WSB1 Involvement in Prostate Cancer Progression.

Prostate cancer (PC) is polygenic disease involving many genes, and more importantly a host of gene-gene interactions, including transcriptional factors. The WSB1 gene is a transcriptional target of numerous oncoproteins, and its dysregulation can contribute to tumor progression by abnormal activation of targeted oncogenes. Using data from the Cancer Genome Atlas, we tested the possible involvement of WSB1 in PC progression. A multi-dimensional scaling (MDS) model was applied to clarify the association of WSB1 expression with other key genes, such as c-myc, ERG, Enhancer of Zeste 1 and 2 (EHZ1 and EZH2), WNT10a, and WNT 10b. An increased WSB1 expression was associated with higher PC grades and with a worse prognosis. It was also positively related to EZH1, EZH2, WNT10a, and WNT10b. Moreover, MDS showed the central role of WSB1 in influencing the other target genes by its central location on the map. Our study is the first to show a link between WSB1 expression and other genes involved in PC progression, suggesting a novel role for WSB1 in PC progression. This network between WSB1 and EZH2 through WNT/ß-catenin may have an important role in PC progression, as suggested by the association between high WSB1 expression and unfavorable prognosis in our analysis.

Steroid Hormones as Modulators of Emotional Regulation in Male Urogenital Cancers.

BACKGROUND: Tumors develop within an organism operating in a specific social and physical environment. Cortisol and dehydroepiandrosterone (DHEA), two of the most abundant steroid hormones in humans, are involved in both emotional regulation and the tumor progression. Several studies reported preclinical findings that DHEA can have preventive and therapeutic efficacy in treating major age-associated diseases, including cancer, although the mechanisms of action are not yet defined. The main aim of current study was to investigate the relationship between psychological and physiological emotional regulation and cancer development. METHOD: This study assessed the quality of life of urogenital cancer male patients using several validated tools, including the Functional Assessment of Cancer Therapy-General and the Profile of Mood States. Saliva samples were collected to monitor peripheral activity of both cortisol and DHEA. It was hypothesized that patients with a better quality of life would have higher levels of the DHEA/cortisol ratios. RESULTS: We found that the quality of life was positively related to DHEA, but not cortisol levels. Negative mood increases were related to lower levels of DHEA. Logistic regression of the predictors of metastases indicated three main independent factors involved: DHEA, age, and cortisol. In other words, the higher the DHEA levels in comparison to cortisol levels, controlling for age, the lower the probability of metastases. CONCLUSION: Our results appear to support the hypothesis that emotional dysregulation mediated by DHEA/cortisol activity is a key factor in the probability of metastasis in urogenital cancers.

Proposal for a New Diagnostic Histopathological Approach in the Evaluation of Ki-67 in GEP-NETs.

INTRODUCTION: Studies have shown that the Ki-67 index is a valuable biomarker for the diagnosis, and classification of gastro-entero-pancreatic neuroendocrine tumors (GEP-NETs). We re-evaluated the expression of Ki-67 based on the intensity of the stain, basing our hypothesis on the fact that the Ki-67 protein is continuously degraded. BACKGROUND: The aim was to evaluate whether a new scoring method would be more effective in classifying NETs by reducing staining heterogeneity. METHODS: Patients with GEP-NET (n = 87) were analyzed. The classification difference between the two methods was determined. RESULTS: The classification changed significantly when the Ki-67 semiquantal index was used. The percentage of G1 patients increased from 18.4% to 60.9%, while the G2 patients decreased from 66.7% to 29.9% and the G3 patients also decreased from 14.9% to 9.2%. Moreover, it was found that the traditional Ki-67 was not significantly related to the overall survival (OS), whereas the semiquantal Ki-67 was significantly related to the OS. CONCLUSIONS: The new quantification was a better predictor of OS and of tumor classification. Therefore, it could be used both as a marker of proliferation and as a tool to map tumor dynamics that can influence the diagnosis and guide the choice of therapy.

Multi-Dimensional Scaling Analysis of Key Regulatory Genes in Prostate Cancer Using the TCGA Database.

