DDX11 was recently identified as a cause of Warsaw breakage syndrome (WABS). However, the functional mechanism of DDX11 and the contribution of clinically described mutations to the pathogenesis of WABS are elusive. Here, we show that DDX11 is a novel nucleolar protein that preferentially binds to hypomethylated active ribosomal DNA (rDNA) gene loci, where it interacts with upstream binding factor (UBF) and the RNA polymerase I (Pol I). DDX11 knockdown changed the epigenetic state of rDNA loci from euchromatic structures to more heterochromatic structures, reduced the activity of UBF, decreased the recruitment of UBF and RPA194 (a subunit of Pol I) to rDNA promoter, suppressed rRNA transcription and thereby inhibited growth and proliferation of HeLa cells. Importantly, two indentified WABS-derived mutants, R263Q and K897del, and a Fe-S deletion construct demonstrated significantly reduced binding abilities to rDNA promoters and lowered DNA-dependent ATPase activities compared with wild-type DDX11. Knockdown of the zebrafish ortholog of human DDX11 by morpholinos resulted in growth retardation and vertebral and craniofacial malformations in zebrafish, concomitant with the changes in histone epigenetic modifications at rDNA loci, the reduction of Pol I recruitment to the rDNA promoter and a significant decrease in nascent pre-RNA levels. These growth disruptions in zebrafish in response to DDX11 reduction showed similarities to the clinically described developmental abnormalities found in WABS patients for the first time in any vertebrate. Thus, our results indicate that DDX11 functions as a positive regulator of rRNA transcription and provides a novel insight into the pathogenesis of WABS.

Oxidative stress is a major risk factor for neurodegenerative disease. The Kelch-like ECH-associated protein 1 (Keap1)−nuclear factor erythroid 2 related factor 2 (Nrf2)−antioxidant response element (ARE) pathway is one of the most potent defensive systems against oxidative stress. Selenepezil, a selenium-based compound, was previously found to exhibit excellent acetylcholinesterase (AChE) inhibition, to mimic endogenous glutathione peroxidase (GPx) activity, and to exhibit scavenging activity for hydrogen peroxide in vitro. However, none of these activities have been evaluated in a cellular model, and detailed molecular mechanisms are not elucidated. Moreover, whether selenepezil ameliorates memory deficits in vivo remains unknown. This study validated the cytoprotective effect of selenepezil against 6-hydroxydopamine (6-OHDA)-or H 2 O 2 -induced SH-SY5Y cell damage via alleviation or neutralization of intracellular microtubule disorder, reactive oxygen species (ROS) accumulation, mitochondrial dysfunction, and cell apoptosis. Our study clearly demonstrated that selenepezil pretreatment exhibited remarkable cytoprotective effect in a Nrf2-dependent manner via activating the Keap1−Nrf2−ARE pathway and stimulating the transcription of Nrf2−ARE-regulated cytoprotective genes. Moreover, selenepezil•HCl exerts neuroprotective effect via attenuating Aβ-induced cognitive impairment in Alzheimer's disease (AD) rat and was more active than the reference drug donepezil. In summary, selenepezil deserves further consideration for AD therapy.

No abstract

Objective: We aimed to describe the 25-hydroxyvitamin D (25(OH)D) status of southern Chinese individuals by a high-accuracy liquid chromatography tandem mass spectrometry (LC-MS/MS) method which can trace to reference measurement procedure. Materials and methods: From January 2018 to June 2019, a total of 4775 southern Chinese individuals were evaluated in our study. The serum levels of parathyroid hormone (PTH) were detected simultaneously in 162 cases. 25(OH)D was determined by LC-MS/MS, and PTH was detected using routine automated analysers. The distribution of the concentration, prevalence and seasonal variability of 25(OH)D in males and females of different age groups were studied. Results: The mean 25(OH)D concentration in our study was 32.57 ng/mL (4.20-101.40 ng/mL). The global 25(OH)D concentration in males was higher than that in females of different age group. The prevalence of vitamin D deficiency (< 20 ng/mL) in females (16.65%) was higher than that in males (6.83%). The prevalence of vitamin D deficiency (< 20 ng/mL) was most common in winter (22.98% of all women and 15.49% of all men). 25(OH)D concentrations were higher in those from whom blood samples were collected in summer and autumn than in winter and spring. 25(OH)D 2 was detected in 672 serum samples (14.07%). In addition, there was a negative correlation between the concentrations of 25(OH)D and serum PTH (r = − 0.149, P < 0.05). Conclusion: Our study demonstrated that the average serum 25(OH)D concentration in southern Chinese individuals was higher than that in other Chinese cohorts by a high-accuracy LC-MS/MS method. The global 25(OH)D concentration in males was higher than that in females of different ages, and the prevalence of vitamin D deficiency in females was higher than that in males. Seasonal change was an important aspect of 25(OH)D concentration in young and middle-aged people but became less relevant for that in older subjects. 25(OH)D 2 detection was of minor practical significance in our study. In addition, we also found that there was a negative correlation between the serum levels of 25(OH)D and PTH in southern Chinese individuals.

