Spatial distribution and health risks assessment of heavy metals in e-waste dumping sites from Pakistan.

The present study focused on to determine the concentration and health risk of heavy metals (Cu, Pb, Zn, Cd, Hg, Cr) in e-waste contaminated soils collected from different provinces of Pakistan. Further, the impact of heavy metals on soil enzyme activities and microbial community was also investigated. The concentration (mg/kg) of Hg, Zn, Fe, Cu, Pb, Cd, and Cr ranged between 0-0.258, 2.284-6.587, 3.005-40.72, 8.67-36.88, 12.05-35.03, 1.03-2.43, and 33.13-60.05, respectively. The results revealed that Lahore site of Punjab province indicated more concentration of heavy metals as compared to other sites. The level of Cr at all sites whereas Hg at only two sites exceeds the World Health Organization standards (WHO) for soil. Soil enzyme activity exhibited dynamic trend among the sites. Maximum enzyme activity was observed for urease followed by phosphatase and catalase. Contamination factor (Cf), Pollution load index (PLI), and geo-accumulation index (Igeo) results showed that all the sites are highly contaminated with Cu, Cd, and Pb. Hazard index (HI) was less than 1 for children and adults suggesting non-carcinogenic health risk. Principle component analysis results depicted relation among Cr, Fr, catalase, and actinomycetes; Cd, OM, urease, and bacteria, and Pb, Cu, Zn, Hg, and phosphatase, suggesting soil enzymes and microbial community profiles were influenced by e-waste pollution. Therefore, there is a dire need to introduce sustainable e-waste recycling techniques as well as to make stringent e-waste management policies to reduce further environmental contamination.

Predicting ambient PM2.5 concentrations via time series models in Anhui Province, China.

Due to rapid expansion in the global economy and industrialization, PM2.5 (particles smaller than 2.5 µm in aerodynamic diameter) pollution has become a key environmental issue. The public health and social development directly affected by high PM2.5 levels. In this paper, ambient PM2.5 concentrations along with meteorological data are forecasted using time series models, including random forest (RF), prophet forecasting model (PFM), and autoregressive integrated moving average (ARIMA) in Anhui province, China. The results indicate that the RF model outperformed the PFM and ARIMA in the prediction of PM2.5 concentrations, with cross-validation coefficients of determination R2, RMSE, and MAE values of 0.83, 10.39 µg/m3, and 6.83 µg/m3, respectively. PFM achieved the average results (R2 = 0.71, RMSE = 13.90 µg/m3, and MAE = 9.05 µg/m3), while the predicted results by ARIMA are comparatively poorer (R2 = 0.64, RMSE = 15.85 µg/m3, and MAE = 10.59 µg/m3) than RF and PFM. These findings reveal that the RF model is the most effective method for predicting PM2.5 and can be applied to other regions for new findings.

Endoscopic pilonidal sinus treatment (EPSiT) for sacrococcygeal pilonidal sinus disease: A prospective long-term study.

Objectives: To assess short-term and long-term outcomes of endoscopic pilonidal sinus treatment for pilonidal sinus disease. METHODS: The prospective study was conducted at Shifa International Hospital, Islamabad, Pakistan, from July 2015 to July 2021, and comprised all pilonidal sinus cases undergoing minimal invasive endoscopic pilonidal sinus treatment who were treated by a single surgical team. The primary outcomes were duration of healing, post-operative morbidities, persistence of discharge and recurrence at 1-7 years. The secondary outcomes were operative time, return to work, cosmetic results and patient satisfaction. The patients were observed for wound healing and discharge on follow-up in the out-patient department at 1, 3, 6 and 24 weeks. They were further followed up every year through telephonic survey for persistence or recurrence of symptoms. Patient satisfaction was assessed using the 36-item Short Form Survey questionnaire filled at admission and then at 6 weeks post-surgery. Data was analysed using SPSS 23. RESULTS: Of the 67 patients, 55(82%) were males and 12(18%) were females. The overall mean age was 25.69±8.305 years. There were 13(19.4%) patients with a history of recurrent disease and previous procedures for pilonidal sinus, while 54(80.6%) had no previous surgery. The median operative time was 35 minutes (interquartile range: 20-45 minutes). Complete wound healing was achieved in 60(89.6%) patients, while recurrence was seen in 7(10.4%). The median time off work was 2.5 days (interquartile range: 1-3 days). Patient satisfaction with the procedure was significantly high (p<0.05). CONCLUSIONS: Endoscopic pilonidal sinus treatment appeared to be a good minimally invasive surgical technique for the treatment of pilonidal sinus disease in terms of both short-term and long-term outcomes.

