Leucyl-tRNA Synthetase Contributes to Muscle Weakness through Mammalian Target of Rapamycin Complex 1 Activation and Autophagy Suppression in a Mouse Model of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD), caused by loss-of-function mutations in the dystrophin gene, results in progressive muscle weakness and early fatality. Impaired autophagy is one of the cellular hallmarks of DMD, contributing to the disease progression. Molecular mechanisms underlying the inhibition of autophagy in DMD are not well understood. In the current study, the DMD mouse model mdx is used for the investigation of signaling pathways leading to suppression of autophagy. Mammalian target of rapamycin complex 1 (mTORC1) is found to be hyperactive in the DMD muscles, accompanying muscle weakness and autophagy impairment. Surprisingly, Akt, a well-known upstream regulator of mTORC1, is not responsible for mTORC1 activation or the dystrophic muscle phenotypes. Instead, leucyl-tRNA synthetase (LeuRS) is found to be overexpressed in mdx muscles compared with the wild type. LeuRS is known to activate mTORC1 in a noncanonical mechanism that involves interaction with RagD, an activator of mTORC1. Disrupting LeuRS interaction with RagD by the small-molecule inhibitor BC-LI-0186 reduces mTORC1 activity, restores autophagy, and ameliorates myofiber damage in the mdx muscles. Furthermore, inhibition of LeuRS by BC-LI-0186 improves dystrophic muscle strength in an autophagy-dependent manner. Taken together, our findings uncover a noncanonical function of the housekeeping protein LeuRS as a potential therapeutic target in the treatment of DMD.

Spontaneous Rise of Hydrogen Microbubbles in Interfacial Gas Evolution Reaction.

Liquid organic hydrogen carrier is a promising option for the transport and storage of hydrogen as a clean energy source. This study examines the stability and behavior of organic drops immobilized on a substrate during an interfacial hydrogen-evolution reaction (HER) at the drop surface and its surrounding aqueous solution. Hydrogen microbubbles form within the drop and rise to the drop apex. The growth rate of the hydrogen in-drop bubble increases with the concentration of the reactant in the surrounding medium. The drop remains stable till the buoyancy acting on the in-drop bubble is large enough to overcome the capillary force and the external viscous drag. The bubble spontaneously rises and carries a portion drop liquid to the solution surface. These spontaneous rising in-drop bubbles are detected in measurements using a high-precision sensor placed on the upper surface of the aqueous solution, reversing the settling phase from phase separation in the reactive emulsion. The finding from this work provides new insights into the behaviors of drops and bubbles in many interfacial gas evolution reactions in clean technologies.

Glucose Oxidase Activity Colorimetric Assay Using Redox-Sensitive Electrochromic Nanoparticle-Functionalized Paper Sensors.

Glucose oxidase (GOx) activity assays are vital for various applications, including glucose metabolism estimation and fungal testing. However, conventional methods involve time-consuming and complex procedures. In this study, we present a colorimetric platform for in situ GOx activity measurement utilizing redox-sensitive electrochromic nanoparticles based on polyaniline (PAni). The glucose-adsorbed colorimetric paper sensor, herein termed Glu@CPS, is created by immobilizing ferrocene and glucose onto paper substrates that have been functionalized with PAni nanoparticles. Glu@CPS not only demonstrated rapid detection (within 5 min) but also exhibited remarkable selectivity for GOx and a limit of detection as low as 1.25 µM. Moreover, Glu@CPS demonstrated consistent accuracy in the measurement of GOx activity, exhibiting no deviations even after being stored at ambient temperature for a duration of one month. To further corroborate the effectiveness of this method, we applied Glu@CPS in the detection of GOx activity in a moldy red wine. The results highlight the promising potential of Glu@CPS as a convenient and precise platform for GOx activity measurement in diverse applications including food quality control, environmental monitoring, and early detection of fungal contamination.

Microfluidic Detection and Analysis of Microplastics Using Surface Nanodroplets.

