Retrograde signaling from autophagy modulates stress responses.

Macroautophagy is a process in which cytoplasmic components, including whole organelles, are degraded within lysosomes. Basally, this process is essential for homeostasis and is constitutively functional in most cells, but it can also be implemented as part of stress responses. We discuss findings showing that autophagy proteins can modulate and amplify the activities of transcription factors involved in stress responses, such as those in the p53, FOXO, MiT/TFE, Nrf2, and NFκB/Rel families. Thus, transcription factors not only amplify stress responses and autophagy but are also subject to retrograde regulation by autophagy-related proteins. Physical interactions with autophagy-related proteins, competition for activating intermediates, and "signalphagy," which is the role autophagy plays in the degradation of specific signaling proteins, together provide powerful tools for implementing negative feedback or positive feed-forward loops on the transcription factors that regulate autophagy. We present examples illustrating how this network interacts to regulate metabolic and physiologic responses.

Adherence to Disease Modifying Drugs among Patients with Multiple Sclerosis in Germany: A Retrospective Cohort Study.

BACKGROUND: Long-term therapies such as disease modifying therapy for Multiple Sclerosis (MS) demand high levels of medication adherence in order to reach acceptable outcomes. The objective of this study was to describe adherence to four disease modifying drugs (DMDs) among statutorily insured patients within two years following treatment initiation. These drugs were interferon beta-1a i.m. (Avonex), interferon beta-1a s.c. (Rebif), interferon beta-1b s.c. (Betaferon) and glatiramer acetate s.c. (Copaxone). METHODS: This retrospective cohort study used pharmacy claims data from the data warehouse of the German Institute for Drug Use Evaluation (DAPI) from 2001 through 2009. New or renewed DMD prescriptions in the years 2002 to 2006 were identified and adherence was estimated during 730 days of follow-up by analyzing the medication possession ratio (MPR) as proxy for compliance and persistence defined as number of days from initiation of DMD therapy until discontinuation or interruption. FINDINGS: A total of 52,516 medication profiles or therapy cycles (11,891 Avonex, 14,060 Betaferon, 12,353 Copaxone and 14,212 Rebif) from 50,057 patients were included into the analysis. Among the 4 cohorts, no clinically relevant differences were found in available covariates. The Medication Possession Ratio (MPR) measured overall compliance, which was 39.9% with a threshold MPR≥0.8. There were small differences in the proportion of therapy cycles during which a patient was compliant for the following medications: Avonex (42.8%), Betaferon (40.6%), Rebif (39.2%), and Copaxone (37%). Overall persistence was 32.3% at the end of the 24 months observation period, i.e. during only one third of all included therapy cycles patients did not discontinue or interrupt DMD therapy. There were also small differences in the proportion of therapy cycles during which a patient was persistent as follows: Avonex (34.2%), Betaferon (33.4%), Rebif (31.7%) and Copaxone (29.8%). CONCLUSIONS: Two years after initiating MS-modifying therapy, only 30-40% of patients were adherent to DMDs.

ATG5: a distinct role in the nucleus.

Both apoptotic and autophagic pathways are activated in cells during anticancer treatment using DNA-damaging agents. Thus, the outcome is balanced between apoptotic cell death and enhanced autophagy, with the possibility of prolonged cell survival. It seems intuitively obvious that this survival mechanism might interfere with the desired tumor cell killing. We addressed this question by tipping the balance in favor of autophagy, using etoposide or cisplatin at low, sublethal doses. Over 4 days, only a little apoptosis was observed, but both drugs sharply increased autophagic flux. Surprisingly, cells underwent a cell cycle arrest at G 2/M, followed later by mitotic catastrophe with formation of multipolar spindles, missegregated chromosomes, or enlarged, irregular, sometimes multiple nuclei. Why? The answer is that even a low level of DNA damage not only upregulates autophagy, but also provokes the recruitment of an autophagy-related protein, ATG5, to the nucleus, where it binds BIRC5/survivin, thereby interfering with correct assembly of the chromosome passenger complex needed for cytokinesis.