Co-delivery of hesperidin and clarithromycin in a nanostructured lipid carrier for the eradication of Helicobacter pylori in vitro.

Effective and precise eradication of Helicobacter pylori (H. pylori) is the most promising approach to avoid H. pylori-related gastrointestinal disorders. The present study was conducted to demonstrate the efficacy of the co-delivery of hesperidin (Hesp) and clarithromycin (CLR) in nanostructured lipid carriers (NLCs) against H. pylori. We have produced a new delivery system by combining bioflavonoid Hesp and CLR NLCs to address the failure in single antibiotic therapies. Briefly, a blend of solid lipid, liquid lipid, and surfactant was used. Homogeneous NLCs with all the formulations showed a nano size and surface-negative charge and presented high in vitro stability and slow release of the drug even after 24 h. Bioimaging studies by scanning electron microscopy, transmission electron microscopy, and imaging flow cytometry indicated that NLCs interacted with the membrane by adhering to the outer cell membrane and disrupted the membrane that resulted in the leakage of cytoplasmic contents. The prepared NLCs provide sustained and controlled drug release that can be used to increase the rate of H. pylori eradication.

Effect of smoking on periodontal health and validation of self-reported smoking status with serum cotinine levels.

OBJECTIVE: To investigate whether self-reported smoking and serum cotinine levels associate with periodontal pocket development and to determine the accuracy of self-reported smoking using serum cotinine. MATERIALS AND METHODS: This 4-year prospective cohort study included data from 294 dentate adults, aged ≥30 years, who participated in both the Health 2000 Survey and the Follow-up Study of Finnish Adults' Oral Health. Subjectively reported smoking status (daily smokers n = 62, occasional smokers n = 12, quitters n = 49, and never-smokers n = 171), serum cotinine levels, demographic factors, education level, dental behaviours and medical history were collected at baseline. The outcome measure was the number of teeth with periodontal pocketing ≥4 mm over 4 years. RESULTS: Self-reported daily smokers had 1.82 (95% CI: 1.32-2.50) higher incidence of deepened periodontal pockets than never-smokers. A positive association was observed between serum cotinine (≥42.0 µg/L) and the development of periodontal pockets. The misclassification rate of self-reported smoking was 6%. CONCLUSIONS: Both self-reported daily smoking and higher serum cotinine were associated with periodontal pocket development. Self-reported smoking was fairly accurate in this study. However, higher cotinine levels among a few self-reported never-smokers indicated misreporting or passive smoking. Thus, self-reports alone are not enough to assess the smoking-attributable disease burden.

Mucocoele of the appendix: A case report.

Mucinous cystadenoma is a rare tumour of the appendix. It accounts for only 0.4% of the gastrointestinal tract malignancies and is reported rarely in the literature. Therefore, surgical management is not yet established. Here we report the case of a 65-year-old female who presented with a dragging sensation and a feeling of a mass in the right iliac fossa. Her computed tomography (CT) suggested formation of an abscess in the parietal peritoneum. She was scheduled for laparotomy and upon exploration, a mass was found arising from the tip of the retroperitoneal appendix. The whole of the appendix was studded with mucoid material. Right hemicolectomy was performed and histopathology of the appendix showed mucinous cystadenoma with no evidence of malignant changes. The recovery was uneventful and the patient was discharged on the fourth post-operative day. The unusual presentation of retroperitoneal pseudomyxoma without any intraperitoneal pathology prompted us to report this case.

Educational interventions for preventing lead poisoning in workers.

