Prevalence of metabolic syndrome in the United States National Health and Nutrition Examination Survey 2011-18.

PURPOSE: To estimate the prevalence of metabolic syndrome (MetS) in the US National Health and Nutrition Examination Survey (NHANES) 2011-18. METHODS: This study included 8183 eligible nonpregnant participants aged ≥20 years from the NHANES 2011-18. MetS was defined as the presence of at least three of the following components: central obesity, reduced high-density lipoprotein cholesterol, elevated triglycerides, elevated blood pressure, and elevated fasting blood glucose. The prevalence of MetS was estimated taking into account the complex sampling. The time trend was evaluated using logistic regression. RESULTS: The total prevalence of MetS increased from 37.6% [95% confidence interval (CI): 34.0%-41.4%] in 2011-12 to 41.8% (95% CI: 38.1%-45.7%) in 2017-18 (P for trend = .028). Among the MetS components, the prevalence of elevated glucose increased from 48.9% (95% CI: 45.7%-52.5%) in 2011-12 to 64.7% (95% CI: 61.4%-67.9%) in 2017-18 (P for trend <.001). The prevalence of MetS in participants with low educational attainment increased from 44.4% (95% CI: 38.8%-50.1%) in 2011-12 to 55.0% (95% CI: 50.8%-59.1%) in 2017-18 (P for trend = .01). CONCLUSION: The prevalence of MetS increased during 2011-18, notably in participants with low educational attainment. Lifestyle modification is needed to prevent MetS and the associated risks of diabetes and cardiovascular disease. Key messages What is already known on this topic: Prevalence of metabolic syndrome is an index of the cardiometabolic health of a population. What this study adds: The prevalence of metabolic syndrome in US adults increased during 2011-18, notably in participants with low educational attainment. How this study might affect research, practice, or policy: Lifestyle modification is needed to prevent metabolic syndrome and the associated risks of diabetes and cardiovascular disease.

Economic evaluation of diagnostic sleep studies for obstructive sleep apnoea in the adult population: a systematic review.

Evidence regarding the cost-effectiveness of limited channel tests compared to laboratory and home polysomnography (PSG) in diagnosing obstructive sleep apnoea (OSA) is unclear. Eligible studies were systematically sought across the following databases: MEDLINE, PsychINFO, SCOPUS, CINAHL, Cochrane, Emcare, Web of Science and ProQuest. Title and abstracts were screened before full-text review. Only full and partial economic evaluations reporting at least one economic outcome were included. A standardised template was used for critical appraisal and data extraction. Relevant findings were summarised using a qualitative approach adhering to economic reporting standards. Literature searches identified 999 non-duplicate abstracts, where 85 studies were retrieved for full-text review. The number of studies that met eligibility criteria and were included in the final analyses was 35, of which 31 investigated Level 3 and four assessed Level 4 tests. Based on the dominance ranking framework, both Level 3 and Level 4 tests were cost-effective compared to PSG. Although study designs and methodologies differ broadly, the findings indicated that using limited channel diagnostic sleep tests for OSA is associated with lower costs and non-inferior health outcomes relative to PSG. Limited channel tests also resulted in shorter waiting times and improved access to diagnostic services for patients with OSA. PROSPERO REGISTRATION NUMBER: CRD42020150130.

In vitro assessment of pathogenicity and virulence encoding gene profiles of avian pathogenic Escherichia coli strains associated with colibacillosis in chickens.

BACKGROUND: Avian pathogenic Escherichia coli (APEC) strains have been associated with various disease conditions in avian species due to virulence attributes associated with the organism. AIMS: This study was carried out to determine the in vitro pathogenic characteristics and virulence encoding genes found in E. coli strains associated with colibacillosis in chickens. METHODS: Fifty-two stock cultures of E. coli strains isolated from chickens diagnosed of colibacillosis were tested for their ability to produce haemolysis on blood agar and take up Congo red dye. Molecular characterization was carried out by polymerase chain reaction (PCR) amplification of virulence encoding genes associated with APEC. RESULTS: Eleven (22%) and 41 (71%) were positive for haemolysis on 5% sheep red blood agar and Congo red agar, respectively. Nine virulence-associated genes were detected as follows: FimH (96%), csgA (52%), iss (48%), iut (33%), tsh (21%), cva (15%), kpsII (10%), pap (2%), and felA (2%). CONCLUSION: The APEC strains exhibited virulence properties and harbored virulence encoding genes which could be a threat to the poultry population and public health. The putative virulence genes were diverse and different in almost all isolate implying that pathogenesis was multi-factorial and the infection was multi-faceted which could be a source of concern in the detection and control of APEC infections.