ATG5 is induced by DNA-damaging agents and promotes mitotic catastrophe independent of autophagy.

Anticancer drug therapy activates both molecular cell death and autophagy pathways. Here we show that even sublethal concentrations of DNA-damaging drugs, such as etoposide and cisplatin, induce the expression of autophagy-related protein 5 (ATG5), which is both necessary and sufficient for the subsequent induction of mitotic catastrophe. We demonstrate that ATG5 translocates to the nucleus, where it physically interacts with survivin in response to DNA-damaging agents both in vitro and in carcinoma tissues obtained from patients who had undergone radiotherapy and/or chemotherapy. As a consequence, elements of the chromosomal passenger complex are displaced during mitosis, resulting in chromosome misalignment and segregation defects. Pharmacological inhibition of autophagy does not prevent ATG5-dependent mitotic catastrophe, but shifts the balance to an early caspase-dependent cell death. Our data suggest a dual role for ATG5 in response to drug-induced DNA damage, where it acts in two signalling pathways in two distinct cellular compartments, the cytosol and the nucleus.

Socioepidemiology of cigarette smoking among Cambodian Americans in Long Beach, California.

We examined the prevalence of cigarette smoking among Cambodian Americans in Long Beach, California. A stratified random sample of 1,414 adult respondents was selected from 15 census tracts with high concentrations of Cambodian Americans. The prevalence of current smokers was 13.0%; the sex-specific prevalence of smoking was 24.4% for men and 5.4% for women. The mean age of the sample was 50.5 years; about 60% of the respondents were women. The survey response rate was 90.5% among households in which respondents self-identified as Cambodian American, and at least one person completed the survey. Significant covariates of current smoking were gender, age, education, marital status, and health status. The prevalence of smoking among Cambodian men was higher than among other males in California. Culturally-tailored interventions should consider demographic characteristics of the target population as well as the high level of respect given to religion, elders, and peers.

Expression, localization, and functional model of cholesterol transporters in lactating and nonlactating mammary tissues of murine, bovine, and human origin.

Members of the ATP-binding cassette (ABC) transporters play a pivotal role in cellular lipid efflux. To identify candidate cholesterol transporters implicated in lipid homeostasis and mammary gland (MG) physiology, we compared expression and localization of ABCA1, ABCG1, and ABCA7 and their regulatory genes in mammary tissues of different species during the pregnancy-lactation cycle. Murine and bovine mammary glands (MGs) were investigated during different functional stages. The abundance of mRNAs was determined by quantitative RT-PCR. Furthermore, transporter proteins were localized in murine, bovine, and human MGs by immunohistochemistry. In the murine MG, ABCA1 mRNA abundance was elevated during nonlactating compared with lactating stages, whereas ABCA7 and ABCA1 mRNA profiles were not altered. In the bovine MG, ABCA1, ABCG1, and ABCA7 mRNAs abundances were increased during nonlactating stages compared with lactation. Furthermore, associations between mRNA levels of transporters and their regulatory genes LXRalpha, PPARgamma, and SREBPs were found. ABCA1, ABCG1, and ABCA7 proteins were localized in glandular MG epithelial cells (MEC) during lactation, whereas during nonlactating stages, depending on species, the proteins showed distinct localization patterns in MEC and adipocytes. Our results demonstrate that ABCA1, ABCG1, and ABCA7 are differentially expressed between lactation and nonlactating stages and in association with regulatory genes. Combined expression and localization data suggest that the selected cholesterol transporters are universal MG transporters involved in transport and storage of cholesterol and in lipid homeostasis of MEC. Because of the species-specific expression patterns of transporters in mammary tissue, mechanisms of cholesterol homeostasis seem to be differentially regulated between species.

Initiation to methamphetamine use in a binational sample of women at the US-Mexico border.

