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Abnormal mitochondrial metabolism has been described in the Alzheimer's disease (AD) brain. However, the relationship between AD pathophysiology and key mitochondrial processes remains elusive. The purpose of this study was to investigate whether mitochondrial complex I dysfunction is associated with amyloid aggregation or glucose metabolism and brain atrophy in patients with mild AD using positron emission tomography (PET).Amyloid- and tau-positive symptomatic AD patients with clinical dementia rating 0.5 or 1 (N = 30; mean age ± standard deviation: 71.8 ± 7.6 years) underwent magnetic resonance imaging and PET scans with [[In symptomatic cases, although mitochondrial complex I reduction is linked to a wide range of downstream neurodegenerative processes such as hypometabolism, atrophy, and cognitive decline, a link to amyloid was not observable. The data presented here support [

Large numbers of cells are generally required for quantitative global proteome profiling due to surface adsorption losses associated with sample processing. Such bulk measurement obscures important cell-to-cell variability (cell heterogeneity) and makes proteomic profiling impossible for rare cell populations (e.g., circulating tumor cells (CTCs)). Here we report a surfactant-assisted one-pot sample preparation coupled with mass spectrometry (MS) method termed SOP-MS for label-free global single-cell proteomics. SOP-MS capitalizes on the combination of a MS-compatible nonionic surfactant, n-Dodecyl-β-D-maltoside, and hydrophobic surface-based low-bind tubes or multi-well plates for ‘all-in-one’ one-pot sample preparation. This ‘all-in-one’ method including elimination of all sample transfer steps maximally reduces surface adsorption losses for effective processing of single cells, thus improving detection sensitivity for single-cell proteomics. This method allows convenient label-free quantification of hundreds of proteins from single human cells and ~1200 proteins from small tissue sections (close to ~20 cells). When applied to a patient CTC-derived xenograft (PCDX) model at the single-cell resolution, SOP-MS can reveal distinct protein signatures between primary tumor cells and early metastatic lung cells, which are related to the selection pressure of anti-tumor immunity during breast cancer metastasis. The approach paves the way for routine, precise, quantitative single-cell proteomics., Tsai, Zhang, Scholten et al. develop a surfactant- assisted one-pot sample preparation coupled with mass spectrometry method (SOP-MS) for label-free global single-cell proteomics. This method allows researchers to measure hundreds of proteins from single human cells, suggesting its utility for quantitative single-cell proteomics.

Chaperone-mediated autophagy (CMA) is a protein degradation mechanism through lysosomes. By targeting the KFERQ motif of the substrate, CMA is responsible for the degradation of about 30% of cytosolic proteins, including a series of proteins associated with neurodegenerative diseases (NDs). The fact that decreased activity of CMA is observed in NDs, and ND-associated mutant proteins, including alpha-synuclein and Tau, directly impair CMA activity reveals a possible vicious cycle of CMA impairment and pathogenic protein accumulation in ND development. Given the intrinsic connection between CMA dysfunction and ND, enhancement of CMA has been regarded as a strategy to counteract ND. Indeed, genetic and pharmacological approaches to modulate CMA have been shown to promote the degradation of ND-associated proteins and alleviate ND phenotypes in multiple ND models. This review summarizes the current knowledge on the mechanism of CMA with a focus on its relationship with NDs and discusses the therapeutic potential of CMA modulation for ND.

Our previous findings have shown that the chlorophyllides composites have anticancer activities to breast cancer cell lines (MCF-7 and MDA-MB-231). In the present study, microarray gene expression profiling was utilized to investigate the chlorophyllides anticancer mechanism on the breast cancer cells lines. Results showed that chlorophyllides composites induced upregulation of 43 and 56 differentially expressed genes (DEG) in MCF-7 and MDA-MB-231 cells, respectively. In both cell lines, chlorophyllides composites modulated the expression of annexin A4 (ANXA4), chemokine C-C motif receptor 1 (CCR1), stromal interaction molecule 2 (STIM2), ethanolamine kinase 1 (ETNK1) and member of RAS oncogene family (RAP2B). Further, the KEGG annotation revealed that chlorophyllides composites modulated DEGs that are associated with the endocrine system in MCF-7 cells and with the nervous system in MDA-MB-231 cells, respectively. The expression levels of 9 genes were validated by quantitative reverse transcription PCR (RT-qPCR). The expression of CCR1, STIM2, ETNK1, MAGl1 and TOP2A were upregulated in both chlorophyllides composites treated-MCF-7 and MDA-MB-231 cells. The different expression of NLRC5, SLC7A7 and PKN1 provided valuable information for future investigation and development of novel cancer therapy.

Background: Approximately 85% of the U.S. military active duty population is male and less than 50 years of age, with elevated levels of known risk factors for oropharyngeal squamous cell carcinoma (OPSCC), including smoking, excessive use of alcohol, and greater numbers of sexual partners, and elevated prevalence of human papilloma virus (HPV). Given the recent rise in incidence of OPSCC related to the HPV, the Department of Defense Serum Repository provides an unparalleled resource for longitudinal studies of OPSCC in the military for the identification of early detection biomarkers. Methods: We identified 175 patients diagnosed with OPSCC with 175 matched healthy controls and retrieved a total of 978 serum samples drawn at the time of diagnosis, 2 and 4 years prior to diagnosis, and 2 years after diagnosis. Following immunoaffinity depletion, serum samples were analyzed by targeted proteomics assays for multiplexed quantification of a panel of 146 candidate protein biomarkers from the curated literature. Results: Using a Random Forest machine learning approach, we derived a 13-protein signature that distinguishes cases versus controls based on longitudinal changes in serum protein concentration. The abundances of each of the 13 proteins remain constant over time in control subjects. The AUC for the derived Random Forest classifier was 0.90. Conclusions: This 13-protein classifier is highly promising for detection of OPSCC prior to overt symptoms. Impact: Use of longitudinal samples has significant potential to identify biomarkers for detection and risk stratification.

To translate the International Consultation of Incontinence Questionnaire (ICIQ)-bladder diary into Chinese and validate it among Chinese women with lower urinary tract symptoms. After receiving permission to translate and validate the ICIQ-bladder diary from Bristol Urological Institute, a Chinese ICIQ-bladder diary was developed through translation, re-translation and cultural adaption. Subsequently, we tested its reliability, validity and responsiveness and evaluated its optimal record duration among females with lower urinary tract symptoms. A total of 146 participants were recruited in the study. We interviewed eight women about the content and format of the ICIQ-bladder diary, and they all thought it was clear and easy to use. Nineteen women completed the ICIQ-bladder diary twice with a 2-week interval. The resulting agreement of each item fluctuated between 0.582 and 0.940. A total of 11 urologists and nurses evaluated its content validity, and the experts’ authority was 0.94 ± 0.06. The item content validity index of the ICIQ-bladder diary was 0.909–1.000, and the scale content validity index was 0.974. Another 22 women recorded their information in the ICIQ-bladder diary pre- and post-treatment, which showed a significant difference (P

The therapeutic effect of CAR-T is often accompanied by sCRS, which is the main obstacle to the promotion of CAR-T therapy. The JAK1/2 inhibitor ruxolitinib has recently been confirmed as clinically effective in maintaining control over sCRS, however, its mechanism remains unclear. In this study, we firstly revealed that ruxolitinib significantly inhibited the proliferation of CAR-T cells without damaging viability, and induced an efficacy-favored differentiation phenotype. Second, ruxolitinib reduced the level of cytokine release not only from CAR-T cells, but also from other cells in the immune system. Third, the cytolytic activity of CAR-T cells was restored once the ruxolitinib was removed; however, the cytokines released from the CAR-T cells maintained an inhibited state to some degree. Finally, ruxolitinib significantly reduced the proliferation rate of CAR-T cells in vivo without affecting the therapeutic efficacy after withdrawal at the appropriate dose. We demonstrated pre-clinically that ruxolitinib interferes with both CAR-T cells and the other immune cells that play an important role in triggering sCRS reactions. This work provides useful and important scientific data for clinicians on the question of whether ruxolitinib has an effect on CAR-T cell function loss causing CAR-T treatment failure when applied in the treatment of sCRS, the answer to which is of great clinical significance.