Economic evaluation of cognitive behavioural therapy for insomnia (CBT-I) for improving health outcomes in adult populations: A systematic review.

Cognitive behavioural therapy for insomnia (CBT-I) is a promising intervention with established efficacy, yet evidence of its cost-effectiveness remains unclear. Systematic searches were conducted in Medline, Psychinfo, ProQuest, Cochrane, Scopus, CINAHL, Web of Science and Emcare. Titles and abstracts were screened against eligibility criteria, and studies reporting full economic evaluations of CBT-I in adult populations were included and examined in detail. Study characteristics were extracted using a standardised template. Quantitative measures and relevant findings were summarised using a qualitative approach following recommended reporting standards. 1,168 non-duplicate articles were identified, of which 44 were selected for full-text review. Seven full economic evaluations of CBT-I in adult populations met the inclusion criteria and were incorporated in the final synthesis. Using the dominance ranking framework to compare cost and outcomes, CBT-I was cost-effective compared to pharmacotherapy or no treatment. The limited number of studies included in this review implies that caution should be exercised when interpreting these results. Future studies are encouraged to employ longer time-horizons and larger sample sizes to enable better determination of sustained cost and outcomes changes. Prospero registration number: CRD42019133554.

Effects of drying on total polyphenols content and antioxidant properties of Carica papaya leaves.

BACKGROUND: Papaya is widely grown in Malaysia and normally only the fruits are consumed. Other parts of the plant such as leaves, roots, bark, peel, seeds and pulp are also known to have medicinal properties and have been used to treat various diseases. Papaya leaves also contain flavonoids, alkaloids phenolic compounds and cynogenetic compounds, and are also reported to be able to treat dengue fever. RESULTS: Studies were carried out on drying of papaya leaves using hot air (60, 70 and 80 °C), shade and freeze drying. Effective diffusivities were estimated ranging from 2.09 × 10-12 to 2.18 × 10-12 m2 s-1 from hot air drying, which are within the order of magnitudes reported for most agricultural and food products. The activation energy to initiate drying showed a relatively low value (2.11 kJ mol-1 ) as a result of the thin leave layer that eased moisture diffusion. In terms of total polyphenols content and antioxidant activities, freeze-dried sample showed a significantly higher (P < 0.05) total polyphenols content [2158 mg gallic acid equivalent 100 g dry weight-1 ] and antioxidant activities [2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) = 571 mg TE 100 g DW-1 and 2,2'-diphenyl-1-picrylhydrazyl (DPPH) = 215 µg mg-1 ] compared to hot air and shade dried samples. CONCLUSION: Freeze dried sample retained the most total polyphenols content and showed the highest antioxidant activities in both ABTS and DPPH antioxidant assays. Hot air and shade drying are not conducive with repect to preserving the antioxidants as a result of possible thermal degradation at elevated temperatures and oxidations under prolonged drying condition. © 2020 Society of Chemical Industry.

Efficacy of Phacoemulsification Alone vs Phacoemulsification With Goniosynechialysis in Patients With Primary Angle-Closure Disease: A Randomized Clinical Trial.