This article examines comparative risk behaviors associated with methamphetamine use in a binational sample of women in the border cities of Tijuana, Mexico, and San Diego, California. Specifically, the study examined the differences and similarities in drug use and sexual risk behavior and the patterns of initiation to methamphetamine use. The binational pilot sample consisted of 70 adult women in Tijuana and 55 women in San Diego. Although there were important differences in the presentation of risk behavior and patterns of initiation between the two binational samples, women on both sides of the US-Mexico border also showed remarkable similarities in their risk profile. Results from this study suggest that despite significant cultural and socioeconomic differences between the study cities, certain specific substance abuse patterns (e.g., methamphetamine use) in border regions with an increasing demographic exchange and integration are emerging as an "equalizer" of risk, capable of dissolving context-based differentiating factors, and creating a more homogenous subpopulation of substance users.

Treadmill training for patients with Parkinson's disease.

BACKGROUND: Treadmill training is used in rehabilitation and is described as improving gait parameters of patients with Parkinson's disease. OBJECTIVES: To assess the effectiveness of treadmill training in improving the gait function of patients with Parkinson's disease and the acceptability and safety of this type of therapy. SEARCH STRATEGY: We searched the Cochrane Movement Disorders Group Specialised Register (see Review Group details for more information) (last searched March 2009), Cochrane Central Register of Controlled Trials (The Cochrane Library 2009, Issue 2), MEDLINE (1950 to March 2009), and EMBASE (1980 to March 2009).We also handsearched relevant conference proceedings, searched trials and research registers, and checked reference lists (last searched March 2009). We contacted trialists, experts and researchers in the field and manufacturers of commercial devices. SELECTION CRITERIA: We included randomised controlled trials comparing treadmill training with no treadmill training in patients with Parkinson's disease. DATA COLLECTION AND ANALYSIS: Two review authors independently selected trials for inclusion, assessed trial quality and extracted data. We contacted the trialists for additional information. We analysed the results as standardised mean differences (SMDs) and mean differences (MDs) for continuous variables and relative risk differences (RD) for dichotomous variables. MAIN RESULTS: We included eight trials (203 participants) in this review. Treadmill training improved gait speed (SMD 0.50; 95% confidence interval (CI) 0.17 to 0.84; P = 0.003; I(2) = 0%) (fixed-effect model), stride length (SMD 0.42; 95% CI 0.00 to 0.84; P = 0.05; I(2) = 0%), walking distance (MD = 358 metres; 95% CI 289 to 426; P < 0.0001; I(2) = 30%), but cadence did not improve (MD 1.06; 95% CI -4.32 to 6.44; P = 0.70; I(2) = 0%) at the end of study. Treadmill training did not increase the risk of patients dropping out (RD -0.07; 95% CI -0.18 to 0.05; P = 0.26; I(2) = 51%) (random-effects model). Adverse events were not reported. AUTHORS' CONCLUSIONS: Patients with Parkinson's disease who receive treadmill training are more likely to improve their impaired gait hypokinesia. However, the results must be interpreted with caution because there were variations between the trials in patient characteristics, the duration and amount of training, and types of treatment. Additionally, it is not known how long these improvements may last.

Getting tougher for the fourth quarter: boomers and physical activity.

Baby Boomers might not consider themselves as growing old but are starting to reach the last quarter of average life spans. This article asks how Boomers prepare for their fourth quarters through physical activity. Three years (1999-2001) of National Health Interview Survey data yielded 96,501 adult respondents. Dependent variables were moderate, vigorous, and strengthening activity. Old boomers (1946-1955) and young boomers (1956-1965) were compared to respondents born before 1926, after 1975, and 10-year cohorts between. SUDAAN multiple logistic regression adjusted for complex sampling structure and multiply imputed income. Age-adjusted, older cohorts showed greater likelihood of activity than younger cohorts, offsetting moderate-activity declines with age until sharp decreases at advanced age: a plateau across Boomer and younger-aged cohorts. Interventions should promote activity at intensities and frequencies to which Boomers are most receptive.