Exosomes are a promising noninvasive tumor biomarker for cancer diagnosis and classification. However, efficient capture and precise analysis of exosomes in complex biological samples remain challenging. Here, sensitive profiling of exosomes with an integrated separation-detection strategy of 37 min is performed based on boronic acid-directed coupling immunoaffinity. The modification of g-C3N4 nanosheets with boronic acid (BCNNS) contributes to antibody binding under physiological conditions, which is accompanied by fluorescence enhancement. When exosomes are captured by an antibody equipped with BCNNS, a decrease in fluorescence can be induced; moreover, using the dispersion property of BCNNS, the exosomes can be separated by a simple centrifugation step. The protocol shows a favorable sensitivity with a detection limit of 2484 particles/mL. By changing only the fused antibody, exosome phenotype information profiling can be achieved, and exosomes derived from four different cell lines (HeLa, HepG2, MCF-7, and MCF-10A) can be successfully distinguished. More significantly, the positive prediction accuracy results reach 100% for serum samples from different individuals and have the advantage of multiple parameters; thus, the method has great potential in noninvasive diagnosis and point-of-care testing.

As the cost of diabetes treatment continues to grow, it is critical to accurately predict the medical costs of diabetes. Most medical cost studies based on convolutional neural networks (CNNs) ignore the importance of multi-granularity information of medical concepts and time interval characteristics of patients' multiple visit sequences, which reflect the frequency of patient visits and the severity of the disease. Therefore, this paper proposes a new end-to-end deep neural network structure, MST-CNN, for medical cost prediction. The MST-CNN model improves the representation quality of medical concepts by constructing a multi-granularity embedding model of medical concepts and incorporates a time interval vector to accurately measure the frequency of patient visits and form an accurate representation of medical events. Moreover, the MST-CNN model integrates a channel attention mechanism to adaptively adjust the channel weights to focus on significant medical features. The MST-CNN model systematically addresses the problem of deep learning models for temporal data representation. A case study and three comparative experiments are conducted using data collected from Pingjiang County. Through experiments, the methods used in the proposed model are analyzed, and the super contribution of the model performance is demonstrated.

Tau in Alzheimer's disease (AD) is assessed via cerebrospinal fluid (CSF) and Positron emission tomography (PET). Novel methods to detect phosphorylated tau (pTau) in blood have been recently developed. We aim to investigate agreement of tau status as determined by [We assessed cognitively unimpaired young, cognitively unimpaired, mild cognitive impairment and AD individuals with [The threshold for positivity was 15.085 pg/mL for plasma pTau181 and 17.652 pg/mL for plasma pTau231. Most individuals had concordant statuses, however, 18% of plasma181/PET, 26% of plasma231/PET and 25% of the pTau231/pTau181 were discordant. Positivity to at least one biomarker was often accompanied by diagnosis of cognitive impairment, Aβ positivity, APOEε4 carriership, higher levels of [Plasma pTau181, pTau231 and [Moreover, this study was supported by Weston Brain Institute, Canadian Institute of Health Research and Fonds de Recherche du Québec.

G protein‐coupled estrogen receptor‐1 (GPER), a member of the G protein‐coupled receptor (GPCR) superfamily, mediates estrogen‐induced proliferation of normal and malignant breast epithelial cells. However, its role in breast cancer stem cells (BCSCs) remains unclear. Here we showed greater expression of GPER in BCSCs than non‐BCSCs of three patient‐derived xenografts of ER−/PR+ breast cancers. GPER silencing reduced stemness features of BCSCs as reflected by reduced mammosphere forming capacity in vitro, and tumor growth in vivo with decreased BCSC populations. Comparative phosphoproteomics revealed greater GPER‐mediated PKA/BAD signaling in BCSCs. Activation of GPER by its ligands, including tamoxifen (TMX), induced phosphorylation of PKA and BAD‐Ser118 to sustain BCSC characteristics. Transfection with a dominant‐negative mutant BAD (Ser118Ala) led to reduced cell survival. Taken together, GPER and its downstream signaling play a key role in maintaining the stemness of BCSCs, suggesting that GPER is a potential therapeutic target for eradicating BCSCs., What's new? G protein‐coupled estrogen receptor‐1 (GPER) mediates estrogen‐induced proliferation of normal and malignant breast epithelial cells. However, the role of GPER in breast cancer stem cells (BCSC) biology remains unclear. Here, using patient‐derived xenografts of ER–/PR+ breast cancer, the authors found higher expression of GPER in BCSCs than non‐BCSCs. Moreover, the results indicated that stemness features were sustained via GPER‐mediated PKA/BAD phosphorylation. Stimulation by the GPER ligand tamoxifen enhanced BCSC cell viability and population and BAD phosphorylation. The findings revealed a vital role of GPER‐mediated signaling pathways in BCSC survival, suggesting GPER as a potential therapeutic target for eradicating BCSCs.

Aberrant phospho-signaling is a hallmark of cancer. We investigated kinase-substrate regulation of 33,239 phosphorylation sites (phosphosites) in 77 breast tumors and 24 breast cancer xenografts. Our search discovered 2134 quantitatively correlated kinase-phosphosite pairs, enriching for and extending experimental or binding-motif predictions. Among the 91 kinases with auto-phosphorylation, elevated EGFR, ERBB2, PRKG1, and WNK1 phosphosignaling were enriched in basal, HER2-E, Luminal A, and Luminal B breast cancers, respectively, revealing subtype-specific regulation. CDKs, MAPKs, and ataxia-telangiectasia proteins were dominant, master regulators of substrate-phosphorylation, whose activities are not captured by genomic evidence. We unveiled phospho-signaling and druggable targets from 113 kinase-substrate pairs and cascades downstream of kinases, including AKT1, BRAF and EGFR. We further identified kinase-substrate-pairs associated with clinical or immune signatures and experimentally validated activated phosphosites of ERBB2, EIF4EBP1, and EGFR. Overall, kinase-substrate regulation revealed by the largest unbiased global phosphorylation data to date connects driver events to their signaling effects.

VEGF stimulates the formation of new blood vessels by inducing endothelial cell (EC) proliferation and migration. Brefeldin A (BFA)-inhibited guanine nucleotide-exchange protein (BIG)1 and 2 accelerate the replacement of bound GDP with GTP to activate ADP-ribosylation factor (Arf)1, which regulates vesicular transport between the Golgi and plasma membrane. Although it has been reported that treating cells with BFA interferes with Arf1 activation to inhibit VEGF secretion, the role of BIG1 and BIG2 in VEGF trafficking and expression, EC migration and proliferation, and vascular development remains unknown. Here, we found that inactivation of Arf1 reduced VEGF secretion but did not affect the levels of VEGF protein. Interestingly, however, BIG1 and BIG2 knockdown significantly decreased the levels of VEGF mRNA and protein in glioblastoma U251 cells and HUVECs. Furthermore, depletion of BIG1 and BIG2 inhibited HUVEC angiogenesis by diminishing cell migration. Angioblast migration and intersegmental vessel sprouting were also impaired when the BIG2 homolog, Arf guanine nucleotide exchange factor (arfgef)2, was knocked down in zebrafish with endothelial expression of green fluorescent protein (GFP). Depletion of arfgef2 by clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) also caused defects in vascular development of zebrafish embryos. Taken together, these data reveal that BIG1 and BIG2 participate in endothelial cell angiogenesis.-Lu, F.-I., Wang, Y.-T., Wang, Y.-S., Wu, C.-Y., Li, C.-C. Involvement of BIG1 and BIG2 in regulating VEGF expression and angiogenesis.