IMPORTANCE: The effectiveness of intraocular pressure (IOP) lowering phacoemulsification combined with goniosynechialysis (GSL) compared with phacoemulsification without GSL remains unknown. OBJECTIVE: To compare the IOP outcome after 1 year in patients with synechial primary angle-closure disease (PACD) and cataract who underwent phacoemulsification with intraocular lens implantation (PEI) alone compared with PEI with GSL (PEI-GSL). DESIGN, SETTING, AND PARTICIPANTS: A multicenter randomized clinical trial was conducted from September 29, 2011, to March 16, 2015; data analysis was performed from April 1, 2015, to March 4, 2019. Patients with PACD, defined as primary angle closure or primary angle-closure glaucoma, and at least 90° peripheral anterior synechiae (PAS) with cataract were included. Patients were randomized to undergo PEI alone or PEI-GSL. Patients were followed up for 1 year with standardized evaluations. Intention-to-treat analysis was performed. INTERVENTIONS: Phacoemulsification with intraocular lens implantation alone or with GSL. MAIN OUTCOMES AND MEASURES: Successful control of IOP at 12 months, defined as IOP 21 mm Hg or lower without use of topical IOP-lowering medications and a decrease in IOP of 20% or more from baseline IOP. RESULTS: Data from 78 patients (78 eyes) were analyzed. Of these, 37 patients were Chinese (47.4%) and 54 were women (69.2%); mean (SD) age was 67.7 (8.9) years. Mean deviation (SD) at baseline was -13.5 dB (9.4 dB). Forty patients were randomized to the PEI group and 38 to the PEI-GSL group. The mean (SD) IOP at baseline was 22.3 (8.5) mm Hg for the PEI group and 22.9 (5.3) mm Hg for the PEI-GSL group. At 1 year, the mean IOP was 14.3 (5.0) mm Hg for the PEI group and 15.9 (4.5) mm Hg for the PEI-GSL group. Successful control at 1 year occurred in 21 patients (52.5%) in the PEI group and 22 patients (57.9%) in the PEI-GSL group (mean difference, 5.4%; 95% CI, -18.0% to 28.2%; P = .63). In eyes that achieved successful control, mean IOP at 1 year was 12.5 (2.7) mm Hg (range, 7.0-19.0) for the PEI group and 13.6 (2.4) mm Hg (range, 9.0-18.0) for the PEI-GSL group. The number of medications at baseline and 1 year decreased from a mean of 2.2 (0.8) to 0.5 (0.9) in the PEI group and 1.9 (0.9) to 0.6 (1.2) in the PEI-GSL group (P < .001 for each), with a mean change difference of 0.4% (95% CI, -0.02% to 0.9%; P = .06). There were 3 postoperative complications (7.5%) in the PEI group and 3 (7.9%) in the PEI-GSL group. These included IOP spike (IOP≥30 mm Hg) (n = 3), excessive anterior chamber inflammation (n = 1), and posterior capsule opacification (n = 2). CONCLUSIONS AND RELEVANCE: This randomized clinical trial was unable to show that PEI-GSL added additional IOP lowering compared with PEI alone in patients with PACD. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT02376725.

The prevalence and characteristics of obstructive sleep apnea in hospitalized patients with type 2 diabetes in China.

Data on the prevalence of obstructive sleep apnea in subjects with type 2 diabetes mellitus in China is scarce. We conducted a multi-centre, cross-sectional study involving 12 hospitals from six regional cities to investigate the prevalence of obstructive sleep apnea in hospitalized patients with type 2 diabetes mellitus and to explore the association between obstructive sleep apnea and related risk factors, diabetic complications and comorbidities in China. Each hospital recruited at least 70 consecutive patients with type 2 diabetes mellitus who were admitted to the endocrinology ward. A total of 880 participants were enrolled and administered overnight sleep monitoring with a portable monitor (ApneaLink™, ResMed, San Diego, CA, USA); other information was collected from medical charts and a standardized questionnaire. In this study, 60.0% (95% confidence interval: 56.8%, 63.2%) of hospitalized patients in China with type 2 diabetes mellitus had comorbid obstructive sleep apnea (apnea-hypopnea index ≥ 5). Only 1.5% (eight of 528) of the patients with both conditions had been diagnosed previously with obstructive sleep apnea. The prevalence of moderate-severe (apnea-hypopnea index ≥ 15) and severe obstructive sleep apnea (apnea-hypopnea index ≥ 30) was estimated to be 25.6% (22.7, 28.5%) and 10.3% (8.3, 12.4%), respectively. Age, sex, body mass index, snoring, reported breath-holding in sleep or gasping or choking arousal, sleepiness, diabetes duration, hypertension, diabetic nephropathy and cardiovascular diseases history were correlated significantly with the severity of obstructive sleep apnea. In China, the prevalence of obstructive sleep apnea in hospitalized patients with type 2 diabetes mellitus is high. Routine screening for and treatment of obstructive sleep apnea is an important, but often neglected, part of the management of diabetes.

