At short intervals after the intravenous administration of oestradiol-17beta, diethylstilboestrol, testosterone or saline control solution to ovariectomized rats, highly purified lysosome samples were prepared in substantial yield from preputial glands, sex accessory organs rich in these organelles. The preparations were essentially devoid of mitochondrial contamination. Exposure in vivo to doses of these hormones varying from 0.1 to 5mug/100g body wt. provoked dose-dependent labilization of the lysosomal membrane surface, as evidenced by significantly diminished structural latency of several characteristic acid hydrolases, including acid phosphatase, beta-glucuronidase and acid ribonuclease II, when such preparations were subsequently challenged in vitro with autolytic conditions, detergent or mechanical stress. Enhanced lytic susceptibility induced by hormone pretreatment was occasionally detectable in the initial preparation without further provocative stimuli in vitro. Comparable results were obtained with the corresponding fractions of uterus, despite the more limited concentration of lysosomes in this steroidal target organ. By the present criteria oestradiol-17alpha was essentially inert, even in a dose 25 times that effective for its active beta-epimer (<0.1mug/100g body wt.). Pretreatment with diethylstilboestrol exerted substantial membrane-destabilizing influence in preputial-gland lysosome samples from orchidectomized rats. Moreover, administration of testosterone to gonadectomized animals resulted in essentially equivalent dose-dependent augmentation of lysosomal enzyme release in preputial-gland preparations of either sex. The membrane stability of lysosome-enriched preparations from uterus, on the other hand, was unaffected by testosterone pretreatment. The sensitivity, specificity and selectivity of the lysosomal response to sex steroids provide evidence for the physiological significance of this phenomenon as a general mechanism for mediation of secondary biochemical transformations in the hormone-stimulated target cell.
The present investigation was designed to determine whether the rapid intracellular redistribution of lysosomes to a perinuclear position in reproductive target cells after administration of specific gonadal hormones in vivo was accompanied by evidence of penetration of the nucleoplasm by lysosomal marker enzymes. Nuclei were prepared by conventional isolation and purification procedures from preputial glands or uteri that were excised from gonadectornized rats within 2-15 min after intravenous injection of physiological doses of estradiol-17/3, testosterone or saline control solutions. Control and experimental preparations were essentially "pure", as judged by routine phasecontrast microscopic criteria in general use. However, nuclei from target cells of animals given hormone contained substantial activities of representative lysosomal hydrolases, including acid phosphatase, /3-glucuronidase and acid ribonuclease 11. In contrast, the samples originating from the target tissue of saline-treated controls or from non-target tissues contained minimal concentrations of these enzyme activities. After further purification of the nuclei by removal of the outer nuclear membrane through brief exposure to very low concentrations of Triton X-100 in the cold, the resultant 'ultrapurified' nuclei retained significant concentrations of structurally latent lysosomal marker enzymes after hormonal pretreatment without evidence of appreciable contamination by enzymes of mitochondria1 origin. Similar results were obtained with nuclei isolated by non-aqueous procedures. These combined observations appear to exclude adsorption artifacts. The total activities of lysosomal enzymes in the unfractionated homogenates and the MgZ+-dependent ribonuclease activity indigenous to the nuclear compartment were unaffected by prior hormone treatment. These results are consistent with the hypothesis that invasion of the nucleoplasm of specific target cells by lysosomal products may serve as a mechanism for triggering genic derepression in steroid hormone-induced growth.
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