Addictions, both substance and behavioral, have been conceptualized as involving similar biopsychosocial processes with different opportunistic expressions. A maladaptive stress response in combination with craving or urges to engage in the addictive behavior may be among the underlying factors common to behavioral and substance addictions. The current study compared the neuroendocrine (cortisol) and subjective responses to stress of gamblers and smokers to healthy controls. We assessed if participants responded differently to smoking or gambling cues before and after a psychosocial stressor. To this end, the subjective urges/cravings of all participants were measured in response to smoking or gambling cues versus neutral cues, once under normal conditions and again after exposure to a stressor. Salivary cortisol was measured prior to, immediately following, and 10 minutes after conclusion of the stressor. Smokers and gamblers showed a similar blunted cortisol response to the acute stressor that differed from the control group's response. Following a stressor, subjective craving in smokers increased whereas gamblers' urges decreased. In smokers, a blunted cortisol and subjective stress response were related to increased urges. In contrast, for gamblers, changes in cortisol levels were unrelated to urges, and higher subjective stress was associated with decreased urges. In conclusion, individuals with a substance and a behavioral addiction share common patterns of reactivity to stress. However, while the stressor enhanced cue-related craving in smokers, it generally had the opposite effect on gamblers. Further research is necessary to elucidate the complicated patterns of similarities and differences that underlie substance and behavioral addictions.

No abstract

A candidate reference measurement procedure (RMP) for measurement of unconjugated estriol in human serum has been developed and validated. The proposed method is highly reliable and uses isotope dilution coupled with liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) and requires no derivatization. An appropriate amount of serum was accurately weighed and spiked with an isotopically labeled internal standard. Unconjugated estriol and its internal standard were extracted from serum matrix using liquid-liquid extraction prior to reversed-phase LC-MS/MS. Calibrator bracketing was used to give higher specificity and accuracy for assigning serum level. The accuracy of the candidate RMP was validated by split-sample comparison to established RMPs. The lowest limit of detection (LLoD) and lowest limit of quantification (LLoQ) for developed RMP was estimated to be 0.14 nmol/L and 0.35 nmol/L, respectively. Both intra- and inter-assay imprecisions were ≤2.19% at 1.39, 17.34 and 69.35 nmol/L, respectively. Recoveries were 98.54% to 100.34% and linear response ranged from 0.35 to 173.38 nmol/L. No interference was observed. Biases were 5.6% and 2.8% against the targets of RELA2015A (3.87 nmol/L) and RELA2015B (40.62 nmol/L), respectively. Moreover, the candidate RMP was successfully applied to measure level of unconjugated estriol in serum samples of pregnant women (n = 3) and compared with two immunoassays in clinical laboratory. Our developed method is simple, accurate, and can be used as a candidate RMP to determine total unconjugated estriol level in human serum. Further improvement of certain immunoassays in accuracy and precision is needed. Graphical abstract Selected ion chromatograms by LC-MS/MS using a C18 column for uE from a serum sample.

PICT-1 is an essential ribosome biogenesis factor whose loss induces p53 accumulation and apoptosis. Here, we show that DNA damage changes PICT-1 localization and decreases PICT-1 protein levels via the proteasome pathway. Two important phosphatidylinositol 3-kinase-like kinases (PIKKs), ataxia-telangiectasia mutated (ATM) and the Ku70 subunit of DNA-dependent protein kinase (DNA-PK), co-localize and interact with PICT-1 in the nucleolus. Computational prediction of phosphorylation sites and detection using an anti-phospho-substrate antibody suggest that PICT-1 might be a substrate of PIKKs. PICT-1 S233 and T289 were identified as the key phosphorylation sites in this pathway, as mutating both to alanine abolished UVB-induced increase of PICT-1 phosporylation. Inhibition of PIKKs or ATM (with wortmannin and KU55933, respectively) prevented the agglomeration and degradation of PICT-1, suggesting that ATM is a key regulator of PICT-1. PICT-1(S233A, T289A) demonstrated marked resistance to DNA damage-induced agglomeration and loss of PICT-1. Phosphomimetic PICT-1 (S233D, T289D) showed a different nuclear distribution and was more rapidly degraded after DNA damage than wild-type PICT-1. Furthermore, both phosphorylation and degradation of PICT-1 released RPL11 from the nucleolus to the nucleoplasm, increased binding of RPL11 to MDM2, and promoted p53 accumulation and apoptosis in an ATM-dependent manner after DNA damage. These data indicate that PICT-1 is a major nucleolar sensor of the DNA damage repair response and an important upstream regulator of p53 via the RPL11-MDM2-p53 pathway.