Learning clinical procedures through Internet visual resources: a qualitative study amongst undergraduate students.

OBJECTIVE: Acquiring competency in performing clinical procedures is central to professional education of healthcare providers. Internet visual resources (IVR), defined as visual materials openly accessible on public websites, provides a new channel to learn clinical procedures. This qualitative study aimed to profile the experience and opinions of undergraduate students (in dentistry, medicine and nursing) in learning clinical procedures through IVR. METHODS: From clinical degree programmes (Bachelor of Dental Surgery, Bachelor of Medicine, Bachelor of Surgery, and Bachelor of Nursing) of University of Hong Kong, 31 students were recruited to join six focus group discussions, which were transcribed and subjected to thematic analysis using inductive method, in which themes emerge from data. FINDINGS: Students actively looked for IVRs through various means and used them for pre-clinical preparation, post-clinical revision, learning simple and advanced procedures, exploring alternative and updated techniques, and benchmarking against international peers. IVRs were valued for their visual stimulation, inclusion of a wide variety of real-life cases, convenience in access, user-friendliness and time-saving features. Students tended to share and discuss IVRs with their peers rather than with tutors, even when contents deviated from school teaching or faculty's e-learning materials. When doubts persisted, they chose to follow faculty guidelines for examination purpose. Students were frustrated sometimes by difficulties in judging the scientific quality, lack of immediate interactive discussions and loosely structured presentations in some IVRs. Teachers' attitudes towards IVR appeared to vary greatly. CONCLUSION: Despite the wide spectrum of experience and opinions, IVR was generally viewed by undergraduates from across clinical faculties as enhancing their clinical confidence and self-perceived competency, enriching their learning experience and serving as an important supplement to formal learning in the planned curriculum.

Single-molecule analysis reveals human UV-damaged DNA-binding protein (UV-DDB) dimerizes on DNA via multiple kinetic intermediates.

How human DNA repair proteins survey the genome for UV-induced photoproducts remains a poorly understood aspect of the initial damage recognition step in nucleotide excision repair (NER). To understand this process, we performed single-molecule experiments, which revealed that the human UV-damaged DNA-binding protein (UV-DDB) performs a 3D search mechanism and displays a remarkable heterogeneity in the kinetics of damage recognition. Our results indicate that UV-DDB examines sites on DNA in discrete steps before forming long-lived, nonmotile UV-DDB dimers (DDB1-DDB2)2 at sites of damage. Analysis of the rates of dissociation for the transient binding molecules on both undamaged and damaged DNA show multiple dwell times over three orders of magnitude: 0.3-0.8, 8.1, and 113-126 s. These intermediate states are believed to represent discrete UV-DDB conformers on the trajectory to stable damage detection. DNA damage promoted the formation of highly stable dimers lasting for at least 15 min. The xeroderma pigmentosum group E (XP-E) causing K244E mutant of DDB2 found in patient XP82TO, supported UV-DDB dimerization but was found to slide on DNA and failed to stably engage lesions. These findings provide molecular insight into the loss of damage discrimination observed in this XP-E patient. This study proposes that UV-DDB recognizes lesions via multiple kinetic intermediates, through a conformational proofreading mechanism.

De novo ocular hypertension after Descemet stripping endothelial keratoplasty: comparative 3-year incidence, risk factors, and outcomes.