Markers of cancer stemness predispose patients to tumor aggressiveness, drug and immunotherapy resistance, relapse, and metastasis. DDX17 is a cofactor of the Drosha–DGCR8 complex in miRNA biogenesis and transcriptional coactivator and has been associated with cancer stem-like properties. However, the precise mechanism by which DDX17 controls cancer stem-like features remains elusive. Here, we show that the E3 ligase HectH9 mediated K63-polyubiquitination of DDX17 under hypoxia to control stem-like properties and tumor-initiating capabilities. Polyubiquitinated DDX17 disassociated from the Drosha–DGCR8 complex, leading to decreased biogenesis of anti-stemness miRNAs. Increased association of polyubiquitinated DDX17 with p300-YAP resulted in histone 3 lysine 56 (H3K56) acetylation proximal to stemness-related genes and their subsequent transcriptional activation. High expression of HectH9 and six stemness-related genes (BMI1, SOX2, OCT4, NANOG, NOTCH1, and NOTCH2) predicted poor survival in patients with head and neck squamous cell carcinoma and lung adenocarcinoma. Our findings demonstrate that concerted regulation of miRNA biogenesis and histone modifications through posttranslational modification of DDX17 underlies many cancer stem-like features. Inhibition of DDX17 ubiquitination may serve as a new therapeutic venue for cancer treatment. Significance: Hypoxia-induced polyubiquitination of DDX17 controls its dissociation from the pri-miRNA–Drosha–DCGR8 complex to reduce anti-stemness miRNA biogenesis and association with YAP and p300 to enhance transcription of stemness-related genes.

Background Mitochondrial electron transport chain abnormalities have been reported in postmortem pathological specimens of Alzheimer’s disease (AD). However, it remains unclear how amyloid and tau are associated with mitochondrial dysfunction in vivo. The purpose of this study is to assess the local relationships between mitochondrial dysfunction and AD pathophysiology in mild AD using the novel mitochondrial complex I PET imaging agent [18F]BCPP-EF. Methods Thirty-two amyloid and tau positive mild stage AD dementia patients (mean age ± SD: 71.1 ± 8.3 years) underwent a series of PET measurements with [18F]BCPP-EF mitochondrial function, [11C]PBB3 for tau deposition, and [11C] PiB for amyloid deposition. Age-matched normal control subjects were also recruited. Inter and intrasubject comparisons of levels of mitochondrial complex I activity, amyloid and tau deposition were performed. Results The [18F]BCPP-EF uptake was significantly lower in the medial temporal area, highlighting the importance of the mitochondrial involvement in AD pathology. [11C]PBB3 uptake was greater in the temporo-parietal regions in AD. Region of interest analysis in the Braak stage I-II region showed significant negative correlation between [18F]BCPP-EF SUVR and [11C]PBB3 BPND (R = 0.2679, p = 0.04), but not [11C] PiB SUVR. Conclusions Our results indicated that mitochondrial complex I is closely associated with tau load evaluated by [11C]PBB3, which might suffer in the presence of its off-target binding. The absence of association between mitochondrial complex I dysfunction with amyloid load suggests that mitochondrial dysfunction in the trans-entorhinal and entorhinal region is a reflection of neuronal injury occurring in the brain of mild AD.

In recent years, α-glucosidase inhibitors (AGIs) have played a significant role in the treatment of type II diabetes (T2D), so it is necessary to develop a reliable and sensitive method to find new AGIs. Herein, we establish a novel method based on fluorescent carbon nitride nanoparticles (CNNPs) for the sensitive detection of the activity of α-glucosidase (α-glu) and the screening of its inhibitors. A CNNP-based fluorescent probe is synthesized from green raw materials, urea and lysine, by a one-pot method. In the presence of α-glu, the substrate 4-nitrophenyl-α-D-glucopyranoside (pNPG) is hydrolyzed to generate 4-nitrophenol (pNP), leading to the fluorescence (FL) quenching of CNNPs due to the inner filter effect (IFE). On the other hand, the activity of α-glu is inhibited after the addition of AGIs, which turns on the FL of CNNPs. In this way, the detection of α-glu activity and the screening of AGIs are achieved. The linear range is 1.25–10.00 U L−1 with a limit of detection as low as 0.17 U L−1 and the IC50 values of two typical inhibitors (gallic acid and acarbose) are 813 μM and 465 μM, respectively. The CNNP probe is further applied for the determination of α-glu activity in human serum samples with satisfactory results.

Dysregulation of phosphorylation-mediated signaling drives the initiation and progression of many diseases. A substrate-specific kinase assay capable of quantifying the altered site-specific phosphorylation of its phenotype-dependent substrates provides better specificity to monitor a disease state. We report a sensitive and rapid substrate-specific kinase assay by integrating site-specific peptide reporter and multiple reaction monitoring (MRM)-MS platform for relative and absolute quantification of substrate-specific kinase activity at the sensitivity of nanomolar kinase and nanogram cell lysate. Using non-small cell lung cancer as a proof-of-concept, three substrate peptides selected from constitutive phosphorylation in tumors (HDGF-S165, RALY-S135, and NRD1-S94) were designed to demonstrate the feasibility. The assay showed good accuracy (15% nominal deviation) and reproducibility (15% CV). In PC9 cells, the measured activity for HDGF-S165 was 3.2 ± 0.2 fmol μg

BackgroundTo investigate the association of plasma pTau181, assessed with a new immunoassay, with neurodegeneration of white matter and gray matter cross-sectionally and longitudinally, in aging and Alzheimer’s disease.MethodsObservational data was obtained from the Alzheimer’s Disease Neuroimaging Initiative, in which participants underwent plasma assessment and magnetic resonance imaging. Based on their clinical diagnosis, participants were classified as cognitively unimpaired and cognitively impaired. Linear regressions and linear mixed-effect models were used to test the cross-sectional and longitudinal associations between baseline plasma pTau181 and neurodegeneration using voxel-based morphometry.ResultsWe observed a negative correlation at baseline between plasma pTau181 and gray matter volume in cognitively unimpaired individuals. In cognitively impaired individuals, we observed a negative association between plasma pTau181 and both gray and white matter volume. In longitudinal analyses conducted in the cognitively unimpaired group, plasma pTau181 was negatively correlated with gray matter volume, starting 36 months after baseline assessments. Finally, in cognitively impaired individuals, plasma pTau181 concentrations were negatively correlated with both gray and white matter volume as early as 12 months after baseline, and neurodegeneration increased in an incremental manner until 48 months.ConclusionsHigher levels of plasma pTau181 correlate with neurodegeneration and predict further brain atrophy in aging and Alzheimer’s disease. Plasma pTau181 may be useful in predicting AD-related neurodegeneration, comparable to positron emission tomography or cerebrospinal fluid assessment with high specificity for AD neurodegeneration.

