Identification of KIN (KIN17), a human gene encoding a nuclear DNA-binding protein, as a novel component of the TP53-independent response to ionizing radiation.

Ionizing radiation elicits a genetic response in human cells that allows cell survival. The human KIN (also known as KIN17) gene encodes a 45-kDa nuclear DNA-binding protein that participates in the response to UVC radiation and is immunologically related to the bacterial RecA protein. We report for the first time that ionizing radiation and bleomycin, a radiomimetic drug, which produce single- and double-strand breaks, increased expression of KIN in human cells established from tumors, including MeWo melanoma, MCF7 breast adenocarcinoma, and ATM+ GM3657 lymphoblast cells. KIN expression increased rapidly in a dose-dependent manner after irradiation. Under the same conditions, several genes controlled by TP53 were induced with kinetics similar to that of KIN. Using the CDKN1A gene as a marker of TP53 responsiveness, we analyzed the up-regulation of KIN and showed that is independent of the status of TP53 and ATM. In contrast, the presence of a dominant mutant for activating transcription factor 2 (ATF2) completely abolished the up-regulation of KIN. Our results suggest a role for ATF2 in the TP53-independent increase in KIN expression after gamma irradiation.

Regulatory influence of germ cells on sertoli cell function in the pre-pubertal rat after acute irradiation of the testis.

While germ cell regulation of Sertoli cells has been extensively explored in adult rats in vivo, in contrast, very little is known about germ cell influence on Sertoli cell function at the time when spermatogenesis begins and develops. In the present study various Sertoli cell parameters (number, testicular androgen binding protein (ABP) and testin, serum inhibin-B and, indirectly, follicle-stimulating hormone (FSH)) were investigated after the exposure of 19-day-old rats to a low dose of 3 Grays of gamma-rays. Differentiated spermatogonia were the primary testicular targets of the gamma-rays, which resulted in progressive maturation depletion, sequentially and reversibly affecting all germ cell classes. Testicular weight declined to a nadir when pachytene spermatocytes and spermatids were depleted from the seminiferous epithelium and complete or near complete recovery of spermatogenesis and testicular weight was observed at the end of the experiment. Blood levels of FSH and ABP were normal during the first 11 days after irradiation, when spermatogonia and early spermatocytes were depleted. While the number of Sertoli cells was not significantly affected by the irradiation, from days 11-66 after gamma-irradiation, ABP production declined and FSH levels increased when pachytene spermatocytes and spermatids were depleted and the recovery of these parameters was only observed when spermatogenesis was fully restored. Comparison of the pattern of change in serum levels of inhibin-B and testicular levels of testin and of germ cell numbers strongly suggest a relationship between the disappearance of spermatocytes and spermatids from the seminiferous epithelium and the decrease in levels of inhibin-B and increase in levels of testin from 7 to 36 days post-irradiation. Levels of testin and inhibin-B were restored before spermatogenesis had totally returned to normal. In conclusion, this in vivo study shows that pre-pubertal Sertoli cell function is under the complex control of various germ cell classes. This control presents clear differences when compared with that previously observed in adult animals and depends on the Sertoli cell parameter of interest, as well as on the germ cell type.

Molecular cloning and characterization of the human KIN17 cDNA encoding a component of the UVC response that is conserved among metazoans.

We describe the cloning and characterization of the human KIN17 cDNA encoding a 45 kDa zinc finger nuclear protein. Previous reports indicated that mouse kin17 protein may play a role in illegitimate recombination and in gene regulation. Furthermore, overproduction of mouse kin17 protein inhibits the growth of mammalian cells, particularly the proliferation of human tumour-derived cells. We show here that the KIN17 gene is remarkably conserved during evolution. Indeed, the human and mouse kin17 proteins are 92.4% identical. Furthermore, DNA sequences from fruit fly and filaria code for proteins that are 60% identical to the mammalian kin17 proteins, indicating conservation of the KIN17 gene among metazoans. The human KIN17 gene, named (HSA)KIN17, is located on human chromosome 10 at p15-p14. The (HSA)KIN17 RNA is ubiquitously expressed in all the tissues and organs examined, although muscle, heart and testis display the highest levels. UVC irradiation of quiescent human primary fibroblasts increases (HSA)KIN17 RNA with kinetics similar to those observed in mouse cells, suggesting that up-regulation of the (HSA)KIN17 gene after UVC irradiation is a conserved response in mammalian cells. (HSA)kin17 protein is concentrated in intranuclear focal structures in proliferating cells as judged by indirect immunofluorescence. UVC irradiation disassembles (HSA)kin17 foci in cycling cells, indicating a link between the intranuclear distribution of (HSA)kin17 protein and the DNA damage response.