BACKGROUND: Occupational lead exposure can lead to serious health effects that range from general symptoms (depression, generalised ache, and digestive signs, such as loss of appetite, stomach ache, nausea, diarrhoea, and constipation) to chronic conditions (cerebrovascular and cardiovascular diseases, cognitive impairment, kidney disease, cancers, and infertility). Educational interventions may contribute to the prevention of lead uptake in workers exposed to lead, and it is important to assess their effectiveness. OBJECTIVES: To assess the effect of educational interventions for preventing lead uptake in workers exposed to lead. SEARCH METHODS: We searched CENTRAL, MEDLINE, Embase, CINAHL, and OSH UPDATE to 5 June 2020, with no language restrictions. SELECTION CRITERIA: We sought randomised controlled trials (RCT), cluster-RCTs (cRCT), interrupted time series (ITS), controlled before-after studies (CBA) and uncontrolled before-after studies that examined the effects of an educational intervention aimed at preventing lead exposure and poisoning in workers who worked with lead, for which effectiveness was measured by lead levels in blood and urine, blood zinc protoporphyrin levels and urine aminolevulinic acid levels. DATA COLLECTION AND ANALYSIS: Two review authors independently screened the search results, assessed studies for eligibility, and extracted data using standard Cochrane methods. We used the ROBINS-I tool to assess the risk of bias, and GRADE methodology to assess the certainty of the evidence. MAIN RESULTS: We did not find any RCT, cRCT, ITS or CBA studies that met our criteria. We included four uncontrolled before-after studies studies, conducted between 1982 and 2004. Blood lead levels Educational interventions may reduce blood lead levels, but the evidence is very uncertain. In the short-term after the educational intervention, blood lead levels may decrease (mean difference (MD) 9.17 µg/dL, 95% confidence interval (CI) 4.14 to 14.20; one study with high baseline blood lead level, 18 participants; very low-certainty evidence). In the medium-term, blood lead levels may decrease (MD 3.80 µg/dL, 95% CI 1.48 to 6.12; one study with high baseline blood lead level, 34 participants; very low-certainty evidence). In the long-term, blood lead levels may decrease when the baseline blood lead levels are high (MD 8.08 µg/dL; 95% CI 3.67 to 12.49; two studies, 69 participants; very low-certainty evidence), but not when the baseline blood lead levels are low (MD 1.10 µg/dL, 95% CI -0.11 to 2.31; one study, 52 participants, very low-certainty evidence). Urine lead levels In the long-term, urinary lead levels may decrease after the educational intervention, but the evidence is very uncertain (MD 42.43 µg/L, 95% CI 29.73 to 55.13; one study, 35 participants; very low-certainty evidence). Behaviour change The evidence is very uncertain about the effect of educational intervention on behaviour change. At medium-term follow-up after the educational intervention, very low-certainty evidence from one study (89 participants) found inconclusive results for washing before eating (risk ratio (RR) 1.71, 95% CI 0.42 to 6.91), washing before drinking (RR 1.37, 95% CI 0.61 to 3.06), and not smoking in the work area (RR 1.04, 95% CI 0.74 to 1.46). Very low-certainty evidence from one study (21 participants) suggested that employers may improve the provision of fit testing for all respirator users (RR 1.87, 95% CI 1.16 to 3.01), and prohibit eating, drinking, smoking, and other tobacco use in the work area (RR 4.25, 95% CI 1.72 to 10.51), however, the results were inconclusive for the adequate provision of protective clothing (RR 1.40, 95% CI 0.82 to 2.40). At long-term follow-up, very low-certainty evidence from one study (89 participants) suggested that workers may improve washing before drinking (RR 3.24, 95% CI 1.09 to 9.61), but results were inconclusive for washing before eating (RR 11.71, 95% CI 0.66 to 208.33), and for not smoking in the work area (RR 1.56, 95% CI 0.98 to 2.50). Very low-certainty evidence from one study (21 participants) suggested that employers may improve the provision of fit testing for all respirator users (RR 1.70, 95% CI 1.09 to 2.63), may provide adequate protective clothing (RR 2.80, 95% CI 1.23 to 6.37), and may prohibit eating, drinking, smoking, and other tobacco use in the work area (RR 2.13, 95% CI 1.19 to 3.81). Improved knowledge or awareness of the adverse health effects of lead The evidence is very uncertain about the effect of educational intervention on workers' knowledge. At medium-term follow-up, questionnaires found that workers' knowledge may improve (MD 5.20, 95% CI 3.29 to 7.11; one study, 34 participants; very low-certainty evidence). At long-term follow-up, there may be an improvement in workers' knowledge (MD 5.80, 95% CI 3.89 to 7.71; one study, 34 participants; very low-certainty evidence), but results were inconclusive for employers' knowledge (RR 1.67, 95% CI 0.74 to 3.75; one study, 21 participants; very low-certainty evidence). None of the studies measured the other outcomes of interest: blood zinc protoporphyrin levels, urine aminolevulinic acid levels, air lead levels, and harms. One study provided the costs of each component of the intervention. AUTHORS' CONCLUSIONS: Educational interventions may prevent lead poisoning in workers with high baseline blood lead levels and urine lead levels but this is uncertain. Educational interventions may not prevent lead poisoning in workers with low baseline blood lead levels but this is uncertain.

