Leydig Cell tumors (LCT) are a rare group of endocrine tumors in the testicular interstitium. Between 1 and 3% of testicular malignances in adults and 4% in prepubertal children belong to LCT. An increasing incidence of this type of neoplasia has been reported recently all around the world. Particularly, a strong relationship between LCT and the use of anabolic steroids (which are commonly used nowadays) has been reported recently. In prepubertal boys, symptoms include feminization or virilization, depending on the major circulating steroid (estradiol or testosterone respectively). Adult patients show loss of libido, penile dysfunction, infertility and/or gynecomastia. Although the etiology still is unknown, several studies indicate that tumoral Leydig cells have an excessive production of insulin-like growth factor (IGF-1), as well as aromatase (CYP19) overexpression, which causes an enormous amount of estrogens (particularly estradiol, E2), and both factors play an important role in tumorigenesis. While usually benign, when LCT became malignant in adults they respond poorly to radio and chemotherapy. Likewise, it has been reported that both therapies increase the incidence of several tumors. All these data imply the need of new therapeutic targets to avoid the chirurgical dissection of the testes and the consequences of the hormonal therapies associated, which implicate not only the loss in reproductive function, but also psychological disorders. Several publications have reported that histidine decarboxylase (HDC), the only enzyme capable of catalyzing the conversion from L-histidine to histamine (HA) in mammals, has an important role in the development of several types of tumors, such as colorectal, breast and melanoma. At the same time, in our laboratory we have reported that HA induces cell proliferation of murine Leydig cells, and complementary, this cell proliferation decreases when inhibiting selectively HDC, as well as steroid synthesis (progesterone and E2). Also, we observed a higher expression of HDC in pediatric LCT (n = 3) than normal controls corresponding to different stages of sexual maturation (n = 9). It has been described that HDC knock out mice have an
incomplete angiogenesis, and also that MA-10 Leydig cells HDC expression correlates with vascular endothelial growth factor (VEGF). The aim of this study is to improve our knowledge about the role of HDC in LCT biology, particularly, the angiogenesis modulation. We used the R2C Leydig cell line, the most used model for in vitro studies of Leydigioma, because it overexpresses CYP19 and constitutively produces high levels of IGF-1 and E2, as well as human LCT. R2C and pediatric LCT angiogenic capability was evaluated in vitro by measuring proliferation of human umbilical vein endothelial cells (HUVEC). In addition, we verified R2C
cells angiogenic capability in vivo, using quail embryo vasculature (chorioallantoic membrane assay). Both models have been validated for the study of angiogenesis. Conditioned medium obtained from R2C cell culture stimulated angiogenesis in vitro (p <0.001) as well as in vivo (p <0.001). The in vitro effect was reverted with a previous treatment on the R2C cell culture using α-methyl-DL-histidine hydrochloride (α-MHD, 10 μM), a specific HDC activity inhibitor (p <0.001). Finally, human conditioned medium from pediatric LCT increased HUVEC proliferation (p <0.01). In the same way, the analyzed patients showed higher testosterone and estradiol levels than normal serum concentrations, which was in concordance to phenotypical features
observed in presence of LCT. Our results indicate that tumoral Leydig cells (TLC) produce HA, as well as other angiogenic factors, and it could be stimulating the vascular endothelium. The selective inhibition of HDC attenuates the pro-angiogenic capability in TLC. Considering all these results and previous observations of our laboratory, specific inhibitors of HDC could be used, in the future, as a potential therapeutic target for the treatment of LCT.