SFRP-4 abrogates Wnt-3a-induced beta-catenin and Akt/PKB signalling and reverses a Wnt-3a-imposed inhibition of in vitro mammary differentiation.

BACKGROUND: Conserved Wnt ligands are critical for signalling during development; however, various factors modulate their activity. Among these factors are the Secreted Frizzled-Related Proteins (SFRP). We previously isolated the SFRP-4 gene from an involuting rat mammary gland and later showed that transgenic mice inappropriately expressing SFRP-4 during lactation exhibited a high level of apoptosis with reduced survival of progeny. RESULTS: In order to address the questions related to the mechanism of Wnt signalling and its inhibition by SFRP-4 which we report here, we employed partially-purified Wnt-3a in a co-culture model system. Ectopic expression of SFRP-4 was accomplished by infection with a pBabepuro construct. The co-cultures comprised Line 31E mouse mammary secretory epithelial cells and Line 30F, undifferentiated, fibroblast-like mouse mammary cells. In vitro differentiation of such co-cultures can be demonstrated by induction of the beta-casein gene in response to lactogenic hormones.We show here that treatment of cells with partially-purified Wnt-3a initiates Dvl-3, Akt/PKB and GSK-3beta hyperphosphorylation and beta-catenin activation. Furthermore, while up-regulating the cyclin D1 and connexin-43 genes and elevating transepithelial resistance of Line 31E cell monolayers, Wnt-3a treatment abrogates differentiation of co-cultures in response to the lactogenic hormones prolactin, insulin and glucocorticoid. Cells which express SFRP-4, however, are largely unaffected by Wnt-3a stimulation. Since a physical association between Wnt-3a and SFRP-4 could be demonstrated with immunoprecipitation/Western blotting experiments, this interaction, presumably owing to the Frizzled homology region typical of all SFRPs, explains the refractory response to Wnt-3a which was observed. CONCLUSION: This study demonstrates that Wnt-3a treatment activates the Wnt signalling pathway and interferes with in vitro differentiation of mammary co-cultures to beta-casein production in response to lactogenic hormones. Similarly, in another measure of differentiation, following Wnt-3a treatment mammary epithelial cells could be shown to up-regulate the cyclin D1 and connexin-43 genes while phenotypically they show increased transepithelial resistance across the cell monolayer. All these behavioural changes can be blocked in mammary epithelial cells expressing SFRP-4. Thus, our data illustrate in an in vitro model a mechanism by which SFRP-4 can modulate a differentiation response to Wnt-3a.

Asymmetry in scientific method and limits to cross-disciplinary dialogue: toward a shared language and science policy in pharmacogenomics and human disease genetics.

Pharmacogenomics is a hybrid field of experimental science at the intersection of human disease genetics and clinical pharmacology sharing applications of the new genomic technologies. But this hybrid field is not yet stable or fully integrated, nor is science policy in pharmacogenomics fully equipped to resolve the challenges of this emerging hybrid field. The disciplines of human disease genetics and clinical pharmacology contain significant differences in their scientific practices. Whereas clinical pharmacology originates as an experimental science, human disease genetics is primarily observational in nature. The result is a significant asymmetry in scientific method that can differentially impact the degree to which gene-environment interactions are discerned and, by extension, the study sample size required in each discipline. Because the number of subjects enrolled in observational genetic studies of diseases is characteristically viewed as an important criterion of scientific validity and reliability, failure to recognize discipline-specific requirements for sample size may lead to inappropriate dismissal or silencing of meritorious, although smaller-scale, craft-based pharmacogenomic investigations using an experimental study design. Importantly, the recognition that pharmacogenomics is an experimental science creates an avenue for systematic policy response to the ethical imperative to prospectively pursue genetically customized therapies before regulatory approval of pharmaceuticals. To this end, we discuss the critical role of interdisciplinary engagement between medical sciences, policy, and social science. We emphasize the need for development of shared standards across scientific, methodologic, and socioethical epistemologic divides in the hybrid field of pharmacogenomics to best serve the interests of public health.

