Levine E, Cupp A S, Miyashiro L, Skinner M K
Center for Reproductive Biology, Department of Genetics and Cell Biology, Washington State University, Pullman, Washington 99164-4231, USA.
Biol Reprod. 2000 Mar;62(3):477-90. doi: 10.1095/biolreprod62.3.477.
Embryonic testis development requires the morphogenesis of cords and growth of all cell populations to allow organ formation. It is anticipated that coordination of the growth and differentiation of various cell types involves locally produced growth factors. The current study was an investigation of the hypothesis that transforming growth factor-alpha (TGF-alpha) is involved in regulating embryonic testis growth. TGF-alpha has previously been shown to function in the postnatal testis. TGF-alpha and other members of the epidermal growth factor (EGF) family act through the epidermal growth factor receptor (EGFR) to stimulate cell proliferation and tissue morphogenesis. To understand the potential actions of TGF-alpha in the embryonic testis, general cell proliferation was investigated. Characterization of cell proliferation in the rat testis throughout embryonic and postnatal development indicated that each cell type has a distinct pattern of proliferation. Germ cell growth was transiently suppressed around birth. Interstitial cell growth was high embryonically and decreased to low levels around birth. A low level of Sertoli cell proliferation was observed at the onset of testis cord formation. Sertoli cell proliferation in early embryonic development was low; the levels were high later in embryonic development and remained high until the onset of puberty. Both TGF-alpha and the EGFR were shown to be expressed in the embryonic and postnatal rat and mouse testis. Perturbation of TGF-alpha function using neutralizing antibodies to TGF-alpha on testis organ cultures dramatically inhibited the growth of both embryonic and neonatal testis. TGF-alpha antibodies had no effect on cord formation. The TGF-alpha antibody was found to be specific for TGF-alpha in Western blots when compared to EGF and heregulin. Testis growth was also inhibited by perturbation of EGFR signaling using an EGFR kinase inhibitor. Therefore, TGF-alpha appears to influence embryonic testis growth but not morphogenesis (i.e., cord formation). Treatment of embryonic testis organ cultures with exogenous TGF-alpha also perturbed development, leading to an increased proliferation of unorganized cells. Testis from EGFR and TGF-alpha knockout mice were analyzed for testis morphology. TGF-alpha knockout mice had no alterations in testis phenotype, while EGFR knockout mice had a transient decrease in the relative amount of interstitial cells before birth. Observations suggest that there may be alternate or compensatory factors that allow testis growth to occur in the apparent absence of TGF-alpha actions in the mutant mice. In summary, the results obtained suggest that TGF-alpha is an important factor in the regulation of embryonic testis growth, but other factors will also be involved in the process.
胚胎睾丸发育需要索状结构的形态发生以及所有细胞群体的生长，以实现器官形成。预计各种细胞类型的生长和分化协调涉及局部产生的生长因子。本研究旨在调查转化生长因子-α（TGF-α）参与调节胚胎睾丸生长这一假说。此前已证明TGF-α在出生后睾丸中发挥作用。TGF-α和表皮生长因子（EGF）家族的其他成员通过表皮生长因子受体（EGFR）发挥作用，以刺激细胞增殖和组织形态发生。为了解TGF-α在胚胎睾丸中的潜在作用，对一般细胞增殖进行了研究。对大鼠睾丸在整个胚胎期和出生后发育过程中的细胞增殖特征进行分析表明，每种细胞类型都有独特的增殖模式。生殖细胞生长在出生前后短暂受到抑制。间质细胞生长在胚胎期较高，出生前后降至低水平。在睾丸索形成开始时观察到支持细胞增殖水平较低。早期胚胎发育中支持细胞增殖水平较低；在胚胎发育后期水平较高，并一直保持到青春期开始。TGF-α和EGFR在胚胎期和出生后的大鼠及小鼠睾丸中均有表达。在睾丸器官培养物上使用抗TGF-α的中和抗体干扰TGF-α功能，显著抑制了胚胎期和新生期睾丸的生长。TGF-α抗体对索状结构形成没有影响。与EGF和神经调节蛋白相比，在蛋白质印迹中发现TGF-α抗体对TGF-α具有特异性。使用EGFR激酶抑制剂干扰EGFR信号传导也抑制了睾丸生长。因此，TGF-α似乎影响胚胎睾丸生长，但不影响形态发生（即索状结构形成）。用外源性TGF-α处理胚胎睾丸器官培养物也扰乱了发育，导致无组织细胞的增殖增加。对来自EGFR和TGF-α基因敲除小鼠的睾丸进行了形态学分析。TGF-α基因敲除小鼠的睾丸表型没有改变，而EGFR基因敲除小鼠在出生前间质细胞的相对数量有短暂减少。观察结果表明，在突变小鼠中，可能存在其他替代或补偿性因子，使得在明显缺乏TGF-α作用的情况下睾丸仍能生长。总之，所得结果表明TGF-α是调节胚胎睾丸生长的重要因子，但该过程中也会涉及其他因子。