Prostate cancer (PC) is a polygenic disease with multiple gene interactions. Therefore, a detailed analysis of its epidemiology and evaluation of risk factors can help to identify more accurate predictors of aggressive disease. We used the transcriptome data from a cohort of 243 patients from the Cancer Genome Atlas (TCGA) database. Key regulatory genes involved in proliferation activity, in the regulation of stress, and in the regulation of inflammation processes of the tumor microenvironment were selected to test a priori multi-dimensional scaling (MDS) models and create a combined score to better predict the patients' survival and disease-free intervals. Survival was positively correlated with cortisol expression and negatively with Mini-Chromosome Maintenance 7 (MCM7) and Breast-Related Cancer Antigen2 (BRCA2) expression. The disease-free interval was negatively related to the expression of enhancer of zeste homolog 2 (EZH2), MCM7, BRCA2, and programmed cell death 1 ligand 1 (PD-L1). MDS suggested two separate pathways of activation in PC. Within these two dimensions three separate clusters emerged: (1) cortisol and brain-derived neurotrophic factor BDNF, (2) PD-L1 and cytotoxic-T-lymphocyte-associated protein 4 (CTL4); (3) and finally EZH2, MCM7, BRCA2, and c-Myc. We entered the three clusters of association shown in the MDS in several Kaplan-Meier analyses. It was found that only Cluster 3 was significantly related to the interval-disease free, indicating that patients with an overall higher activity of regulatory genes of proliferation and DNA repair had a lower probability to have a longer disease-free time. In conclusion, our data study provided initial evidence that selecting patients with a high grade of proliferation and DNA repair activity could lead to an early identification of an aggressive PC with a potentials for metastatic development.

Gastrin-Releasing Peptide Receptor in Low Grade Prostate Cancer: Can It Be a Better Predictor Than Prostate-Specific Membrane Antigen?

The aim of the present study was to evaluate whether prostate cancer (PC) patients can be accurately classified on the bases of tissue expression of gastrin-releasing peptide receptor (GRPR) and prostate-specific membrane antigen (PSMA). This retrospective study included 28 patients with PC. Formalin-fixed paraffin-embedded samples were used for diagnosis. Immunohistochemistry staining techniques were used to evaluate PSMA and GRPR expression (both number of cells expressed and % of area stained). To assess the independent associations among selected variables, a multi-dimensional scaling (MDS) analysis was used. It was found that the PSMA expression was inversely correlated with GRPR expression. Only the number of cells expressing GRPR was significantly related to the Gleason score. Both the percentage of area expressing GRPR and the number of cells expressing PSMA were close to reaching significance at the 0.05 level. MDS provided a map of the overall, independent association confirming that GRPR and PSMA represent inversely correlated measures of the same dimension. In conclusion, our data showed that GRPR expression should be evaluated in prostate biopsy specimens to improve our ability to detect PC with low grades at the earliest phases of development. Considering that GRPRs appear to be directly involved in the mechanisms of tumor proliferation, advancements in nuclear medicine radiotherapy can focus on this receptor to improve the therapeutic approach to PC. Further studies in our laboratory will investigate the molecular mechanisms of activation based on GRPR.

Immunomodulatory Effects of Stress and Environmental Enrichment in Long-Evans Rats (Rattus norvegicus).

Stress can influence the secretion of neuroendocrine mediators, thereby exposing immune cells to altered signaling and interactions. Here we investigated the synergetic effect of stress and environmental enrichment on the immune response of Long-Evans rats. Subjects (n = 46) were assigned to 5 treatment groups: acute compared with chronic stress with or without environmental enrichment, plus an unmanipulated control group. Animals also were classified as active, passive, and flexible copers according to back-test assessment. Rats were exposed to enrichment in an open-field containing objects in different areas for 30 min 3 times each week, thus modeling the effects of a temporary increase in environmental stimuli. Animals assigned to chronic stress groups were exposed to predator sound stressors for 30 min daily, whereas animals assigned to acute stress groups were exposed once each week. After 7 wk, a dermal punch biopsy was administered to activate the immune response, after which rats were challenged through a forced swim test. Biologic samples were collected to measure corticosterone, dehydroepiandrosterone (DHEA), oxytocin, testosterone, and the cytokines IL6 and IL10. Rats exposed to chronic stress had lower DHEA:corticosterone ratios, suggesting increased allostatic load. Enrichment exposure modulated these effects, lowering overall corticosterone and testosterone levels and increasing DHEA and oxytocin levels in animals exposed to the predator sound. The immune response was decreased in rats exposed to chronic stress, but the effect of environmental enrichment helped to mitigate the negative influence on cells producing IL6. Combining acute stress and exposure to an enriched environment returned a healthier profile in terms of both immune activation and stress regulation. By using a multidimensional scaling model, we found that a combination of 'good' stress and exposure to brief sessions of enriching stimuli can reliably predict health in Long-Evans rats.

Natural-enriched environments lead to enhanced environmental engagement and altered neurobiological resilience.