OBJECTIVE: To compare the 3-year incidence of de novo ocular hypertension (OHT) after Descemet stripping automated endothelial keratoplasty (DSAEK) and penetrating keratoplasty (PK). For DSAEK, to evaluate predictors for OHT and 2-year outcomes after OHT development. METHODS: This was a review of the prospective Singapore Corneal Transplant Study at a single tertiary referral center. Consecutive DSAEKs and PKs for Fuchs' endothelial dystrophy (FED) and pseudophakic bullous keratopathy (PBK) in eyes without pre-existing glaucoma were analyzed. OHT incidence after DSAEK and PK were compared using Kaplan-Meier survival analysis, and OHT risk factors identified using Cox proportional regression. OHT was defined: intraocular pressure (IOP) ≥ 24 mmHg or ≥ 10 mmHg from baseline. Secondary outcomes 2 years after OHT development in DSAEK were rates of glaucoma medical therapy failure, IOP success, graft failure and rejection, and best-spectacle corrected visual acuity (BSCVA). RESULTS: There were 108 (96.4%) DSAEKs and 216 (96%) PKs. The 1-, 2- and 3-year de novo OHT incidence was not significantly different between DSAEK (36.1%, 47.2%, 47.2%, respectively) and PK (35.7%, 44.9%, 45.8%, respectively; P = 0.914). OHT incidence did not differ in subgroup analyses of multiple clinical variables (P > 0.1). OHT predictors after DSAEK were: fellow eye glaucoma (hazard ratio [HR] 3.20, P = 0.004), age <60 years (HR 2.41, P = 0.016), concurrent goniosynechiolysis (HR 3.29, P = 0.021), post-graft complications or procedures (HR 2.85, P = 0.006). Two years after OHT onset, 29.7% of DSAEKs failed glaucoma medical therapy requiring trabeculectomy. Complete and qualified IOP success was achieved in 23.5% and 76.5%, respectively. Graft failure developed in 9.8% and graft rejection in 5.9%. At 6 months, 1, and 2 years from OHT onset, 86.3%, 88.3%, and 92.1% achieved BSCVA 20/40, respectively. CONCLUSION: DSAEK and PK have comparable OHT risks. A significant 30% of DSAEK eyes with OHT require filtration surgery. Effective IOP control and good graft and visual outcomes are achieved with treatment.

Activation of transient receptor potential vanilloid 1 decreases endothelial nitric oxide synthase phosphorylation at Thr497 by protein phosphatase 2B-dependent dephosphorylation of protein kinase C.

AIMS: We investigated the effects and underlying molecular mechanism of transient receptor potential vanilloid 1 (TRPV1), a calcium (Ca(2+) )-permeable non-selective cation channel, on phosphorylation of endothelial nitric oxide synthase (eNOS) at threonine 497 (Thr497) in bovine aortic endothelial cells (BAECs) and in mice. METHODS: Western blotting and immunoprecipitation were used for the evaluation of protein phosphorylation; protein phosphatase 2B (PP2B) activity was assessed by convention kit; Griess assay was for NO production; tube formation and Matrigel plug assay were used for angiogenesis. RESULTS: In BAECs, treatment with the TRPV1 ligand evodiamine decreased the phosphorylation of eNOS at Thr497, protein kinase Cα (PKCα) at Serine 657 (Ser657) and PKCß2 at Ser660. Evodiamine increased protein phosphatase 2B (PP2B) activity and promoted the formation of a PP2B-PKC complex. Inhibition of TRPV1 activation by the pharmacological antagonists, removal of extracellular Ca(2+) or pharmacological inhibition of PI3K/Akt/calmodulin-dependent protein kinase II/AMP-activated protein kinase signalling pathway abolished the evodiamine-induced alterations in phosphorylation of eNOS at Thr497, PKCα at Ser657, PKCß2 at Ser660 and PP2B activity, as well as the formation of a PP2B-PKC complex. Inhibition of PP2B activation partially reduced the evodiamine-induced NO bioavailability and tube formation in endothelial cells (ECs) and angiogenesis in mice. Moreover, evodiamine decreased the phosphorylation of eNOS at Thr497, PKCα at Ser657 and PKCß2 at Ser660 in apolipoprotein E (ApoE)-deficient mouse aortas but not TRPV1-deficient or ApoE/TRPV1 double-knockout mice. CONCLUSION: TRPV1 activation in ECs may elicit a Ca(2+) -dependent effect on PP2B-PKC signalling, which leads to dephosphorylation of eNOS at Thr497 in ECs and in mice.

Identification of human-selective analogues of the vascular-disrupting agent 5,6-dimethylxanthenone-4-acetic acid (DMXAA).