Short term pretreatment with medroxyprogesterone and testosterone may potentiate irradiation damage to spermatogenesis in rats.

BACKGROUND: Hormonal treatments lasting 2-6 months inhibit spermatogenesis in men and have been proposed as germ cell protection against anticancer therapy. Because it is unthinkable to delay anticancer treatments, the authors investigated the protection afforded against irradiation of rats by 22 days of hormonal pretreatment. METHODS: Adult Sprague-Dawley rats were assigned to an untreated control group (C) or to one of 5 treatments: medroxyprogesterone acetate plus testosterone only (M), 3 or 5 gray of irradiation (R3 and R5), or hormonal treatment prior to 3 or 5 gray of irradiation (MR3 and MR5). Mating trials were conducted 1, 24, 45, 65, 86, and 109 days after treatment. At 122 days, genital organ weights, testis histology, and epididymal spermatozoa were evaluated. RESULTS: Irradiation reduced sperm production and had a clastogenic effect on postmeiotic germ cells. No protective effect of steroid treatment was observed. Moreover, testis weight, tubule diameter, the repopulating index, and the sperm head count decreased more in the MR5 group than in the R5 group. Mating tests showed decreases in positive vaginal smears and fertility at both 45 and 65 days, and an increase in resorption at 109 days. CONCLUSIONS: These results indicate that hormonal pretreatment potentiates irradiation damage to germ cells, especially stem cells, as regards survival and genomic alterations, probably because of increased lipoperoxidation of late spermatids.

The nuclear concentration of kin17, a mouse protein that binds to curved DNA, increases during cell proliferation and after UV irradiation.

UV-irradiation induces, in mammalian cells, the expression of a set of genes known as the 'UV-response', which may be reminiscent of the bacterial response, called SOS system. The multifunctional protein RecA controls the expression of the SOS genes. We report the expression profile of a mouse gene conserved among mammals, called Kin17, that codes a DNA-binding protein of undetermined biochemical activity and which shares epitopes with the bacterial RecA protein. We demonstrate that the level of Kin17 RNA was 5-fold higher in mid-S phase of serum-stimulated BALB/c 3T3 fibroblasts than in quiescent cells. Cells in S-phase displayed a high level of kin17 protein with a marked nuclear localisation. The maximal level of Kin17 RNA was observed 18 h after serum stimulation, indicating that Kin17 gene is a new member of the late growth-related genes. The accumulation of kin17 protein during cell proliferation follows the increase in Kin17 RNA and correlates with DNA synthesis, which suggests a possible role of kin17 protein in a transaction related to DNA-replication. In quiescent fibroblasts, a 3-fold increase in Kin17 RNA was seen 13 h after UV irradiation. In parallel, kin17 protein accumulated in the nucleus, which suggests that it might be required after the stress produced by UV irradiation.

Overexpression of kin17 protein forms intranuclear foci in mammalian cells.

We used antibodies against E coli RecA protein to identify in mouse cells a 45-kDa DNA-binding protein called kin17, which has an active zinc finger and a nuclear localisation signal. Kin17 protein produced in E coli binds preferentially to the curved DNA of a bacterial promoter in vivo and in vitro, suggesting a transcriptional regulation activity. The fact that in rodent cells kin17 protein levels increase after gamma-irradiation suggests its participation in a cellular response to ionising radiation. We raised polyclonal antibodies against the whole kin17 protein and against its derived synthetic peptides. We report the detection of kin17 protein and of truncated forms of the protein by Western blot or by immunocytochemistry after transient overexpression in cultured human cells. Our results indicate that the cross-reactivity with the anti-RecA antibodies is due to an antigenic determinant located in the core of kin17 protein, between residues 129 and 228. The kin17 protein is located in the nucleus and is concentrated in small nuclear dot-like structures throughout the nucleoplasm. The RecA homologous region seems to play an essential role in the localisation of kin17 protein since the deletion of this particular region dramatically changes the form and the distribution of the intranuclear foci. We hypothesise that these dot-like structures reflect nuclear metabolism compartmentalization.

