Identification of G-quadruplex-interacting proteins in living cells using an artificial G4-targeting biotin ligase.

G-quadruplexes (G4s) are noncanonical nucleic acid structures pivotal to cellular processes and disease pathways. Deciphering G4-interacting proteins is imperative for unraveling G4's biological significance. In this study, we developed a G4-targeting biotin ligase named G4PID, meticulously assessing its binding affinity and specificity both in vitro and in vivo. Capitalizing on G4PID, we devised a tailored approach termed G-quadruplex-interacting proteins specific biotin-ligation procedure (PLGPB) to precisely profile G4-interacting proteins. Implementing this innovative strategy in live cells, we unveiled a cohort of 149 potential G4-interacting proteins, which exhibiting multifaceted functionalities. We then substantiate the directly binding affinity of 7 candidate G4-interacting-proteins (SF3B4, FBL, PP1G, BCL7C, NDUV1, ILF3, GAR1) in vitro. Remarkably, we verified that splicing factor 3B subunit 4 (SF3B4) binds preferentially to the G4-rich 3' splice site and the corresponding splicing sites are modulated by the G4 stabilizer PDS, indicating the regulating role of G4s in mRNA splicing procedure. The PLGPB strategy could biotinylate multiple proteins simultaneously, which providing an opportunity to map G4-interacting proteins network in living cells.

Inert Pepper aptamer-mediated endogenous mRNA recognition and imaging in living cells.

The development of RNA aptamers/fluorophores system is highly desirable for understanding the dynamic molecular biology of RNAs in vivo. Peppers-based imaging systems have been reported and applied for mRNA imaging in living cells. However, the need to insert corresponding RNA aptamer sequences into target RNAs and relatively low fluorescence signal limit its application in endogenous mRNA imaging. Herein, we remolded the original Pepper aptamer and developed a tandem array of inert Pepper (iPepper) fluorescence turn-on system. iPepper allows for efficient and selective imaging of diverse endogenous mRNA species in live cells with minimal agitation of the target mRNAs. We believe iPepper would significantly expand the applications of the aptamer/fluorophore system in endogenous mRNA imaging, and it has the potential to become a powerful tool for real-time studies in living cells and biological processing.

The Transcriptome-Wide Mapping of 2-Methylthio-N6-isopentenyladenosine at Single-Base Resolution.

Hundreds of modified bases have been identified and enzymatically modified to transfer RNAs (tRNAs) to regulate RNA function in various organisms. 2-Methylthio-N6-isopentenyladenosine (ms2i6A), a hypermodified base found at tRNA position 37, exists in both prokaryotes and eukaryotes. ms2i6A is traditionally identified by separating and digesting each tRNA from total RNA using RNA mass spectrometry. A transcriptome-wide and single-base resolution method that enables absolute mapping of ms2i6A along with analysis of its distribution in different RNAs is lacking. Here, through chemoselective methylthio group bioconjugation, we introduce a new approach (redox activated chemical tagging sequencing, ReACT-seq) to detect ms2i6A transcriptome-wide at single-base resolution. Using the chemoselectivity between the methylthio group and oxaziridine group, ms2i6A is bio-orthogonally tagged with an azide group without interference of canonical nucleotides, advancing enrichment of methylthio group modified RNAs prior to sequencing. ReACT-seq was demonstrated on nine known tRNAs and proved to be highly accurate, and the reverse transcription stop (RT-stop) character enables ReACT-seq detection at single-base resolution. In addition, ReACT-seq identified that the modification of ms2i6A is conservative and may not exist in other RNAs.

Antibody-Free Fluorine-Assisted Metabolic Sequencing of RNA N4-Acetylcytidine.

N4-Acetylcytidine (ac4C) has been found to affect a variety of cellular and biological processes. For a mechanistic understanding of the roles of ac4C in biology and disease, we present an antibody-free, fluorine-assisted metabolic sequencing method to detect RNA ac4C, called "FAM-seq". We successfully applied FAM-seq to profile ac4C landscapes in human 293T, HeLa, and MDA cell lines in parallel with the reported acRIP-seq method. By comparison with the classic ac4C antibody sequencing method, we found that FAM-seq is a convenient and reliable method for transcriptome-wide mapping of ac4C. Because this method holds promise for detecting nascent RNA ac4C modifications, we further investigated the role of ac4C in regulating chemotherapy drug resistance in chronic myeloid leukemia. The results indicated that drug development or combination therapy could be enhanced by appreciating the key role of ac4C modification in cancer therapy.