Stochastic DNA walkers capable of traversing on three-dimensional (3D) tracks have received great deal of attention. However, DNA walker-based biosensors exhibit limited amplification efficiency because of their slow walking kinetics and low processivity. Herein, by taking advantage of the high processivity of a DNA rolling machine, a sensitive ratiometric DNA nanomachine biosensor is designed. The biosensor is constructed with hairpin-loaded Au nanoparticles (NPs) (hpDNA@AuNPs) as a DNA walker and AgNCs-decorated magnetic NPs (AgNCs@MNPs) as a DNA rolling machine. In the presence of target DNA, exonuclease III (Exo III)-powered DNA walker is activated to accomplish first-stage amplification via a burnt-bridge mechanism, generating a great deal of toehold-loaded AuNPs (Toehold@AuNPs) to hybridize with magnetic nanoparticles loaded with silver-nanoclusters-labeled DNA (AgNCs@MNPs) with the assistance of Exo III. These trigger rapid rolling of AuNPs on the AgNCs@MNPs surface and release free AgNCs, converting the biological signal into a mass spectrometric signal ratio (107Ag/197Au) with detection by ICP-MS. A linear range of 0.5-500 fmol L-1 is achieved with a detection limit of 119 amol L-1 for the p53 gene. The practical applicability of the biosensor has been demonstrated in the accurate assay of the p53 gene in the human blood.

The objective was to investigate the long-term efficacy and patient satisfaction of Le Fort colpocleisis for the treatment of severe pelvic organ prolapse.This was a retrospective study of patients who underwent Le Fort colpocleisis from January 2007 to August 2018 in our hospital. Follow-up was conducted via outpatient visits or the telephone. Records were reviewed for anatomical recurrence, complications, urinary and intestinal symptoms post-operation, reoperation rate, patient satisfaction, Patient Global Impression of Improvement (PGI-I) score, regret rate etc. RESULTS: A total of 208 patients underwent follow-up. The follow-up time was 60.7 ± 34.18 (12-140) months. There were no intraoperative complications. Postoperative urinary retention occurred in 3.8% of patients (8 out of 208). There was no anatomical recurrence. New or more severe urinary symptoms occurred in 8.7% of patients (18 out of 208); new or more severe intestinal symptoms occurred in 1.9% of patients (4 out of 208). The reoperation rate was 1.44% (3 out of 208). Three cases of reoperation occurred for the following reasons: a case of severe stress urinary incontinence, a case of abscess in the vaginal septum, and a case of uterine malignancy after 2 years of colpocleisis. Patient satisfaction was as follows: 98.6% (205 out of 208) of patients were very satisfied. The PGI-I score was very much improved or improved in 99.5% (207 out of 208) of patients. A total of 0.96% (2 out of 208) of patients regretted undergoing colpocleisis.The long-term follow-up results showed that Le Fort colpocleisis was a safe and effective surgical procedure associated with high satisfaction. There was a very low regret rate, but the procedure should be taken seriously.

Objective To assess the incidence rate and risk of ankylosing spondylitis (AS) in patients with previous human papillomavirus (HPV) infection compared with those without HPV infection. Methods All patients with HPV infection (n = 66,314) in the NHIRD (2003–2013) were individually matched with up to four control subjects without HPV infection by age and sex (n = 265,256). All of the patients were tracked until an AS event was noted. Chi-square test was used to analyze the distribution of sociodemographic characteristics in the HPV cohort and non-HPV cohort. Cox proportional hazards regression was used to calculate the HRs for the development of AS, adjusting for age, sex, urbanization, length of hospital stay, medications, and comorbidities adjustment. The Kaplan-Meier method was used to plot the cumulative incidence curves. Results The HPV cohort had a 1.329 (95% C.I. = 1.138–1.552) times higher risk of AS than that of the non-HPV cohort after adjusting for sex, age, urbanization, length of hospital stay, comorbidities, and medications. Additionally, we applied propensity score weighting to reconfirm the accuracy of our analysis, and the results showed a 1.348 (95% C.I. = 1.153–1.575) times greater risk of AS in the HPV cohort compared with the non-HPV cohort. The cumulative incidence curves plotted by the Kaplan-Meier method revealed that after 120 follow-up months, the HPV cohort displayed a higher cumulative incidence of AS than that of the non-HPV cohort. (Log-rank test p Conclusions Patients with HPV infection had a higher risk of developing AS compared with non-HPV patients.

The effects of triterpenic acids-enriched fraction from Cyclocarya paliurus (CPT) on nonalcoholic fatty liver disease (NAFLD) were investigated using in vivo and in vitro models. In high fat diet-induced Wister rats, CPT significantly increased superoxide dismutase (SOD) activity and glutathione/oxidized glutathione (GSH/GSSG) ratio, reduced malondialdehyde (MDA) and protein carbonyl (PCO) levels. Moreover, CPT restored mitochondrial membrane potential dysfunction, decreased cytochrome P450 enzyme 2E1 (CYP2E1) activity, improved nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2-mediated antioxidant enzyme heme oxygenase1 (HO-1) expression. In free fatty acids-induced HepG2 cells, CPT dramatically decreased ROS content, increased mitochondrial NADH dehydrogenase (Complex I) and mitochondrial cytochrome C oxidase (Complex IV) levels. Furthermore, CPT could upregulate HO-1, quinine oxidoreductase 1 (NQO1) expression, and increase Nrf2 translocation from cytoplasm-to-nucleus. The results indicated CPT could protect mitochondria function and improve oxidative stress by activating Nrf2. Therefore, it can be inferred that CPT may be a potential agent against NAFLD.

A synergy between β-amyloid (Aβ) and tau appears to occur in Alzheimer disease (AD), but the mechanisms of interaction, and potential locations, are little understood. This study investigates the possibility of such interactions within the cortical synaptic compartments of APP/PS1 mice. We used label-free quantitative mass spectrometry to study the phosphoproteome of synaptosomes, covering 2400 phosphopeptides and providing an unbiased survey of phosphorylation changes associated with amyloid pathology. Hyperphosphorylation was detected on 36 synaptic proteins, many of which are associated with the cytoskeleton. Importantly, tau is one of the most hyperphosphorylated proteins at the synapse, upregulated at both proline-directed kinase (PDK) sites (S199/S202, S396/S404) and nonPDK sites (S400). These PDK sites correspond to well-known pathological tau epitopes in AD patients, recognized by AT8 and PHF-1 antibodies, respectively. Hyperphosphorylation at S199/S202, a rarely examined combination, was further validated in patient-derived human synaptosomes by immunoblotting. Global surveys of upregulated phosphosites revealed 2 potential kinase motifs, which resemble those of cyclin-dependent kinase 5 (CDK5, a PDK) and casein kinase II (CK2, a nonPDK). Our data demonstrate that, within synaptic compartments, amyloid pathology is associated with tau hyperphosphorylation at disease-relevant epitopes. This provides a plausible mechanism by which Aβ promotes the spreading of tauopathy.

Chlorophyllides can be found in photosynthetic organisms. Generally, chlorophyllides have a-, b-, c-, d-, and f-type derivatives, and all chlorophyllides have a tetrapyrrole structure with a Mg ion at the center and a fifth isocyclic pentanone. Chlorophyllide a can be synthesized from protochlorophyllide a, divinyl chlorophyllide a, or chlorophyll. In addition, chlorophyllide a can be transformed into chlorophyllide b, chlorophyllide d, or chlorophyllide f. Chlorophyllide c can be synthesized from protochlorophyllide a or divinyl protochlorophyllide a. Chlorophyllides have been extensively used in food, medicine, and pharmaceutical applications. Furthermore, chlorophyllides exhibit many biological activities, such as anti-growth, antimicrobial, antiviral, antipathogenic, and antiproliferative activity. The photosensitivity of chlorophyllides that is applied in mercury electrodes and sensors were discussed. This article is the first detailed review dedicated specifically to chlorophyllides. Thus, this review aims to describe the definition of chlorophyllides, biosynthetic routes of chlorophyllides, purification of chlorophyllides, and applications of chlorophyllides.

