Dysregulation of IGF-I signaling in uterine leiomyoma.

IGF-I expression has been observed in human uterine leiomyomas. To examine whether autocrine IGF-I signaling plays a role in the growth of these tumors, we used an animal model of uterine leiomyoma (the Eker rat) to investigate regulation of IGF-I and the IGF-I receptor (IGF-IR) expression in tumors and normal myometrium. During the normal estrous cycle, myometrial IGF-I expression peaked on the day of proestrus when the rate of proliferation in this tissue is greatest. In leiomyomas, the expression of IGF-I was increased 7.5-fold compared with the age-matched normal tissue. The level of IGF-IR mRNA in both tumor and non-tumor tissues was found to inversely correlate with that of IGF-I. Changes observed in IGF-I signaling components correlated with the activation state of the signal-transducing protein insulin receptor substrate-1 (IRS-1). During diestrus and proestrus when IGF-I levels were increasing, tyrosine phosphorylation of IRS-1 was increased up to 5.7-fold in the normal myometrium relative to estrus, when IGF-I levels were the lowest. Additionally, IRS-1 phosphorylation was 4-fold greater in leiomyomas relative to age-matched normal myometrium. Autocrine stimulation of the IGF-IR may, therefore, play a role in regulating the normal growth of the myometrium, and dysregulation of IGF-I signaling could contribute to the neoplastic growth of uterine leiomyomas.

Potential dietary exposure to p-nonylphenol from food-contact use of tris(nonylphenyl)phosphite (TNPP).

The potential exposure to p-nonylphenol from the use of tris(nonylphenyl)phosphite (TNPP) in food-contact materials was calculated. The calculation was made on the basis of migration data from moulded plaques prepared from linear low-density polyethylene (LLDPE), films prepared from LLDPE and polyvinyl chloride (PVC) films. The data were obtained using the customary expermental procedures developed by the US Food and Drug Administration (FDA). The potential migration of p-nonylphenol to food from the use of TNPP as a component of food-contact articles was determined using food-simulating solvents and time and temperature conditions recommended by the FDA. Using the data obtained from these studies, along with procedures based on the FDA's conventional method for estimating potential dietary exposure using food simulating migration data, the potential dietary exposure to p-nonylphenol from the use of TNPP was determined to be approximately 25.5 parts per billion (ppb). Because the conditions of the migration tests exaggerate actual use conditions, this value overstates the potential dietary exposure.

Binge drinking, sensible drinking, and abstinence after alcohol-related vehicular crashes: the role of intervention versus screening.

Brief interventions have been shown to reduce problem drinking in a variety of populations and settings. The hypothesis for our randomized trial was that individuals injured in alcohol-related crashes who received a more intensive intervention (brief counseling) would have reduced binge drinking as compared to those with a less intensive intervention (simple advice) and controls. Non-alcohol dependent, seriously injured individuals (N = 186) were enrolled in the protocol. At baseline, mean binges/month (b/m) were 5.88 and at 12 months were 2.02 b/m. Although there was no significant difference by condition, at 12 months the brief counseling group had the lowest rate of binge drinking (1.97 b/m). Whether these drinking patterns were a result of the crash, injury, screening for alcohol use, or combination of these factors is difficult to determine.

Validity of self-reported alcohol consumption in nondependent drinkers with unintentional injuries.

BACKGROUND: Self-report has become an anchor for alcohol assessment in the acute and primary care populations. The purpose of the study was to determine the validity of self-reported alcohol consumption after unintentional injuries in hospitalized, nondependent drinkers. METHODS: Non-alcohol-dependent subjects 18 years of age and older with unintentional injuries (n = 209) were enrolled in the study and were interviewed if they had either an admitting blood alcohol concentration (BAC) > or = 10 mg/dl (0.01 g/dl) or a positive screen for a history of problem drinking. The self-reported number of standard drinks, time that drinking commenced, sex, and weight were used to calculate estimated blood alcohol concentration (EBAC), which was then compared to the admission BAC. RESULTS: We had data to calculate the EBAC on 141 of the 209 subjects. Seven men and no women with positive (> or = 10 mg/dl) BAC denied drinking. Of the 134 subjects for whom we had data to calculate EBAC and who acknowledged drinking, mean BAC was 147.06 mg/dl and mean EBAC was 68.66 mg/dl. For women (n = 30), mean BAC was 149.53 mg/dl and mean EBAC was 114.67 mg/dl; for men (n = 104), mean BAC was 146.35 mg/dl and mean EBAC was 55.38 mg/dl. The Spearman's p between laboratory BAC and EBAC was 0.461 (p < 0.001) for all subjects (n = 134), 0.275 (NS) for women (n = 30), and 0.532 (p < 0.001) for men (n = 104). For women and men separately, multiple regression analyses were performed to predict BAC by using weight and reported number of drinks. For women, weight and number of drinks accounted for 3% of the variance in laboratory BAC [r = 0.181, F(2,47) = 0.797,p = NS]. In contrast, for men these same predictors accounted for 34% of the variance [r = 0.585, F(2,135) = 35.203,p < 0.001). CONCLUSIONS: Most nondependent patients with unintentional injury acknowledged drinking before injury. After injury, women and men have different patterns of reporting their drinking, with men more frequently underreporting but reporting more accurately and women more random in their self-reports.