Sonographic morphological and qualitative deficits in the elbow ulnar collateral ligament and ulnohumeral joint in throwing arms of asymptomatic collegiate baseball pitchers.

OBJECTIVE: The ulnar collateral ligament (UCL) supports the medial elbow against valgus torque and is commonly injured in baseball pitchers. Changes in UCL morphology and pathology occur with long-term pitching, with more severe findings at higher competition levels. We examined the bilateral differences and the relationship between UCL morphology, pathology, and ulnohumeral joint laxity in asymptomatic collegiate pitchers using ultrasound. MATERIALS AND METHODS: Division I college pitchers (n = 41) underwent ultrasound scans of their bilateral medial elbows, both at rest and in a valgus-stressed position. The presence of enthesopathy, calcifications, and degeneration was assessed qualitatively. UCL thickness and ulnohumeral joint gap were measured with online calipers. The bilateral differences were analyzed using paired t-tests and chi-square analysis, and the relationships between thickness, gapping, and degenerative changes were analyzed using regression analyses. RESULTS: The throwing arm demonstrated greater distal UCL thickness (mean difference (MD) = 0.2 mm (95%CI = 0.1-0.3), p < 0.01), resting and stressed gap (MD = 0.3 mm (95%CI = 0.0-0.7), p = 0.04; MD = 0.4 (95%CI = 0.0-0.9), p = 0.02), and greater prevalence of degeneration and enthesopathy (p = 0.03) compared bilaterally. Enthesopathy and calcifications predicted increased distal UCL thickness (p = 0.04; p = 0.02). Degenerative scores predicted increased stressed-resting ulnohumeral joint gap (p < 0.01). CONCLUSION: In the throwing arms of collegiate pitchers, ultrasound demonstrated UCL thickening, enthesopathy/intra-ligamentous calcification, and greater laxity of the ulnohumeral joint relative to the non-throwing arm. Degeneration of the UCL, not thickness, was related to greater elbow joint gapping. This study demonstrates the utility of ultrasound for examining sonographic characteristics of the UCL in a sample of college pitchers.

Band Structure Engineering within Two-Dimensional Borocarbonitride Nanosheets for Surface-Enhanced Raman Scattering.

Herein, with two-dimensional (2D) borocarbonitride (BCN) as a metal- and plasmon-free surface-enhanced Raman scattering (SERS) platform, we demonstrate a band structure engineering strategy to facilitate the charge transfer process for an enhanced SERS response. Especially, when the conduction band of the BCN substrate is tuned to align with the LUMO of the target molecule, remarkable SERS performance is achieved, ascribed to the borrowing effect from the vibronic coupling of resonances through the Herzberg-Teller coupling term. Meanwhile, fluorescence quenching is achieved due to the efficient charge transfer between the BCN substrate and target molecule. Consequently, BCN can accurately detect 20 kinds of trace chemical and bioactive analytes. Moreover, BCN exhibits excellent thermal and chemical stability, which can not only withstand high-temperature (300 °C) heating in the air but also resist long-term corrosion in harsh acid (pH = 0, HCl) and base (pH = 14, NaOH). This work provides new insight into band structure engineering in promoting the SERS performance of plasmon- and metal-free semiconductor substrates.

Photoactive G-Quadruplex Ligand Identifies Multiple G-Quadruplex-Related Proteins with Extensive Sequence Tolerance in the Cellular Environment.

