arbitrary units; pixel × 10⁵; means ± SEM). A, alveolar lumen; CM, cardiomyocytes; E, endothelial cell; I and Z, I and Z bands, respectively; ID, intercalated disk; IS, intercellular spaces; M, mitochondria; P1 and P2, type 1 and 2 pneumocytes, respectively. Nineteen-month-old females of each genotype were used, 10 for histological analysis and 5 for alveolar surface measurement; 16-month-old males of each genotype were used, 8 for evaluation of cardiomyocytes size and left ventricle wall surface, 5 for f-actin staining and 3 for electron microscopy. (Scale bar: a and b, 40 μm; c–f, 100 μm; g and h, 2 μm.)