DES-treated Females Research Articles

Mice lacking membrane estrogen receptor 1 are protected from reproductive pathologies resulting from developmental estrogen exposure†.

Both membrane and nuclear fractions of estrogen receptor 1 (ESR1) mediate 17β-estradiol (E2) actions. Mice expressing nuclear (n)ESR1 but lacking membrane (m)ESR1 (nuclear-only estrogen receptor 1 [NOER] mice) show reduced E2 responsivity and reproductive abnormalities culminating in adult male and female infertility. Using this model, we investigated whether reproductive pathologies caused by the synthetic estrogen diethylstilbestrol (DES) are mitigated by mESR1 ablation. Homozygous and heterozygous wild-type (WT and HET, respectively) and NOER male and female mice were subcutaneously injected with DES (1 mg/kg body weight [BW]) or vehicle daily from postnatal day (PND) 1-5. Uterine histology was assessed in select DES-treated females at PND 5, whereas others were ovariectomized at PND 60 and treated with E2 (10 μg/kg BW) or vehicle 2 weeks later. Neonatal DES exposure resulted in ovary-independent epithelial proliferation in the vagina and uterus of WT but not NOER females. Neonatal DES treatment also induced ovary-independent adult expression of classical E2-induced transcripts (e.g., lactoferrin [Ltf] and enhancer of zeste homolog 2 [Ezh2]) in WT but not NOER mice. At PND 90, DES-treated WT and HET males showed smaller testes and a high incidence of bacterial pyogranulomatous inflammation encompassing the testes, epididymis and occasionally the ductus deferens with spread to lumbar lymph nodes; such changes were largely absent in NOER males. Results indicate that male and female NOER mice are protected from deleterious effects of neonatal DES, and thus mESR1 signaling is required for adult manifestation of DES-induced reproductive pathologies in both sexes.

Prenatal diethylstilbestrol induces malformation of the external genitalia of male and female mice and persistent second-generation developmental abnormalities of the external genitalia in two mouse strains

Potential trans-generational influence of diethylstilbestrol (DES) exposure emerged with reports of effects in grandchildren of DES-treated pregnant women and of reproductive tract tumors in offspring of mice exposed in utero to DES. Accordingly, we examined the trans-generational influence of DES on development of external genitalia (ExG) and compared effects of in utero DES exposure in CD-1 and C57BL/6 mice injected with oil or DES every other day from gestational days 12 to 18. Mice were examined at birth, and on 5–120 days postnatal to evaluate ExG malformations. Of 23 adult (>60 days) prenatally DES-exposed males, features indicative of urethral meatal hypospadias (see text for definitions) ranged from 18% to 100% in prenatally DES-exposed CD-1 males and 31% to 100% in prenatally DES-exposed C57BL/6 males. Thus, the strains differed only slightly in the incidence of male urethral hypospadias. Ninety-one percent of DES-exposed CD-1 females and 100% of DES-exposed C57BL/6 females had urethral–vaginal fistula. All DES-exposed CD-1 and C57BL/6 females lacked an os clitoris. None of the prenatally oil-treated CD-1 and C57BL/6 male and female mice had ExG malformations. For the second-generation study, 10 adult CD-1 males and females, from oil- and DES-exposed groups, respectively, were paired with untreated CD-1 mice for 30 days, and their offspring evaluated for ExG malformations. None of the F1 DES-treated females were fertile. Nine of 10 prenatally DES-exposed CD-1 males sired offspring with untreated females, producing 55 male and 42 female pups. Of the F2 DES-lineage adult males, 20% had exposed urethral flaps, a criterion of urethral meatal hypospadias. Five of 42 (11.9%) F2 DES lineage females had urethral–vaginal fistula. In contrast, all F2 oil-lineage males and all oil-lineage females were normal. Thus, prenatal DES exposure induces malformations of ExG in both sexes and strains of mice, and certain malformations are transmitted to the second-generation.