G-Quadruplex (G4) is a noncanonical nucleic acid secondary structure with multiple biofunctions. Identifying G4-related proteins (G4RPs) is important for understanding the roles of G4 in biology. Current methods to identify G4RPs include discovery from specific biological processes or in vitro pull-down assays with specific G4 sequences. Here, we report an in vivo strategy used to identify G4RPs with extensive sequence tolerance based on G4 ligand-mediated cross-linking. Applying this method, we identified 114 and 281 G4RPs in SV589 and MM231 cells, respectively. The results successfully overlapped with all the pull-down assay literature. Through the electrophoretic mobility shift assay (EMSA), we identified some new G4-binding proteins. Moreover, enhanced cross-linking and immunoprecipitation (eCLIP) confirmed that one newly identified G4-binding protein, SERBP1, interacts with G4 in the cellular environment. The method we developed provides a new strategy for identifying proteins that interact with nucleic secondary structures in cells and benefit the study of their biological roles.

A Methyl Scan of the Pyrrolidinium Ring of Nicotine Reveals Significant Differences in Its Interactions with α7 and α4ß2 Nicotinic Acetylcholine Receptors.

The two major nicotinic acetylcholine receptors (nAChRs) in the brain are the α4ß2 and α7 subtypes. A "methyl scan" of the pyrrolidinium ring was used to detect differences in nicotine's interactions with these two receptors. Each methylnicotine was investigated using voltage-clamp and radioligand binding techniques. Methylation at each ring carbon elicited unique changes in nicotine's receptor interactions. Replacing the 1'-N-methyl with an ethyl group or adding a second 1'-N-methyl group significantly reduced interaction with α4ß2 but not α7 receptors. The 2'-methylation uniquely enhanced binding and agonist potency at α7 receptors. Although 3'- and 5'-trans-methylations were much better tolerated by α7 receptors than α4ß2 receptors, 4'-methylation decreased potency and efficacy at α7 receptors much more than at α4ß2 receptors. Whereas cis-5'-methylnicotine lacked agonist activity and displayed a low affinity at both receptors, trans-5'-methylnicotine retained considerable α7 receptor activity. Differences between the two 5'-methylated analogs of the potent pyridyl oxymethylene-bridged nicotine analog A84543 were consistent with what was found for the 5'-methylnicotines. Computer docking of the methylnicotines to the Lymnaea acetylcholine binding protein crystal structure containing two persistent waters predicted most of the changes in receptor affinity that were observed with methylation, particularly the lower affinities of the cis-methylnicotines. The much smaller effects of 1'-, 3'-, and 5'-methylations and the greater effects of 2'- and 4'-methylations on nicotine α7 nAChR interaction might be exploited for the design of new drugs based on the nicotine scaffold. SIGNIFICANCE STATEMENT: Using a comprehensive "methyl scan" approach, we show that the orthosteric binding sites for acetylcholine and nicotine in the two major brain nicotinic acetylcholine receptors interact differently with the pyrrolidinium ring of nicotine, and we suggest reasons for the higher affinity of nicotine for the heteromeric receptor. Potential sites for nicotine structure modification were identified that may be useful in the design of new drugs targeting these receptors.

Cost-effectiveness of MR Imaging-guided Strategies for Detection of Prostate Cancer in Biopsy-Naive Men.

Purpose To evaluate the cost-effectiveness of multiparametric diagnostic magnetic resonance (MR) imaging examination followed by MR imaging-guided biopsy strategies in the detection of prostate cancer in biopsy-naive men presenting with clinical suspicion of cancer for the first time. Materials and Methods A decision-analysis model was created for biopsy-naive men who had been recommended for prostate biopsy on the basis of abnormal digital rectal examination results or elevated prostate-specific antigen levels (age groups: 41-50 years, 51-60 years, and 61-70 years). The following three major strategies were evaluated: (a) standard transrectal ultrasonography (US)-guided biopsy; (b) diagnostic MR imaging followed by MR imaging-targeted biopsy, with no biopsy performed if MR imaging findings were negative; and (c) diagnostic MR imaging followed by MR imaging-targeted biopsy, with a standard biopsy performed when MR imaging findings were negative. The following three MR imaging-guided biopsy strategies were further evaluated in each MR imaging category: (a) biopsy with cognitive guidance, (b) biopsy with MR imaging/US fusion guidance, and (c) in-gantry MR imaging-guided biopsy. Model parameters were derived from the literature. The primary outcome measure was net health benefit (NHB), which was measured as quality-adjusted life-years (QALYs) gained or lost by investing resources in a new strategy compared with a standard strategy at a willingness-to-pay (WTP) threshold of $50 000 per QALY gained. Probabilistic sensitivity analysis was performed by using Monte Carlo simulations. Results Noncontrast MR imaging followed by cognitively guided MR biopsy (no standard biopsy if MR imaging findings were negative) was the most cost-effective approach, yielding an additional NHB of 0.198 QALY compared with the standard biopsy approach. Noncontrast MR imaging followed by in-gantry MR imaging-guided biopsy (no standard biopsy if MR imaging findings were negative) led to the highest NHB gain of 0.251 additional QALY compared with the standard biopsy strategy. All MR imaging strategies were cost-effective in 94.05% of Monte Carlo simulations. Analysis by age groups yielded similar results. Conclusion MR imaging-guided strategies for the detection of prostate cancer were cost-effective compared with the standard biopsy strategy in a decision-analysis model. © RSNA, 2017 Online supplemental material is available for this article.