Lead affects steroidogenesis in rat Leydig cells in vivo and in vitro.

Lead is known to impede the male reproductive function, however, the mechanisms through which the adverse effects are mediated are not clearly elucidated. In order to get insight into those mechanisms, we have examined the effects of lead on the biosynthesis of steroid hormones by Leydig cells in the rat. To determine whether lead has a direct action on Leydig cells, we have compared the concentrations of testosterone secreted by Leydig cells in ex vivo experiments after animals had been injected with high doses of lead and in vitro experiments with Leydig cells from normal rats maintained in culture in presence or absence of lead. In ex vivo experiments male Spargue-Dawley rats were injected i.p. with lead acetate (8 mg lead/kg/day, 5 days a week for 5 weeks) or with sodium acetate. Testosterone production by Leydig cells isolated and maintained in culture for 48 h was then assessed under basal conditions or after stimulation by human chorionic gonadotrophin (hCG). Both basal and hCG-stimulated testosterone production dropped by 59% and 37%, respectively, with Leydig cells from lead-exposed rats. For in vitro experiments, cultures of Leydig cells from control rats were exposed to various concentrations of lead acetate for different periods. Dose and time-dependent reductions of testosterone level were observed in the culture medium. The effective doses of hCG for maximal and half-maximal testosterone production did not change, indicating that the sensitivity of Leydig cells to hCG was not impaired by exposure to lead in vitro. Progesterone production was also decreased after this exposure. The negative effect of lead on testosterone and progesterone production was correlated with the lower expression of the enzymes cytochromes P450scc (CYP11A1) and P450c17 (CYP17) and 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) involved in steroid hormone biosynthesis, as shown by immunohistochemistry. Ultrastructural alterations of the smooth endoplasmic reticulum observed after lead administration might be correlated with the lower expression of the microsomal enzymes P450c17 and 3 beta-HSD. Our results indicate that lead can adversely affect the Leydig cell function by impairing directly steroidogenesis.

Reproductive toxicity of chronic lead exposure in male and female mice.

The reproductive toxicity of lead was investigated in NMRI mice exposed to 0.5% lead acetate in drinking water from day 1 of intra-uterine life until 60 days after birth. Compared with control mice, the weights of lead-exposed fetuses and subsequently of the lead-exposed weaned pups, male and female, diminished by 11 and 13% respectively. The lead-exposed male and female offspring of lead-exposed dams were mated with unexposed females and males, to examine the effect of lead exposure on reproductive function. Male fertility was not affected but reduced female fertility was observed: litters were smaller and a smaller number of implantation sites was found in lead-exposed females. In lead-exposed males, the weights of the body, testes and epididymes diminished by about 13%, and seminal vesicle and ventral prostate weights, by about 29%. Testicular histology and the number and morphology of epididymal spermatozoa were normal. The levels of plasma FSH, LH and testosterone, and of testicular testosterone, were not modified. These results suggest that the hypothalamic-pituitary-testicular axis is not adversely affected by the above lead exposure, and that therefore the decreased seminal vesicle and ventral prostate weights might not be the consequence of reduced testosterone levels. The hypothesis that lead has a direct effect on these organs as well as a secondary effect resulting from possibly reduced food consumption by lead-exposed mice cannot be excluded. Consequently, in male NMRI mice, exposure to lead might affect reproductive function by acting directly and/or indirectly on accessory sex organs.

Impairment of testicular endocrine function after lead intoxication in the adult rat.

To clarify the mechanism of the action of lead on male reproductive function, adult male rats were injected intraperitoneally (i.p.) with lead acetate (8 mg/kg/day of lead), 5 days a week for 35 days. Despite this high dose, germ cells and Sertoli cells did not appear to be major targets of lead. However, lead determination in the reproductive organs showed that the accessory sex glands are such a target. Epididymal function was unchanged. In lead-exposed rats, plasma and testicular testosterone dropped by about 80%, but plasma luteinizing hormone (LH) only dropped by 32%. After luteinizing hormone releasing hormone (LHRH) stimulation of the pituitary, the plasma LH level reached the control one, but plasma testosterone remained significantly reduced by 37%. The sharp decrease in the testosterone:LH ratio in lead-exposed rats, combined with the significant reduction of intertubular tissue volume in the testes, indicate impaired Leydig cell function.

Translocation of plutonium from rat and monkey lung after inhalation of industrial plutonium oxide and mixed uranium and plutonium oxide.