Detection of microplastics from water is crucial for various reasons, such as food safety monitoring, monitoring of the fate and transport of microplastics, and development of preventive measures for their occurrence. Currently, microplastics are detected by isolating them using filtration, separation by centrifugation, or membrane filtration, subsequently followed by analysis using well-established analytical methods, such as Raman spectroscopy. However, due to their variability in shape, color, size, and density, isolation using the conventional methods mentioned above is cumbersome and time-consuming. In this work, we show a surface-nanodroplet-decorated microfluidic device for isolation and analysis of small microplastics (diameter of 10 µm) from water. Surface nanodroplets are able to capture nearby microplastics as water flows through the microfluidic device. Using a model microplastic solution, we show that microplastics of various sizes and types can be captured and visualized by using optical and fluorescence microscopy. More importantly, as the surface nanodroplets are pinned on the microfluidic channel, the captured microplastics can also be analyzed using a Raman spectroscope, which enables both physical (i.e., size and shape) and chemical (i.e., type) characterization of microplastics at a single-particle level. The technique shown here can be used as a simple, fast, and economical detection method for small microplastics.

Determination of Trace Organic Contaminant Concentration via Machine Classification of Surface-Enhanced Raman Spectra.

Surface-enhanced Raman spectroscopy (SERS) has been well explored as a highly effective characterization technique that is capable of chemical pollutant detection and identification at very low concentrations. Machine learning has been previously used to identify compounds based on SERS spectral data. However, utilization of SERS to quantify concentrations, with or without machine learning, has been difficult due to the spectral intensity being sensitive to confounding factors such as the substrate parameters, orientation of the analyte, and sample preparation technique. Here, we demonstrate an approach for predicting the concentration of sample pollutants from SERS spectra using machine learning. Frequency domain transform methods, including the Fourier and Walsh-Hadamard transforms, are applied to spectral data sets of three analytes (rhodamine 6G, chlorpyrifos, and triclosan), which are then used to train machine learning algorithms. Using standard machine learning models, the concentration of the sample pollutants is predicted with >80% cross-validation accuracy from raw SERS data. A cross-validation accuracy of 85% was achieved using deep learning for a moderately sized data set (∼100 spectra), and 70-80% was achieved for small data sets (∼50 spectra). Performance can be maintained within this range even when combining various sample preparation techniques and environmental media interference. Additionally, as a spectral pretreatment, the Fourier and Hadamard transforms are shown to consistently improve prediction accuracy across multiple data sets. Finally, standard models were shown to accurately identify characteristic peaks of compounds via analysis of their importance scores, further verifying their predictive value.

Technical Note on Simplified Free Gingival Graft Using Tack Fixation (sFGG).

Free gingival graft (FGG) is the gold standard procedure for the reliable augmentation of lost keratinized mucosa (KM) around dental implants. This conventional surgical approach has its drawbacks, including limitations in manipulation, the requirement for suturing, postoperative discomfort, and pain. This case report aimed to evaluate the efficacy of a simplified free gingival graft (sFGG) in addressing the issue of inadequate keratinized mucosa around dental implants. Fixation tacks were used to perform the sFGG procedure. Initially, a partial-thickness flap was created and apically repositioned. The gingival graft was harvested from the palate with a narrow profile and securely affixed to the recipient site using 5 mm long fixation tacks. Significant gains in keratinized mucosa were achieved and successfully maintained within 1 year. Consequently, the sFGG technique emerges as a simple and reliable treatment approach for managing inadequate keratinized mucosa around dental implants.

Volumetric evaluation of effects of platelet-rich fibrin and concentrated growth factor on early bone healing after endodontic microsurgery: a randomized controlled trial.

BACKGROUND: This randomized controlled clinical trial compared the effects of platelet-rich fibrin (PRF) and concentrated growth factor (CGF) on early bone healing after endodontic microsurgery. METHODS: Eighteen patients with an isolated periapical lesion < 10 mm in the maxillary anterior region were randomly assigned to three groups: control, PRF, or CGF. Endodontic microsurgery was performed and PRF or CGF membranes were placed over the bone defects in the experimental groups. The volume of the bone defect at postoperative one week, three months, and six months was evaluated using cone-beam computed tomography and Mimics software. The results were statistically analyzed using the Kruskal-Wallis test and post-hoc Mann-Whitney U test with Bonferroni correction. RESULTS: At the three-month follow-up, the PRF and CGF groups showed significantly greater bone healing compared with the control group (p > 0.05). However, no significant difference was observed between the PRF and CGF groups. At the six-month follow-up, no significant differences were observed between the groups. CONCLUSIONS: These results suggested that PRF and CGF promote early bone healing after endodontic microsurgery.

