Discovery Small-Molecule p300 Inhibitors Derived from a Newly Developed Indazolone-Focused DNA-Encoded Library.

The DNA-encoded library (DEL) is a robust tool for chemical biology and drug discovery. In this study, we developed a DNA-compatible light-promoted reaction that is highly efficient and plate-compatible for DEL construction based on the formation of the indazolone scaffold. Employing this high-efficiency approach, we constructed a DEL featuring an indazolone core, which enabled the identification of a novel series of ligands specifically targeting E1A-binding protein (p300) after DEL selection. Taken together, our findings underscore the feasibility of light-promoted reactions in DEL synthesis and unveil promising avenues for developing p300-targeting inhibitors.

Regulation of biofilm gene expression by DNA replication in Bacillus subtilis.

Bacillus subtilis relies on biofilms for survival in harsh environments. Extracellular polymeric substance (EPS) is a crucial component of biofilms, yet the dynamics of EPS production in single cells remain elusive. To unveil the modulation of EPS synthesis, we built a minimal network model comprising the SinI-SinR-SlrR module, Spo0A, and EPS. Stochastic simulations revealed that antagonistic interplay between SinI and SinR enables EPS production in bursts. SlrR widens these bursts and increases their frequency by stabilizing SinR-SlrR complexes and depleting free SinR. DNA replication and chromosomal positioning of key genes dictate pulsatile changes in the slrR:sinR gene dosage ratio (gr) and Spo0A-P levels, each promoting EPS production in distinct phases of the cell cycle. As the cell cycle lengthens with nutrient stress, the duty cycle of gr pulsing decreases, whereas the amplitude of Spo0A-P pulses elevates. This coordinated response facilitates keeping a constant proportion of EPS-secreting cells within colonies across diverse nutrient conditions. Our results suggest that bacteria may 'encode' eps expression through strategic chromosomal organization. This work illuminates how stochastic protein interactions, gene copy number imbalance, and cell-cycle dynamics orchestrate EPS synthesis, offering a deeper understanding of biofilm formation.

SENP3-regulated Nodal signaling plays a potential role in cardiac left-right asymmetry development.

Congenital heart disease (CHD) is a type of major defect that occurs during embryonic development. Although significant advances have been made in the treatment of CHD, its etiology and molecular mechanism remain unclear. To identify the critical role of SUMOylation in cardiac development, we generated SENP3 knockout mice and showed that SENP3 knockout mice die on embryonic day 8.5 with an open neural tube and reversed left-right cardiac asymmetry. Moreover, SENP3 knockout promoted apoptosis and senescence of H9C2 cells. Further studies showed that Nodal, a critical gene that forms left-right asymmetry, is regulated by SENP3 and that SENP3 regulates cell apoptosis and senescence in a Nodal-dependent manner. Furthermore, Nodal was hyper-SUMOylated after SENP3 knockout, and SUMOylation of Nodal inhibited its ubiquitination and ubiquitin-proteasome degradation pathway. Nodal overexpression enhanced cell apoptosis and senescence; however, the mutation at the SUMOylation site of Nodal reversed its effect on the apoptosis and senescence of H9C2 cells. More importantly, the SENP3-Nodal axis regulates cell senescence by inducing cell autophagy. These results suggest that the SENP3-Nodal signaling axis regulates cardiac senescence-autophagy homeostasis, which in turn affects cardiac development and results in the occurrence of CHD.

Harnessing Cooperative Multivalency in Thioguanine for Surface-Enhanced Raman Scattering (SERS)-Based Differentiation of Polyfunctional Analytes Differing by a Single Functional Group.

Small-molecule receptors are increasingly employed to probe various functional groups for (bio)chemical analysis. However, differentiation of polyfunctional analogs sharing multiple functional groups remains challenging for conventional mono- and bidentate receptors because their insufficient number of binding sites limits interactions with the least reactive yet property-determining functional group. Herein, we introduce 6-thioguanine (TG) as a supramolecular receptor for unique tridentate receptor-analyte complexation, achieving ≥97 % identification accuracy among 16 polyfunctional analogs across three classes: glycerol derivatives, disubstituted propane, and vicinal diols. Crucially, we demonstrate distinct spectral changes induced by the tridentate interaction between TG's three anchoring points and all the analyte's functional groups, even the least reactive ones. Notably, hydrogen bond (H-bond) networks formed in the TG-analyte complexes demonstrate additive effects in binding strength originating from good bond linearity, cooperativity, and resonance, thus strengthening complexation events and amplifying the differences in spectral changes induced among analytes. It also enhances spectral consistency by selectively forming a sole configuration that is stronger than the respective analyte-analyte interaction. Finally, we achieve 95.4 % accuracy for multiplex identification of a mixture consisting of multiple polyfunctional analogs. We envisage that extension to other multidentate non-covalent interactions enables the development of interference-free small molecule-based sensors for various (bio)chemical analysis applications.