A novel zwitterionic polymer ionic liquid functionalized magnetic nanospheres, shortly as Fe3O4@PCL-PILs, is synthesized by grafting ionic liquid VimCOOHBr onto polymer e-caprolactone (PCL) modified magnetic nanospheres via esterification and surface-initiated free radical polymerization. This established synthesis strategy offers the obtained magnetic nanospheres with well-defined core-corona structure, compact grafting layer, favorable zwitterionic and negative-charged surface, and high magnetic susceptibility. The as-prepared Fe3O4@PCL-PILs nanospheres exhibit typical "zwitterionic hydrophilic interaction liquid chromatography (ZIC-HILIC)" behaviors toward protein binding, and selectively adsorption of glycoprotein is achieved. The adsorption capacity of the magnetic nanospheres toward Immunoglobulin G is high up to 1136.4 mg g-1, and the captured Immunoglobulin G could be efficiently recovered by using 0.5% NH3 H2O (v/v) as stripping reagent, providing a recovery of 80.5%. Fe3O4@PCL-PILs nanospheres are then employed as sorbent for the selective isolation of Immunoglobulin G from human whole blood, obtaining high-purity Immunoglobulin G as demonstrated by polyacrylamide gel electrophoresis assays.

Aminoacyl-tRNA synthetase cofactors play important roles in coordinating aminoacylation and translation. In this study, we describe an additional function of the fission yeast aminoacyl-tRNA synthetase cofactor 1 (Asc1) in translation. We found that Asc1 directly binds and stabilizes the interaction between small ribosomal protein Rps0A/uS2 and eukaryotic initiation factor 3a (eIF3a). In the absence of Asc1, the interaction between eIF3a and Rps0A/uS2 was compromised. The interaction between Rps0A/uS2 and eIF3a mediated the 40S ribosomal subunit binding of eIF3 in 43S preinitiation complex formation to stimulate translation initiation. Keeping with this idea, in an asc1 mutant, the association of mRNA with the 40S ribosomal subunit was defective and protein synthesis was compromised. To show that Asc1 is directly involved in translation, we demonstrate that the addition of recombinant Asc1 is able to rescue the translation defect of the asc1 mutant in a cell-free system. Furthermore, this function of Asc1 is likely to be evolutionarily conserved, as a similar interaction with eIF3a and Rps0A/uS2 could be identified in the budding yeast Saccharomyces cerevisiae and human aminoacyl-tRNA synthetase cofactors. Together, these results identify a function of aminoacyl-tRNA synthetase cofactors in translation preinitiation complex formation, which adds significantly to the expanded functions associated with aminoacyl-tRNA synthetases and their cofactors.

Nanozymes have emerged as promising alternatives to overcome the high cost and low stability issues of natural enzymes. Particularly, those with peroxidase-like activities have been extensively studied to construct versatile biosensors. In this article, we demonstrate that the modification of the graphitic carbon nitride nanosheets (g-C3N4 nanosheets) by plasmonic gold nanoparticles (AuNPs) greatly enhances its catalytic performance as peroxidase mimetic. In the presence of H2O2, the AuNPs@g-C3N4 nanosheets can catalyze the redox reaction of 3,3′,5,5′- tetramethylbenzidine (TMB) to produce a blue color. Based on the observation, a colorimetric sensing method for glucose is further developed with the assistance of glucose oxidase (GOx). The linear range for glucose is from 5 to 100 μmol L−1 (R2 = 0.9967) and the limit of detection (LOD) is 1.2 μmol L−1. The LOD can be further lowered down to 0.75 μmol L−1 by using H2SO4 as termination agent and measuring the absorbance of the yellow product at λ = 451 nm. Moreover, the practical usefulness of AuNPs@g-C3N4 nanosheets as a peroxidase nanozyme for glucose determination in human serum and urine is also demonstrated.

Purpose of Review Abnormal accumulation of tau protein is the main hallmark of tauopathies and is closely associated with neurodegeneration and cognitive impairment, whereas the advance in PET imaging provides a non-invasive detection of tau inclusions in the brain. In this review, we discuss the potential of PET imaging as a biomarker in tauopathies, the latest development of novel tau tracers with new clinical information that has been disclosed, and the opportunities for improving diagnosis and designing clinical trials in the future. Recent Findings In recent years, several first-generation tau PET tracers including [11C]PBB3, [18F]THK-5117, [18F]THK-5351 and [18F]AV-1451 have been developed and succeeded in imaging neurofibrillary pathology in vivo. Due to the common off-target binding and subcortical white matter uptake seen in the first-generation tracers, several research institutes and pharmaceutical companies have been working on developing second-generation tau PET tracers which exhibit higher binding affinity and selectivity. Summary Tau PET imaging is promising to serve as a biomarker to support differential diagnosis and monitor disease progression in many neurodegenerative diseases.

Two new sesquiterpenoids were isolated from Stellera chamaejasme L., known as the traditional Chinese herb 'Rui Xiang Lang Du'. The compounds were elucidated as stelleraguaianone B (1) and C (2) by comprehensive spectroscopic analysis, including 1D and 2D NMR as well as HRESIMS, and by comparing their NMR data with known compounds. In addition, the structure of 1 was further confirmed by single-crystal X-ray diffraction analysis. Both the compounds were evaluated for their cytotoxicity on common tumour cell lines in vitro, which revealed that compound 1 exhibits cytotoxic activity on A549 cells, while 2 has no activity.

The purity of chlorophylls plays one of the key role for the production of chlorophyllides. We have designed a facile method for chlorophyll purification by twice solvent extraction. Twice extraction causes the loss of chlorophylls, but the purity of total chlorophylls can be enhanced 182%. Then, the purified chlorophylls can be converted to relatively pure chlorophyllides facilely. The results show that higher purity of chlorophyllides could be obtained when purified chlorophylls (ethanol-hexane extract) was used as starting materials than that of crude chlorophylls (ethanol-only extract). In biocompatibility test, the results showed that the prepared chlorophyllides can be applied as biomaterials. When the prepared chlorophyllides were applied to anticancer tests, they were active both in MCF7 and MDA-MB-231 (multidrug resistant breast cancer cells) cell lines. In addition, the results suggested that the prepared chlorophyllides could be a potential candidate of combination therapy with doxorubicin to breast cancers.

Cellular supply of deoxythymidine triphosphate (dTTP) is crucial for DNA replication and repair. Thymidylate kinase (TMPK) catalyzes the conversion of thymidine monophosphate to thymidine diphosphate, which is an essential step for dTTP synthesis. Despite their major cellular localization in cytosol, TMPK and ribonucleotide reductase (RNR) are detected at DNA damage sites for local dNDP formation. Because deoxyuridine diphosphate is synthesized by RNR, the simultaneous recruitment of TMPK and RNR to DNA damage sites is critical for preventing deoxyuridine triphosphate-mediated toxic repair. This study investigates the mechanism responsible for the recruitment of TMPK to DNA damage sites. Our data demonstrate the requirement of ataxia telangiectasia mutated (ATM) kinase activity for TMPK recruitment to DNA lesion sites. Moreover, we find that TMPK is able to form the complex with histone acetyltransferase Tip60 and RNR. Inhibition of ATM kinase reduces the complex formation and TMPK phosphorylation. Our analysis further shows the presence of TMPK phosphorylation at serine 88, which is an ATM kinase consensus site. A phosphorylation-defective mutation at this site suppresses TMPK recruitment to DNA damage sites and the complex formation with Tip60. Finally, we provide evidence that this site is critical for the function of TMPK in DNA repair but not for catalytic activity. Together, these findings suggest that Tip60-ATM signaling has a functional contribution to the recruitment of TMPK to DNA damage sites, thereby increasing local dTTP synthesis for DNA repair.-Hu, C.-M., Tsao, N., Wang, Y.-T., Chen, Y.-J., Chang, Z.-F. Thymidylate kinase is critical for DNA repair via ATM-dependent Tip60 complex formation.