BACKGROUND: Species selectivity of DMXAA (5,6-dimethylxanthenone-4-acetic acid, Vadimezan) for murine cells over human cells could explain in part the recent disappointing phase III trials clinical results when preclinical studies were so promising. To identify analogues with greater human clinical potential, we compared the activity of xanthenone-4-acetic acid (XAA) analogues in murine or human cellular models. METHODS: Analogues with a methyl group systematically substituted at different positions of the XAA backbone were evaluated for cytokine induction in cultured murine or human leukocytes; and for anti-vascular effects on endothelial cells on matrigel. In vivo antitumour activity and cytokine production by stromal or cancer cells was measured in human A375 and HCT116 xenografts. RESULTS: Mono-methyl XAA analogues with substitutions at the seventh and eighth positions were the most active in stimulating human leukocytes to produce IL-6 and IL-8; and for inhibition of tube formation by ECV304 human endothelial-like cells, while 5- and 6-substituted analogues were the most active in murine cell systems. CONCLUSION: Xanthenone-4-acetic acid analogues exhibit extreme species selectivity. Analogues that are the most active in human systems are inactive in murine models, highlighting the need for the use of appropriate in vivo animal models in selecting clinical candidates for this class of compounds.

The capillary dysfunction hypothesis of Alzheimer's disease.

It is widely accepted that hypoperfusion and changes in capillary morphology are involved in the etiopathogenesis of Alzheimer's disease (AD). This is difficult to reconcile with the hyperperfusion observed in young high-risk subjects. Differences in the way cerebral blood flow (CBF) is coupled with the local metabolic needs during different phases of the disease can explain this apparent paradox. This review describes this coupling in terms of a model of cerebral oxygen availability that takes into consideration the heterogeneity of capillary blood flow patterns. The model predicts that moderate increases in heterogeneity requires elevated CBF in order to maintain adequate oxygenation. However, with progressive increases in heterogeneity, the resulting low tissue oxygen tension will require a suppression of CBF in order to maintain tissue metabolism. The observed biphasic nature of CBF responses in preclinical AD and AD is therefore consistent with progressive disturbances of capillary flow patterns. Salient features of the model are discussed in the context of AD pathology along with potential sources of increased capillary flow heterogeneity.

Essential role of transient receptor potential vanilloid type 1 in evodiamine-mediated protection against atherosclerosis.

AIM: We investigated whether transient receptor potential vanilloid type 1 (TRPV1) was involved in the therapeutic effect of evodiamine, a main bioactive component in the fruit of Evodiae rutaecarpa, on the development of atherosclerosis in apolipoprotein E-deficient (ApoE(-/-)) mice and ApoE(-/-)TRPV1(-/-) mice. METHODS: Histopathology was examined by haematoxylin and eosin staining, levels of cytokines and mediators were evaluated by ELISA kits, and protein expression was determined by Western blotting. RESULTS: Chronic administration with evodiamine (10 mg kg(-1) body weight) reduced the size of atherosclerotic lesions and alleviated the hyperlipidaemia and systemic inflammation, as well as hepatic macrovesicular steatosis, in ApoE(-/-) mice. Treating ApoE(-/-) mice with evodiamine enhanced hepatic cholesterol clearance, as revealed by upregulation of hepatic low-density lipoprotein receptor and ATP-binding cassette (ABC) transporters ABCG5, ABCG8 and cholesterol 7α-hydrolase. Genetic deletion of TRPV1 in ApoE(-/-) mice promoted the progression of atherosclerosis; elevated the serum levels of cholesterol, cytokines and chemokines; and exacerbated hepatic macrovesicular steatosis. Moreover, genetic deletion of TRPV1 abrogated the evodiamine-evoked atheroprotection but not anti-obesity effect in ApoE(-/-) mice. CONCLUSION: Evodiamine may confer novel TRPV1-dependent atheroprotection and TRPV1-independent anti-obesity action.

Fish in throat: An Unusual Foreign Body.

A 19 year-old man was presented to us in a state of respiratory distress with history of alleged accidentally swallowed the live fish. Flexible nasopharyngolaryngoscope showed a big live fish impacted in the laryngopharynx. Attempts to remove the fish orally were futile as the fish was impacted. We resorted to tracheostomy under local anaesthesia, followed by direct laryngoscopy and removal of the fish under general anaesthesia. The literature review of such rare incidence and approach to such case are discussed.

Damaged DNA induced UV-damaged DNA-binding protein (UV-DDB) dimerization and its roles in chromatinized DNA repair.