This study was designed to compare the translocation from lung of the Pu contained in the pure and mixed industrial oxides PuO2 and (U,Pu)O2. The latter had a Pu content of 20% w/w. For this purpose, young adult male rats and male and female baboons were exposed to a single inhalation of these oxides. Two baboons were exposed to the reference PuO2, i.e. 239PuO2. Rats were killed under anaesthesia 1, 15, 30, 90 and 180 days after exposure, and baboons, also under anaesthesia, 1 year thereafter. The results indicate that lung retention of Pu was independent of the oxide inhaled, but was smaller in rat (12-15% of the initial pulmonary burden, 6 months after exposure) than in baboon (56-80% of this burden, 1 year after exposure). In rat, Pu translocation kinetics were similar for the two industrial oxides, but as from day 15 after inhalation until 6 months thereafter, measurement of Pu deposits in the liver and skeleton showed that translocation of Pu from the mixed oxide was 2-3 times greater than that from the industrial Pu oxide. In baboon, the largest amounts of Pu were retained in the lung and thoracic lymph nodes for the three oxides inhaled. Pu translocation to the liver, skeleton and kidneys, and also urinary Pu excretion, were greater after inhalation of the mixed oxide than after inhalation of the industrial and reference Pu oxides. Nevertheless, the amount of mixed oxide Pu translocated to these sites and excreted in urine remained under 3% of the initial pulmonary burden.

Effects of lead poisoning of rats during pregnancy on the reproductive system and fertility of their offspring.

1. The effects of lead poisoning during pregnancy were tested on female Sprague-Dawley rats that inhaled 5 mg m-3 lead oxide for 13 days during gestation. At the end of gestation, the respective blood lead levels of dams and fetuses were 71.1 and 83.2 micrograms 100 ml-1, indicating lead poisoning. 2. In the 90 day-old male offspring of the exposed dams, testis weight and histology, and epididymal weight and sperm reserve, were all similar to those of control males. Spermatozoa mobility and morphology were normal. 3. Also similar to control values were the pituitary weight in these male offspring, their plasma FSH, LH and testosterone levels, and the weight of their ventral prostate and seminal vesicles, the targets of the sexual hormones. 4. When male and female offspring of exposed dams were mated, their fertility was normal, with no increase in prenatal death or malformations, and no changes in the size or sex ratio of litters. 5. These results indicate that, under our experimental conditions, lead oxide inhalation by rats during pregnancy did not perturb reproductive function in their male offspring.

Effect of ingestion and inhalation of lead on the reproductive system and fertility of adult male rats and their progeny.

Ninety-day-old Sprague-Dawley rats were intoxicated for 70 d with lead, given either as 0.3% lead acetate in drinking water or by inhalation as 5 mg m-3 lead oxide. Direct or transmitted lead toxicity for the male reproductive system was assessed in the rats and their offspring from pituitary and genital organ weights after exposure, the numbers of Sertoli and germ cells, the number, motility and morphology of epididymal spermatozoa, the levels of plasma testosterone, LH and FSH and fertility tests. Whole blood lead levels were similar after lead ingestion and after inhalation (58.0 +/- 1.7 micrograms dl-1 vs. 51.1 +/- 1.8 micrograms dl-1). Lead acetate ingestion did not affect the reproductive system or fertility of rats. Inhalation of lead oxide did not affect fertility either, but seminal vesicle weight dropped significantly, which might suggest an alteration in the pattern of testosterone secretion. In the male progeny of sires that inhaled lead, the number of epididymal spermatozoa decreased but this did not interfere with fertility. Our results show that for the doses studied, lead inhalation and lead ingestion do not produce strikingly different effects on the male rat's reproductive system. Differences between the present findings and those of others might be due to difference of rat strain or of age at exposure.

Effect of an acute exposure of rat testes to gamma rays on germ cells and on Sertoli and Leydig cell functions.