Cyclin-dependent kinase 2 (CDK2) plays a key role in eukaryotic cell cycle progression which could facilitate the transition from G1 to S phase. The dysregulation of CDK2 is closely related to many cancers. CDK2 is utilized as one of the most studied kinase targets in oncology. In this article, 24 benzamide derivatives were designed, synthesized and investigated for the inhibition activity against CDK2. Our results revealed that the compound 25 is a potent CDK2 inhibitor exhibiting a broad spectrum anti-proliferative activity against several human breast cancer cells. Additionally, compound 25 could block cell cycle at G0 or G1 and induce significant apoptosis in MDA-MB-468 cells. These findings highlight a rationale for further development of CDK2 inhibitors to treat human breast cancer.

Irritable bowel syndrome (IBS) is a highly prevalent chronic functional gastrointestinal disorder. Recent studies have showed increasing important role of gut microbiota in the pathophysiological changes of IBS. Our study aims to elaborate the association between intestinal flora with the genesis and the development of IBS.Illumina high-throughput sequencing technology was applied to investigate microbial communities of IBS patients and healthy donors. Stool specimens from the IBS-D patients were equally premixed and implanted into germ free C57B/6 mice to construct IBS animal model, and the normal group was also transplanted with normal premixed feces. The post-transplant defecation and intra-epithelial lymphocyte counts were evaluated. Microbial communities were also checked by the illumina high-throughput sequencing technology.Fifteen genuses significantly different were found expressed in the gut flora of IBS patients, and six genuses showed significantly different abundances between the stool specimens of mice of IBS group and normal group. Among these differences, Parasutterella expression was remarkably different in both screening and validation experiments and also related to chronic intestinal inflammation; therefore, Parasutterella expression is considered in association with the development and progression of IBS.Parasutterella may be related with the genesis and development of IBS and also associated with chronic intestinal inflammation in IBS patients.

Unified staging systems for pelvic organ prolapse (POP) have been established. We examined the application of the POP quantification (POP-Q) system in China by examining its use in scientific journal articles.Relevant articles were identified by searching the Sinomed database using the terms: uterus prolapse, cystocele, proctocele, prolapse, and pelvic floor; limited to Chinese core journals in obstetrics and gynecology, from January 2004 to December 2014. We analyzed systems for grading POP severity and the adoption of POP-Q in different article categories and hospitals of different levels. For the last decade, with two 5-year groups (2005-2009; 2010-2014), the χ(2) test was used to evaluate inter-group differences.In a total of 429 articles, 331 included a staging system, 70.7% of which used POP-Q. The POP-Q system first appeared in 2004 in China, was reported in 50% of articles in 2007, and its highest use occurred in 2012 (89.5%). In 234 POP-Q system-utilizing reports, operative treatment and basic research accounted for 73.1% and 14.0% respectively. POP-Q usage increased from 2005-2009 to 2010-2014 in surgery-related articles (54.2% vs 85.2%; P = 0.000). The proportion of reports using POP-Q in level I, II, and III hospitals was 20.0%, 35.4%, and 77.8% respectively.The POP-Q system, first used in 2004 in China, is now the most commonly used grading system, with surgery reports and level III hospitals accounting for the largest proportion of POP-Q applications.

// Chia-Ming Yeh 1,2,* , Pi-Che Chen 3,* , Hsiao-Yen Hsieh 2,5 , Yeong-Chin Jou 3 , Chang-Te Lin 3 , Ming-Hsuan Tsai 1 , Wen-Yu Huang 1,2 , Yi-Ting Wang 1,2 , Ru-Inn Lin 1,6 , Szu-Shan Chen 1,2 , Chun-Liang Tung 4 , Shu-Fen Wu 1,2 , De-Ching Chang 1,2 , Cheng-Huang Shen 3 , Cheng-Da Hsu 5 and Michael W.Y. Chan 1,2,* 1 Department of Life Science, National Chung Cheng University, Min-Hsiung, Chia-Yi, Taiwan 2 Institute of Molecular Biology, National Chung Cheng University, Min-Hsiung, Chia-Yi, Taiwan 3 Department of Urology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chia-Yi, Taiwan 4 Department of Pathology, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chia-Yi, Taiwan 5 Department of Medical Research, Ditmanson Medical Foundation Chia-Yi Christian Hospital, Chia-Yi, Taiwan 6 Departments of Radiation Oncology, Buddhist Dalin Tzu Chi General Hospital, Chia Yi, Taiwan * These authors have contributed equally to this work Correspondence to: Michael W.Y. Chan, email: // Keywords : urothelial carcinoma, DNA methylation, urine, ZNF671 Received : April 03, 2015 Accepted : July 16, 2015 Published : July 22, 2015 Abstract The molecular mechanism underlying the lethal phenomenon of urothelial carcinoma (UC) tumor recurrence remains unresolved. Here, by methylation microarray, we identified promoter methylation of the zinc-finger protein gene, ZNF671 in bladder UC tumor tissue samples, a finding that was independently validated by bisulphite pyrosequencing in cell lines and tissue samples. Subsequent assays including treatment with epigenetic depressive agents and in vitro methylation showed ZNF671 methylation to result in its transcriptional repression. ZNF671 re-expression in UC cell lines, via ectopic expression, inhibited tumor growth and invasion, in possible conjunction with downregulation of cancer stem cell markers (c-KIT, NANOG, OCT4). Clinically, high ZNF671 methylation in UC tumor tissues (n=96; 63 bladder, 33 upper urinary tract) associated with tumor grade and poor locoregional disease-free survival. Quantitative MSP analysis in a training (n=97) and test (n=61) sets of voided urine samples from bladder UC patients revealed a sensitivity and specificity of 42%-48% and 89%-92.8%, respectively, for UC cancer detection. Moreover, combining DNA methylation of ZNF671 and 2 other genes ( IRF8 and sFRP1 ) further increased the sensitivity to 96.2%, suggesting a possible three-gene UC biomarker. In summary, ZNF671 , an epigenetically silenced novel tumor suppressor, represents a potential predictor for UC relapse and non-invasive biomarker that could assist in UC clinical decision-making.

Background Ribonuclease-L (RNase-L) was known to be a ubiquitous enzyme involved in several cellular functions, especially innate immunity. It was recently shown to participate in adipogenesis in rodents. Here, we developed a method to measure serum levels of RNase-L and analyzed the relationship between RNase-L and metabolic syndrome (MetS). Methods A total of 396 subjects were recruited from a health check-up program. An in-house RNase-L immunoassay was developed. The serum RNase-L levels of these subjects were measured, and the association of MetS-related factors with RNase-L levels was assessed. Results The mean serum level of RNase-L of the subjects with MetS were lower than those without (16.5 ± 6.4 vs. 18.4 ± 8.0 μg/ml, P = 0.018). The subjects with central obesity, elevated blood pressure, or impaired fasting glucose also had lower serum RNase-L levels in comparison to those without. In multivariate linear regression analysis, diastolic blood pressure (β = −0.129, P = 0.024) and high-density lipoprotein cholesterol (HDL-C) (β = 0.127, P = 0.036) were related to serum RNase-L. For every 5 μg/ml increase in serum RNase-L levels, it is associated with a reduced risk of MetS (OR 0.83, 95% CI 0.71–0.98, P = 0.028), central obesity (OR 0.82, 95% CI 0.71–0.94, P = 0.005), or low HDL-C (OR 0.86, 95% CI 0.74–1.00, P = 0.042). Moreover, age is inversely related to serum RNase-L levels in various analyses. Conclusions The serum RNase-L levels were inversely associated with MetS, unfavorable metabolic profiles, and age.