Endobronchial Metastasis of Renal Cancer Presented as Atelectasis.

The most common primary tumors associated with endobronchial metastasis (EBM) are colorectal, breast, and renal. When EBM is present, respiratory symptoms such as shortness of breath or hemoptysis accompanied by coughing usually appear. Herein, we report a case of atelectasis caused by EBM of renal cell carcinoma (RCC) in a 53-year-old man who underwent laparoscopic radical nephrectomy for RCC 5 years ago. The patient's primary RCC stage was pT1b, and the histological cell type was clear cell RCC with Fuhrman nuclear grade 3/4. At the time of EBM diagnosis, the patient was classified as "favorable" according to IMDC (International mRCC Database Consortium) risk calculator. The patient refused surgical treatment and received targeted therapy with sunitinib. A tumor mass spontaneously detached and came out through the airway during targeted therapy. Subsequently, the patient's respiratory symptoms were alleviated, and his atelectasis disappeared. This case shows that when there is atelectasis due to EBM of RCC, the obstructed bronchus may be reopened with targeted therapy without any interventional treatment.

Sex differences in skeletal muscle size, function, and myosin heavy chain isoform expression during post-injury regeneration in mice.

Skeletal muscle regeneration is an essential process to restore muscle function after injury and is influenced by various factors. Despite the known importance of sex hormones in muscle regeneration, whether and what sex difference exists in this process is still unclear. In this study, we provide evidence for a clear sex difference in muscle regeneration in mice. At 7 and 14 days after barium chloride-induced muscle injury, female mice showed a faster recovery of muscle fiber size than males. Consistently, muscle force in female mice was restored faster than in males after injury, and this functional difference was maintained at 14 months of age when regenerative capacity declined. Myosin heavy chain isoform profiling and fatigability test revealed dynamic remodeling of myosin heavy chain isoform expression including a type IIB to IIA/X MHC transition and reduced fatigability in regenerated muscles compared to uninjured muscles. A significant sex difference was detected in myosin heavy chain IIX content, although this did not lead to different fatigability. Together, our results suggest that sex is an important determinant of the recovery of regenerating skeletal muscle and is partially involved in the remodeling of myosin heavy chain isoforms during muscle regeneration.

Effects of Unripe Black Raspberry Extract Supplementation on Male Climacteric Syndrome and Voiding Dysfunction: A Pilot, Randomized, Double-Blind, Placebo-Controlled Trial.

Male climacteric syndrome (MCS) is a medical condition that can affect middle-aged men whose testosterone levels begin to decline considerably. These symptoms may include fatigue, decreased libido, mood swings, and disturbed sleep. MCS can be managed with lifestyle modifications and testosterone replacement. However, testosterone therapy may cause number of side effects, including an increased risk of cardiovascular issues. This study aims to evaluate the efficacy and safety of unripe black raspberry extract (BRE) against MCS and voiding dysfunction in men with andropause symptoms. A total of 30 subjects were enrolled and randomly assigned to the BRE group (n = 15) or the placebo group (n = 15). Participants were supplemented with 4800 mg BRE or placebo twice daily for 12 weeks. The impact of BRE was assessed using the Aging Male's Symptoms (AMS scale), International Prostate Symptom Score (IPSS) and the IPSS quality of life index (IPSS-QoL). Additionally, male sex hormones, lipid profiles, and anthropometric indices were assessed 6 and 12 weeks after treatment. The AMS scores did not differ significantly between the two groups. In the BRE group, the total IPSS and IPSS-QoL scores decreased significantly after 12 weeks compared to baseline (p < 0.05), but there was no significant difference compared to the placebo group. However, a significant difference was observed in the IPSS voiding symptoms sub-score compared to the placebo group. Furthermore, LDL-C and TC levels were also significantly lower in the BRE group than in the placebo group (p < 0.05). Collectively, the study provides strong evidence supporting the safety of BRE as a functional food and its supplementation potentially enhances lipid metabolism and alleviates MCS and dysuria symptoms, limiting the development of BPH.