The mammalian brain has evolved in close synchrony with the natural environment; consequently, trends toward disengagement from natural environments in today's industrialized societies may compromise adaptive neural responses and lead to psychiatric illness. Investigations of rodents housed in enriched environments indicate enhanced neurobiological complexity; yet, the origin of these stimuli, natural vs. manufactured, has not been sufficiently explored. In the current study, groups of rats were exposed to one of three environments: (1) a standard environment with only food and water, (2) an artificial-enriched environment with manufactured stimuli and (3) a natural-enriched environment with natural stimuli. Results indicated that, during the dark phase, natural-enriched animals exhibited longer durations interacting with objects than the artificial-enriched group; further, the natural-enriched group engaged in more social behavior than the other two groups. Both enriched groups exhibited less anxiety in response to a novel object but the natural-enriched rats exhibited less anxiety-typical behavior in response to a predator odor than the other groups. Less fos activation in the amygdala was observed in both enriched groups following a water escape task whereas an increase in fos activation in the nucleus accumbens was observed in the natural-enriched animals. Thus, the current findings indicate the potential importance of exposure to complex environments, especially natural-like habitats, in the maintenance of emotional health, perhaps providing a buffer against the emergence of anxiogenic responses.

Effort-Based Reward (EBR) training enhances neurobiological efficiency in a problem-solving task: insights for depression therapies.

Effort-Based Reward (EBR) training strengthens associations between effort and rewards, leading to increased persistence in an unsolvable task when compared to control animals. EBR training involves placing animals in a test apparatus in which they are trained to dig in mounds to retrieve froot loop rewards (contingent group); these animals are compared to control animals that are given the same number of rewards, regardless of expended effort (noncontingent group). In the current study, the effect of EBR training on performance in a spatial task (Dry Land Maze) was explored to determine cognitive resilience during behavioral testing. Additionally, animals received BrdU injections during training to assess the role of neurogenesis on subsequent behavioral performance. Following the probe test, animals were perfused so that fos-immunoreactive (ir) cells in the hippocampus and cortical areas could be assessed. Behavioral results indicated that contingent rats were approximately 50% more efficient in locating and interacting with the previous baited well during the probe test than noncontingent animals, recruiting approximately 20% less c-fos ir-cells in the insular cortex, retrosplenial cortex, and dentate gyrus. A multidimensional scaling analysis grouped noncontingent animals together in a quadrant characterized by high latencies to find the previous baited well and higher ir-cell activation in the aforementioned areas. Thus, our data support the hypothesis that the EBR training enhances both cognitive functioning and emotional regulation during challenging events. Considering the ongoing controversy about the efficacy of pharmacological interventions in treating depression, the EBR model provides a valuable alternative for the investigation of the neurobiology of mood disorders.

Characteristic neurobiological patterns differentiate paternal responsiveness in two Peromyscus species.

Rodent paternal models provide unique opportunities to investigate the emergence of affiliative social behavior in mammals. Using biparental and uniparental Peromyscus species (californicus and maniculatus, respectively) we assessed paternal responsiveness by exposing males to biological offspring, unrelated conspecific pups, or familiar brothers following a 24-hour separation. The putative paternal circuit we investigated included brain areas involved in fear/anxiety [cingulate cortex (Cg), medial amygdala (MeA), paraventricular nucleus of the hypothalamus (PVN), and lateral septum (LS)], parental motivation [medial preoptic area (MPOA)], learning/behavioral plasticity (hippocampus), olfaction [pyriform cortex (PC)], and social rewards (nucleus accumbens). Paternal experience in californicus males reduced fos immunoreactivity (ir) in several fear/anxiety areas; additionally, all californicus groups exhibited decreased fos-ir in the PC. Enhanced arginine vasopressin (AVP) and oxytocin (OT)-ir cell bodies and fibers, as well as increased neuronal restructuring in the hippocampus, were also observed in californicus mice. Multidimensional scaling analyses revealed distinct brain activation profiles differentiating californicus biological fathers, pup-exposed virgins, and pup-naïve virgins. Specifically, associations among MPOA fos, CA1 fos, dentate gyrus GFAP, CA2 nestin-, and PVN OT-ir characterized biological fathers; LS fos-, Cg fos-, and AVP-ir characterized pup-exposed virgins, and PC-, PVN-, and MeA fos-ir characterized pup-naïve virgins. Thus, whereas fear/anxiety areas characterized pup-naïve males, neurobiological factors involved in more diverse functions such as learning, motivation, and nurturing responses characterized fatherhood in biparental californicus mice. Less distinct paternal-dependent activation patterns were observed in uniparental maniculatus mice. These data suggest that dual neurobiological circuits, leading to the inhibition of social-dependent anxiety as well as the activation of affiliative responses, characterize the transition from nonpaternal to paternal status in californicus mice.