In this study we identified Pdc2, the fission yeast ortholog of human Pat1b protein, which forms a complex with Lsm1-7 and plays a role in coupling deadenylation and decapping. The involvement of Pdc2 in RNA degradation and P-body function was also determined. We found that Pdc2 interacts with Dcp2 and is required for decapping in vivo. Although not absolutely essential for P-body assembly, overexpression of Pdc2 enhanced P-body formation even in the absence of Pdc1, the fission yeast functional homolog of human Edc4 protein, indicating that Pdc2 also plays a role in P-body formation. Intriguingly, in the absence of Pdc2, Lsm1 was found to accumulate in the nucleus, suggesting that Pdc2 shuttling between nucleus and cytoplasm plays a role in decreasing the nuclear concentration of Lsm1 to increase Lsm1 in the cytoplasm. Furthermore, unlike other components of P-bodies, the deadenylase Ccr4 did not accumulate in P-bodies in cells growing under favorable conditions and was only recruited to P-bodies after deprivation of glucose in a Pdc2-Lsm1-dependent manner, indicating a function of Pdc2 in cellular response to environmental stress. In supporting this idea, pdc2 mutants are defective in recovery from glucose starvation with a much longer time to re-enter the cell cycle. In keeping with the notion that Pat1 is a nucleocytoplasmic protein, functioning also in the nucleus, we found that Pdc2 physically and genetically interacts with the nuclear 5′–3′ exonuclease Dhp1. A function of Pdc2-Lsm1, in concert with Dhp1, regulating RNA by promoting its decapping/destruction in the nucleus was suggested.

The transcriptional repressor Slug is best known to control epithelial-mesenchymal transition (EMT) and promote cancer invasion/metastasis. In this study, we demonstrate that Slug is temporally regulated during cell cycle progression. At G1/S transition, cyclin E-cyclin-dependent kinase 2 mediates the phosphorylation of Slug at Ser-54 and Ser-104, resulting in its ubiquitylation and degradation. Non-phosphorylatable Slug is markedly stabilized at G1/S transition compared with wild-type Slug and greatly leads to downregulation of DNA synthesis and checkpoint-related proteins, including TOP1, DNA Ligase IV and Rad17, reduces cell proliferation, delays S-phase progression and contributes to genome instability. Our results indicate that Slug has multifaceted roles in cancer progression by controlling both EMT and genome stability.

TDP-43 is an important pathological protein that aggregates in the diseased neuronal cells and is linked to various neurodegenerative disorders. In normal cells, TDP-43 is primarily an RNA-binding protein; however, how the dimeric TDP-43 binds RNA via its two RNA recognition motifs, RRM1 and RRM2, is not clear. Here we report the crystal structure of human TDP-43 RRM1 in complex with a single-stranded DNA showing that RRM1 binds the nucleic acid extensively not only by the conserved β-sheet residues but also by the loop residues. Mutational and biochemical assays further reveal that both RRMs in TDP-43 dimers participate in binding of UG-rich RNA or TG-rich DNA with RRM1 playing a dominant role and RRM2 playing a supporting role. Moreover, RRM1 of the amyotrophic lateral sclerosis-linked mutant D169G binds DNA as efficiently as the wild type; nevertheless, it is more resistant to thermal denaturation, suggesting that the resistance to degradation is likely linked to TDP-43 proteinopathies. Taken together all the data, we suggest a model showing that the two RRMs in each protomer of TDP-43 homodimer work together in RNA binding and thus the dimeric TDP-43 recognizes long clusters of UG-rich RNA to achieve high affinity and specificity.

We performed the first proteogenomic study on a prospectively collected colon cancer cohort. Comparative proteomic and phosphoproteomic analysis of paired tumor and normal adjacent tissues produced a catalog of colon cancer-associated proteins and phosphosites, including known and putative new biomarkers, drug targets, and cancer/testis antigens. Proteogenomic integration not only prioritized genomically inferred targets, such as copy-number drivers and mutation-derived neoantigens, but also yielded novel findings. Phosphoproteomics data associated Rb phosphorylation with increased proliferation and decreased apoptosis in colon cancer, which explains why this classical tumor suppressor is amplified in colon tumors and suggests a rationale for targeting Rb phosphorylation in colon cancer. Proteomics identified an association between decreased CD8 T cell infiltration and increased glycolysis in microsatellite instability-high (MSI-H) tumors, suggesting glycolysis as a potential target to overcome the resistance of MSI-H tumors to immune checkpoint blockade. Proteogenomics presents new avenues for biological discoveries and therapeutic development.

Methodologies to enrich heterogeneous types of phosphopeptides are critical for comprehensive mapping of the under-explored phosphoproteome. Taking advantage of the distinct binding affinities of Ga(3+) and Fe(3+) for phosphopeptides, we designed a metal-directed immobilized metal ion affinity chromatography for the sequential enrichment of phosphopeptides. In Raji B cells, the sequential Ga(3+)-Fe(3+)-immobilized metal affinity chromatography (IMAC) strategy displayed a 1.5-3.5-fold superior phosphoproteomic coverage compared to single IMAC (Fe(3+), Ti(4+), Ga(3+), and Al(3+)). In addition, up to 92% of the 6283 phosphopeptides were uniquely enriched in either the first Ga(3+)-IMAC (41%) or second Fe(3+)-IMAC (51%). The complementary properties of Ga(3+) and Fe(3+) were further demonstrated through the exclusive enrichment of almost all of 1214 multiply phosphorylated peptides (99.4%) in the Ga(3+)-IMAC, whereas only 10% of 5069 monophosphorylated phosphopeptides were commonly enriched in both fractions. The application of sequential Ga(3+)-Fe(3+)-IMAC to human lung cancer tissue allowed the identification of 2560 unique phosphopeptides with only 8% overlap. In addition to the above-mentioned mono- and multiply phosphorylated peptides, this fractionation ability was also demonstrated on the basic and acidic phosphopeptides: acidophilic phosphorylation sites were predominately enriched in the first Ga(3+)-IMAC (72%), while Pro-directed (85%) and basophilic (79%) phosphorylation sites were enriched in the second Fe(3+)-IMAC. This strategy provided complementary mapping of different kinase substrates in multiple cellular pathways related to cancer invasion and metastasis of lung cancer. Given the fractionation ability and ease of tip preparation of this Ga(3+)-Fe(3+)-IMAC, we propose that this strategy allows more comprehensive characterization of the phosphoproteome both in vitro and in vivo.

Background: TDP-43 forms aggregates in various neurodegenerative disorders. Results: The C-terminal-truncated RRM2 of TDP-43 forms non-amyloid fibrils in vitro and plays a dominant role in forming inclusions in vivo. Conclusion: The proteolytic cleavage of TDP-43 that removes the N-terminal dimerization domain may produce unassembled truncated RRM2 fragments for aggregation. Significance: This result provides a new direction for the prevention and treatment of TDP-43-associated diseases., TDP-43 is the major pathological protein identified in the cellular inclusions in amyotrophic lateral sclerosis and frontotemporal lobar degeneration. The pathogenic forms of TDP-43 are processed C-terminal fragments containing a truncated RNA-recognition motif (RRM2) and a glycine-rich region. Although extensive studies have focused on this protein, it remains unclear how the dimeric full-length TDP-43 is folded and assembled and how the processed C-terminal fragments are misfolded and aggregated. Here, using size-exclusion chromatography, pulldown assays, and small angle x-ray scattering, we show that the C-terminal-deleted TDP-43 without the glycine-rich tail is sufficient to form a head-to-head homodimer primarily via its N-terminal domain. The truncated RRM2, as well as two β-strands within the RRM2, form fibrils in vitro with a similar amyloid-negative staining property to those of TDP-43 pathogenic fibrils in diseases. In addition to the glycine-rich region, the truncated RRM2, but not the intact RRM2, plays a key role in forming cytoplasmic inclusions in neuronal cells. Our data thus suggest that the process that disrupts the dimeric structure, such as the proteolytic cleavage of TDP-43 within the RRM2 that removes the N-terminal dimerization domain, may produce unassembled truncated RRM2 fragments with abnormally exposed β-strands, which can oligomerize into high-order inclusions.