Germ cells and Sertoli and Leydig cell functions were studied from 7 to 180 days after an acute exposure of 2-month-old rat testes to 9 Gy of gamma rays. Body weight, testis and epididymal weights were recorded. Sertoli cell parameters (androgen-binding protein, ABP, in caput epididymis and plasma follicle stimulating hormone, FSH) and Leydig cell parameters (plasma luteinizing hormone, LH, testosterone and prostate and seminal vesicle weights) were determined together with the number of germ cells and Sertoli cells. Irradiation did not affect body weight but significantly reduced testicular and epididymal weights from day 7 and day 15 post-irradiation respectively. The cells killed by irradiation were mainly spermatogonia and preleptotene spermatocytes engaged in replicating their DNA at the time of exposure, but all spermatocytes seemed damaged as they gave abnormal descendent cells. By day 34, only elongated spermatids remained in a few tubules and thereafter very little regeneration of the seminiferous epithelium occurred, except for one rat which showed a better regeneration. Levels of ABP decreased by day 15 when the germ cell depletion had reached the pachytene spermatocytes, whereas FSH and LH levels rose when the number of elongated spermatids decreased. Levels of testosterone and the weight of the seminal vesicles did not change; occasionally, the prostate weight was slightly reduced. These results support our hypothesis that pachytene spermatocytes and elongated spermatids are involved in influencing some aspects of Sertoli cell function in the adult rat.

Effect of mass, oxidation state and duration of chronic ingestion on plutonium absorption in fed rats.

The effects of time, mass and oxidation state on plutonium gastrointestinal absorption and tooth adsorption were studied during and after chronic ingestion of plutonium-238 (IV) or (VI) (1.55-15.60 kBq/ml) in 6.5 mM bicarbonate medium by fed rats via drinking water for 8 days to 3 months. Animals were killed during the ingestion to follow the kinetics of whole-body storage and clearance of plutonium. At 1.55 kBq/ml the amount of plutonium retained in the skeleton increased continuously during the 85 days of ingestion and reached a plateau thereafter. This plutonium retention was therefore dependent on the total mass administered but not proportional to this mass, as the fraction of administered plutonium retained decreased during the first 22 days of ingestion and then stabilized. This is reflected by the gastrointestinal transfer (f1), which had risen to (3.80 +/- 0.82) x 10(-5) on Day 3 of ingestion and then decreased to a stabilized value of (1.07 +/- 0.06) x 10(-5) from Day 30 to the end of the ingestion period. In the liver, the amount of plutonium retained reached a plateau, which lasted from Day 30 to the end of ingestion. The kidneys and spleen were also found to be retention sites. By Day 3 of ingestion, for a mass ingested of 5 x 10(-7) g/kg of body mass, the maximum mean value of f1 we found was smaller than the 10(-4) recommended by ICRP Report 30. The oxidation state had no effect on f1. Large plutonium deposition was observed on the teeth. For both oxidation states (IV) and (VI), about 0.10% of the administered dose was deposited on the teeth after 3 days of ingestion, whatever the plutonium concentration administered. However, whereas the amount of plutonium (IV) deposited did not change throughout the ingestion period, tooth deposition of plutonium (VI) decreased.

Assessment of testicular function after acute and chronic irradiation: further evidence for an influence of late spermatids on Sertoli cell function in the adult rat.

To study cell to cell communications within the testis of adult Sprague-Dawley rats, we used acute whole body neutron plus gamma-irradiation (0.99 Gray of neutron and 0.24 Gray of gamma-rays, 3 min; Exp A) over 7-121 days postirradiation and chronic whole body gamma-irradiation (7 cGy/day 60Co gamma-rays; Exp B) over 14-84 days of irradiation and 7-86 days postirradiation. Neither irradiation protocol had an effect on the body weight of the animals. Neutron plus gamma-rays induced dramatic damages to spermatogonia, preleptotene spermatocytes, spermatozoa, and, to a lesser extent, pachytene spermatocytes. In contrast, gamma-rays induced a selective destruction of spermatogonia. Subsequently, in both experiments a maturation-depletion process led to a marked decrease in all germ cell types. A complete or near complete recovery of the different germ cell types and spermatozoa took place during the two postirradiation periods. Under both irradiation protocols Sertoli cells number was unchanged. Androgen-binding protein and FSH levels were normal in spite of the disappearance of most germ cells from spermatogonia to early spermatids. However, the decline of androgen-binding protein as well as the rise of FSH and their subsequent recovery were highly correlated to the number of late spermatids and spermatozoa. Moreover, it appeared that spermatocytes may also interfere with the production of inhibin (Exp B). With neither irradiation was Leydig cell function altered, except in Exp B in which elevated LH levels were temporarily observed. Correlation analysis suggested a relationship between preleptotene spermatocytes and Leydig cell function. In conclusion, this study establishes that chronic gamma-irradiation is particularly useful in the study of intratesticular paracrine regulation in vivo and provides further support to the concept that late spermatids play a major role in controlling some aspects of Sertoli cell function in the adult rat.