Alcohol use and periodontal pocket development: findings from a 4-yr longitudinal study.

This study investigated whether alcohol use influences periodontal pocket development during a 4-yr follow-up period. The study included those participants who took part in both the Health 2000 Survey and the Follow-up Study on Finnish Adults' Oral Health. The participants at baseline were aged ≥30 yr, periodontally healthy, and did not have diabetes or rheumatoid arthritis. The development of periodontal pockets at follow-up was measured as the number of teeth with periodontal pockets and the presence of periodontal pockets. Alcohol use at baseline was measured as g/wk, frequency, and use over the risk limit. Incidence rate ratios with 95% CI were estimated using negative binomial regression models and Poisson regression models with a robust variance estimator. No consistent association was found between any of the alcohol variables and periodontal pocket development in the total population or among non-smokers. Among smokers, a positive association was found with the frequency of alcohol use. In general, risk estimates were slightly higher for women than for men. In summary, light-to-moderate alcohol use appears not to be consistently associated with the development of periodontal pockets. The adverse effects on the periodontium seem, to some extent, to be dependent on gender and smoking.

Associations of Cardiorespiratory Fitness and Adiposity With Arterial Stiffness and Arterial Dilatation Capacity in Response to a Bout of Exercise in Children.

Purpose: To investigate the associations of directly measured peak oxygen uptake ( V˙O2peak ) and body fat percentage (BF%) with arterial stiffness and arterial dilatation capacity in children. Methods: Findings are based on 329 children (177 boys and 152 girls) aged 8-11 years. V˙O2peak was assessed by a maximal cardiopulmonary exercise test on a cycle ergometer and scaled by lean body mass (LM). BF% and LM were measured by bioelectrical impedance. Stiffness index (measure of arterial stiffness) and change in reflection index (ΔRI, measure of arterial dilatation capacity) were assessed by pulse contour analysis. Data were analyzed by linear regression models. Results:V˙O2peak/LM was positively associated with ΔRI in boys adjusted for age and BF% (ß = 0.169, P = .03). Further adjustments for systolic blood pressure, heart rate, and the study group had no effect on this association, but additional adjustment for clinical puberty attenuated it (ß = 0.171, P = .07). BF% was inversely related to ΔRI in boys adjusted for age and V˙O2peak/LM (ß = -0.171, P = .03). V˙O2peak or BF% was not associated with ΔRI in girls or with stiffness index in either boys or girls. Conclusion: Increasing cardiorespiratory fitness and decreasing adiposity may improve arterial health in childhood, especially among boys.

Identification of proteins responding to pathogen-infection in the red alga Pyropia yezoensis using iTRAQ quantitative proteomics.

BACKGROUND: Pyropia yezoensis is an important marine crop which, due to its high protein content, is widely used as a seafood in China. Unfortunately, red rot disease, caused by Pythium porphyrae, seriously damages P. yezoensis farms every year in China, Japan, and Korea. Proteomic methods are often used to study the interactions between hosts and pathogens. Therefore, an iTRAQ-based proteomic analysis was used to identify pathogen-responsive proteins following the artificial infection of P. yezoensis with P. porphyrae spores. RESULTS: A total of 762 differentially expressed proteins were identified, of which 378 were up-regulated and 384 were down-regulated following infection. A large amount of these proteins were involved in disease stress, carbohydrate metabolism, cell signaling, chaperone activity, photosynthesis, and energy metabolism, as annotated in the KEGG database. Overall, the data showed that P. yezoensis resists infection by inhibiting photosynthesis, and energy and carbohydrate metabolism pathways, as supported by changes in the expression levels of related proteins. The expression data are available via ProteomeXchange with the identifier PXD009363. CONCLUSIONS: The current data provide an overall summary of the red algae responses to pathogen infection. This study improves our understanding of infection resistance in P. yezoensis, and may help in increasing the breeding of P. porphyrae-infection tolerant macroalgae.