Gene transfer of hepatocyte growth factor by electroporation reduces bleomycin-induced lung fibrosis.

Abnormal alveolar wound repair contributes to the development of pulmonary fibrosis after lung injury. Hepatocyte growth factor (HGF) is a potent mitogenic factor for alveolar epithelial cells and may therefore improve alveolar epithelial repair in vitro and in vivo. We hypothesized that HGF could increase alveolar epithelial repair in vitro and improve pulmonary fibrosis in vivo. Alveolar wound repair in vitro was determined using an epithelial wound repair model with HGF-transfected A549 alveolar epithelial cells. Electroporation-mediated, nonviral gene transfer of HGF in vivo was performed 7 days after bleomycin-induced lung injury in the rat. Alveolar epithelial repair in vitro was increased after transfection of wounded epithelial monolayers with a plasmid encoding human HGF, pCikhHGF [human HGF (hHGF) gene expressed from the cytomegalovirus (CMV) immediate-early promoter and enhancer] compared with medium control. Electroporation-mediated in vivo HGF gene transfer using pCikhHGF 7 days after intratracheal bleomycin reduced pulmonary fibrosis as assessed by histology and hydroxyproline determination 14 days after bleomycin compared with controls treated with the same vector not containing the HGF sequence (pCik). Lung epithelial cell proliferation was increased and apoptosis reduced in hHGF-treated lungs compared with controls, suggesting increased alveolar epithelial repair in vivo. In addition, profibrotic transforming growth factor-beta1 (TGF-beta1) was decreased in hHGF-treated lungs, indicating an involvement of TGF-beta1 in hHGF-induced reduction of lung fibrosis. In conclusion, electroporation-mediated gene transfer of hHGF decreases bleomycin-induced pulmonary fibrosis, possibly by increasing alveolar epithelial cell proliferation and reducing apoptosis, resulting in improved alveolar wound repair.

Sociocultural determinants of tobacco use among Cambodian Americans.

The objectives of this study included the following: obtaining qualitative information on tobacco use among Cambodian Americans, identifying cultural factors that influence tobacco use and acquiring information for the development of effective smoking prevention and cessation strategies. Data were collected by using demographic and behavioral questionnaires and focus group interviews. A total of 14 focus group interviews that covered cultural practices associated with smoking were administered. Statistical analyses included univariate frequency distributions and cross-tabulations. The subjects (n = 119) were Cambodian American volunteers who participated in social services programs offered by a community service organization. All subjects were 18 years of age or older and resided in the city of Long Beach. The principal outcomes measured were cigarette smoking and tobacco use. Other variables included reasons for smoking, traditional uses of tobacco, stress factors related to smoking and the perceived health effects of smoking. Predisposing, reinforcing and enabling factors associated with tobacco-use behaviors included peer group influences, smoking adopted as a coping method, tobacco used for medicinal purposes and smoking practiced within cultural traditions. The frequency of smoking was four times higher among males than among females. Smokers (n = 29) in comparison with non-smokers (n = 90) tended to be men (79% versus 33%), not married (68% versus 49%) and unemployed (79% versus 54%), and had attained somewhat lower levels of education. The role of cultural factors needs to be considered when designing appropriate smoking cessation strategies for Cambodian Americans.

Lactoferrin (Lf): Retinoid interactions in the mammary glands of transgenic mice overexpressing human Lf.