Diethylstilbesterol has Gender-Specific Effects on Weight Gain and Bone Development in Mice

Neonatal exposure to diethylstilbesterol (DES) in female mice programs estrogen-sensitive tissues, resulting in greater body weight gain and positive effects on bone architecture at adulthood. Using the CD-1 mouse model, the objective of the present study was to examine how short-term neonatal exposure to DES modulates weight gain as well as bone mineral density (BMD), bone strength, and bone microarchitecture in both males and females at adulthood. Male and female offspring (n = 8–12 pups/treatment/gender) were randomized to DES (2 mg/kg bw/d) or control (corn oil) from postnatal day 1 to 5 (subcutaneous injection, once daily) and sacrificed at 4 mo of age. Body weight was measured weekly, while bone mineral, strength, and microarchitecture were measured at 4 mo of age. DES treatment resulted in significantly higher body weight in females but lower weight in males at 4 mo of age. In DES-treated females, markedly higher BMD of lumbar vertebrae (LV1–LV3) was translated into significantly stronger LV2 that was more resistant to fracture; similar effects were observed at the femur midpoint. At the spine, males had a markedly lower BMD and peak load, suggesting an adverse effect. Microstructural analyses demonstrated that functional changes in femurs, i.e., peak load, were primarily due to modulation of cortical bone. In conclusion, neonatal exposure to DES exerted gender-specific effects on body weight gain and bone health.

Diethylstilbestrol (DES) has gender‐specific effects on body weight and bone development in mice

Neonatal exposure to DES in female mice programs estrogen‐sensitive tissues, resulting in greater body weight and positive effects on bone metabolism at adulthood. Because DES can be used as a positive control when investigating effects of dietary estrogens in early life, the study objective was to determine how short‐term neonatal exposure to DES modulates body weight, as well as bone mineral, strength and microarchitecture in both male and female mice at adulthood. Male and female offspring (n=8–12 pups/treatment/gender) were randomized to DES (2 μg/d) or control (corn oil) from postnatal day 1 to 5 (subcutaneous injection) and killed at 4 months of age. Body weight was measured weekly. Bone mineral, strength and microarchitecture were measured at 4 months of age. DES treatment resulted in higher (p<0.05) body weight among females but lower (p<0.05) body weight among males at 4 months of age. Among DES‐treated females, higher (p<0.05) BMD of LV1‐LV3 was translated into stronger (p<0.05) LV2 that was more resistant to fracture with similar effects at the femur midpoint. At the spine, males had lower BMD (p<0.05) and peak load (p<0.05) suggesting an adverse effect. Microstructural analyses demonstrated that functional changes in femurs (i.e. peak load) were primarily due to changes in cortical bone. In conclusion, early exposure to DES has gender‐specific effects on body weight and bone health.

NEONATAL ESTROGEN TREATMENT AND ITS CONSEQUENCES FOR THYMUS DEVELOPMENT, SERUM LEVEL OF AUTOANTIBODIES TO CARDIOLIPIN, AND THE DELAYED-TYPE HYPERSENSITIVITY RESPONSE

Eight-week-old female and male NMRI mice treated neonatally with the synthetic estrogen diethylstilbestrol (DES), estradiol-17 b , or tamoxifen displayed an enlarged thymus when compared with controls (approximately 1.5-fold). In control females, either ovariectomy or adrenalectomy increased thymus weight to the level characteristic for DEStreated females, but these endocrine ablations had no significant effect in DES females. The serum estrogen levels were similar in intact DES, ovariectomized DES, and ovariectomized female controls; serum corticosterone was similar in controls and DES females. The expression of the Thy1.2+ marker and the percentages of CD4+CD8+ DP and CD4+ and CD8+ SP cell subsets were similar in thymocyte populations from 8-wk-old controls and DES females; the CD4+ and CD8+ SP subsets were similar in splenocyte populations. The levels of serum immunoglobulin (Ig) G and IgM autoantibodies to cardiolipin showed age-dependent fluctuations but were similar in controls and DES females; however, the IgG autoantibodies in DES females were qualitatively different from those in controls with respect to sensitivity to bovine serum (a source of Beta2-glycoprotein I). Contrary to females, DES-treated males had higher levels of autoantibodies than controls. The delayed-type hypersensitivity (DTH) response to oxazolone was similar in controls and DES animals at 8 wk, increased in DES females and males at 6 mo, but was reduced in DES females at 1 yr. Thus, even though adult mice with thymus enlargement after neonatal estrogen treatment do not differ from controls with respect to the expression of the Thy1.2 marker or percentages of CD4+/CD8+ DP or SP subsets in thymus and spleen, qualitative and quantitative differences occur in immune parameters (autoantibodies to cardiolipin) and a T-cell-dependent immune response (DTH).