High-throughput sperm DNA analysis at the single-cell and population levels.

Clinical semen quality assessment is critical to the treatment of infertility. Sperm DNA integrity testing provides critical information that can steer treatment and influence outcomes and offspring health. Flow cytometry is the gold standard approach to assess DNA integrity, but it is not commonly applied at the clinical level. The sperm chromatin dispersion (SCD) assay provides a simpler and cheaper alternative. However, SCD is low-throughput and non-quantitative - sperm assessment is serial, manual and suffers inter- and intra-observer variations. Here, an automated SCD analysis method is presented that enables quantitative sperm DNA quality assessment at the single-cell and population levels. Levering automated optical microscopy and a chromatin diffusion-based analysis, a sample of thousands of sperm that would otherwise require 5 hours is assessed in under 10 minutes - a clinically viable workflow. The sperm DNA diffusion coefficient (DDNA) measurement correlates (R2 = 0.96) with DNA fragmentation index (DFI) from the cytometry-based sperm chromatin structure assay (SCSA). The automated measurement of population-level sperm DNA fragmentation (% sDF) prevents inter-observer variations and shows a good agreement with the SCSA % DFI (R2 = 0.98). This automated approach standardizes and accelerates SCD-based sperm DNA analysis, enabling the clinical application of sperm DNA integrity assessment.

RhoA/ROCK signalling activated by ARHGEF3 promotes muscle weakness via autophagy in dystrophic mdx mice.

BACKGROUND: Duchenne muscular dystrophy (DMD), caused by dystrophin deficiency, leads to progressive and fatal muscle weakness through yet-to-be-fully deciphered molecular perturbations. Emerging evidence implicates RhoA/Rho-associated protein kinase (ROCK) signalling in DMD pathology, yet its direct role in DMD muscle function, and related mechanisms, are unknown. METHODS: Three-dimensionally engineered dystrophin-deficient mdx skeletal muscles and mdx mice were used to test the role of ROCK in DMD muscle function in vitro and in situ, respectively. The role of ARHGEF3, one of the RhoA guanine nucleotide exchange factors (GEFs), in RhoA/ROCK signalling and DMD pathology was examined by generating Arhgef3 knockout mdx mice. The role of RhoA/ROCK signalling in mediating the function of ARHGEF3 was determined by evaluating the effects of wild-type or GEF-inactive ARHGEF3 overexpression with ROCK inhibitor treatment. To gain more mechanistic insights, autophagy flux and the role of autophagy were assessed in various conditions with chloroquine. RESULTS: Inhibition of ROCK with Y-27632 improved muscle force production in 3D-engineered mdx muscles (+25% from three independent experiments, P < 0.05) and in mice (+25%, P < 0.001). Unlike suggested by previous studies, this improvement was independent of muscle differentiation or quantity and instead related to increased muscle quality. We found that ARHGEF3 was elevated and responsible for RhoA/ROCK activation in mdx muscles, and that depleting ARHGEF3 in mdx mice restored muscle quality (up to +36%, P < 0.01) and morphology without affecting regeneration. Conversely, overexpressing ARHGEF3 further compromised mdx muscle quality (-13% vs. empty vector control, P < 0.01) in GEF activity- and ROCK-dependent manner. Notably, ARHGEF3/ROCK inhibition exerted the effects by rescuing autophagy which is commonly impaired in dystrophic muscles. CONCLUSIONS: Our findings uncover a new pathological mechanism of muscle weakness in DMD involving the ARHGEF3-ROCK-autophagy pathway and the therapeutic potential of targeting ARHGEF3 in DMD.

Phacoemulsification with and without iStent: A Systematic Review and Meta-Analysis of Comparative Studies.