Changes in androgen binding protein (ABP) production following continuous low dose gamma irradiation (IR) of adult rat.

Continuous low dose gamma irradiation induces a progressive degeneration of germ cells with a concomittant increase in blood FSH; however, the Sertoli cell function is not too much altered since serum ABP level is normal and it is likely that the decrease of epididymal ABP content is the consequence of a reduction in seminiferous tubule fluid excretion. Obviously, spermatids seems to be involved in the regulation of Sertoli cell ABP synthesis.

Localization of plutonium retention in the small intestine of the neonatal rat, guinea pig, baboon and macaca after Pu-citrate ingestion.

The retention of Pu-citrate in the gastrointestinal wall was compared at similar post ingestion times after ingestion at 2 days of age by rats and guinea pigs and at 1 to 34 days by neonatal primates. The small intestine was the main site of the Pu retention in all species. In rats and primates, most of the Pu was retained in the distal ileum, whereas in guinea pigs it was more homogeneously distributed. In the rats, Pu was retained in the epithelial cells on villi, but in the guinea pigs and primates it was confined to the macrophages under the epithelial cells in the lacteal region.

Influence of germ cells upon Sertoli cells during continuous low-dose rate gamma-irradiation of adult rats.

The effects of continuous gamma-irradiation of adult rats at two low-dose rates (7 cGy and 12 cGy/day; up to a total dose of 9.1 Gy and 10.69 Gy 60Co gamma-ray, respectively) were investigated. Over a period of 3-131 days of irradiation, groups of experimental and control animals were killed. Body weight, testis, epididymis, prostate and seminal vesicle weights, the number of germ cells and Sertoli cells, tubular ultrastructure, epididymal and testicular levels of biologically active androgen-binding protein (ABP), and the plasma concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone were monitored. Irradiation had no effect on body weight, whereas testicular and epididymal weight began to decrease following 35 and 50 days of irradiation at 7 and 12 cGy, respectively. At 7 cGy the target cells of the gamma-rays were essentially A spermatogonia, whereas at 12 cGy A spermatogonia and preleptotene spermatocytes were primarily affected. This resulted in a progressive and sequential dose-related reduction in the number of pachytene spermatocytes, round spermatids and late spermatids (LS). Under both irradiation procedures the Sertoli cell number remained unchanged whereas partial (7 cGy) or no change (12 cGy) was seen at the Leydig cell level. Whatever the irradiation protocol, from the time LS numbers decreased, vacuolisation of the Sertoli cell cytoplasm progressively occurred, followed by thickening and folding of the peritubular tissue. Moreover, in parallel to the drop in the number of these germ cell types, ABP production fell whereas FSH levels rose. A highly significant positive correlation was found between LS numbers and these Sertoli cell parameters. This study supports our previous concept of a control of certain important aspects of Sertoli cell function by late spermatids in the adult rat.

Effect of continuous low-dose gamma-irradiation on rat Sertoli cell function.

Continuous low-dose gamma-irradiation of mature rats induced a progressive degeneration of the germ cells. Blood FSH increased by 127, 176 and 214%, respectively, after 55, 70 and 85 days of treatment when compared to FSH levels in control rats (8.50 +/- 0.60 ng/ml); conversely, serum LH and testosterone levels were unchanged. The Sertoli cell function was affected by the treatment from 70 days on, as attested by androgen binding protein (ABP) and transferrin secretions which diminished 35-40%. Serum ABP levels were not altered, whatever the duration of irradiation, even though epididymal ABP contents (as well as concentrations) diminished 34-60% when compared to those of the controls. Moreover, in purified Leydig cells, LH-stimulated intracellular cAMP levels, which were decreased by seminiferous tubule medium (STM) from control rats, were enhanced in presence of STM from treated animals. Testosterone output was stimulated 9-fold in presence of oLH and further increased (46-76%) from stages XIV-V by STM prepared from control and irradiated rats, respectively. After 85 days the STM effects on both cAMP and testosterone syntheses were zero. These results demonstrate a probable alteration of Sertoli cell function after irradiation, but also a role of the germ cells in the regulation of the synthesis of ABP, transferrin and Sertoli cell paracrine factors.