Effect of neonatal androgenization on positive feedback in female mice

Exposure of female mice to androgens within 5 days of birth impairs fertility. Such treatment in rats results in a post-pubertal acyclic state of persistent vaginal cornification and in an inability, when ovariectomized, to show normal positive feedback on luteinizing hormone (LH) release in response to steroid challenge. In the present study, we explored whether neonatally androgenized mice demonstrate positive feedback. Female mice were administered 100 micrograms of testosterone propionate (TP) on either Day 1 (TP1) or Day 5 (TP5) after birth, or vehicle on Day 1 (SO1). Androgen-treated mice had a statistically significant advance in onset of vaginal opening as compared with vehicle-treated mice. All mice that received TP entered constant vaginal estrus, whereas those given vehicle showed variable cytology. All mice were ovariectomized at 7 wk of age and received Silastic capsules containing a priming dose of 17 beta-estradiol. When all mice were challenged 1 wk later with sequential administration of estradiol benzoate and progesterone, a significant increase in plasma LH level was present only in the vehicle-treated mice. We conclude that neonatal androgenization defeminizes the neuroendocrine mechanisms controlling gonadotropin release.     

Effect of neonatal androgenization on uterine sarcomagenesis induced by 1,2-dimethylhydrazine in female CBA mice

A single injection of testosterone propionate to newborn female CBA mice provoked earlier puberty and formation of permanent estrous in the majority of animals. This led to an abrupt reduction of the latent period and the increased incidence of 1,2-dimethylhydrazine-induced uterine sarcomas (90 versus 9% upon DMH injection to intact mice).     

Modifying action of neonatal androgenization on 1,2-dimethylhydrazine-induced carcinogenesis in male CBA-strain mice

Newborn male CBA mice received a single treatment with 0.5 mg testosterone propionate. Weekly subcutaneous injections of 1,2-dimethylhydrazine (DMH) were given to 2-month-old mice. The incidence of pararenal angiosarcomas and colonic tumors in neonatally androgenized mice reached 78.5 and 71.0%, respectively by the 35th week after the DMH treatment was commenced. In DMH-treated control mice, the incidence of the above tumors amounted to 25 and 32%, respectively.     

Inhibition by dehydroepiandrosterone of aggression responses of spayed female mice towards lactating intruders is potentiated by neonatal androgenization

Female mice of an inbred Swiss strain were injected subcutaneously (s.c.) with either testosterone propionate or oil-vehicle on the day of birth. Once adult (8 week-old), they were spayed, and then treated either with dehydroepiandrosterone (80 micrograms/day s.c. for 15 days) or with solvent. The decrease of biting attacks by dehydroepiandrosterone was definitely more prominent in neonatally imprinted females than in controls.     

Reversal of reproductive deficiency in the hpg male mouse by neonatal androgenization.

Some aspects of reproductive function in the GnRH-deficient hypogonadal (hpg) mutant mouse can be restored by transplanting normal fetal brain tissue containing GnRH cells into the central nervous system of adult hpg mice. However, hpg males showing physiological response to the graft fail to display sexual behavior and are infertile. We hypothesized that the reproductive deficit of these males is due to insufficient perinatal exposure to testicular androgens as a consequence of the GnRH deficiency. To test this hypothesis we androgenized hpg males by giving them neonatal injections of testosterone propionate (TP). Controls consisted of hpg males not androgenized neonatally and of normal males. All three groups received a TP implant in adulthood, and their copulatory behavior and reproductive capability were recorded. In addition, other hpg males, not androgenized neonatally, received fetal brain transplants containing GnRH neurons and were also tested for copulatory behavior and reproductive capability before and after receiving a TP implant. Three of 8 neonatally androgenized hpg males expressed the full repertoire of male sexual behavior, including intromission and ejaculation, and sired several litters. Three of 7 control hpg males that were not androgenized neonatally but received TP implants in adulthood also displayed mounting and intromission, but there was no evidence of ejaculation, and these males failed to impregnate normal females. Of the 8 hpg males that responded to a fetal transplant with testicular growth, only 1 displayed mounting behavior. However, when given a TP implant, 4 of 8 hpg males with grafts displayed mounting and intromissions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Monoclonal antibodies against human alpha-fetoprotein more reactive to cell-surface alpha-fetoprotein than to free alpha-fetoprotein.