Induction of protein expression in a tissue-specific manner by gene transfer over-expression techniques has been one means to define the function of a protein in a biological paradigm. Studies with retinoid reporter constructs transfected in mammary cell lines suggests that lactoferrin (Lf) affects retinoid signaling pathways and alters apoptosis. We tested the effects and interactions of over-expressed mammary-specific human lactoferrin (hLf) and dietary retinol palmitate on lactation and mammary gland development in mice. Increased retinol palmitate in the diet increased daily retinol equivalents (RE) to 2.6-fold over the normal mouse control diet. Transgene (Tg) expression in the dam fed control diet depressed pup weight gain. Severe depression of pup weight gain was observed when homozygote TgTg dams were fed the RE diet. Normal weight gain was restored when pups were placed with a wild type dam fed the RE diet; conversely, normal growing pups from the wild type dams showed declining weight gains when fostered to the TgTg RE-fed dams. Northern analysis of mammary tissue extracts showed a reduction in WAP and an increase in IGFBP-3 mRNA that was associated with the presence of the transgene. Histological evaluation of 3 days lactating mammary tissue showed mammary epithelial cells from TgTg animals contained excessive secretory products, suggesting a block in cellular secretion mechanisms. In addition, the mammary cells displayed a cellular apical membrane puckering that extended into the alveoli lumens. These studies demonstrate an in vivo interaction of Tg-hLf expression and dietary retinoids in mouse mammary glands. While normal mammary gland physiology may not be representative by these experiments because high Lf concentrations during early lactation are abnormal, the demonstrated biological interaction suggests that typical periods of high Lf concentrations may have impact upon developing and involuting mammary glands.

The link between obesity and the built environment. Evidence from an ecological analysis of obesity and vehicle miles of travel in California.

AIMS: Obesity and physical inactivity are known to be risk factors for many chronic diseases including hypertension, coronary artery disease, diabetes, and cancer. We sought to explore the association between an indicator of transportation data (Vehicle Miles of Travel, VMT) at the county level as it relates to obesity and physical inactivity in California. METHODS: Data from the California Health Interview Survey 2001 (CHIS 2001), the US 2000 Census, and the California Department of Transportation were merged to examine ecological correlations between vehicle miles of travel, population density, commute time, and county indicators of obesity and physical inactivity. Obesity was measured by body mass index (BMI). Physical inactivity was based on self-reported behaviors including walking, bicycling, and moderate to vigorous activity. The unit of analysis was the county. Thirty-three counties in California with population size greater than 100,000 persons per county were retained in the analyses. RESULTS: CHIS 2001 statewide obesity prevalence ranged from 11.2% to 28.5% by county. Physical inactivity ranged from 13.4% to 35.7%. Daily vehicle miles of travel ranged from 3.3 million to 183.8 million per county. By rank bivariate correlation, obesity and physical inactivity were significantly associated (p<0.01). Furthermore, by rank analysis of variance, the highest mean rank obesity was associated with the highest rank of VMT (p<0.01). Similar rank patterns were observed between obesity and physical inactivity and commute time. Associations between VMT and physical inactivity were examined but failed to reach statistical significance. CONCLUSION: This analysis adds to the growing evidence supporting the association between VMT (a measure of automobile transportation) and obesity. An urban design characterized by over dependence on motorized transportation may be related to adverse health effects.

Cloning and characterization of a novel zinc finger protein (rZFP96) in the rat corpus luteum.

The corpus luteum (CL) is a temporary organ involved in the maintenance of pregnancy. In the course of its life-cycle, the CL undergoes two distinct and consecutive processes for its inevitable removal through apoptosis: functional and structural luteolysis. We isolated a gene encoding for a novel rat zinc finger protein (ZFP), named rat ZFP96 (rZFP96) from an ovarian lambda cDNA library. Sequence analysis revealed close sequence and structural similarity to mouse ZFP96 and human zinc finger protein 305 (ZNF305). Quantitative reverse transcription-polymerase chain reaction analysis revealed a positive correlation with the end of pregnancy, that is, the onset of structural luteolysis of the CL. Messenger RNA levels increased 3-fold (P < 0.01) between days 13 and 22 of pregnancy and 8-fold (P < 0.01) between day 13 of pregnancy and day 1 post-partum. In addition, we detected rZFP96 expression in mammary, placenta, heart, kidney and skeletal muscle. Sequence analysis predicted that rZFP96 has a high probability of localizing to the nuclear compartment. The presence of both a perfect consensus TGEKP linker sequence between zinc fingers 2 and 3 as well as several similar sequences between the other zinc fingers suggests physical interaction with DNA. Speculatively, rZFP96 may therefore function as a transcription factor, switching-off pro-survival genes and/or upregulating pro-apoptotic genes and thereby contributing to the demise of the CL.