Real-time MRI-guided percutaneous sclerotherapy of low-flow head and neck lymphatic malformations in the pediatric population - a stepwise approach.

Real-time MRI-guided percutaneous sclerotherapy is a novel and evolving treatment for congenital lymphatic malformations in the head and neck. We elaborate on the specific steps necessary to perform an MRI-guided percutaneous sclerotherapy of lymphatic malformations including pre-procedure patient work-up and preparation, stepwise intraprocedural interventional techniques and post-procedure management. Based on our institutional experience, MRI-guided sclerotherapy with a doxycycline-gadolinium-based mixture as a sclerosant for lymphatic malformations of the head and neck region in children is well tolerated and effective.

Andrographolide Inhibits Static Mechanical Pressure-Induced Intervertebral Disc Degeneration via the MAPK/Nrf2/HO-1 Pathway.

Purpose: To explore the molecular mechanism by which andrographolide (ADR) inhibits static mechanical pressure-induced apoptosis in nucleus pulposus cells (NPCs) and to assess the role of ADR in inhibiting IDD. Methods: Hematoxylin-eosin (HE), toluidine blue, and immunofluorescence staining were used to identify NPCs. An NPC apoptosis model was constructed using a homemade cell pressurization device. The proliferation activity, reactive oxygen species (ROS) content, and apoptosis rate were detected using kits. The expression of related proteins was detected using Western blot. A rat tailbone IDD model was constructed using a homemade tailbone stress device. HE staining and safranine O-fast green FCF cartilage staining were used to observe the degeneration degree of the intervertebral disk. Results: ADR inhibits static mechanical pressure-induced apoptosis and ROS accumulation in NPCs and improves cell viability. ADR can promote the expression of Heme oxygenase-1 (HO-1), p-Nrf2, p-p38, p-Erk1/2, p-JNK, and other proteins, and its effects can be blocked by inhibitors of the above proteins. Conclusion: ADR can inhibit IDD by activating the MAPK/Nrf2/HO-1 signaling pathway and suppressing static mechanical pressure-induced ROS accumulation in the NPCs.

Relationships between Perceived Discrimination and Suicidal Ideation among Impoverished Chinese College Students: The Mediating Roles of Social Support and Loneliness.

We explored the mediating effect of social support and loneliness in the relationships between perceived discrimination and suicidal ideation among impoverished Chinese college students. Using the convenience cluster sampling method, we chose a total of 964 impoverished college students from a central province of China. Students completed the cross-sectional survey using the Perceived Discrimination Questionnaire, the Social Support Rating Scale, the University of California at Los Angeles Loneliness Scale, and the Beck Scale for Suicidal Ideation. Correlation analysis and structural equation modeling analysis were conducted to clarify the relationships between study variables. Correlation analysis showed that perceived discrimination, loneliness, and suicidal ideation were positively correlated with each other; social support was negatively correlated with perceived discrimination, loneliness, and suicidal ideation. In addition, structural equation modeling analysis indicated that perceived discrimination had a direct positive effect on suicidal ideation; social support and loneliness partially mediated the relationship between perceived discrimination and suicidal ideation. Specifically, perceived discrimination was positively associated with suicidal ideation via social support and loneliness separately, and had a serial association through both social support and loneliness. Thus, perceived discrimination may have influenced suicidal ideation through both social support and loneliness.

