Progesterone Receptors in the Developing Genital Tubercle: Implications for the Endocrine Disruptor Hypothesis as the Etiology of Hypospadias

Abstract

In fetal mice genital tubercles the ontogenetic expression of progesterone receptors and the effect of in utero estrogen and testosterone exposure were investigated. To evaluate ontogenetic progesterone receptor expression genital tubercles from untreated fetuses at gestational days 12, 14, 16 and 18, and newborn pups were prepared for real-time reverse transcriptase-polymerase chain reaction or immunohistochemistry. To evaluate estrogen and testosterone effects pregnant dams were gavaged once daily with corn oil (vehicle), ethinyl estradiol or testosterone propionate from gestational days 12 through 17. At gestational day 19 the genital tubercles of delivered fetuses were harvested for morphological examination and then pooled for real-time reverse transcriptase-polymerase chain reaction. Progesterone receptor protein was first detected at gestational day 12 in the urethral plate and mesenchyma. At later stages staining intensity increased with a greater progesterone receptor signal, especially in the urethra. Progesterone receptor mRNA expression showed different increasing patterns in each sex until birth. However, no difference was noted between male and female genital tubercles in terms of the distribution and quantity of progesterone receptor expression. In utero ethinyl estradiol led to 8.2, 9.7 and 5.2-fold increases in progesterone receptor mRNA in females and in males with and without hypospadias, respectively. Testosterone propionate significantly decreased progesterone receptor mRNA levels in females and males. Progesterone receptors are expressed in developing genital tubercles, suggesting a direct role of progesterone in normal genital tubercle patterning. Their increasing expression until birth also implies increasing sensitivity of the genital tubercles to the effects of estrogenic and progestogenic endocrine disruptors during fetal life. Ethinyl estradiol and testosterone propionate lead to opposing effects on progesterone receptor expression, in addition to their opposing morphological effects on the genital tubercles. These findings expand our knowledge of genital tubercle morphogenesis and provide important information for understanding the effects of endocrine disruptors.