A simple one-step hydrothermal green approach was reported for the preparation of carbon dots (CDs) without any further decoration or modification with papaya powder as natural carbon source. In this economical and eco-friendly system, deionized water or 90% ethanol was used as solvent to produce water-soluble or ethanol-soluble CDs, respectively, termed as W-CDs and E-CDs. The quantum yield (QY) for W-CDs was 18.98%, while that for E-CDs was 18.39%. The potentials of the prepared carbon dots toward diverse applications were thoroughly investigated. W-CDs and E-CDs provide promising probes for fluorescence detection of Fe(3+), offering limits of detection of 0.48μmolL(-1) and 0.29μmolL(-1), respectively. W-CDs was further demonstrated to be a promising probe for fluorescence sensing of Escherichia coli O157: H7, along with a limit of detection of 9.5×10(4)cfumL(-1). Meanwhile, both W-CDs and E-CDs exhibit favorable biocompatibility, and demonstrated to be efficient for Hela cell imaging.

Background A computationally efficient tool is required for a genome-wide gene-gene interaction analysis that tests an extremely large number of single-nucleotide polymorphism (SNP) interaction pairs in genome-wide association studies (GWAS). Current tools for GWAS interaction analysis are mainly developed for unrelated case-control samples. Relatively fewer tools for interaction analysis are available for complex disease studies with family-based design, and these tools tend to be computationally expensive. Results We developed a fast gene-gene interaction test, GCORE-sib, for discordant sib pairs and implemented the test into an efficient tool. We used simulations to demonstrate that the GCORE-sib has correct type I error rates and has comparable power to that of the regression-based interaction test. We also showed that the GCORE-sib can run more than 10 times faster than the regression-based test. Finally, the GCORE-sib was applied to a GWAS dataset with approximately 2,000 discordant sib pairs, and the GCORE-sib finished testing 19,368,078,382 pairs of SNPs within 6 days. Conclusions An efficient gene-gene interaction tool for discordant sib pairs was developed. It will be very useful for genome-wide gene-gene interaction analysis in GWAS using discordant sib pairs. The tool can be downloaded for free at http://gcore-sib.sourceforge.net. Electronic supplementary material The online version of this article (doi:10.1186/s12859-016-1145-z) contains supplementary material, which is available to authorized users.

Patients with dementia differ in their behavioral and psychological symptoms according to their diagnosis with Alzheimer's disease (AD) or vascular dementia (VaD), requiring different symptom-management strategies. This study analyzed data from a sub-sample of a randomized clinical trial to determine the effects of a home-based training program on family caregivers of patients with dementia in northern Taiwan. Our sub-sample comprised patient-caregiver dyads (46 VaD and 68 AD patients) followed for 18 months. Caregivers of AD patients in the intervention group had better competence, preparedness, health-related quality of life, and fewer depressive symptoms than those in the control group. Caregivers of VaD patients in the intervention group had better competence and health-related quality of life than those in the control group. The training program benefited family caregivers of AD patients more than caregivers of VaD patients. Specific training programs need to be developed for family caregivers of VaD patients.

Human polynucleotide phosphorylase (hPNPase) is a 3′-to-5′ exoribonuclease that degrades specific mRNA and miRNA, and imports RNA into mitochondria, and thus regulates diverse physiological processes, including cellular senescence and homeostasis. However, the RNA-processing mechanism by hPNPase, particularly how RNA is bound via its various domains, remains obscure. Here, we report the crystal structure of an S1 domain-truncated hPNPase at a resolution of 2.1 Å. The trimeric hPNPase has a hexameric ring-like structure formed by six RNase PH domains, capped with a trimeric KH pore. Our biochemical and mutagenesis studies suggest that the S1 domain is not critical for RNA binding, and conversely, that the conserved GXXG motif in the KH domain directly participates in RNA binding in hPNPase. Our studies thus provide structural and functional insights into hPNPase, which uses a KH pore to trap a long RNA 3′ tail that is further delivered into an RNase PH channel for the degradation process. Structural RNA with short 3′ tails are, on the other hand, transported but not digested by hPNPase.

Human mesenchymal stromal cells (hMSCs) are promising candidates for cell therapy and tissue regeneration. Knowledge of the molecular mechanisms governing hMSC commitment into osteoblasts is critical to the development of therapeutic applications for human bone diseases. Because protein phosphorylation plays a critical role in signaling transduction network, the purpose of this study is to elucidate the phosphoproteomic changes in hMSCs during early osteogenic lineage commitment. hMSCs cultured in osteogenic induction medium for 0, 1, 3, and 7 days were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). Surprisingly, we observed a dramatic loss of protein phosphorylation level after 1 day of osteogenic induction. Pathways analysis of these reduced phosphoproteins exhibited a high correlation with cell proliferation and protein synthesis pathways. During osteogenic differentiation, differentially expressed phosphoproteins demonstrated the dynamic alterations in cytoskeleton at the early stages of differentiation. The fidelity of our quantitative phosphoproteomic analyses were further confirmed by Western blot analyses, and the changes from protein expression or its phosphorylation level were distinguished. In addition, several ion channels and transcription factors with differentially expressed phosphorylation sites during osteogenic differentiation were identified and may serve as potentially unexplored transcriptional regulators of the osteogenic phenotype of hMSCs. Taken together, our results have demonstrated the dynamic changes in phosphoproteomic profiles of hMSCs during osteogenic differentiation and unraveled potential candidates mediating the osteogenic commitment of hMSCs. The findings in this study may also shed light on the development of new therapeutic targets for metabolic bone diseases such as osteoporosis and osteomalacia.

Although Sp1 is known to undergo posttranslational modifications such as phosphorylation, glycosylation, acetylation, sumoylation, and ubiquitination, little is known about the possible interplay between the different forms of Sp1 that may affect its overall levels. It is also unknown whether changes in the levels of Sp1 influence any biological cell processes. Here, we identified RNF4 as the ubiquitin E3 ligase of Sp1. From in vitro and in vivo experiments, we found that sumoylated Sp1 can recruit RNF4 as a ubiquitin E3 ligase that subjects sumoylated Sp1 to proteasomal degradation. Sp1 mapping revealed two ubiquitination-related domains: a small ubiquitin-like modifier in the N-terminus of Sp1(Lys16) and the C-terminus of Sp1 that directly interacts with RNF4. Interestingly, when Sp1 was phosphorylated at Thr739 by c-Jun NH 2 -terminal kinase 1 during mitosis, this phosphorylated form of Sp1 abolished the Sp1–RNF4 interaction. Our results show that, while sumoylated Sp1 subjects to proteasomal degradation, the phosphorylation that occurs during the cell cycle can protect Sp1 from degradation by repressing the Sp1–RNF4 interaction. Thus, we propose that the interplay between posttranslational modifications of Sp1 plays an important role in cell cycle progression and keeps Sp1 at a critical level for mitosis.