Socialization processes in primates: use of multivariate analyses: II—Influence of sex on social development of captive rhesus monkeys

In a previous study, we demonstrated the importance of social interactions in the development of non-human primate infants. These results confirmed the social network concept. According to this concept, it is assumed that socialization processes would differ in various social environments. However, much variability remained to be explained. In the present study we investigated especially the influence of infants' gender on socialization processes. In relation to the previous results, the influence was tested within groups having the same social structure. At the Wisconsin Regional Primate Research Center, four mother-peer groups of rhesus macaques ( Macaca mulatta) were studied. Therefore no adult male model was available to infants. Twenty infants, including eight males, eight females and four prenatally DES-treated females, were the subjects of this study. We considered six comprehensive developmental parameters to account for the two main socialization processses, the acquisition of the social behavioral repertoire and the establishment of a network of social relationships. We analyzed the variability of these parameters using a new multivariate technique previously described in a companion paper. This new technique is derived from the Principal Components Analysis and Multivariate Analyses of Variance. The model used in this analysis included two intrinsic features of infants, sex and age and two variables related to the social organization of the groups, mother's rank and social group. This 4-variable-model significantly accounted for most of the variability of the developmental parameters. The results showed that the social group and the infant's sex were the most influential variables when infants initiated interactions: male and DES female infants displayed a greater behavioral diversity than non-treated female infants. When infants were recipients in interactions, social organization variables (social group and mother's rank, respectively) had the greatest influence on the socialization processes while infants' gender was less influential. In this case the effect of sex was confounded with the social group effect; male infants then differed from both kind of female infants in being looked at more frequently, in being contacted by a larger variety of partners and in receiving a greater variety of social behaviors. This new example of applying multivariate methods to the study of individual social development confirms the potential of this method to provide new insights into behavioral development.

The Effect of Prenatal Exposure to the Phytoestrogen Genistein on Sexual Differentiation in Rats

Exposure to naturally occurring estrogens during critical periods of development can alter morphologic and physiologic markers of sexual differentiation. The current experiment characterizes the effects of in utero treatment with genistein, an isoflavonoid phytoestrogen, on birth weight, anogenital distance (AGD) at birth. GnRH stimulated luteinizing hormone (LH) secretion, volume of the sexually dimorphic nucleus in the preoptic area of the hypothalamus (SDN-POA), puberty onset, and vaginal cyclicity. Pregnant Charles River CD rats were injected sc daily on gestation day 16-20 with either 25,000 micrograms genistein (G25), 5,000 micrograms genistein (G5), 5 micrograms diethylstillbestrol (DES), 50 micrograms estradiol benzoate (E), or corn oil alone for controls. Birth weights and anogenital distance was taken and exposed progeny were subsequently used in two experiments. In Experiment 1 intra-atrial catheters were placed in adult castrated rats, GnRH was given iv, serial blood samples were drawn and sera were assayed for LH by radioimmunoassay (RIA). Brains obtained by subsequent decapitation were saved for histology. In Experiment 2, females were monitored for timing of vaginal opening as a marker of puberty onset, and vaginal smears were taken to monitor cyclicity. G25-treated females and DES- and E-treated animals of both sexes had decreased weights at birth compared with controls. G5- and E-treated animals of both sexes and DES males had smaller AGD than controls. No significant differences in pituitary responsiveness to GnRH were found among treatment groups. There was a nonsignificant decrease in SDN-POA volume in G5-treated females while DES- and E-treated females had increased SDN-POA volume compared with controls. G5-treated females had delayed puberty onset, and DES-treated females had atypical vaginal cycles in comparison with controls. The results confirm that prenatal exposure to estrogens in the environment can influence sexual differentiation. Our previous experiments have demonstrated that castrate female rats exposed as neonates to genistein have decreased pituitary responsiveness to GnRH challenge and enlarged SDN-POA volume in comparison with controls. Prenatal genistein at these dosages did not significantly alter these markers. However, genistein did mimic other estrogens' effects on AGD and birth weight and had a unique influence on puberty onset. Not only are genistein's effects different from other estrogens, but dosage and timing of exposure during development appear to be important factors in genistein's ability to modify these end points.