Longitudinal relationships between bullying and prosocial behavior: The mediating roles of trauma-related guilt and shame.

The present study examined the relationships between bullying, trauma-related guilt, trauma-related shame, and prosocial behaviors. We investigated 1,322 college students using a longitudinal approach to explore the internal mechanism between bullying, prosocial behaviors, and the probable mediating effects of trauma-related guilt and shame. The results suggested that bullying negatively predicted prosocial behaviors and that trauma-related guilt played a positive mediating role. In contrast, trauma-related shame played a negative mediating role in the relationship between bullying and prosocial behaviors. These findings indicated that trauma-related guilt and shame played adaptive and maladaptive roles after bullying victimization, which also provided a theoretical basis for the relevant intervention.

Lucent Patellar Lesions: A Pictorial Review.

A radiographically lucent patellar lesion may represent a variety of etiologies, ranging from more commonly seen degenerative, metabolic, infectious, developmental, posttraumatic, postoperative causes to rarer benign and malignant neoplasms. Clinical symptoms, surgical history, laboratory values, and radiographic features may help narrow the differential. In addition, radiographic features such as circumscribed borders and sharply delineated margins favor benign lesions while ill-defined margins suggest malignant etiologies. This case series illustrates the imaging findings and explores relevant clinical findings in a variety of interesting lucent patellar lesions.

Wogonin inhibits inflammation and apoptosis through STAT3 signal pathway to promote the recovery of spinal cord injury.

Spinal cord injury (SCI) is a complex central traumatic disease. STAT3 signal transduction pathway plays an important role in SCI. Wogonin has been reported to exhibit neuroprotection. However, the molecular mechanism of its potential therapeutic effect after SCI remains unclear. In this study, rats were divided into the following groups: Sham; SCI; SCI + wogonin; and SCI + wogonin + colivelin (Colivelin is an effective activator of the STAT3 pathway). Motor function was evaluated by Basso Beattie Bresnahan (BBB) score. Histomorphological changes in the spinal cords were observed by Hematoxylin-eosin (HE) staining and Nissl staining. Western blot, Transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining, and immunofluorescence were used to detect changes in the neuronal inflammation, apoptosis, and STAT3 signal pathway. Western blot and immunofluorescence techniques were also performed to detect the regulatory effect and the underlying mechanism of wogonin on the inflammation and apoptosis of PC12 cells. Experimental results in vivo and in vitro showed that wogonin could promote the recovery of motor function, improve the histopathological morphology, inhibit the activation of the STAT3 signal pathway, and reduce the neuronal inflammation and apoptosis in the rats with SCI. Activation of the STAT3 signal pathway by colivelin reversed the therapeutic effect of wogonin. Therefore, wogonin could inhibit inflammation and apoptosis by inhibiting the STAT3 signal pathway and promote the functional recovery of rats with SCI.

Utility of Large Diameter Visible Trephine in Percutaneous Endoscopic Lumbar Interbody Fusion: A Technical Report.

PURPOSE: In this study, a large diameter visible trephine was designed and used in percutaneous endoscopic lumbar interbody fusion to increase endoscopic bone decompression efficiency. Large diameter visible trephine-related technical notes and preliminary clinical experience are described. METHODS: A large diameter visible trephine was designed with normal diameter visible trephine as template. A total of 38 patients with lumbar degenerative diseases who underwent single-level percutaneous endoscopic lumbar interbody fusion with large or normal diameter visible trephine were included into a retrospective study. Operation time, bone decompression time, blood loss, intraoperative fluoroscopy, bone decompression fluoroscopy, and dura or nerve injury cases were recorded and analyzed statistically. Visual Analog Scale (VAS) scores and Oswestry Disability Index (ODI) were used to analyze the clinical outcomes of the 2 groups. RESULTS: The baseline data of the 2 groups were statistically similar. There was no significant difference in postoperative VAS and ODI scores between the 2 groups. Operation time and bone decompression time of large diameter visible trephine group were significantly shorter than that of normal diameter visible trephine group (P < 0.05). Intraoperative fluoroscopy times and bone decompression fluoroscopy times of large diameter visible trephine group were significantly more than that of normal diameter visible trephine group (P < 0.05). Blood loss of the 2 groups were not statistically different. There were no dura or nerve injury cases in the 2 groups. CONCLUSIONS: For percutaneous endoscopic lumbar interbody fusion, the large diameter visible trephine is a safe and efficient endoscopic bone decompression tool under fluoroscopic guidance.

