HORMONE CONTROL OF SPERMATOGONIAL ARREST IN MUTANT MICE

幼年精原细胞缺失（jsd）小鼠在青春期启动一波精子发生，但是尽管a精原细胞继续存在，精子分化却停止了。虽然机制尚不清楚，但我们发现睾酮(T)抑制精原细胞分化，可以用GnRH拮抗剂或雌二醇（E2）恢复。我们假设jsd小鼠精原细胞分化的抑制是由于T对特定体细胞（支持细胞、间质细胞、小管周围肌样细胞或小动脉平滑肌）基因表达的作用。我们利用小鼠遗传学的力量来阐明T介导jsd小鼠精原细胞“阻滞”的机制，并开始识别涉及以下目标的细胞类型和激素调节基因：(1)为了确定雄激素而非FSH的特定作用，我们将完成使用携带雄激素受体（AR）和FSH β基因突变的jsd小鼠的研究。为了确定E2的作用方式和部位，我们将比较gnrh -拮抗剂和E2治疗在jsd小鼠和携带雌激素受体(ER)- α或erβ突变的jsd小鼠中恢复精子发生的情况。（II）为了确定激素通过直接作用于睾丸和/或精管来影响精原细胞的分化，我们将在体外培养jsd小鼠的睾丸组织和精小管中检测它们对精原细胞的影响。（III）为了确定激素抑制精原细胞分化的靶细胞，我们将在jsd小鼠和AR缺陷jsd小鼠之间移植小管，并使用由不同体细胞特异性启动子驱动的crer -recombinase，对具有loxP位点的AR基因进行细胞型特异性消除。(4)为了确定相关基因，我们将使用微阵列技术在jsd小鼠靶细胞中差异筛选候选激素调节基因。一个好的候选基因必须在一个方向上被GnRH拮抗剂改变，在相反的方向上被T改变。在jsd小鼠中阐明这种精原细胞“阻滞”及其逆转的机制可能适用于基因决定的男性不育病例。由于其与毒物处理和衰老大鼠精原细胞分化失败的显著相似性，这些结果也适用于生殖毒物引起的无精子症和随着年龄的增长精子发生能力下降。

Juvenile spermatogonial depletion (jsd) mice initiate a wave of spermatogenesis at puberty, but then sperm differentiation ceases despite the continued presence of A spermatogonia. Although the mechanism is not known, we showed that testosterone (T) inhibits spermatogonial differentiation, which can be restored with GnRH antagonist or estradiol (E2). We hypothesize that inhibition of spermatogonial differentiation in jsd mice is due to the action of T on gene expression in a specific somatic cell (Sertoli, Leydig, peritubular myoid, or arteriolar smooth muscle). We utilize the power of mouse genetics to elucidate the mechanism by which T mediates spermatogonial "arrest" in jsd mice and begin identifying the cell type and hormonally regulated gene(s) involved in the following Aims: (I) To establish the specific role of androgen and not FSH, we will complete studies using jsd mice also carrying mutations in the androgen receptor (AR) and FSHbeta genes. To determine the mode and the site of action of E2, the restoration of spermatogenesis with GnRH-antagonist and E2 treatment will be compared in jsd mice with that in jsd mice carrying mutations for estrogen receptor (ER)-alpha or for ERbeta. (II) To establish that the hormones affect spermatogonial differentiation by direct action on the testis and/or seminiferous tubules, we will examine their effects on spermatogonia in in vitro cultures of testicular tissue and tubules from jsd mice. (III) To identify the cell that is the target for hormonal inhibition of spermatogonial differentiation, we will transplant tubules between jsd mice and AR-deficient jsd mice and use cell-type specific elimination of an AR gene with loxP sites using Cre-recombinase driven by promoters specific for the different somatic cells. (IV) To identify the responsible gene, we will differentially screen for candidate hormone-regulated genes in the target cell of jsd mice using microarrays. A good candidate gene must have its level changed by GnRH antagonist in one direction and by T in the opposite direction. Elucidation of the mechanism of this spermatogonial "arrest" and its reversal in the jsd mouse could apply to cases of genetically determined male infertility. Because of its remarkable similarity to the failure of spermatogonial differentiation in toxicant-treated and aging rats, these results could also apply to azoospermia induced by reproductive toxicants and the decline in spermatogenesis with aging.