Forward-predictive SERS-based chemical taxonomy for untargeted structural elucidation of epimeric cerebrosides.

Achieving untargeted chemical identification, isomeric differentiation, and quantification is critical to most scientific and technological problems but remains challenging. Here, we demonstrate an integrated SERS-based chemical taxonomy machine learning framework for untargeted structural elucidation of 11 epimeric cerebrosides, attaining >90% accuracy and robust single epimer and multiplex quantification with <10% errors. First, we utilize 4-mercaptophenylboronic acid to selectively capture the epimers at molecular sites of isomerism to form epimer-specific SERS fingerprints. Corroborating with in-silico experiments, we establish five spectral features, each corresponding to a structural characteristic: (1) presence/absence of epimers, (2) monosaccharide/cerebroside, (3) saturated/unsaturated cerebroside, (4) glucosyl/galactosyl, and (5) GlcCer or GalCer's carbon chain lengths. Leveraging these insights, we create a fully generalizable framework to identify and quantify cerebrosides at concentrations between 10-4 to 10-10 M and achieve multiplex quantification of binary mixtures containing biomarkers GlcCer24:1, and GalCer24:1 using their untrained spectra in the models.

Creating 3D Nanoparticle Structural Space via Data Augmentation to Bidirectionally Predict Nanoparticle Mixture's Purity, Size, and Shape from Extinction Spectra.

Nanoparticle (NP) characterization is essential because diverse shapes, sizes, and morphologies inevitably occur in as-synthesized NP mixtures, profoundly impacting their properties and applications. Currently, the only technique to concurrently determine these structural parameters is electron microscopy, but it is time-intensive and tedious. Here, we create a three-dimensional (3D) NP structural space to concurrently determine the purity, size, and shape of 1000 sets of as-synthesized Ag nanocubes mixtures containing interfering nanospheres and nanowires from their extinction spectra, attaining low predictive errors at 2.7-7.9 %. We first use plasmonically-driven feature enrichment to extract localized surface plasmon resonance attributes from spectra and establish a lasso regressor (LR) model to predict purity, size, and shape. Leveraging the learned LR, we artificially generate 425,592 augmented extinction spectra to overcome data scarcity and create a comprehensive NP structural space to bidirectionally predict extinction spectra from structural parameters with <4 % error. Our interpretable NP structural space further elucidates the two higher-order combined electric dipole, quadrupole, and magnetic dipole as the critical structural parameter predictors. By incorporating other NP shapes and mixtures' extinction spectra, we anticipate our approach, especially the data augmentation, can create a fully generalizable NP structural space to drive on-demand, autonomous synthesis-characterization platforms.

Protectin D1 ameliorates non-compressive lumbar disc herniation through SIRT1-mediated CGRP signaling.

Background. Neuro-inflammatory response promotes the initiation and sustenance of lumbar disc herniation (LDH). Protectin D1 (PD1), as a new type of specialized pro-resolving mediator (SPM), can improve the prognosis of various inflammatory diseases. Recent studies have shown that over representation of calcitonin gene-related peptides (CGRP) may activate nociceptive signaling following nerve injury. Silent information regulator 1 (SIRT1) is ubiquitously expressed in the dorsal horn of the spinal cord and plays a role in the pathogenesis of LDH. In this study, we investigated the analgesic effects of PD1 and elucidated the impact of neurogenic inflammation in the pathogenesis of neuropathic pain induced by non-compressive lumbar disc herniation (NCLDH) in a rat model. Methods. NCLDH models were established by applying protruding autologous nucleus pulposus to the L5 Dorsal root ganglion (DRG). PD1, SIRT1 antagonist or agonist, CGRP or antagonist were administered as daily intrathecal injections for three consecutive days postoperatively. Behavioral tests were conducted to assess mechanical and thermal hyperalgesia. The ipsilateral lumbar (L4-6) segment of the spinal dorsal horn was isolated for further analysis. Alterations in the release of SIRT1 and CGRP were explored using western blot and immunofluorescence. Results. Application of protruded nucleus (NP) materials to the DRG induced mechanical and thermal allodynia symptoms, and deregulated the expression of pro-inflammatory and anti-inflammatory cytokines in rats. Intrathecal delivery of PD1 significantly reversed the NCLDH-induced imbalance in neuro-inflammatory response and alleviated the symptoms of mechanical and thermal hyperalgesia. In addition, NP application to the DGRs resulted the spinal upregulation of CGRP and SIRT1 expression, which was almost restored by intrathecal injection of PD1 in a dose-dependent manner. SIRT1 antagonist or agonist and CGRP or antagonist treatment further confirmed the result. Conclusion. Our findings indicate PD1 has a potent analgesic effect, and can modulate neuro-inflammation by regulating SIRT1-mediated CGRP signaling in NCLDH.