Expression and localization of secreted frizzled-related protein-4 in the rodent ovary: evidence for selective up-regulation in luteinized granulosa cells.

Secreted frizzled-related protein-4 (sFRP-4) belongs to a family of soluble proteins that have a Frizzled-like cysteine-rich domain and function as modulators of Wnt-Frizzled (Fz) signals. As several Wnts and Fz are expressed at defined stages of follicular development in rodent ovaries, these studies were undertaken to evaluate the hormone-regulated expression and localization of sFRP-4. In the mouse ovary, the expression of sFRP-4 mRNA was up-regulated in granulosa cells of large antral follicles after human chorionic gonadotropin administration and was also elevated in corpora lutea, as determined by RT-PCR and in situ hybridization analyses. In hypophysectomized rat ovaries, sFRP-4 expression was similarly induced by human chorionic gonadotropin and further up-regulated by PRL. PRL also stimulated the secretion of sFRP-4 protein from luteinized rat granulosa cells in culture. Therefore, regulation of sFRP-4 by LH and PRL may be important for modulating Fz-1, which is known to be expressed in periovulatory follicles, and Wnt-4/Fz-4, which are expressed in corpora lutea.

Association chain graphs: modelling etiological pathways.

Multiple time-dynamic and interrelated risk factors are usually involved in the complex etiology of disorders. This paper presents a strategy to explore and display visually the relative importance of different association pathways for the onset of disorder over time. The approach is based on graphical chain models, a tool that is powerful but still under-utilized in most fields. Usually, the results of these models are displayed using directed acyclic graphs (DAGs). These draw an edge between a pair of variables whenever the assumption of conditional independence given variables on an earlier or equal temporal footing is violated to a statistically significant extent. In the present paper, the graphs are modified in that confidence intervals for the strengths of associations (statistical main effects) are visualized. These new graphs are called association chain graphs (ACGs). Statistical interactions cause 'edges' between the respective variables within the DAG framework (because the assumption of conditional independence is violated). In contrast they are represented as separate graphs within the subsample where the different association chains may work within the ACG framework. With this new type of graph, more specific information can be displayed whenever the data are essentially described only with statistical main- and two-way interaction effects.

Regulated expression of inhibitor of apoptosis protein 3 in the rat corpus luteum.

We sought to investigate the role inhibitor of apoptosis proteins (IAPs) play in the life cycle of the corpus luteum (CL) of the rat. We isolated two clones with amino acid homology to rat IAP2 (BIRC 3) and three to rat IAP3 (rIAP3; BIRC 4). The expression of rIAP3 mRNA was examined in the rat CL during and after pregnancy, in Day 8 pregnant rats after 24-h treatment of gonadotropin-releasing hormone-agonist (GnRH-Ag), and in a CL organ culture model of spontaneous apoptosis in the absence of tropic support with and without superoxide dismutase. We used real-time RT-PCR to quantitate rIAP3 mRNA expression. Interestingly, a significant reduction in rIAP3 levels was seen at the time of CL regression in the course of natural pregnancy and the GnRH-Ag model. Surprisingly, rIAP3 mRNA levels in the CL organ culture model of spontaneous apoptosis failed to show significant changes, although TUNEL (terminal deoxynucleotide transferase-mediated dUTP nick end-labeling) reaction showed 30%-40% of the cells undergoing DNA fragmentation after 2 h in culture. In situ hybridization revealed that rIAP3 expression was localized to the cytoplasm of luteal and granulosa cells. These data clearly demonstrate both the presence of IAPs in the rat CL and the regulation of rIAP3 during in vivo apoptotic cell death, indicating a role for IAPs in the maintenance of CL function and demise.