Attribution of injury to alcohol involvement in young adults seriously injured in alcohol-related motor vehicle crashes.

BACKGROUND: Approximately 40% of all traffic fatalities are associated with the use of alcohol. Hospitalization for serious injury after a motor vehicle crash related to use of alcohol may be an opportunity to change drinking behaviors in non-alcohol-dependent drinkers, thereby reducing the risk for future disability and death. OBJECTIVES: To determine the degree to which non-alcohol-dependent adults aged 18 to 45 years with alcohol-related vehicular trauma attributed their injury to use of alcohol. METHODS: During hospitalization, 132 subjects involved in alcohol-related motor vehicle crashes were interviewed. The interviews included the question, "To what extent do you believe your alcohol consumption was responsible for this injury?" Responses were measured on a 7-point scale ranging from 1 (not at all) to 7 (totally). RESULTS: In response to the question about attribution of injury to alcohol, 37.8% of subjects responded "not at all," 24.3 responded "somewhat," and 37.9% responded "mostly" or "totally." Spearman rank correlation between attribution of injury to alcohol involvement and blood alcohol content at admission was r = 0.440 (P < .001). CONCLUSIONS: More than 60% of patients injured in alcohol-related motor vehicle crashes attributed their injury partly or totally to use of alcohol. When alcohol-free, hospitalized patients with higher blood levels of alcohol on admission were more likely than those with lower levels to attribute their injury to alcohol. Hospitalization for a motor vehicle crash related to use of alcohol provides an opportunity for interventions to decrease drinking.

Potential exposure to bisphenol A from food-contact use of polycarbonate resins.

The potential exposure to bisphenol A (BPA) from the use of consumer products or packages made from bisphenol A-derived polycarbonate resins was calculated. The calculation was made on the basis of migration data from moulded discs prepared from a composite of polycarbonate resins. The data were obtained using the customary experimental procedures developed by the United States Food and Drug Administration (FDA). The potential migration of BPA to food from its contact with articles made from polycarbonate resins was determined using food-simulating solvents and time and temperature conditions recommended by FDA. The study demonstrates that no detectable BPA was found in the extracts obtained under FDA's most severe default testing condition using a method sensitive to 5 parts per billion (ppb) in the food simulants. Using these data, along with FDA's conventional procedure for estimating potential dietary exposure using food simulating migration data, the potential dietary exposure to bisphenol A from use of polycarbonate resins was determined to be less than 0.25 ppb.

Regulation of apoptosis in uterine leiomyomata.

Tumors developing from hormone-dependent tissues, such as the breast and prostate, have been successfully treated in the clinic by methods of hormone ablation, and the resulting tumor regression has been shown to occur at least in part by the process of apoptosis. The growth of leiomyomas arising from uterine smooth muscle cells is similarly modulated by circulating steroid hormones and has been associated with periods of increased estrogen secretion. The inhibition of ovarian hormone production by endocrine therapy often results in the regression of these tumors, but the role of apoptosis in this process has not been elucidated. Using cell lines derived from the Eker rat model of uterine leiomyoma, we have investigated the mechanism of growth inhibition by estrogen deprivation. Estrogen-depleted medium and the antiestrogen tamoxifen significantly reduced cell numbers in culture and arrested cell proliferation, but did not induce apoptosis. However, the presence of an intact apoptotic pathway was demonstrated in these cells by serum starvation. In vivo data were in agreement with in vitro results, which showed that tamoxifen treatment does not change the apoptotic rate of leiomyoma tissues. Therefore, growth modulation of leiomyomas by hormone deprivation occurs via mechanisms independent of apoptosis, indicating a fundamental difference in the response of leiomyomas to hormone deprivation from that of tumors of the breast and prostate. These data suggest that creation of a hypoestrogenic milieu within leiomyomas reduces tumor volume without inducing a concomitant increase in the rate of apoptosis, which may be responsible for the limited effectiveness of currently available medicinal therapies.