Four new hypogean species of the genus Triplophysa (Osteichthyes, Cypriniformes, Nemacheilidae) from Guizhou Province, Southwest China, based on molecular and morphological data.

Recently described cave species of the genus Triplophysa have been discovered in southwestern China, suggesting that the diversity of the genus is severely underestimated and that there may be many undescribed species. In this work, four new species of the genus Triplophysa are described from southwestern Guizhou Province, China, namely Triplophysacehengensis Luo, Mao, Zhao, Xiao & Zhou, sp. nov. and Triplophysarongduensis Mao, Zhao, Yu, Xiao & Zhou, sp. nov. from Rongdu Town, Ceheng County, Guizhou, Triplophysapanzhouensis Yu, Luo, Lan, Xiao & Zhou, sp. nov. from Hongguo Town, Panzhou City, Guizhou, and Triplophysaanlongensis Song, Luo, Lan, Zhao, Xiao & Zhou, sp. nov. from Xinglong Town, Anlong County, Guizhou. These four new species can be distinguished from all recognized congeners by a combination of morphological characteristics and significant genetic divergences. The discovery of these species increases the number of known cave species within the genus Triplophysa to 39, making the genus the second most diverse group of cave fishes in China after the golden-line fish genus Sinocyclocheilus. Based on the non-monophyletic relationships of the different watershed systems in the phylogenetic tree, this study also discusses the use of cave species of the genus Triplophysa to determine the possible historical connectivity of river systems.

Surface-Enhanced Raman Scattering-Based Surface Chemotaxonomy: Combining Bacteria Extracellular Matrices and Machine Learning for Rapid and Universal Species Identification.

Rapid, universal, and accurate identification of bacteria in their natural states is necessary for on-site environmental monitoring and fundamental microbial research. Surface-enhanced Raman scattering (SERS) spectroscopy emerges as an attractive tool due to its molecule-specific spectral fingerprinting and multiplexing capabilities, as well as portability and speed of readout. Here, we develop a SERS-based surface chemotaxonomy that uses bacterial extracellular matrices (ECMs) as proxy biosignatures to hierarchically classify bacteria based on their shared surface biochemical characteristics to eventually identify six distinct bacterial species at >98% classification accuracy. Corroborating with in silico simulations, we establish a three-way inter-relation between the bacteria identity, their ECM surface characteristics, and their SERS spectral fingerprints. The SERS spectra effectively capture multitiered surface biochemical insights including ensemble surface characteristics, e.g., charge and biochemical profiles, and molecular-level information, e.g., types and numbers of functional groups. Our surface chemotaxonomy thus offers an orthogonal taxonomic definition to traditional classification methods and is achieved without gene amplification, biochemical testing, or specific biomarker recognition, which holds great promise for point-of-need applications and microbial research.

Apple consumption affects cecal health by regulating 12S-hydroxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S)-HETE) levels through modifying the microbiota in rats.

