CCAAT/Enhancer Binding Protein-delta (C/EBP-delta) regulates cell growth, migration and differentiation

<p>Abstract</p> <p>Background</p> <p>CCAAT/enhancer binding protein-delta (C/EBP-delta) is a member of the highly conserved C/EBP family of basic region leucine zipper transcription factors. C/EBP family members regulate cell growth and differentiation and "loss of function" alterations in C/EBPs have been reported in a variety of human cancers. C/EBP-delta gene expression is upregulated by G₀growth arrest, IL-6 family cytokines and endotoxin treatments. C/EBP-delta exhibits properties of a tumor suppressor gene, including reduced expression and promoter methylation-induced silencing in transformed cell lines and primary tumors. In addition, C/EBP-delta gene expression is repressed by c-Myc, an oncogene that is over-expressed in a wide range of human cancers. "ChIP-chip" studies demonstrated that C/EBP-delta functions as a transcriptional activator of target genes that function in intracellular signal transduction, transcription, DNA binding/repair, cell cycle control, cell adhesion, and apoptosis. Despite progress in determining the biochemical functions of C/EBP-delta, the specific cellular defects that are induced by C/EBP-delta "loss of function" alterations are poorly understood. This study investigated the impact of C/EBP-delta "loss of function" alterations on growth arrest, migration/invasion and differentiation in nontransformed mouse mammary epithelial cells (MECs) and primary mouse embryo fibroblasts (MEFs).</p> <p>Results</p> <p>C/EBP-delta siRNA transfected MECs exhibited ~90% reduction in C/EBP-delta mRNA and protein levels. C/EBP-delta siRNA treatment resulted in defective growth arrest as demonstrated by persistently elevated BrdU labeling, ³H-thymidine incorporation and cyclin D1 levels in response to growth arrest treatments. C/EBP-delta siRNA treatment also resulted in increased migration/invasion and defective differentiation. C/EBP-delta knockout MEFs exhibited defective growth arrest and increased proliferation/migration. Re-introduction of C/EBP-delta expression restored the growth arrest response of C/EBP-delta knockout MEFs. Finally, deletion of the C/EBP-delta DNA binding domain or the C/EBP-delta bZIP domain resulted in the loss of C/EBP-delta growth inhibition in clonogenic assays.</p> <p>Conclusions</p> <p>This study demonstrates that C/EBP-delta functions in the regulation of critical cell fate determining programs such as growth arrest, migration, and differentiation. These results support the tumor suppressor function of C/EBP-delta and identify potential mechanisms in which "loss of function" alterations in C/EBP-delta could promote cell transformation and tumorigenesis.</p

Myc interacts with Max and Miz1 to repress C/EBPδ promoter activity and gene expression

<p>Abstract</p> <p>Background</p> <p>"Loss of function" alterations in CCAAT/Enhancer Binding Proteinδ (C/EBPδ) have been reported in a number of human cancers including breast, prostate and cervical cancer, hepatocellular carcinoma and acute myeloid leukemia. C/EBPδ gene transcription is induced during cellular quiescence and repressed during active cell cycle progression. C/EBPδ exhibits tumor suppressor gene properties including reduced expression in cancer cell lines and tumors and promoter methylation silencing.</p> <p>We previously reported that C/EBPδ expression is inversely correlated with c-Myc (Myc) expression. Aberrant Myc expression is common in cancer and transcriptional repression is a major mechanism of Myc oncogenesis. A number of tumor suppressor genes are targets of Myc transcriptional repression including C/EBPα, p15^{<it>INK</it>4}, p21^{<it>CIP</it>1}, p27^{<it>KIP</it>1}and p57^{<it>KIP</it>2}. This study investigated the mechanisms underlying Myc repression of C/EBPδ expression.</p> <p>Results</p> <p>Myc represses C/EBPδ promoter activity in nontransformed mammary epithelial cells in a dose-dependent manner that requires Myc Box II, Basic Region and HLH/LZ domains. Chromatin Immunoprecipitation (ChIP) assays demonstrate that Myc, Miz1 and Max are associated with the C/EBPδ promoter in proliferating cells, when C/EBPδ expression is repressed. EMSAs demonstrate that Miz1 binds to a 30 bp region (-100 to -70) of the C/EBPδ promoter which contains a putative transcription initiator (Inr) element. Miz1 functions exclusively as a repressor of C/EBPδ promoter activity. Miz1 siRNA expression or expression of a Miz1 binding deficient Myc (MycV394D) construct reduces Myc repression of C/EBPδ promoter activity. Max siRNA expression, or expression of a Myc construct lacking the HLH/LZ (Max interacting) region, also reduces Myc repression of C/EBPδ promoter activity. Miz1 and Max siRNA treatments attenuate Myc repression of endogenous C/EBPδ expression. Myc Box II interacting proteins RuvBl1 (Pontin, TIP49) and RuvBl2 (Reptin, TIP48) enhances Myc repression of C/EBPδ promoter activity.</p> <p>Conclusion</p> <p>Myc represses C/EBPδ expression by associating with the C/EBPδ proximal promoter as a transient component of a repressive complex that includes Max and Miz1. RuvBl1 and RuvBl2 enhance Myc repression of C/EBPδ promoter activity. These results identify protein interactions that mediate Myc repression of C/EBPδ, and possibly other tumor suppressor genes, and suggest new therapeutic targets to block Myc transcriptional repression and oncogenic function.</p