Unusual presentation of infective endocarditis during pregnancy: A case report.

Infective endocarditis (IE) is uncommon but a very serious infection during pregnancy. In most cases, the disease tends to run a subacute course and involves the mitral valve. We present the case of a 25-year old pregnant female who developed shortness of breath and fever 2 weeks prior to parturition. The symptoms did not subside after her delivery, which was carried out via c-section. Based on vegetations attached to aortic valve cusps and positive blood culture for staphylococcus Aureus, the diagnosis of infective endocarditis was made.

Non-nuclear estrogen receptor alpha activation in endothelium reduces cardiac ischemia-reperfusion injury in mice.

Steroid hormone receptors including estrogen receptors (ER) classically function as ligand-regulated transcription factors. However, estrogens also elicit cellular effects through binding to extra-nuclear ER (ERα, ERß, and G protein-coupled ER or GPER) that are coupled to kinases. How extra-nuclear ER actions impact cardiac ischemia-reperfusion (I/R) injury is unknown. We treated ovariectomized wild-type female mice with estradiol or an estrogen-dendrimer conjugate (EDC), which selectively activates extra-nuclear ER, or vehicle interventions for two weeks. I/R injury was then evaluated in isolated Langendorff perfused hearts. Two weeks of treatment with estradiol significantly decreased infarct size and improved post-ischemic contractile function. Similarly, EDC treatment significantly decreased infarct size and increased post-ischemic functional recovery compared to vehicle-treated hearts. EDC also caused an increase in myocardial protein S-nitrosylation, consistent with previous studies showing a role for this post-translational modification in cardioprotection. In further support of a role for S-nitrosylation, inhibition of nitric oxide synthase, but not soluble guanylyl cyclase blocked the EDC mediated protection. The administration of ICI182,780, which is an agonist of G-protein coupled estrogen receptor (GPER) and an antagonist of ERα and ERß, did not result in protection; however, ICI182,780 significantly blocked EDC-mediated cardioprotection, indicating participation of ERα and/or ERß. In studies determining the specific ER subtype and cellular target involved, EDC decreased infarct size and improved functional recovery in mice lacking ERα in cardiomyocytes. In contrast, protection was lost in mice deficient in endothelial cell ERα. Thus, extra-nuclear ERα activation in endothelium reduces cardiac I/R injury in mice, and this likely entails increased protein S-nitrosylation. Since EDC does not stimulate uterine growth, in the clinical setting EDC-like compounds may provide myocardial protection without undesired uterotrophic and cancer-promoting effects.

Knowledge of scientific misconduct in publication among medical students.

BACKGROUND: Publication is a central element in research dissemination and scientific misconduct in publication is relatively ignored in biomedical research. This study is to assess the knowledge of scientific misconduct in publication among private and public sector medical students. METHODS: A questionnaire-based cross-sectional study was carried in four (two public and two private) medical colleges of Karachi in 2015. After ethical approval, data were collected through convenient sampling and analyzed in SPSS 16.0. Descriptive statistics was used to summarize the data and Chi-square test was used for cross tabulation with sex, type of medical colleges, and knowledge of scientific misconduct in publication. RESULTS: A total of 592 medical students participated with mean age of 22.2 ± 1.47 years. The majority (491, 79%) of medical students had heard about the word "publication ethics," higher among public sector students than from private sector (P < 0.001). Only 78 (13.2%) reported to had published original articles, and 64 (10.8%) and 53 (9%) medical students had heard of "ICMJE authorship criteria" and "COPE," respectively. Knowledge about fabrication of data and scientific misconduct in publication was found to be statistically significant (P < 0.05) among males than female students. Statistically significant differences were also observed between public and private medical students for knowledge regarding salami slicing, ghost author, fabrication, and photomanipulation (P < 0.001) and for plagiarism (P < 0.005). Participants from public sector colleges scored significantly better in all above variables than private medical colleges except knowledge about salami slicing in which participants from latter performed significantly better than public sector students. DISCUSSION: The study demonstrates deficiencies in knowledge regarding several aspects of publication ethics among medical students of both public and private medical colleges in Karachi. There is a need to increase the awareness of research and publication ethics among students during their academic years.