Apples are rich in many nutrients and functional components. However, the mechanism of the effect of fresh apple consumption on rats remains unclear. In the present study, fresh apples (10 g kg-1) were added to the diet of Wistar rats, and changes in the microbiota and metabolite content of the cecum were analyzed after 28 days of feeding, and changes in the 12S-hydroxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S)-HETE) content and indicators related to inflammation, oxidative stress, and apoptosis were detected. Subsequently, a fecal microbiota transplantation (FMT) protocol was designed and carried out to verify the relationship between the microbiota and 12(S)-HETE, the cecal structure, and inflammatory factors. The results show that apple consumption significantly reduced the serum levels of alanine aminotransferase (ALT) and immunoglobulin G (IgG), altered the cecal histomorphology, and significantly upregulated the gene expression of claudin-1 and zonula occludens-1 (ZO-1), which encode tight junction proteins. Apple consumption also changed the structure of the cecal microbiota, increasing the abundance of some species (such as Shuttleworthia) and decreasing the abundance of others (such as Alphaproteobacteria). Metabolomic screening identified 64 significantly different metabolites. The FMT results showed that apple consumption reduced 12(S)-HETE metabolite levels in the cecal contents, improved the intestinal structure, and reduced the levels of proinflammatory factor expression by altering the cecal microbiota. In conclusion, this study provides further insight into the effects of apples on animals using rats as experimental animals. It provides basic data for future exploration of the mechanisms of the effect of apple consumption on humans.

SUMOylation of Smad2 mediates TGF-ß-regulated endothelial-mesenchymal transition.

Endothelial-mesenchymal transition (EndoMT) is a complex biological process in which endothelial cells are transformed into mesenchymal cells, and dysregulated EndoMT causes a variety of pathological processes. Transforming growth factor beta (TGF-ß) signaling effectively induces the EndoMT process in endothelial cells, and Smad2 is the critical protein of the TGF-ß signaling pathway. However, whether small ubiquitin-like modifier modification (SUMOylation) is involved in EndoMT remains unclear. Here, we show that Smad2 is predominantly modified by SUMO1 at two major SUMOylation sites with PIAS2α as the primary E3 ligase, whereas SENP1 (sentrin/SUMO-specific protease 1) mediates the deSUMOylation of Smad2. In addition, we identified that SUMOylation significantly enhances the transcriptional activity and protein stability of Smad2, regulating the expression of downstream target genes. SUMOylation increases the phosphorylation of Smad2 and the formation of the Smad2-Smad4 complex, thus promoting the nuclear translocation of Smad2. Ultimately, the wildtype, but not SUMOylation site mutant Smad2 facilitated the EndoMT process. More importantly, TGF-ß enhances the nuclear translocation of Smad2 by enhancing its SUMOylation and promoting the EndoMT process. These results demonstrate that SUMOylation of Smad2 plays a critical role in the TGF-ß-mediated EndoMT process, providing a new theoretical basis for the treatment and potential drug targets of EndoMT-related clinical diseases.

Achieving Molecular Recognition of Structural Analogues in Surface-Enhanced Raman Spectroscopy: Inducing Charge and Geometry Complementarity to Mimic Molecular Docking.

Molecular recognition of complex isomeric biomolecules remains challenging in surface-enhanced Raman scattering (SERS) spectroscopy due to their small Raman cross-sections and/or poor surface affinities. To date, the use of molecular probes has achieved excellent molecular sensitivities but still suffers from poor spectral specificity. Here, we induce "charge and geometry complementarity" between probe and analyte as a key strategy to achieve high spectral specificity for effective SERS molecular recognition of structural analogues. We employ 4-mercaptopyridine (MPY) as the probe, and chondroitin sulfate (CS) disaccharides with isomeric sulfation patterns as our proof-of-concept study. Our experimental and in silico studies reveal that "charge and geometry complementarity" between MPY's binding pocket and the CS sulfation patterns drives the formation of site-specific, multidentate interactions at the respective CS isomerism sites, which "locks" each CS in its analogue-specific complex geometry, akin to molecular docking events. Leveraging the resultant spectral fingerprints, we achieve > 97 % classification accuracy for 4 CSs and 5 potential structural interferences, as well as attain multiplex CS quantification with < 3 % prediction error. These insights could enable practical SERS differentiation of biologically important isomers to meet the burgeoning demand for fast-responding applications across various fields such as biodiagnostics, food and environmental surveillance.

Modular Click Assembly DNA-Encoded Glycoconjugate Libraries with on-DNA Functional Group Transformations.

Carbohydrates are an important class of naturally active products and play vital roles in regulating various physiological activities. To meet the demand for carbohydrate-based libraries used for the identification of potential drug candidates for pharmaceutical-related targets, we developed a set of on-DNA protocols to construct the DNA-encoded glycoconjugates, including Seyferth-Gilbert homologation, anomeric azidation, and CuAAC cyclization. These on-DNA chemistries enable the generation and modification of DNA-linked glycosyl compounds with good conversions and broad substrate scope. Finally, three DNA-linked glycoconjugate libraries were successfully generated to demonstrate their applicability and feasibility in library preparation.

