Perinatal estrogen exposure: later repercussion on the fertility of rats

The aim of the present study was to investigate the effects of perinatal estrogen exposure in the fertility of rats. Thus, rats were treated with estrogen on the 21st or 22nd day of intra-uterine life or treated with estrogen immediately after birth. It was observed that the testicular descent of males and beginning of puberty of females were advanced in all estrogen-treated groups. The females from estrogen-treated groups showed reduced frequency of estrous in 15 consecutive days of study, and there was an increase in estrous duration. Their fertility also were impaired and a reduction in the number of alive fetuses, as well as enhancement of pre- and postimplantation loss, mainly in the group treated with estrogen on the 21st day of intra-uterine life. However, the alterations observed in the fertility of estrogen-treated male rats were slighter and only females mated with male rats from the group treated with estrogen immediately after birth showed enhanced preimplantation loss. We suggest that the reproductive function is impaired by exposure to estrogen in the perinatal life of rats, and that the mechanisms involved in this effect are distinct for males and females.     

Bcl-2 is overexpressed and alters the threshold for apoptosis in a cholangiocarcinoma cell line

Cholangiocarcinoma is a malignant neoplasm originating from cholangiocytes. The mechanisms responsible for oncogenesis of cholangiocytes are unknown. Resistance to apoptosis, especially by altered expression of B-cell lymphoma/leukemia 2 (Bcl-2) family members, has been implicated as a mechanism contributing to malignant transformation. Thus, our aim was to test the hypothesis that altered expression of Bcl-2 family members by cholangiocarcinoma cells renders them resistant to apoptosis. We compared the apoptotic threshold and expression of the Bcl-2 protein family members, Bcl-2, Bcl-XL, and Bax, in two human cell lines: 1) nonmalignant human cholangiocytes immortalized by transfection with the simian virus 40 (SV 40) large T antigen; and 2) a malignant human cholangiocarcinoma cell line. Apoptosis was induced pharmacologically using beauvericin. Bcl-2, Bcl-x long, and Bax protein expression were evaluated by immunoblot analysis, and Bcl-2 expression was modulated using antisense technology. The cholangiocyte and malignant/nonmaligant phenotype of both cell lines was verified using both in vitro and in vivo approaches. Beauvericin induced apoptosis of nonmalignant cholangiocytes in a concentration- (0 to 25 micromol/L) and time- (0 to 6 hours) dependent manner. In contrast, malignant cholangiocytes were resistant to apoptosis. Although expression of Bcl-x long and Bax protein were similiar in the two cell lines, Bcl-2 protein expression was 15-fold greater in malignant than in nonmalignant cholangiocytes. An 18 mer bcl-2 antisense oligonucleotide reduced expression of Bcl-2 protein by 50% and increased the rate of beauvericin-induced apoptosis more than threefold in the malignant cells. Our results support the hypothesis that resistance to apoptosis by overexpression of Bcl-2 may be a feature of cholangiocarcinoma.     

Hypoxic activation of nuclear factor-kappa B is mediated by a Ras and Raf signaling pathway and does not involve MAP kinase (ERK1 or ERK2)

We have previously shown that hypoxia causes the activation of nuclear factor-kappa B (NF-kappa B), and the phosphorylation of its inhibitory subunit, I kappa B alpha, on tyrosine residues. With the use of dominant negative mutants of Ha-Ras and Raf-1, we investigated some of the early signaling events leading to the activation of NF-kappa B by hypoxia. Both dominant negative alleles of Ha-Ras and Raf-1 inhibited NF-kappa B induction by hypoxia, suggesting that the hypoxia-induced pathway of NF-kappa B induction is dependent on Ras and Raf-1 kinase activity. Furthermore, although conditions of low oxygen can also activate mitogen-activated protein kinases (ERK1 and ERK2), these kinases do not appear to be involved in regulating NF-kappa B by low oxygen conditions, as dominant negative mutants of mitogen-activated protein kinase do not inhibit NF-kappa B activation by hypoxia. Since Ras and Raf-1 have been previously shown to work downstream from membrane-associated tyrosine kinases such as Src, we determined if the Src membrane-associated kinase was also activated by low oxygen conditions. We detected an increase in Src proto-oncogene activity within 15-30 min of cellular exposure to hypoxia. We postulate that Src activation by hypoxia may be one of the earliest events that precedes Ras activation in the signaling cascade which ultimately leads to the phosphorylation and dissociation of the inhibitory subunit of NF-kappa B, I kappa B alpha.     

Socioeconomic status differences in hormone therapy

Socioeconomic status (SES) is a significant sociodemographic correlate of noncontraceptive hormone therapy, yet multiple dimensions of SES have not been examined systematically in previous studies of hormone therapy. This study examined the lifetime incidence of noncontraceptive hormone therapy, how usage varied by type of reproductive organ surgery, and the bivariate and net associations between a large array of SES indicators and the likelihood of ever using hormones by age 53-54 years in a population sample (n = 3,612) of non-Hispanic white female participants in the Wisconsin Longitudinal Study (1957-1993). Approximately half of the women had ever used noncontraceptive hormones; 38.5% were current users. In multivariate logistic regression analyses, the most robust SES predictor of hormone therapy was a woman's husband's occupational status (higher status associated with higher rates of use), after adjustment for all other measured sociologic and biomedical factors (e.g., other SES measures, other health behaviors, menopausal symptoms, age at menopause, health insurance). The association of hormone therapy with education differed between women who underwent hysterectomy and/or oophorectomy (higher odds for less educated women) and those with intact reproductive organs (lower odds for less educated women). Additionally, a woman's own earnings and household net worth were positive net correlates of noncontraceptive hormone therapy.     