FECALITH IN THE ILEUM CAUSING INTESTINAL OBSTRUCTION.

Small bowel obstruction is one of the common conditions presenting in surgical wards, however fecalith is one of the rare causes of bowel obstruction. We present here a case of 65 years old lady, who presented with sub-acute intestinal obstruction. In spite of the initial diagnosis being sigmoid volvulus, exploratory laparotomy revealed a focolith in ileum which was retrieved through an enterotomy and primary closure was done. The patient recovered uneventfully. Thus emphasizing the need of through history and workup which steer us to the correct diagnosis.

Artificial sweeteners: safe or unsafe?

Artificial sweeteners or intense sweeteners are sugar substitutes that are used as an alternative to table sugar. They are many times sweeter than natural sugar and as they contain no calories, they may be used to control weight and obesity. Extensive scientific research has demonstrated the safety of the six low-calorie sweeteners currently approved for use in foods in the U.S. and Europe (stevia, acesulfame-K, aspartame, neotame, saccharin and sucralose), if taken in acceptable quantities daily. There is some ongoing debate over whether artificial sweetener usage poses a health threat .This review article aims to cover thehealth benefits, and risks, of consuming artificial sweeteners, and discusses natural sweeteners which can be used as alternatives.

Prevalence of hepatitis D virus infection in Central Italy has remained stable across the last 2 decades with dominance of subgenotypes 1 and characterized by elevated viral replication.

OBJECTIVES: Here we investigate Hepatitis D virus (HDV)-prevalence in Italy and its fluctuations over time and we provide an extensive characterization of HDV-infected patients. METHODS: The rate of HDV seroprevalence and HDV chronicity was assessed in 1579 hepatitis B surface antigen (HBsAg)+ patients collected from 2005 to 2022 in Central Italy. RESULTS: In total, 45.3% of HBsAg+ patients received HDV screening with an increasing temporal trend: 15.6% (2005-2010), 45.0% (2011-2014), 49.4% (2015-2018), 71.8% (2019-2022). By multivariable model, factors correlated with the lack of HDV screening were alanine-aminotransferase (ALT) less than two times of upper limit of normality (<2ULN) and previous time windows (P <0.002). Furthermore, 13.4% of HDV-screened patients resulted anti-HDV+ with a stable temporal trend. Among them, 80.8% had detectable HDV-ribonucleic acid (RNA) (median [IQR]:4.6 [3.6-5.6] log copies/ml) with altered ALT in 89.3% (median [IQR]:92 [62-177] U/L). Anti-HDV+ patients from Eastern/South-eastern Europe were younger than Italians (44 [37-54] vs 53 [47-62] years, P <0.0001), less frequently nucleos(t)ide analogs (NUC)-treated (58.5% vs 80%, P = 0.026) with higher HDV-RNA (4.8 [3.6-5.8] vs 3.9 [1.4-4.9] log copies/ml, P = 0.016) and HBsAg (9461 [4159-24,532] vs 4447 [737-13,336] IU/ml, P = 0.032). Phylogenetic analysis revealed the circulation of HDV subgenotype 1e (47.4%) and -1c (52.6%). Notably, subgenotype 1e correlated with higher ALT than 1c (168 [89-190] vs 58 [54-88] U/l, P = 0.015) despite comparable HDV-RNA. CONCLUSIONS: HDV-screening awareness is increasing over time even if some gaps persist to achieve HDV screening in all HBsAg+ patients. HDV prevalence in tertiary care centers tend to scarcely decline in native/non-native patients. Detection of subgenotypes, triggering variable inflammatory stimuli, supports the need to expand HDV molecular characterization.

Regulated clearance of histone deacetylase 3 protects independent formation of nuclear receptor corepressor complexes.