Steroid synthesis in ovarian homogenates from hypophysectomized adult female mice treated with diethylstilbestrol in neonatal life.

Eight-week-old female NMRI-mice treated with 5 micrograms diethylstilbestrol (DES) or vehicle (olive oil) from d 1 to 5 after birth were hypophysectomized or sham operated. Thirty days after the operation, homogenates were prepared from the ovaries and incubated at 37 degrees C in the presence of [3H]pregnenolone for 1 h. In parallel experiments ovaries were taken for histology. In control ovaries regressing corpora lutea were still present 4 wk after hypophysectomy (Hx). Ovaries from DES females had no corpora lutea, but showed degeneration of the pre-Hx hypertrophic interstitial tissue. Steroids synthesized during the incubation were extracted and separated in a two-dimensional thin-layer chromatography system. After Hx, the amount of extractable [3H]testosterone, [3H]progesterone, and [3H]androstenedione recovered from homogenates of vehicle-treated females increased, but the amount of recovered [3H]-17 alpha-hydroxyprogesterone was not significantly altered. In contrast, in DES-treated females Hx significantly decreased levels of [3H]androstenedione and [3H]-17 alpha-hydroxyprogesterone, but did not affect [3H]progesterone or [3H]testosterone. The persistent difference in steroid synthetic activity after Hx could be linked to the difference in morphology between different ovarian tissue compartments of control and DES-treated females.

Alterations in hypothalamic and pituitary hormone levels induced by neonatal treatment of female mice with diethylstilbestrol

Neonatal female mice of the NMRI strain were treated with the synthetic estrogen diethylstilbestrol (DES), 5 μg daily for the first 5 days after birth, or with vehicle only. Levels of LH and FSH (pituitary and serum) and LHRH (hypothalamus) were measured by radioimmunoassay (RIA) in 6- to 56-day-old females with 6- to 7-day intervals. Compared to controls, DES-treated females had low levels of LH on days 6, 12, and 21 in the pituitary, and on days 6 and 12 in serum; increased LH levels were seen in both the pituitary and serum on day 42. The FSH levels of DES-treated females were decreased on days 6 and 12 in the pituitary and on day 6 in serum; an increased FSH content occurred on day 21 in the pituitary. In the preoptic area and basal hypothalamus of DES-treated females, levels of LHRH were increased on day 21 and decreased on day 42. On day 56, the serum levels of FSH and LH and the hypothalamic content of LHRH were similar in controls and DES females. A second study including both synthetic and natural estrogen was performed in 12-day-old females. Treatment with 10 −2 μg DES or lower doses or 5 μg estradiol-17β (E 2) on days 1 to 5 after birth had no depressive effect on serum LH. The hypothalamic-pituitary-ovarian feedback system reacted similarly to ovariectomy and E 2 challenge in 15-day-old control and DES-treated females. DES-treated 56-day-old females had a reduced LH response to ovariectomy but increased response to exogenous E 2 compared to controls. These results show that neonatal treatment with DES has pronounced effects on the hypothalamic-pituitary system in the developing female mouse, which may be of importance for the altered ovarian function in these females as adults.

Failure to detect a second-generation effect in female mice after neonatal treatment with an estrogen (diethylstilbestrol).