BACKGROUND: The iStent (Glaukos Corporation; Laguna Hills, CA, USA) is one of the minimally invasive glaucoma devices. It can be inserted at the time of phacoemulsification or as a stand-alone procedure to lower the intraocular pressure (IOP). OBJECTIVE: Our aim was to conduct a systematic review and meta-analysis comparing the effect of iStent insertion at the time of phacoemulsification with phacoemulsification alone in patients with ocular hypertension or open-angle glaucoma. METHODS: We searched EMBASE, MEDLINE (OVID and PubMed), CINAHL, and Cochrane Library for articles published between 2008 and June 2022 (PRISMA 2020 for the checklist). Studies comparing the IOP-lowering effect of iStent with phacoemulsification versus phacoemulsification alone were included. The endpoints were IOP reduction (IOPR) and the mean reduction in the number of glaucoma drops. A quality-effects model was used to compare both surgical groups. RESULTS: Ten studies were included, reporting on 1,453 eyes. Eight hundred fifty three eyes had the combined iStent and phacoemulsification, and 600 eyes underwent phacoemulsification alone. IOPR was higher in the combined surgery at of 4.7 ± 2 mm Hg compared to 2.8 ± 1.9 mm Hg in phacoemulsification alone. A greater decrease in postoperative eye drops was noted in the combined group having a decrease of 1.2 ± 0.3 eye drops versus of 0.6 ± 0.6 drops in isolated phacoemulsification. The quality effect model showed an IOPR weighted mean difference (WMD) of 1.22 mm Hg (confidence interval [CI]: [-0.43, 2.87]; Q = 315.64; p < 0.01; I2 = 97%) and decreased eye drops WMD 0.42 drops (CI: [0.22, 0.62]; Q = 42.6; p < 0.01; I2 = 84%) between both surgical groups. Subgroup analysis shows that the new generation iStent may be more effective in reducing IOP. CONCLUSION: iStent has a synergetic effect with phacoemulsification. The reduction of IOP and glaucoma eye drops was higher when iStent is combined with phacoemulsification compared with isolated phacoemulsification.

Microfluidic formation of surface nanodroplets using green deep eutectic solvents for liquid-liquid nanoextraction and controlled precipitation.

HYPOTHESIS: Surface nanodroplets have recently been employed for in situ chemical analysis leveraging their low volume, e.g. O(10-15 L), that enables rapid analyte extraction and pre-concentration. So far, most surface nanodroplets have been formed using single organic solvents such as 1-octanol, toluene, among others. Designing multicomponent surface nanodroplet with controllable composition is highly desirable for extending their application as extractant. EXPERIMENT: Here, we formed surface nanodroplets using green deep eutectic solvent (gDES) composed of thymol and decanoic acid, both of which are naturally occurring chemicals. The influence of parameters such as flowrate and the composition of deep eutectic solvent on the surface nanodroplet formation were studied. As proof-of-concept, the gDES surface nanodroplets were further used to extract and detect trace amounts of fluorescent rhodamine 6G dye and copper ions from water. FINDINGS: The formation of gDES surface nanodroplets follows the theoretical model which states that the final droplet volume (Vf) scales with the Peclét number (Pe) of the flow during formation by the solvent exchange process, that is Vf âˆ¼ Pe3/4, and the nanodroplets demonstrate excellent ability as extractant for rhodamine 6G and copper ions from water. Surprisingly, the confined volume of gDES surface nanodroplets enables fast and controlled formation of Cu (II)-decanoate crystal.

DMP1 and IFITM5 Regulate Osteogenic Differentiation of MC3T3-E1 on PEO-Treated Ti-6Al-4V-Ca2+/Pi surface.