UV light-induced photoproducts are recognized and removed by the nucleotide-excision repair (NER) pathway. In humans, the UV-damaged DNA-binding protein (UV-DDB) is part of a ubiquitin E3 ligase complex (DDB1-CUL4A(DDB2)) that initiates NER by recognizing damaged chromatin with concomitant ubiquitination of core histones at the lesion. We report the X-ray crystal structure of the human UV-DDB in a complex with damaged DNA and show that the N-terminal domain of DDB2 makes critical contacts with two molecules of DNA, driving N-terminal-domain folding and promoting UV-DDB dimerization. The functional significance of the dimeric UV-DDB [(DDB1-DDB2)(2)], in a complex with damaged DNA, is validated by electron microscopy, atomic force microscopy, solution biophysical, and functional analyses. We propose that the binding of UV-damaged DNA results in conformational changes in the N-terminal domain of DDB2, inducing helical folding in the context of the bound DNA and inducing dimerization as a function of nucleotide binding. The temporal and spatial interplay between domain ordering and dimerization provides an elegant molecular rationale for the unprecedented binding affinities and selectivities exhibited by UV-DDB for UV-damaged DNA. Modeling the DDB1-CUL4A(DDB2) complex according to the dimeric UV-DDB-AP24 architecture results in a mechanistically consistent alignment of the E3 ligase bound to a nucleosome harboring damaged DNA. Our findings provide unique structural and conformational insights into the molecular architecture of the DDB1-CUL4A(DDB2) E3 ligase, with significant implications for the regulation and overall organization of the proteins responsible for initiation of NER in the context of chromatin and for the consequent maintenance of genomic integrity.

Synergistic effect of p53 on TSA-induced stanniocalcin 1 expression in human nasopharyngeal carcinoma cells, CNE2.

Human stanniocalcin 1 (STC1) has recently been identified as a putative protein factor involved in cellular apoptosis. The use of histone deacetylase inhibitor (i.e. trichostatin A (TSA)) and doxorubicin (Dox) is one of the common treatment methods to induce apoptosis in human cancer cells. A study on TSA and Dox-mediated apoptosis may shed light on the regulation and function of STC1 in cancer treatment. In this study, TSA and Dox cotreatment in human nasopharyngeal carcinoma cells (CNE2) elicited synergistic effects on STC1 gene expression and cellular apoptosis. An activation of p53 (TP53) transcriptional activity in Dox- or Dox+TSA-treated cells was revealed by the increased expression levels of p53 mRNA/protein as well as p53-driven luciferase activities. To elucidate the possible involvement of p53 in STC1 gene transcription, a vector expressing wild-type or dominant negative (DN) p53 was transiently transfected into the cells. Both STC1 promoter luciferase constructs and chromatin immunoprecipitation assays did not support the direct role of p53 in STC1 gene transactivation. However, the synergistic effects of p53 on the induction of NF-κB phosphorylation and the recruitment of acetylated histone H3 in STC1 promoter were observed in TSA-cotreated cells. The overexpression of exogenous STC1 sensitized apoptosis in Dox-treated cells. Taken together, this study provides data to show the cross talk of NF-κB, p53, and histone protein in the regulation of STC1 expression and function.

Dissection of stromal and cancer cell-derived signals in melanoma xenografts before and after treatment with DMXAA.

BACKGROUND: The non-malignant cells of the tumour stroma have a critical role in tumour biology. Studies dissecting the interplay between cancer cells and stromal cells are required to further our understanding of tumour progression and methods of intervention. For proof-of-principle of a multi-modal approach to dissect the differential effects of treatment on cancer cells and stromal cells, we analysed the effects of the stromal-targeting agent 5,6-dimethylxanthenone-4-acetic acid on melanoma xenografts. METHODS: Flow cytometry and multi-colour immunofluorescence staining was used to analyse leukocyte numbers in xenografts. Murine-specific and human-specific multiplex cytokine panels were used to quantitate cytokines produced by stromal and melanoma cells, respectively. Human and mouse Affymetrix microarrays were used to separately identify melanoma cell-specific and stromal cell-specific gene expression. RESULTS: 5,6-Dimethylxanthenone-4-acetic acid activated pro-inflammatory signalling pathways and cytokine expression from both stromal and cancer cells, leading to neutrophil accumulation and haemorrhagic necrosis and a delay in tumour re-growth of 26 days in A375 melanoma xenografts. CONCLUSION: 5,6-Dimethylxanthenone-4-acetic acid and related analogues may potentially have utility in the treatment of melanoma. The experimental platform used allowed distinction between cancer cells and stromal cells and can be applied to investigate other tumour models and anti-cancer agents.