Osmotic characteristics and fertility of murine spermatozoa collected in different solutions

Osmotic stress is an important factor that can result in cell damage during cryopreservation. Before ejaculation or collection for cryopreservation, murine spermatozoa are stored in epididymal fluid, a physiologically hyperosmotic environment (w415 mmol/kg). The objectives of this study were to determine the osmotic tolerance limits of sperm motion parameters of ICR and C57BL/6 mouse spermatozoa collected in isosmotic (290 mmol/kg) and hyperosmotic (415 mmol/kg) media, and the effect of the osmolality of sperm collection media on sperm fertility after cryopreservation. Our results indicate that murine spermatozoa collected in media with different osmolalities (290 and 415 mmol/kg Dulbecco’s phosphate buffered saline (DPBS)) appeared to have different osmotic tolerances for the maintenance of sperm motility and other motion parameters in both mouse strains. The hypo- and hyperosmotic treatments decreased motility and affected other motion parameters of spermatozoa collected in 290 mmol/kg DPBS. The extent of the change of motion parameters after treatments corresponded with the levels of osmotic stress. However, for spermatozoa collected in 415 mmol/kg DPBS, exposure to 290 mmol/kg DPBS tended to increase sperm motility and the quality of their motion parameters. The osmolality of sperm collection medium can affect murine sperm fertility. Spermatozoa collected in 415 mmol/kg medium showed higher fertility compared with spermatozoa collected in 290 mmol/kg as assessed by IVF. Results characterizing murine sperm osmotic tolerance collected in media with different osmolalities from different strains and the effect of collection media osmolality on sperm fertility after cryopreservation will be useful in designing cryopreservation protocols

Sensitivity Analysis to Reduce Duplicated Features in ANN Training for District Heat Demand Prediction

Artificial neural network (ANN) has become an important method to model the nonlinear relationships between weather conditions, building characteristics and its heat demand. Due to the large amount of training data required for ANN training, data reduction and feature selection are important to simplify the training. However, in building heat demand prediction, many weather-related input variables contain duplicated features. This paper develops a sensitivity analysis approach to analyse the correlation between input variables and to detect the variables that have high importance but contain duplicated features. The proposed approach is validated in a case study that predicts the heat demand of a district heating network containing tens of buildings at a university campus. The results show that the proposed approach detected and removed several unnecessary input variables and helped the ANN model to reduce approximately 20% training time compared with the traditional methods while maintaining the prediction accuracy. It indicates that the approach can be applied for analysing large number of input variables to help improving the training efficiency of ANN in district heat demand prediction and other applications

Prevalence and Correlates of Missed First Appointments among Outpatients at a Psychiatric Hospital in Nigeria

Background: Missed appointments are common in psychiatric practice. It compromises quality of care, results in poor treatment outcomes and drains financial resources. In Nigeria, where mental health services are poorly organized, missed appointments and its resultant consequences may be burdensome.Aim: This study sought to determine the prevalence and factors (sociodemographic and clinical) associated with missed clinic appointments at a regional psychiatric hospital. Subjects and Methods: A study on a cohort of patients attending the Outpatient Clinics for the first time between June and September 2011 was conducted. We interviewed each participant at their first presentation then tracked through case records to determine adherence to scheduled first clinic appointments after 4 weeks. A questionnaire was used in eliciting sociodemographic characteristics, clinical variables, and patient/caregiversatisfaction with treatment. Descriptive statistics were used to summarize the data and inferential statistics to test associations using the SPSS 16. Results: Three hundred and ten patients were recruited over the study period. The prevalence of missed first appointment was 32.6% (101/310). Participants who were single (P = 0.04), living alone (P &lt; 0.01) or aggressive (P &lt; 0.01) were more likely to miss their first appointment. However, having received previous treatment for a psychiatric illness (P = 0.02) and having comorbidity (P = 0.05) was associated with less likelihood to miss a first appointment. A binary logistic regression analysis showed that having received previous treatment independently predicted a less likelihood to miss first appointment (P = 0.03). Conclusion: Quite a proportion (32.6%) of patients attending outpatient clinics miss scheduled clinic appointments. Receiving previous psychiatriccare predicted adherence to scheduled appointment.Keywords: Missed appointments, Nigeria, Patients, Psychiatr