Despite numerous studies on various surface modifications on titanium and its alloys, it remains unclear what kind of titanium-based surface modifications are capable of controlling cell activity. This study aimed to understand the mechanism at the cellular and molecular levels and investigate the in vitro response of osteoblastic MC3T3-E1 cultured on the Ti-6Al-4V surface modified by plasma electrolytic oxidation (PEO) treatment. A Ti-6Al-4V surface was prepared by PEO at 180, 280, and 380 V for 3 or 10 min in an electrolyte containing Ca2+/Pi ions. Our results showed that PEO-treated Ti-6Al-4V-Ca2+/Pi surfaces enhanced the cell attachment and differentiation of MC3T3-E1 compared to the untreated Ti-6Al-4V control but did not affect cytotoxicity as shown by cell proliferation and cell death. Interestingly, on the Ti-6Al-4V-Ca2+/Pi surface treated by PEO at 280 V for 3 or 10 min, MC3T3-E1 showed a higher initial adhesion and mineralization. In addition, the alkaline phosphatase (ALP) activity significantly increased in MC3T3-E1 on the PEO-treated Ti-6Al-4V-Ca2+/Pi (280 V for 3 or 10 min). In RNA-seq analysis, the expression of dentin matrix protein 1 (DMP1), sortilin 1 (Sort1), signal-induced proliferation-associated 1 like 2 (SIPA1L2), and interferon-induced transmembrane protein 5 (IFITM5) was induced during the osteogenic differentiation of MC3T3-E1 on the PEO-treated Ti-6Al-4V-Ca2+/Pi. DMP1 and IFITM5 silencing decreased the expression of bone differentiation-related mRNAs and proteins and ALP activity in MC3T3-E1. These results suggest that the PEO-treated Ti-6Al-4V-Ca2+/Pi surface induces osteoblast differentiation by regulating the expression of DMP1 and IFITM5. Therefore, surface microstructure modification through PEO coatings with Ca2+/Pi ions could be used as a valuable method to improve biocompatibility properties of titanium alloys.

Ultra-soft and highly stretchable tissue-adhesive hydrogel based multifunctional implantable sensor for monitoring of overactive bladder.

A highly stretchable and tissue-adhesive multifunctional sensor based on structurally engineered islets embedded in ultra-soft hydrogel is reported for monitoring of bladder activity in overactive bladder (OAB) induced rat and anesthetized pig. The use of hydrogel yielded a much lower sensor modulus (1 kPa) compared to that of the bladder (300 kPa), while the strong adhesiveness of the hydrogel (adhesive strength: 260.86 N/m) allowed firm attachment onto the bladder. The change in resistance of printed liquid metal particle thin-film lines under strain were used to detect bladder inflation and deflation; due to the high stretchability and reliability of the lines, surface strains of 200% could be measured repeatedly. Au electrodes coated with Platinum black were used to detect electromyography (EMG). These electrodes were placed on structurally engineered rigid islets so that no interfacial fracture occurs under high strains associated with bladder expansion. On the OAB induced rat, stronger signals (change in resistance and EMG root-mean-square) were detected near intra-bladder pressure maxima, thus showing correlation to bladder activity. Moreover, using robot-assisted laparoscopic surgery, the sensor was placed onto the bladder of an anesthetized pig. Under voiding and filling, bladder strain and EMG were once again monitored. These results confirm that our proposed sensor is a highly feasible, clinically relevant implantable device for continuous monitoring OAB for diagnosis and treatment.

Sigmoidal Auxiliary Tendon-Driven Mechanism Reinforcing Structural Stiffness of Hyper-Redundant Manipulator for Endoscopic Surgery.

The overtube of an endoscopic surgery robot is fixed when performing tasks, unlike those of commercial endoscopes, and this overtube should have high structural stiffness after reaching the target lesion so that sufficient tension can be applied to the lesion tissue with the surgical tool and there are fewer changes in the field of view of the endoscopic camera from this reaction force. Various methods have been proposed to reinforce the structural stiffnesses of hyper-redundant manipulators. However, the safety, rapid response, space efficiency, and cost-effectiveness of these methods should be considered for use in actual clinical environments, such as the gastrointestinal tract. This study proposed a method to minimize the positional changes of the overtube end tip due to external forces using only auxiliary tendons in the optimized path without additional mechanical structures. Overall, the proposed method involved moving the overtube to the target lesion through the main driving tendon and applying tension to the auxiliary tendons to reinforce the structural stiffness. The complete system was analyzed in terms of energy, and the sigmoidal auxiliary tendons were verified to effectively reinforce the structural stiffness of the overtube consisting of rolling joints. In addition, the design guidelines of the overtube for actual endoscopic surgery were proposed considering hollowness, retroflexion, and high structural stiffness. The positional changes due to external forces were confirmed to be reduced by 60% over the entire workspace.