Pt(IV) Prodrugs Designed to Embed in Nanotubes of a Polysaccharide for Drug Delivery.

Cisplatin exhibits a sufficient killing effect on cancer cells; however, it damages normal cells simultaneously. Herein, we developed a prodrug delivery system based on branched ß-(1→3)-d-glucan. This natural biomacromolecule-based polysaccharide nanotube was modified with cisplatin embedded in the hollow cavity (BFCP), showing high anticancer activity and low toxicity in vitro. It is a broad-prospect system, which is based on biocompatible nanomaterials loaded with Pt(IV) prodrugs for cancer cell absorption with subsequent release in tumors by utilizing the intracellular reducibility. BFCP chains adopted a nanotube conformation in water, observed by transmission electron microscopy. In comparison to cisplatin, the Pt(IV) prodrugs not only displayed better antitumor properties but also had significant tumor targeting. A potent natural complex conjugated with redox-responsive platinum prodrugs is a significantly efficient tumor drug demonstrated in vitro and in vivo.

Fast MRI breast cancer screening - Ready for prime time.

OBJECTIVE: The manuscript discusses landmark studies using abbreviated MRI for breast cancer screening. This includes abbreviated dynamic contrast enhanced MRI and diffusion weighted imaging. Our institutional experience with abbreviated MR protocol for breast cancer screening is also described. CONCLUSION: Abbreviated MRI protocols were found to demonstrate value for screening of breast cancer. It has been shown that abbreviated protocol MRI provides similar diagnostic sensitivities to full protocol MRI for breast cancer in women with increased lifetime risk. Our institutional abbreviated MRI protocol for breast cancer offers improved time and workflow efficiencies and has the potential to increase the number of breast cancers detected and the detection of pathologically relevant invasive breast cancer at earlier stages.

Recent Advances in Plasmon-Promoted Organic Transformations Using Silver-Based Catalysts.

Plasmonics has emerged as a promising methodology to promote chemical reactions and has become a field of intense research effort. Ag nanoparticles (NPs) as plasmonic catalysts have been extensively studied because of their remarkable optical properties. This review analyzes the emergence and development of localized surface plasmon resonance (LSPR) in organic chemistry, mainly focusing on the discovery of novel reactions with new mechanisms on Ag NPs. Initially, the basics of LSPR and LSPR-promoted photocatalytic mechanisms are illustrated. Then, the recent advances in plasmonic nanosilver-mediated photocatalysis in organic transformations are highlighted with an emphasis on the related reaction mechanisms. Finally, a proper perspective on the remaining challenges and future directions in the field of LSPR-promoted organic transformations is proposed.

In Situ Growth of Amorphous Fe(OH)3 on Nickel Nitrate Hydroxide Nanoarrays for Enhanced Electrocatalytic Oxygen Evolution.

Development of highly efficient electrocatalyst for the oxygen evolution reaction (OER) is urgently demanded by the clean hydrogen energy. Herein, in order to further boost the OER activity of metal nitrate hydroxide materials, amorphous Fe(OH)3 layer is in situ grown on nickel nitrate hydroxide (NiNH) nanoarrays supported on nickel foam (NF) through an interfacial hydrolysis approach, where the loading amount of the Fe(OH)3 can be simply manipulated by the hydrolysis time. Taking advantage of the synergy of Fe(OH)3 and NiNH, the optimized Fe(OH)3@NiNH/NF sample shows a very promising electrocatalytic OER activity in 1 M KOH solution, requiring a very low overpotential of 212 mV vs. reversible hydrogen electrode (RHE) to deliver a geometrical catalytic current density of 100 mA cm-2 and a low Tafel slope of 49 mV dec-1. This work provides a new strategy for boosting the electrocatalytic activity of metal hydroxide nitrates through the interface engineering.