A method using deep learning to discover new predictors from left-ventricular mechanical dyssynchrony for CRT response.

BACKGROUND: Studies have shown that the conventional parameters characterizing left ventricular mechanical dyssynchrony (LVMD) measured on gated SPECT myocardial perfusion imaging (MPI) have their own statistical limitations in predicting cardiac resynchronization therapy (CRT) response. The purpose of this study is to discover new predictors from the polarmaps of LVMD by deep learning to help select heart failure patients with a high likelihood of response to CRT. METHODS: One hundred and fifty-seven patients who underwent rest gated SPECT MPI were enrolled in this study. CRT response was defined as an increase in left ventricular ejection fraction (LVEF) > 5% at 6 [Formula: see text] 1 month follow up. The autoencoder (AE) technique, an unsupervised deep learning method, was applied to the polarmaps of LVMD to extract new predictors characterizing LVMD. Pearson correlation analysis was used to explain the relationships between new predictors and existing clinical parameters. Patients from the IAEA VISION-CRT trial were used for an external validation. Heatmaps were used to interpret the AE-extracted feature. RESULTS: Complete data were obtained in 130 patients, and 68.5% of them were classified as CRT responders. After variable selection by feature importance ranking and correlation analysis, one AE-extracted LVMD predictor was included in the statistical analysis. This new AE-extracted LVMD predictor showed statistical significance in the univariate (OR 2.00, P = .026) and multivariate (OR 1.11, P = .021) analyses, respectively. Moreover, the new AE-extracted LVMD predictor not only had incremental value over PBW and significant clinical variables, including QRS duration and left ventricular end-systolic volume (AUC 0.74 vs 0.72, LH 7.33, P = .007), but also showed encouraging predictive value in the 165 patients from the IAEA VISION-CRT trial (P < .1). The heatmaps for calculation of the AE-extracted predictor showed higher weights on the anterior, lateral, and inferior myocardial walls, which are recommended as LV pacing sites in clinical practice. CONCLUSIONS: AE techniques have significant value in the discovery of new clinical predictors. The new AE-extracted LVMD predictor extracted from the baseline gated SPECT MPI has the potential to improve the prediction of CRT response.

The efficacy of the combination of radiofrequency ablation and endoscopic metal stent in the treatment of 44 patients with unresectable cholangiocarcinoma / 中华消化杂志

Objective:To explore the efficacy of the combination of radiofrequency ablation(RFA) and endoscopic metal stent in the treatment of patients with unresectable cholangiocarcinoma.Methods:From January 3, 2012 to June 30, 2019, at the Department of Endoscopic of the Third Affiliated Hospital of Naval Medical University, the clinical data of 44 patients with unresectable cholangiocarcinoma who were treated by the combination of RFA and endoscopic metal stent were retrospectively collected, which included age, gender, location of cholangiocarcinoma(hilar cholangiocarcinoma and distal cholangiocarcinoma), etc. Postoperative evaluation was conducted based on the follow-up, including clinical success rate, postoperative complication rate, time of stent patency and overall survival time (OS). The Kaplan-Meier method and log-rank test were used to analyze the difference of OS between patients with hilar cholangiocarcinoma and distal cholangiocarcinoma. Mann-Whitney U test was used for statistical analysis. Results:The age of the 44 patients with cholangiocarcinoma was (70.3±11.6) years old, with 20 males (45.5%). There were 22 patients (50.0%) with hilar cholangiocarcinoma and 22 patients (50.0%) with distal cholangiocarcinoma. The clinical success rate of 44 patients was 93.2%(41/44). A total of 5 patients(11.4%) had postoperative complications, which were all improved by appropriate treatment. The median time of follow-up of the 44 patient was 9.2 months(ranged from 3.1 to 57.6 months), the median time of stent patency was 7.0 months (ranged from 5.8 to 8.2 months). Thirty-two patients (72.7%) died during the follow-up, and the median OS was 10.9 months(ranged from 9.0 to 12.8 months). The median OS of patients with hilar cholangiocarcinoma was 7.8 months(ranged from 4.6 to 11.0 months) and that of patients with distal cholangiocarcinoma was 12.5 months(ranged from 5.7 to 19.4 months), and there was no statistically significant difference( P>0.05). Conclusion:RFA combined with endoscopic metal stent is safe and effective in the treatment of patients with unresectable cholangiocarcinoma.