Endoscopic surgery robot that facilitates insertion of the curved colon and ensures positional stability against external forces: K-COLON.

BACKGROUND: Although various endoscopic surgery robots developed in previous studies are versatile and have high lesion accessibility, they have limitations in terms of reaching the target lesion through the curved path in the large intestine and providing a stable tasking environment for the operator. METHODS: An endoscopic surgery robot was developed for performing surgery in the large intestine. The robot was easily inserted into the target lesion in the curved colon through the mounted soft actuator and demonstrated high structural stiffness through the insertion of the sigmoidal auxiliary tendons. RESULTS: The robot was able to access the target lesion in the curved colon through teleoperation alone. Further, it was confirmed that the high structural stiffness overtube improved the overall task performance in the user test. CONCLUSIONS: The proposed robotic system demonstrated the possibility and potential of performing advanced endoscopic surgery in the large intestine.

Stepwise molecular mechanisms responsible for chemoresistance in bladder cancer cells.

Chemotherapy resistance is an obstacle to cancer therapy and is considered a major cause of recurrence. Thus, understanding the mechanisms of chemoresistance is critical to improving the prognosis of patients. Here, we have established a stepwise gemcitabine-resistant T24 bladder cancer cell line to understand the molecular mechanisms of chemoresistance within cancer cells. The characteristics of the stepwise chemoresistance cell line were divided into 4 phases (parental, early, intermediate, and late phases). These four phase cells showed increasingly aggressive phenotypes in vitro and in vivo experiments with increasing phases and revealed the molecular properties of the biological process from parent cells to phased gemcitabine-resistant cell line (GRC). Taken together, through the analysis of gene expression profile data, we have characterized gene set of each phase indicating the response to anticancer drug treatment. Specifically, we identified a multigene signature (23 genes including GATA3, APOBEC3G, NT5E, MYC, STC1, FOXD1, SMAD9) and developed a chemoresistance score consisting of that could predict eventual responsiveness to gemcitabine treatment. Our data will contribute to predicting chemoresistance and improving the prognosis of bladder cancer patients.

Surface Nanodroplet-Based Extraction Combined with Offline Analytic Techniques for Chemical Detection and Quantification.

Liquid-liquid extraction based on surface nanodroplets can be a green and sustainable technique to extract and concentrate analytes from a sample flow. However, because of the extremely small volume of each droplet (<10 fL, tens of micrometers in base radius and a few or less than 1 µm in height), only a few in situ analytical techniques, such as surface-enhanced Raman spectroscopy, were applicable for the online detection and analysis based on nanodroplet extraction. To demonstrate the versatility of surface nanodroplet-based extraction, in this work, the formation of octanol surface nanodroplets and extraction were performed inside a 3 m Teflon capillary tube. After extraction, surface nanodroplets were collected by injecting air into the tube, by which the contact line of surface droplets was collected by the capillary force. As the capillary allows for the formation of ∼1012 surface nanodroplets on the capillary wall, ≥2 mL of octanol can be collected after extraction. The volume of the collected octanol was enough for the analysis of offline analytical techniques such as UV-vis, GC-MS, and others. Coupled with UV-vis, reliable extraction and detection of two common water pollutants, triclosan and chlorpyrifos, was shown by a linear relationship between the analyte concentration in the sample solution and UV-vis absorbance. Moreover, the limit of detection (LOD) as low as 2 × 10-9 M for triclosan (∼0.58 µg/L) and 3 × 10-9 M for chlorpyrifos (∼1.05 µg/L) could be achieved. The collected surface droplets were also analyzed via gas chromatography (GC) and fluorescence microscopy. Our work shows that surface nanodroplet extraction may potentially streamline the process in sample pretreatment for sensitive chemical detection and quantification by using common analytic tools.