Presence of an insulin-like growth factor I autocrine loop predicts uterine fibroid responsiveness to tamoxifen.

Uterine leiomyoma is an estrogen-responsive tumor, and the present studies examine the ability of the antiestrogen tamoxifen to modulate leiomyoma cell growth. Tamoxifen is an effective form of hormonal therapy for breast cancer, although the mechanism by which tamoxifen inhibits tumor growth is not well understood and may involve mechanisms other than the action of tamoxifen as an estrogen antagonist. Tamoxifen was found to inhibit the proliferation of three of five leiomyoma-derived cell lines (ELT cell lines) in vitro, including an estrogen receptor-negative cell line. The ability of tamoxifen to decrease leiomyoma growth was found to correlate with expression of insulin-like growth factor I (IGF-I) by the tumor cells, suggesting that the inhibitory effects of tamoxifen were associated with expression of this growth factor. The existence of an IGF-I autocrine loop in the cells was investigated, because transcripts for both IGF-I and its cognate receptor were expressed in the tamoxifen-responsive cell lines. An IGF-I RIA demonstrated secreted IGF-I protein in serum-free medium conditioned by the IGF-I-expressing cell line ELT 3, and this same medium supported the growth of IGF-requiring MCF-10A cells, indicating the presence of biologically active IGF-I in the conditioned medium. Exogenous IGF-I stimulated ELT 3 cell proliferation, confirming that this growth factor is mitogenic for leiomyoma cells. IGF-I neutralizing antibody inhibited ELT 3 growth, indicating that the levels of IGF-I produced by the leiomyoma cells were physiologically significant. These data demonstrate the existence of an IGF-I autocrine loop in tamoxifen-sensitive leiomyoma cells, supporting the hypothesis that the presence of an IGF-I autocrine loop predicts uterine fibroid responsiveness to tamoxifen.

Data analytic techniques for treatment outcome studies with pretest/posttest measurements: an extensive primer.

This paper discusses several data analytic technique, for examining treatment efficacy in pretest-posttest control group designs. The following approaches are described: ANOVA on post scores, ANOVA on difference scores, split-plot repeated measures ANOVA, profile analysis, and ANOCOVA with prescore as the co-variate. Guidelines for choosing between available techniques are provided; the primary focus here is on the nature of the null hypothesis, the assumptions underlying the approach, and the power of the procedure. The importance of examining the characteristics of the data set in selecting an analytic technique is illustrated.

Estrogen stimulation and tamoxifen inhibition of leiomyoma cell growth in vitro and in vivo.

Uterine leiomyomas (fibroids) are the most common gynecological neoplasms and may be associated with significant morbidity. Recently, we described a rat model (Eker rat) of fibroid development in which reproductive tract leiomyomas develop spontaneously with high frequency. The present studies describe the estrogen and antiestrogen responsiveness of an Eker rat leiomyoma-derived cell line in vitro and a nude mouse xenograft system in vivo. In this cell line, estradiol stimulated growth in estrogen-depleted medium, whereas the nonsteroidal antiestrogen tamoxifen maximally inhibited cell proliferation in medium containing 10% charcoal-stripped serum. Proliferation was also decreased by the biologically active tamoxifen metabolite 4-hydroxytamoxifen; the metabolite was more effective than the parent compound in exerting this growth inhibition. Compared to placebo-treated controls, estradiol increased the size of tumors that developed in a nude mouse xenograft system, whereas tamoxifen increased tumor latency and decreased tumor size. This study of leiomyoma cells in a well defined system suggests that antiestrogens may prove efficacious in the treatment of this clinically important neoplasm.

Induction and subcellular localization of two major stress proteins in response to copper in the fathead minnow Pimephales promelas.