Inbred female mice of the NMRI strain were treated subcutaneously with 5 micrograms diethylstilbestrol (DES) in olive oil or vehicle only for the first 5 days after birth. One group of DES-treated females was killed at the age of 8-12 weeks, and the uterine cervix and adjacent parts of the vagina and uterine horns prepared for histological studies. In all preparations, the cervical epithelial lining contained regions with heterotopic columnar epithelium (HCE) along 69-100% of the length of the common cervical canal. Ovaries from neonatally DES-treated females were grafted to 8-week-old ovariectomized control hosts and these hosts were mated to control males 2 weeks later. The hosts gave birth to normal-sized litters. The female offspring from these litters had a normal cervical epithelial lining and, in turn, gave birth to normal-sized litters. These results indicate that treatment of neonatal female mice with DES does not affect the female germ cells as far as concerns factors associated with the development of HCE or reduced fertility in the next generation.

The functional importance of the oviduct in neonatally estrogenized mouse females for early embryo survival

Eight-week-old virgin untreated female mice were induced to ovulate using equine chorionic gonadotropin (eCG) and human chorionic gonadotropin (hCG), and were then caged with males overnight. Females with a vaginal plug on the following morning were killed 24 hours later and 2-cell embryos were flushed from the oviduct. These embryos were transferred to the oviduct of 8-week-old control females, to females of the same age treated with 5 micrograms diethylstilbestrol (DES) sc in olive oil for the first 5 days after birth, or to females treated with 1 microgram estradiol-17 beta for 2 days before and 2 days after transfer (estrogen dominated/ED/females). Two days after transfer, a significantly lower number of embryos were recovered from oviducts of DES females compared to control females and a still lower number from ED females. The recovered embryos were cultured in vitro for 4 days testing trophoblast outgrowth ("implantation stage"). The incidence of embryos reaching this stage after development in DES-exposed oviducts was only half of that for embryos passing control oviducts or ED oviducts. It is concluded that the adult oviductal environment in neonatally DES-treated females significantly decreases early embryo developmental potential. The oviductal factor(s) harmful to the embryo may be related to a persistent and possibly increased level of circulating estrogen level in DES females.

Effects of Neonatal Exposure to Diethylstilbestrol on Early Mouse Embryo Development in Vivo and in Vitro1

Eight-week-old female mice of the NMRI strain that had been treated neonatally with diethylstilbestrol (DES, 5 micrograms/day for five days) or not (controls) were treated with gonadotropins to induce ovulation and then were artificially inseminated. Ova or young embryos were recovered from the oviducts on the morning after insemination and on Days 2, 3, and 4. In other experiments, ova were obtained from inseminated females on the morning after ovulation and cultured in vitro. In DES-treated females, a few zygotes developed to the 4-cell stage, but no more advanced stages were seen. Under in vitro conditions, zygotes from DES-treated females developed into blastocysts and to the implantation stage, but the incidence of these stages was lower than with zygotes from controls. Our results point to an abnormal oviductal function in DES-treated females that is not compatible with early embryo survival, even though an additional zygote factor contributing to degeneration of early cleavage stages cannot be excluded.

Ovarian Reproductive Function after Exposure to Diethylstilbestrol in Neonatal Life1

Inbred and random-bred NMRI mice were treated with diethylstilbestrol (DES, 5 micrograms per day) or vehicle (olive oil) on Days 1-5 after birth. At the age of 8 wk, females were treated with saline or eCG and hCG to induce ovulation. Ova never occurred in the ampulla of the uterine tube of saline-treated, DES-treated females when these mice were not mated. After gonadotropin treatment, ova were found in the ampulla of all olive oil-treated females and in approximately 80% of DES-treated females. The number of ovulated ova was similar in both groups. Twenty percent of gonadotropin-treated, DES-treated females had ova in the ampulla and a vaginal plug after being caged with males but none became pregnant. Ovaries from inbred control or DES-treated females were grafted to the ovarian bursa of control or DES-treated ovariectomized hosts. DES-treated hosts, carrying control or DES-exposed ovaries, never became pregnant. Control females, with control ovaries or DES-exposed ovaries, became pregnant; pregnancy rate and litter size were similar for control mice regardless of whether they were supporting DES-exposed or control ovaries. Oocytes from ovaries exposed neonatally to DES can thus give rise to apparently normal offspring. The results also indicate DES-induced nonovarian disturbances, e.g. tubal and/or endometrial function, both of which are important for fertility. In the grafting experiments, a high mortality rate was found in inbred DES-exposed females caged with males. All deaths were associated with vaginal concrements (vaginal stones) and intestinal complications.