An important step in transcriptional regulation by corepressors N-CoR and SMRT is the formation of a stable and active histone deacetylase 3 (HDAC3)-containing complex. Although N-CoR and SMRT are thought to bind HDAC3 competitively, multiple studies have shown that they do not interfere with the function of each other. How this functional independence is sustained under the competitive interaction is unclear. Here, we show that the coupling of corepressor expression with HDAC3 degradation allows cells to maintain a stable level of uncomplexed HDAC3, thereby preventing mutual interference in the assembly of N-CoR and SMRT complexes. The free uncomplexed HDAC3 is highly unstable. Unexpectedly, the rate of HDAC3 degradation is inversely correlated with the expression level of corepressors. Our results indicate that reducing one corepressor accelerates HDAC3 clearance, thus preventing an increase in complex formation between HDAC3 and the other corepressor. In addition, this study also indicates that the formation of a stable and active HDAC3-corepressor complex is a stepwise process in which the C terminus of HDAC3 plays a critical role at late steps of the assembly process.

Key roles for MED1 LxxLL motifs in pubertal mammary gland development and luminal-cell differentiation.

Mediator recently has emerged as a central player in the direct transduction of signals from transcription factors to the general transcriptional machinery. In the case of nuclear receptors, in vitro studies have shown that the transcriptional coactivator function of the Mediator involves direct ligand-dependent interactions of the MED1 subunit, through its two classical LxxLL motifs, with the receptor AF2 domain. However, despite the strong in vitro evidence, there currently is little information regarding in vivo functions of the LxxLL motifs either in MED1 or in other coactivators. Toward this end, we have generated MED1 LxxLL motif-mutant knockin mice. Interestingly, these mice are both viable and fertile and do not exhibit any apparent gross abnormalities. However, they do exhibit severe defects in pubertal mammary gland development. Consistent with this phenotype, as well as loss of the strong ligand-dependent estrogen receptor (ER)alpha-Mediator interaction, expression of a number of known ERalpha-regulated genes was down-regulated in MED1-mutant mammary epithelial cells and could no longer respond to estrogen stimulation. Related, estrogen-stimulated mammary duct growth in MED1-mutant mice was also greatly diminished. Finally, additional studies show that MED1 is differentially expressed in different types of mammary epithelial cells and that its LxxLL motifs play a role in mammary luminal epithelial cell differentiation and progenitor/stem cell determination. Our results establish a key nuclear receptor- and cell-specific in vivo role for MED1 LxxLL motifs, through Mediator-ERalpha interactions, in mammary gland development.

Mechanisms of estrogen receptor antagonism toward p53 and its implications in breast cancer therapeutic response and stem cell regulation.

Estrogen receptor alpha (ERalpha) plays an important role in the onset and progression of breast cancer, whereas p53 functions as a major tumor suppressor. We previously reported that ERalpha binds to p53, resulting in inhibition of transcriptional regulation by p53. Here, we report on the molecular mechanisms by which ERalpha suppresses p53's transactivation function. Sequential ChIP assays demonstrated that ERalpha represses p53-mediated transcriptional activation in human breast cancer cells by recruiting nuclear receptor corepressors (NCoR and SMRT) and histone deacetylase 1 (HDAC1). RNAi-mediated down-regulation of NCoR resulted in increased endogenous expression of the cyclin-dependent kinase (CDK)-inhibitor p21(Waf1/Cip1) (CDKN1A) gene, a prototypic transcriptional target of p53. While 17beta-estradiol (E2) enhanced ERalpha binding to p53 and inhibited p21 transcription, antiestrogens decreased ERalpha recruitment and induced transcription. The effects of estrogen and antiestrogens on p21 transcription were diametrically opposite to their known effects on the conventional ERE-containing ERalpha target gene, pS2/TFF1. These results suggest that ERalpha uses dual strategies to promote abnormal cellular proliferation: enhancing the transcription of ERE-containing proproliferative genes and repressing the transcription of p53-responsive antiproliferative genes. Importantly, ERalpha binds to p53 and inhibits transcriptional activation by p53 in stem/progenitor cell-containing murine mammospheres, suggesting a potential role for the ER-p53 interaction in mammary tissue homeostasis and cancer formation. Furthermore, retrospective studies analyzing response to tamoxifen therapy in a subset of patients with ER-positive breast cancer expressing either wild-type or mutant p53 suggest that the presence of wild-type p53 is an important determinant of positive therapeutic response.