A long term follow-up study of endoscopic radiofrequency ablation for unresectable ampullary carcinoma / 中华消化内镜杂志

Objective:To explore the long-term effect of endoscopic radiofrequency ablation for the treatment of unresectable ampullary carcinoma.Methods:Clinical data of patients with ampullary carcinoma who received endoscopic radiofrequency ablation in the Third Affiliated Hospital of Naval Medical University from January 2012 to May 2019 were retrospectively collected, including basic information, frequency of radiofrequency ablation, the type of biliary stent, postoperative complications, and follow-up. Kaplan-Meier method was used to analyze the survival of patients after endoscopic radiofrequency ablation. Relationship between frequency of radiofrequency ablation, type of biliary stent and overall survival time was analyzed.Results:A total of 50 patients were enrolled, including 31 males and 19 females, aged 73.0±9.7 years. Twenty-five patients (50.0%) underwent 1 radiofrequency ablation treatment, while 25 patients (50.0%) underwent radiofrequency ablation treatments more than twice. Postoperative complications occurred in 6 patients (12.0%), all of which were mild symptoms. The average follow-up was 22.3 months, with a total of 39 (78.0%) deaths, 5 (10.0%) lost to follow-up, and 6 (12.0%) surviving. The median overall survival time was 16.9 (95% CI: 9.1-24.8) months, with cumulative survival rates of 62.0%, 38.5%, 27.0%, and 12.6% at 1, 2, 3, and 5 years, respectively. The median overall survival time of those treated with radiofrequency ablation ≥2 times showed a trend of prolongation compared to patients treated once, but the difference was not statistically significant [26.7 (95% CI: 9.7-43.7) months VS 12.6 (95% CI: 4.9-20.3) months, χ2=3.049, P=0.081]. Plastic stents were used in 32 patients (64.0%) and metal stents in 18 patients (36.0%). There was no significant difference in median overall survival time between patients using metal and plastic stents [17.1 (95% CI: 6.1-28.0) months VS 15.9 (95% CI: 6.9-24.9) months， χ2=0.029， P=0.865]. Conclusion:Endoscopic radiofrequency ablation is a safe treatment for unresectable ampullary carcinoma, and multiple consecutive treatments may increase the survival benefit.

Efficacy of endoscopic stent placement for Bismuth type Ⅳ hilar cholangiocarcinoma: a large-scale retrospective study / 中华消化内镜杂志

Objective:To investigate the efficacy of endoscopic stent placement for patients with Bismuth type Ⅳ hilar cholangiocarcinoma.Methods:Data of 229 patients with unresectable Bismuth type Ⅳ hilar cholangiocarcinoma who successfully underwent endoscopic stent placement at the Department of Endoscopy, the Third Affiliated Hospital of Naval Medical University from January 2002 to January 2019 were retrospectively analyzed. Outcomes included clinical success rate, complication incidence, stent patency period and overall survival time. The patency of stents and overall survival time of patients were estimated by using the Kaplan-Meier method. The independent predictors for stent patency and overall survival of patients were analyzed by a multivariate Cox proportional regression model.Results:The overall clinical success rate was 78.2% (179/229). The incidence of early cholangitis after endoscopic retrograde cholangiopancreatography was 20.5% (47/229). The median stent patency and overall survival time were 5.7 (95% CI: 4.8-6.7) months and 5.1 (95% CI: 4.2-6.0) months, respectively. Further multivariate Cox regression analysis showed that metal stent ( P<0.001, HR=0.452, 95% CI: 0.307-0.666) and bilateral stents with bilateral angiography ( P=0.036, HR=0.644, 95% CI: 0.427-0.971) were independent predictors of stent patency; total bilirubin>200 μmol/L ( P=0.001, HR=1.627, 95% CI: 1.208-2.192), metal stent ( P=0.004, HR=0.636, 95% CI: 0.467-0.866) and antitumor therapy ( P<0.001, HR=0.439, 95% CI:0.308-0.626) were independent predictors of overall survival. Conclusion:There is high incidence of cholangitis in patients with unresectable Bismuth type Ⅳ hilar cholangiocarcinoma treated with endoscopic stenting. Longer stent patency can be achieved with metal stent placement and bilateral drainage. In addition, metal stent for drainage and antitumor therapy can also help increase the survival benefit.