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After 3H testosterone injection into castrated males of the Lizard Lacerta vivipara, the radioactive compound is detected by radioautography of epididymis, femoral glands, gut and liver. Between 1 hr. 30 min. to 12 hrs. of retention the 3H material concentrates progressively into nuclei of the glandular cells of epididymis and femoral organs although no particular concentration occurs in gut and liver cells. This submammalian model is consistent with those previously described for mammals.

In an attempt to understand the mechanism by which estrogens stimulate cell proliferation and mammary carcinogenesis, metastatic human breast cancer cell lines (MCF7, ZR75-1) were found to secrete a 52,000 dalton (52K) protein under estrogen stimulation. Following its purification to homogeneity, the 52K protein was identified as a secreted procathepsin-D-like aspartyl protease bearing mannose-6-phosphate signals. This precursor displays an in vitro autocrine mitogenic activity on estrogen-deprived MCF7 cells and is able to degrade basement membrane and proteoglycans following its autoactivation. The total protease (52K + 48K and 34K) was detected and assayed by monoclonal antibodies and was found to be highly concentrated in proliferative and cystic mastopathies. In breast cancer, its cytosolic concentration appears to be correlated more to tumor invasiveness than to hormone responsiveness. The mRNA of the 52K protease accumulates rapidly following estradiol treatment, as was shown by Northern blot analysis with cloned cDNA. The 52K cathepsin-D-like protease is the first example of a lysosomal protease induced by estrogens in cancer cells. Results obtained using different approaches suggest that two cysteinyl cathepsins are also related to cell transformation and invasiveness. It has been proposed that cathepsin-B is involved in breast cancer and metastatic melanoma, and its regulation by estrogen has been shown in the rat uterus. Cathepsin-L corresponds to the major excreted protein (MEP) whose synthesis and secretion are markedly increased by transformation of NIH 3T3 cells with Ki ras and are regulated by several growth factors. In addition to secreted autocrine growth factors and to other proteases (plasminogen activator, collagenase), lysosomal cathepsins may therefore play an important role in the process of tumor growth and invasion as long as their precursor is secreted abundantly.

Hypophysectomy of the boar resulted in a rapid drop in plasma testosterone level. hCG injections (400 IU/day) considerably elevated testosteronemia (up to 16.41 ng/ml). This effect was obtained either in the days following the operation or several months after (up to 134 days). In the latter case, the effect was temporary because the testosterone level dropped again after 15 days of injections. The sexual target organs (epididymis, seminal vesicle, prostate and Cowper glands) underwent structural change which, in the epithelium, corresponded to fluctuations in testosterone level. These effects were less marked in the cauda epididymis than in other target tissues. In various target organs, the conjunctivomuscular stroma developed after the operation; it did not regress at all, or incompletely, when the epithelia were stimulated indirectly by hCG, and began to hypertrophy when the hCG was no longer effective. Variations in stromal reaction were organ-dependent. When the hCG was administered for more than 14 days, it seemed to have a desensitizing effect; these mechanisms have been discussed.

The growth of MCF 7 human breast cancer cells is stimulated in vitro by estradiol (E2) and we have previously shown that estrogen-regulated glycoproteins released into the culture medium can partly mimic this effect. In this paper, we evaluate the mitogenic activity of the 52 K glycoprotein, which is a major E2-stimulated protein released by MCF 7 cells. The 52 K protein was purified 600-fold by affinity chromatography on Concanavalin A and an anti-52 K monoclonal antibody Sepharose columns. The 99% purified 52 K protein fraction stimulated the growth of estrogen-deprived MCF 7 cells. A mean 1.7-fold increase was obtained with nanomolar concentrations of seven different preparations of 52 K protein. This stimulation represented 40% of the mitogenic effect of E2. Both the 52 K protein and E2 induced microvilli at the cell surface but the effect of the 52 K protein occurred earlier. Other putative growth factors which are also stimulated by E2 and observed by [35S]cysteine labeling did not comigrate with the purified 52 K protein. Finally, the labeled 52 K protein was found to enter MCF 7 cells and to be processed into an immunoreactive 34 K protein. These data indicate that the E2-regulated 52 K glycoprotein is an autocrine mitogen on MCF 7 cells in culture and support the hypothesis that estrogens stimulate the growth of mammary cancer via this (and possibly other) secreted protein(s) acting as autocrine (and paracrine?) growth factors.