Regulation of sexual behaviour in male macaques by sex steroid modulation of the serotonergic system.

The view that androgen action is the primary impetus underlying male-typical behaviour has been irrevocably altered by the profound perturbations in social and sexual behaviour observed in recent models of oestrogen insufficiency in male mice. Evidence is also accumulating for an involvement of oestrogens in the modulation of neural systems that are thought to play important roles in male reproductive functioning. Specifically, the serotonergic system is implicated in diverse autonomic functions, most or all of which are sensitive to oestradiol as well. Although their interaction domains have yet to be examined in male primates, roles have been established for both oestrogen and serotonin in the regulation of male sexual behaviour. We used a blinded, sham-treated and self-controlled, randomized, multitreatment cross-over design to test the hypothesis that male sexual behaviour is regulated by oestrogen modulation of the serotonergic system in intact male Japanese macaques. Regression analysis revealed that oestradiol and whole blood tryptophan, but not testosterone or 5alpha-dihydrotestosterone, had additive, independent effects on male potentia over a range of hormone concentrations, whereas androgens were confirmed to be the primary determinants of sexual motivation. We suggest that modulation of the serotonergic system by 'female hormones' may be fundamental to the regulation of male mating success in higher primates. This might also explain, at least in part, why significant correlations between steroid hormones and male copulatory behaviour have traditionally proven so elusive in this order, thereby warranting a re-evaluation of the current notion that male sexual behaviour has been emancipated from activational hormonal control in higher primates.

Hormonal correlates of changes in interest in unrelated infants across the peripartum period in female baboons (Papio hamadryas anubis sp.).

In past research on human and nonhuman primates, maternal responsiveness and behavior has been thought of as an experiential, cognitive mechanism; however, recent findings have shown that maternal motivation and behavior may not be entirely divorced from the endocrine system. To investigate the relationship between interest in infants and the hormonal changes related to pregnancy, we examined the nature of social interactions across parturition between a large sample (n = 133) of adult female baboons (Papio hamadryas anubis sp.) and unrelated infants. Prepartum data were collected during ten 30-min focal observations for each subject. Each mother-infant pair was then observed through the infant's first 8 weeks of life. A total of 2325 h of observation was recorded. Urine was collected on 65 subjects, starting 5 weeks before the expected date of parturition and ending 4 weeks after parturition. Evidence for a connection between endocrine function and responsiveness toward infants was found. Affiliative behaviors during the prepartum period were positively correlated to the estrogen/cortisol ratio and high dominance rank. In the postpartum period, affiliative behaviors were positively correlated with prepartum progesterone and dominance rank, and negatively correlated with postpartum cortisol levels. Finally, a positive correlation was recorded during the postpartum period between prepartum progesterone and aggression, and a negative correlation between postpartum cortisol and aggression and submission. Our data suggest that the endocrine changes that may help regulate maternal care of offspring also influence the way in which pre- and postpartum female baboons interact with unrelated infants in their social group.

Excretion of urinary steroids in pre- and postpartum female baboons.

Steroid hormones are important regulators of a wide variety of reproductive and behavioral functions. We investigated the ability to track sex steroids and glucocorticoids in urine samples collected noninvasively from pre- and postpartum female baboons. Paired plasma and urine samples were collected every 2 weeks prior to and following birth in 10 females. Changes in concentrations of plasma steroids (estradiol, progesterone, and cortisol) were reflected in changes in urinary metabolite excretion (estrone conjugates, pregnanediol conjugates, and cortisol; r's>0.36, p's<0.001). A low correlation between prepartum plasma and urinary cortisol may reflect late-gestational changes in the production and/or metabolism of glucocorticoids. Steroid excretion profiles in a large sample of females giving birth and caring for healthy infants (n = 108) were compared with profiles obtained from females with poor maternal-fetal outcomes (late-term stillbirth, n = 14) and from females with significant postpartum problems with maternal care (n = 20). Mothers giving birth to stillborn infants had lower prepartum levels of urinary estrone conjugates and cortisol, suggesting reduced placental steroidogenesis. Mothers with postpartum behavioral difficulties had higher concentrations of prepartum estrone excretion, lower cortisol excretion, and elevated E/P ratios throughout the peripartum period. Noninvasive sample collection and enzyme immunoassay, therefore, have predictive utility regarding circulating steroid concentrations and can identify important endocrine correlates of physiological and behavioral abnormalities in baboons.

Differences in the endocrine and behavioral profiles during the peripartum period in macaques.