Hormonal imprinting of the microsomal enzyme system in adults. Microsomal activity change in response to estrogen (DES, AE) treatment during liver regeneration.

Estrogen (diethylstilbestrol-DES or allylestrenol-AE) treatment applied to rats of both sexes during liver regeneration following subtotal hepatectomy had a long lasting influence on the inducibility by phenobarbital of the hepatic microsomal enzyme system of the females. The enzyme activities of the DES-treated females differed hardly from the baseline two weeks after treatment, but increased almost two-fold over control on induction with phenobarbital 5 and 7 weeks later. The AE-treated females showed a smaller although yet significant, enzyme activity increase only at 7 weeks. The influence of estrogens was negligible, and inhibitory rather than stimulatory, in the males. It appears that, in appropriate conditions, enzyme imprinting can also be induced in adult organisms, since, in all probability, availability for imprinting depends not so much on the age of the organism, as on the developmental state of the target cell.

Use of DES-treated rats as an animal model for assessment of pituitary adenoma imaging agents

Prolactin (PRL) secreting pituitary adenomas are the most common type of pituitary tumors. An imaging agent which specifically localized in prolactinomas would be of considerable clinical value for both initial detection and also for monitoring the effects of dopamine agonist therapy. Tritiated spiroperidol ( 3HSp) was selected for initial evaluation as a possible imaging agent based on: (1) demonstrated localization in the pituitary and (2) demonstrated binding to human PRL-secreting tumor tissue. DES was implanted in Fischer F344 rats and induced prolactinoma formation was evidenced by increased pituitary weight, elevated serum PRL levels and by an increase in the proportion of PRL-secreting cells in the pituitary. 3HSp concentrations in pituitary and other tissues of DES-treated rats were assessed in female rats and correlation studies showed that a 5-fold increase in serum PRL was associated with a 6-fold increase in both pituitary weight and % dose/organ accumulation of 3HSp. The number of pituitary D 2 receptors per mg of protein in tissue homogenates was similar in both normal and DES-treated females. A blocking study with (+)-butaclamol demonstrated a D 2 receptor-mediated component to 3HSp localization. In summary, an animal model for prolactinoma was characterized. An assessment of 3HSp accumulation indicates that radiolabelled spiroperidol shows excellent potential for detecting PRL-secreting tumors of the pituitary.

Structural and functional changes in ovaries from adult mice treated with diethylstilboestrol in the neonatal period.

Ovaries from 8-week-old female NMRI mice in different stages of the oestrous cycle, or from females neonatally treated with the synthetic oestrogen diethylstilboestrol (DES; 5-10(-6) micrograms daily for 5 days), were studied histologically and for the ability to synthesize steroids from [3H]pregnenolone in vitro. Daily doses of 10(-4) micrograms DES or higher resulted in absence of corpora lutea. In ovaries lacking corpora lutea, the interstitial tissue dominated and the cells in this compartment were large with a clear cytoplasm. The steroids synthesized in ovarian homogenates were separated with thin-layer chromatography. The homogeneity of the steroids was checked in recrystallization experiments. Daily doses of 5-10(-4) micrograms DES in the neonatal period resulted in pronounced deviations in the pattern of ovarian steroids synthesized as compared with control ovaries. In DES-exposed ovaries, the synthesis of androstenedione and, above all, progesterone was high while the synthesis of 17 alpha-hydroxyprogesterone and testosterone was reduced compared with controls. These results could argue for a difference in activities of 17 alpha-hydroxylase and 17 beta-ol-dehydrogenase in ovaries from DES-treated females compared with controls. After transplantation of DES-exposed ovaries to ovariectomized control females, the steroid pattern changed to that typical for control ovaries. Control ovaries transplanted to DES-treated females had a steroid pattern similar to that of DES-exposed ovaries.