Regulatory roles of complexins in neurotransmitter release from mature presynaptic nerve terminals

Complexins are presynaptic proteins whose functional roles in synaptic transmission are still unclear. In cultured rat hippocampal neurons, complexins are distributed throughout the cell bodies, dendrites and axons, whereas synaptotagmin I and synaptobrevin/VAMP-2, essential proteins for neurotransmitter release, accumulated in the synaptic-releasing sites as early as 1 week in culture. With a maturation of synapses in vitro, complexins also accumulated in the synaptic release sites and co-localized with synaptotagmin I and synaptobrevin/VAMP-2 after 3-4 weeks in culture. Complexins I and II were expressed in more than 90 and 70% of the cultured neurons, respectively; however, they were largely distributed in different populations of synaptic terminals. In the developing rat brain, complexins were distributed in neuronal cell bodies in the early stage of postnatal development, but gradually accumulated in the synapse-enriched regions with development. In mature presynaptic neurons of Aplysia buccal ganglia, injection of anticomplexin II antibody caused a stimulation of neurotransmitter release. Injection of recombinant complexin II and alphaSNAP caused depression and facilitation of neurotransmitter release from nerve terminals, respectively. The effect of complexin was reversed by a subsequent injection of recombinant alphaSNAP, and vice versa. These results suggest that complexins are not essential but have some regulatory roles in neurotransmitter release from presynaptic terminals of mature neurons.     

Determination of stability of Brucella abortus RB51 by use of genomic fingerprint, oxidative metabolism, and colonial morphology and differentiation of strain RB51 from B. abortus isolates from bison and elk

Brucella abortus RB51 and isolates from cattle, bison, and elk were characterized by pulsed-field gel electrophoresis and standard techniques for biotyping Brucella species, which included biochemical, morphological, and antigenic techniques, phage susceptibility, and antibiotic resistance. The objectives were to ascertain the stability of RB51 and to differentiate RB51 from other brucellae. Genomic restriction endonuclease patterns produced by pulsed-field gel electrophoresis demonstrated a unique fingerprint for RB51 relative to other brucellae. Comparisons of the oxidative metabolic profiles of RB51 after time in vivo (14 weeks) and in vitro (75 passages) showed no change in characteristic patterns of oxygen uptake on selected amino acid and carbohydrate substrates. Strain RB51 was biotyped as a typical rough B. abortus biovar 1 (not strain 19) after animal passage or a high number of passages in vitro and remained resistant to rifampin or penicillin and susceptible to tetracycline. No reactions with A or M antiserum or with a monoclonal antibody to the O antigen of Brucella lipopolysaccharides were detected; however, RB51 agglutinated with R antiserum. The results indicate that the genomic fingerprint and rough colonial morphology of RB51 are stable characteristics and can be used to differentiate this vaccine strain from Brucella isolates from cattle, bison, and elk.     

Morphometric and histopathologic analysis of lymphoid depletion in murine spleens following infection with Brucella abortus strains 2308 or RB51 or an htrA deletion mutant

A physician education program altered the consumption of hospital resources by orthopedic surgeons doing total hip replacement. They were presented with verbal and written physician-specific materials during a 7-week period. Xbar and R charts were used to display measurements of resource consumption before and after the educational program. Physicians were provided with surgeon-specific data profiling their own practice and that of their peers. Inferential statistics were used to validate the effects of education. Length of hospital stay for patients receiving total hip replacement dropped from 13.7 days to 9.9 days. Adjusted total charges dropped from $22,103 to $18,607. The variance for length of stay and total charges dropped by one half or more. Physician education programs are effective in reducing consumption of hospital resources. Statistical process control data formatting appears to make the data more usable to these physicians.     

A mutation causing pseudohypoaldosteronism type 1 identifies a conserved glycine that is involved in the gating of the epithelial sodium channel

Pseudohypoaldosteronism type 1 (PHA-1) is an inherited disease characterized by severe neonatal salt-wasting and caused by mutations in subunits of the amiloride-sensitive epithelial sodium channel (ENaC). A missense mutation (G37S) of the human ENaC beta subunit that causes loss of ENaC function and PHA-1 replaces a glycine that is conserved in the N-terminus of all members of the ENaC gene family. We now report an investigation of the mechanism of channel inactivation by this mutation. Homologous mutations, introduced into alpha, beta or gamma subunits, all significantly reduce macroscopic sodium channel currents recorded in Xenopus laevis oocytes. Quantitative determination of the number of channel molecules present at the cell surface showed no significant differences in surface expression of mutant compared with wild-type channels. Single channel conductances and ion selectivities of the mutant channels were identical to that of wild-type. These results suggest that the decrease in macroscopic Na currents is due to a decrease in channel open probability (P(o)), suggesting that mutations of a conserved glycine in the N-terminus of ENaC subunits change ENaC channel gating, which would explain the disease pathophysiology. Single channel recordings of channels containing the mutant alpha subunit (alphaG95S) directly demonstrate a striking reduction in P(o). We propose that this mutation favors a gating mode characterized by short-open and long-closed times. We suggest that determination of the gating mode of ENaC is a key regulator of channel activity.