The present investigation aims to assess the changes in both social interaction and sex steroids excreted in feces of group-living Japanese macaques and rhesus monkeys. By comparing profiles of estrone conjugates (E1C) and pregnanediol-glucuronide (PdG) with the behavioral propensities of two closely related species living in similar environments, we could test the hypothesis that the social behavior of pregnant females shows significant hormonally mediated changes during the late prepartum and early postpartum period. We found a general tendency to withdraw from social life across pregnancy in both species. These behavioral changes were paralleled by endocrine profiles showing a slight prepartum increase in E1C during the last week in the rhesus group, whereas the increase was more marked and continuous in the Japanese macaque group. PdG increased slightly in rhesus macaques, whereas in Japanese macaques the fluctuations were not significant. Postpartum, both hormones dropped to low levels in both species, with no significant variation therein. Consequent to these changes, the E1C/PdG ratio increased significantly in late pregnancy only in the Japanese macaque group. Overall, these results show significant differences in the social behavior and endocrine profiles of two closely related species, thus complementing previous findings and indicating species-specific characteristics of the association between changes in affiliative behaviors and hormonal fluctuations. In particular, the shift between grooming performed and self-grooming, which showed the closest association with variations in the E1C/PdG ratio, could represent a reliable indicator of the change in the internal status of pregnant females, and is probably functional to infant survival.

Peripartum sex steroid changes and maternal style in rhesus and Japanese macaques.

The subtle and complex relationships between the sequential maturation of the endocrine systems during pregnancy and parturition, and the hormonal role in activating the central nervous system to express maternal behavior in primates, are far from being completely understood. Recent studies on the association between sex steroids and maternal behavior have yielded conflicting results in this group. Here we use a comparative approach to assess the correlation between changes in the peripartum endocrine profiles and maternal styles in two closely related macaque species, housed in analogous environments. We included in this study the first seven Japanese macaque and seven rhesus macaque mother-infant pairs born during the birth season of 2001 at the Primate Research Institute, Kyoto University, Japan. We observed each pair 3h/week (six weekly 30-min observation sessions) during the first 12 weeks of lactation. We collected fecal samples twice a week from each mother, starting 4 weeks before parturition and ending 4 weeks after parturition. We tested the hypothesis that neuroendocrine changes during pregnancy and lactation might specifically contribute to the regulation and timing of infant rejection. Despite their biological similarities, we observed a clear difference in maternal style between the two groups concerning rejection rates: rhesus macaque mothers rejected their infants earlier and more frequently throughout the whole 12 weeks of study. On the other hand, protectiveness showed similar patterns and values in the two groups, and maternal warmth was significantly higher in the rhesus group, but it followed a similar pattern over time. We also confirmed an association between maternal rejection and excreted estrogen, but not excreted progesterone, for Japanese macaques. This association was not apparent for the rhesus macaques. This result, coupled with the observation that rhesus mothers are more rejecting than Japanese macaque mothers, tends to support our hypothesis. As a group, rhesus macaques are less inhibited in the rejection of their infants, and this is paralleled by a less marked change in the primacy of estrogen in the last phase of pregnancy. On the contrary, the Japanese group is characterized by higher levels of E(1)C and the E(1)C/PdG ratio. Therefore, according to our hypothesis, their tendency to increase the rejection rate may be suppressed through a feedback loop that enhances maternal motivation and results in a more tolerant outcome toward the infant.

Mother-infant relationships and maternal estrogen metabolites changes in macaques (Macaca fuscata, M. mulatta).

This research assessed mother-infant relationships in rhesus and Japanese macaques living in analogous captive social groups, and monitored changes in the levels of excreted estrogen metabolites during the peripartum period. Each mother-infant pair was focally observed 3 h per week during the first 12 weeks of life of newborns. Fecal samples were collected twice a week from each mother, starting 4 weeks before delivery and ending 4 weeks after delivery. Infant-directed behaviors appeared to be consistently less protective/controlling and more rejecting in rhesus macaques than in Japanese macaques. Estrogen metabolite levels during the perinatal period were, on average, 3-fold higher in Japanese macaques and showed a sharp increase during the last weeks of pregnancy only in the Japanese macaque group. Considering the ecological and behavioral similarities between Japanese and rhesus macaques, the divergence between the two species in the onset and maintenance of maternal behavior was unexpected. This was possibly linked to the difference in the overall body size and life history, and to the striking divergence in estrogen metabolite variation during the peripartum period. Group size, social relationships, and average age of individuals in the two captive groups were not clearly involved in the recorded differences in maternal behavior.