This paper describes the finding of monoclonal antibody (MoAb) more reactive to cell-surface alpha-fetoprotein (AFP) than to free AFP by using a simple in vitro system. Twelve mouse MoAbs, ten IgG1, one IgG2a and one IgG2b, against human AFP from hepatocellular carcinoma were obtained by the cell fusion technique. Each hybridoma supernatant was assayed by enzyme-linked immunosorbent assay (ELISA) to solid-phase AFP. The assay results showed that two MoAbs, 67D and 80G, were most reactive to AFP. 80G had a higher affinity constant than 67D, while the both reactions were similarly difficult to inhibit by free AFP in ELISA. 67D and 80G reacted with AFP on the surface of ethanol-fixed cells from the human hepatoma cell line HuH-7 and this reaction was also difficult to inhibit by free AFP in Cell ELISA. Furthermore, Western blot analysis showed that 67D and 80G were more reactive to membrane-bound AFP than other antibodies. These findings first suggest that there could be anti-AFP MoAbs preferably binding to cell-surface AFP rather than to serum AFP.     

Dynamics of alpha-fetoprotein concentration in mice of different genotypes during the neonatal period]

A comparative assay of the alpha-fetoprotein (alpha-FP) level in mice of various genotypes (CBA, C3H, C57BL/Se/Sn, BALB/c, CC57W, AKR and nude--nu/nu) was conducted in the course of 3 weeks of postnatal period. The concentration of alpha-FP reached the following levels: the first day 2(-10)-2(-9); the 5th day 2(-8); the 8th day 2(-7); the 15th day 2(-4); on the 22nd day the level was zero. Nude mice which showed the alpha-FP concentration of 2(-2) on the 15th day were an exception. A conclusion was drawn that the alpha-FP synthesis was based not on the athymia of nude mice per se, but upon other unknown factors.     

Influence of neonatal androgenization on the testicular steroidogenesis in the adult rat

The in vitro testicular steroidogenesis of male rats, androgenized on the third postnatal day by a single injection of 1 mg testosterone propionate, was investigated when the animals were 100 days old. The neonatal androgenization resulted in a 25% lower testes weight, significantly increased plasma levels of FSH (P less than 0.01) and LH (P less than 0.02), and normal levels of testosterone. Although the testes were hypotrophic, the incubation of the testes pairs yielded the same amounts of testosterone, 7 alpha-hydroxytestosterone and 5 alpha-androstane-(3 alpha + 3 beta), 17 beta-diol as in the control animals. However, the steroidogenic response to an acute hCG stimulation was reduced. From incubations of testes homogenates with various labelled steroid precursors it could be inferred that the activity of the 17 alpha-hydroxylase, the 3 beta-hydroxysteroid dehydrogenase-isomerase and the 17 beta-hydroxysteroid dehydrogenase, expressed per unit of incubated protein, was significantly increased in the testes of the androgenized rats. These data indicate that the basal steroidogenesis in neonatally androgenized male rats is maintained by an increased synthesis per unit of tissue, possibly under influence of an increased gonadotrophic stimulus, but that the maximum steroidogenic capacity is reduced.     

The binding of 3H-labelled androgen-receptor complexes to hypothalamic chromatin of neonatal mice: effect of sex and androgenization

The binding of 3H-labelled androgen-receptor complexes, prepared by (NH4)2SO4 precipitation from the 105,000 g supernatant of hypothalamic cytosol, to hypothalamic chromatin of neonatal mice covalently coupled to cellulose was measured in vitro. Saturation binding was also determined after extraction of histones and the masking of acidic proteins with high molarities of guanidine hydrochloride. This investigation showed the presence of high-affinity, low-capacity acceptor sites for [3H]-testosterone-receptor complexes in male hypothalamic chromatin (Kd value = 0.39 x 10(-10) M and binding sites of 41 fmol per mg of DNA). Acceptor activity seems to be associated with the acidic protein fraction of chromatin. No specific acceptor sites of similar nature were found in chromatin taken from the hypothalami of female mice. On the basis of these results, it is suggested that the androgen-unresponsiveness of female mice is related to the absence of acceptors for the androgen-receptor in female mice hypothalami.     

Early androgenization and aggression pheromone in inbred mice

Three experiments were conducted to investigate the effects of early androgenization on the social interactions in mice. Results of Expt 1 indicated that neonatally testosterone propionate (TP) treated females, when mature, were attacked faster, more frequently, and for longer durations by trained fighters than females treated neonatally with oil. Experiment 2 showed that male castrates coated with urine from neonatally TP-treated females were attacked more often and for longer durations than castrates coated with urine from neonatally oil-treated females. The results implicated a urinary pathway of pheromonal output. Experiment 3 was conducted to investigate the relatively long-term social interactions between a naive male and a female treated at birth with TP or between a naive male and a female treated at birth with oil. The androgenized females showed a greater number of wounds after 5 days of pair housing than the control females. Five out of seven androgenized females were killed. Six control females gave birth, and none of the remaining two androgenized females did.     