3-Methylcholanthrene: Transient Inhibition of the Lytic Step of Mouse Natural Killer Cells<xref ref-type="fn" rid="FN2">2</xref><xref ref-type="fn" rid="FN3">3</xref>

Female mice belonging to NMRI stock were inoculated neonatally with daily doses of 5 micrograms diethylstilbestrol (DES) or olive oil for the first 5 days after birth and in adult life with 20 or 100 micrograms 3-methylcholanthrene (MCA) or the vehicle tricaprylin only. The cytotoxic activity of spleen natural killer (NK) cells against YAC-1 target cells was studied in a 51Cr release assay at 2, 5, 10, or 15 days after the MCA injection. A dose of 20 micrograms MCA to neonatally olive oil-injected females did not influence the NK activity, whereas an injection of 100 micrograms MCA significantly depressed the NK activity to about half the value seen in controls. This suppression was transient, and the normal level was reached again 15 days after the injection. The depressed NK activity could not be related to humoral or cellular suppressor mechanisms. A study at the single-cell level revealed that the inhibition was due to interference with the lytic step of the NK cell without affecting target-binding capacity. In vitro exposure of spleen cells from MCA-treated animals to interferon fully restored the NK activity. Neonatal DES treatment resulted in a depressed NK activity in adult females to a level about half of that seen in olive oil-injected controls. The NK activity in DES-treated females was not influenced by either 20 or 100 micrograms MCA. The MCA-induced suppression of NK activity was discussed in relation to the earlier reported difference in incidence of MCA-induced sarcomas between DES-treated and control females after they were given 20 micrograms MCA in adult life, as well as in relation to the same incidence in the 2 groups treated with 100 micrograms MCA. The results are compatible with a significant role for NK cells during MCA carcinogenesis and indicate that a possible part of the tumorigenic effect of MCA is its early suppressive effect on NK cell activity.

Ovarian influence on mitogen responsiveness of lymphocytes from mice neonatally exposed to diethylstilbestrol.

Treatment of neonatal female mice with diethylstilbestrol (DES) resulted in a reduced response of spleen lymphocytes to concanavalin A (Con A) and lipopolysaccharide (LPS). Ovariectomy or ovariectomy plus estradiol treatment had no influence on the response to Con A in either control or DES-treated females. Ovariectomy reduced the response to LPS in control animals and eliminated the difference between control and DES-treated females. Estradiol treatment enhanced the response of ovariectomized control females to LPS and restored the difference between control and DES-treated animals. Comparison of the mitogen responsiveness during various stages of the estrous cycle showed a tendency for the response to be augmented at proestrus and metestrus, although the differences were not significant.

Neonatal estrogen treatment alters sexual differentiation of hepatic histidase.

Hepatic histidase was used as an enzyme marker for the study of neonatal programming in intact rats. Diethylstilbestrol (DES) or 17 beta-estradiol (E2) treatment for days, 2, 4, and 6 post partum resulted in decreased histidase activities in the adult female, but no effect was seen in prepubertal male and female rats or in adult males. In contrast, similar neonatal doses of testosterone propionate (TP) had no effect on histidase. Dose-response experiments demonstrate a 3-fold greater neonatal sensitivity to DES than to E2. The action of neonatal estrogen treatment is demonstrated to be permanent and irreversible. Neonatal treatment with E2, DES, or TP resulted in decreased uterine wet weights in adult females (E2 less than DES less than TP less than controls). Circulating sera estrogen levels were lower in adult E2- and DES-treated females than in TP-treated and control females. Our results suggest that these alterations may be due to direct toxic effects on the postnatal development of the female reproductive tract and endocrine system and/or to organizational effects on nerve endings in the hypothalamus that result in programming for altered sexual differentiation of hepatic metabolism.