In the present study we characterize the stress response induced by copper in the fathead minnow, Pimephales promelas. The fathead minnow epithelial cell line ATCC CCL 42 was used to examine the induced synthesis and subcellular localization of the two major stress proteins, stress 70 and cpn60. Western blot analysis demonstrated increased stress70 in cells exposed to 400 and 500 microM Cu. Two-dimensional analysis revealed three isoforms of stress70, one of 70 kDa and two of 72 kDa, at the highest Cu concentration. Chaperonin60 abundance did not change over the same range of Cu concentrations. Indirect immunofluorescence microscopy revealed that stress70 localized in the cytoplasm, particularly in the paranuclear region. Chaperonin60 was localized in mitochondria. Further, when we examined the stress response elicited by Cu in fathead minnow larvae in vivo, we found that Cu induced the stress response at nominal Cu concentrations that were more than an order of magnitude lower that in the cell culture. This disparity between the concentration of Cu, which induced the stress response in cells in culture and in vivo, may be the result of differences in Cu complexation that alter its availability, uptake and toxicity.

Rodent model of reproductive tract leiomyomata. Clinical and pathological features.

Mesenchymal tumors of the lower reproductive tract of women are poorly understood at the molecular level as a result in part of the lack of relevant animal models. The present study describes a novel model of gynecological smooth muscle tumors in which these neoplasms arise in Eker rats as part of a familial cancer syndrome. The tumors develop as a result of a germline mutation in the tuberous sclerosis 2 (TSC2) gene, and predisposition to tumor development is inherited in an autosomal dominant fashion. Uterine and/or cervical tumors arise spontaneously as single or multicentric neoplasms and increase in incidence with increasing age. The tumors were classified into three phenotypic variants of leiomyoma/leiomyosarcoma and into stromal cervicovaginal tumors on the basis of cytological and histological features and immunostaining patterns for smooth muscle actin and desmin. Tumors histologically identical to the typical human myometrial leiomyoma arose, as did a subset of atypical leiomyomas having an epithelioid phenotype. Eker rats were found to develop both benign and malignant smooth muscle tumors. The high spontaneous incidence of smooth muscle tumors of uterus and cervix in this rodent model provides a unique opportunity to study the molecular mechanisms underlying the development of these clinically important gynecological neoplasms.

Rodent model of reproductive tract leiomyomata. Establishment and characterization of tumor-derived cell lines.

Uterine myometrial tumors are the most commonly found gynecological neoplasm in women. The underlying causes of uterine leiomyomata are poorly understood, a result in part of the absence of a good animal model system in which to study these tumors. This report describes a novel rat model (Eker rat) in which spontaneous gynecological smooth muscle tumors arise with a high frequency. Leiomyomas are the predominant reproductive tract tumor that arise in these animals, although leiomyosarcomas have also been observed. Cell lines have been established from both the benign and malignant lesions. All of the lines express smooth muscle-specific actin, and leiomyoma-derived cell lines express desmin. Two of the cell lines are tumorigenic in nude mice, and the lines are variable for expression of estrogen and progesterone receptors. These lines are the first rodent tumor-derived lines to be established from leiomyomata and are the only lines available from a hereditary form of these tumors. Together with Eker rats that spontaneously develop leiomyomata, they constitute an in vitro/in vivo model system for gaining insights into the mechanism of transformation of uterine smooth muscle cells and the role of steroid hormones and hormone receptors in myometrial tumorigenesis.

Tissue-specific differences in accumulation of stress proteins in Mytilus edulis exposed to a range of copper concentrations.

This study examines the expression and accumulation of two major stress proteins, stress70 and chaperonin60 (cpn60), in the gill and mantle of blue mussels, Mytilus edulis, which were exposed to a range of Cu concentrations for 7 days. Scope-for-growth (SFG), mortality, and Cu accumulation in gill and mantle tissue were also measured to monitor the physiological effects of Cu exposure in the organisms. In general Cu accumulated to a greater extent in gill relative to mantle tissue. A reduction of SFG index and increased mortality was also observed at the two highest Cu concentrations. We found no significant differences between the two tissues in the expression of cpn60 and stress70 for mussels exposed to Cu ranging from 0 to 10 micrograms/liter Cu (cpn60) and 0 to 32 micrograms/liter Cu (stress70) in sea-water. However, differences in the stress response were observed between the gill and the mantle tissue of mussels exposed to higher Cu concentrations. Chaperonin concentrations were greater than an order of magnitude higher in the gill than in the mantle for these mussels. Further, although the accumulation of stress70 was similar between the two tissues, two additional proteins reacted with antibody to stress70 in gill, but not mantle tissue, of mussels exposed to 100 micrograms/litter Cu. This study suggests that the physiological processes involved in contaminant uptake, distribution, and detoxification may affect the tissue-level expression of the stress response in multicellular organisms. Further, the intensity of the stress response and relative concentrations of chaperonin and stress70 among tissues may help identify tissues which are the most vulnerable to damage caused by a particular environmental stressor.