The effects of neonatal androgenization of male rats on testosterone metabolism by the hypothalamus-pituitary-gonadal axis

Male rats were androgenized on the third postnatal day by a single injection of 1 mg testosterone propionate. The in vitro metabolism of [4-14C]testosterone by pituitary and hypothalamus homogenates was investigated at the age of 90 days. The pituitary and hypothalamus homogenates from control and neonatally androgenized animals converted [4-14C]testosterone to the same metabolites, mainly 5 alpha-reduced derivatives; the quantitative yield of 5 alpha-reduced metabolites was much higher in the pituitary homogenates of androgenized rats. The hypothalamic homogenates showed no differences. In the androgenized rats a very significant increase of the plasma FSH levels was measured while the LH levels were also augmented. The plasma levels of testosterone were not different from the values in control rats, notwithstanding a 25% reduction in testes weight. The present experiments appear to indicate that the neonatal androgenization results in an accentuation of the sexual dimorphism which normally exists in the pituitary of adult rats for the 5 alpha-reductase activity.     

Can neonatal beta-adrenergic stimulation prevent the effects of androgenization in female rats?

A 25 micrograms dose of testosterone propionate injected at 4 days of age induced 90% anovulation at 100 days of age. The systemic administration of orciprenaline (8 or 16 micrograms) or yohimbine (100 micrograms) did not prevent androgenization. Twenty-five or fifty micrograms of orciprenaline injected intraventricularly reduced only partially (to 54 and 67% respectively) the effectiveness of androgenization. We concluded that beta-adrenergic receptor stimulation had a very limited ability to prevent androgenization, since the beta-stimulation obtained directly with orciprenaline prevented androgenization to a very limited extent, while the possible indirect stimulation through an increase in norepinephrine endogenous release by alpha-2 receptor blocker yohimbine was ineffective.     

Rotavirus vp7 antigen produced by Lactococcus lactis induces neutralizing antibodies in mice

AIMS: To determine if live recombinant Lactococcus lactis strains expressing rotavirus VP7 antigen are immunogenic in mice. METHODS AND RESULTS: Using the food-grade lactic acid bacterium L. lactis as a carrier, we expressed VP7, the major rotavirus outer shell protein and one of the main components of the infective particle, as a cytoplasmic, secreted or cell wall anchored forms. Our results showed that recombinant L. lactis strains secreting VP7 proved to be more immunogenic than strains containing the antigen in the cytoplasm or anchored to the cell wall. CONCLUSIONS: This is the first demonstration that recombinant L. lactis producing VP7 can induce the production of a neutralizing antibody response against rotavirus by the intragastric route. SIGNIFICANCE AND IMPACT OF THE STUDY: Rotaviruses are the single most important aetiological agents of severe diarrhoea of infants and young children worldwide and have been estimated to be responsible for 650 000-800 000 deaths per year of children younger than 5 years old in development countries. Thus, the development of a safe and effective vaccine has been a global public health goal. Although two of five mice orally inoculated with L. lactis strains secreting VP7 elicited a specific-antibody response, these strains could be very useful to be used as a prototype to develop a new generation of protective rotavirus vaccines.     

B cell dependent T cell function blocked by perinatal exposure of mice to anti-IgM antibodies

Mice born to mothers deprived of B lymphocytes by their chronic treatment with anti-IgM antibodies (Su/N) do not possess naturally occurring anti-ids (present in sera of normal mice at 2 weeks of age) up to 10 weeks, despite the presence of normal levels of B cells and serum Ig (in these animals). Su/N mice of the same age also lack a T cell subset which together with anti-ids are thought to participate in an antisuppressor regulatory pathway. It is suggested that early development of these T cells may be linked and be dependent on the presence of these anti-ids synthesized early in ontogeny, providing one explanation for a selective T cell deficiency of B cell-deprived mice.     

Decreased and disproportionate T-cell population in adult mice after neonatal exposure to diethylstilbestrol

Female Balb/c mice neonatally treated with diethylstilbestrol show persistent impairment of several immunological parameters. The distribution of different classes of lymphocytes in the spleen has been determined at 6, 12, and 18 weeks of age. DES resulted in a decreased percentage of T cells in spleen while the number of B cells was normal. Utilizing Lyt antisera the T-cell subpopulations were found to be imbalanced with an increase in Lyt 123 cells and a concomitant decrease in Lyt 1 cells. Ovariectomy did not influence the diethylstilbestrol-induced alterations in the T-lymphocyte population.