Monoubiquitinated histone H2A destabilizes photolesion-containing nucleosomes with concomitant release of UV-damaged DNA-binding protein E3 ligase.

How the nucleotide excision repair (NER) machinery gains access to damaged chromatinized DNA templates and how the chromatin structure is modified to promote efficient repair of the non-transcribed genome remain poorly understood. The UV-damaged DNA-binding protein complex (UV-DDB, consisting of DDB1 and DDB2, the latter of which is mutated in xeroderma pigmentosum group E patients, is a substrate-recruiting module of the cullin 4B-based E3 ligase complex, DDB1-CUL4B(DDB2). We previously reported that the deficiency of UV-DDB E3 ligases in ubiquitinating histone H2A at UV-damaged DNA sites in the xeroderma pigmentosum group E cells contributes to the faulty NER in these skin cancer-prone patients. Here, we reveal the mechanism by which monoubiquitination of specific H2A lysine residues alters nucleosomal dynamics and subsequently initiates NER. We show that DDB1-CUL4B(DDB2) E3 ligase specifically binds to mononucleosomes assembled with human recombinant histone octamers and nucleosome-positioning DNA containing cyclobutane pyrimidine dimers or 6-4 photoproducts photolesions. We demonstrate functionally that ubiquitination of H2A Lys-119/Lys-120 is necessary for destabilization of nucleosomes and concomitant release of DDB1-CUL4B(DDB2) from photolesion-containing DNA. Nucleosomes in which these lysines are replaced with arginines are resistant to such structural changes, and arginine mutants prevent the eviction of H2A and dissociation of polyubiquitinated DDB2 from UV-damaged nucleosomes. The partial eviction of H3 from the nucleosomes is dependent on ubiquitinated H2A Lys-119/Lys-120. Our results provide mechanistic insight into how post-translational modification of H2A at the site of a photolesion initiates the repair process and directly affects the stability of the human genome.

Gray matter nulled and vascular space occupancy dependent fMRI response to visual stimulation during hypoxic hypoxia.

Two cerebral blood volume (CBV)-weighted fMRI techniques, gray matter nulled (GMN) and vascular space occupancy (VASO)-dependent techniques at spatial resolution of 2 × 2 × 5 mm(3), were compared in the study investigating functional responses in the human visual cortex to stimulation in normoxia (inspired O(2) = 21%) and mild hypoxic hypoxia (inspired O(2) = 12%). GMN and VASO signals and T(2)* were quantified in activated voxels. While the CBV-weighted signal changes in voxels activated by visual stimulation were similar in amplitude in both fMRI techniques in both oxygenation conditions, the number of activated voxels during hypoxic hypoxia was significantly reduced by 72 ± 22% in GMN fMRI and 66 ± 23% in VASO fMRI. T(2)* prolonged in GMN and VASO activated voxels in normoxia by 1.6 ± 0.5 ms and 1.7 ± 0.5 ms, respectively. In hypoxia, however, T(2)* shortened in GMN-activated voxels by 0.7 ± 0.6 ms (p < 0.001 relative to normoxia), but prolonged in VASO-activated ones by 1.1 ± 0.6 ms (p < 0.05 relative to normoxia). The data show that the hemodynamic responses to visual stimulation were not affected by hypoxic hypoxia, but T(2)* increases by both CBV-weighted fMRI techniques were smaller in activated voxels in hypoxia. The mechanisms influencing GMN fMRI signal in both oxygenation conditions were explored by simulating effects of the oxygen extraction fraction (OEF) and partial voluming with cerebral spinal fluid (CSF) and white matter in imaging voxels. It is concluded that while GMN fMRI data point to increased, rather than decreased OEF during visual stimulation in hypoxia, partial voluming by CSF is likely to affect the CBV quantification by GMN fMRI under the experimental conditions used.