Effect of diesel exhaust particles on RANK/RANKL expression in in vivo and in vitro models of middle ear inflammation.

OBJECTIVE: Increasing evidence suggests a link between middle ear inflammation and the development of diesel exhaust particles (DEPs). Chronic middle ear inflammation can lead to bone damage and remodeling. This study aimed to explore the impact of DEPs on the expression of interleukin (IL)-6 and RANKL under conditions of middle ear inflammation. METHODS: DEPs were collected by burning fuel in a diesel engine at the Gwangju Institute of Science and Technology. Human middle ear epithelial cells were cultured to 70-80% confluence in culture plates and then treated with DEPs at concentrations of 0, 5, 10, 20, 40, and 80 µg/mL for 24 h. Cell viability was assessed manually. B6.SJL mice, aged 9 weeks, were exposed to DEPs at a concentration of 200 µg/m3 for 1 h daily over a period of 28 days. The expression levels of IL-6, tumor necrosis factor α, RANKL, and RANK were evaluated using hematoxylin and eosin staining and western blot analysis of the harvested middle ear samples. RESULTS: The viability of human middle ear epithelial cells was found to decrease in a dose-dependent manner after 24 h. The mRNA expression level of IL-6 exhibited the most significant increase at the 48-h mark. In contrast, the mRNA expression levels of RANKL and RANK showed a marked increase as early as 6 h post-exposure, with both genes subsequently displaying a time-dependent decrease. Histological analysis revealed that the middle ear mucosa was thicker in the group exposed to DEPs compared to the control group. Additionally, the protein expression levels of IL-6 and RANKL were elevated in the DEP-exposed group relative to the normal control group. CONCLUSIONS: We confirmed the expression of osteoclast-related proteins in the mouse middle ear. These results imply that air pollutants might affect RANKL/RANK signaling, which is associated with bone remodeling.

Anticancer Evaluation of Novel Benzofuran-Indole Hybrids as Epidermal Growth Factor Receptor Inhibitors against Non-Small-Cell Lung Cancer Cells.

The epidermal growth factor receptor (EGFR), also known as ErbB1 and HER1, belongs to the receptor tyrosine kinase family. EGFR serves as the primary driver in non-small-cell lung cancer (NSCLC) and is a promising therapeutic target for NSCLC. In this study, we synthesized a novel chemical library based on a benzofuran-indole hybrid scaffold and identified 8aa as a potent and selective EGFR inhibitor. Interestingly, 8aa not only showed selective anticancer effects against NSCLC cell lines, PC9, and A549, but it also showed significant inhibitory effects against the double mutant L858R/T790M EGFR, which frequently occurs in NSCLC. In addition, in PC9 and A549 cells, 8aa potently blocked the EGFR signaling pathway, cell viability, and cell migration. These findings suggest that 8aa, a benzofuran-indole hybrid derivative, is a novel EGFR inhibitor that may be a potential candidate for the treatment of NSCLC patients with EGFR mutations.

Curcumol metabolized by rat liver S9 fraction and orally administered in mouse suppressed the proliferation of colon cancer in vitro and in vivo.

Following 3R (reduction, refinement, and replacement) principles, we employed the rat liver S9 fraction to mimic liver metabolism of curcumol having high in vitro IC50 on cancer cells. In HCT116 and HT29 colon cancer cells, the metabolites of curcumol by S9 fraction exerted more enhanced activity in inducing cell cycle arrest and apoptosis via regulating the expression of cyclin D1, CDK1, p21, PARP and Bcl-2 than curcumol. In addition, oral administration of curcumol at 4 mg/kg BW significantly suppressed the development of colon tumor induced by azoxymethane/dextran sulfate sodium, and induced cell cycle arrest and apoptosis in tumor tissues. In mass analysis, curcumenol and curzerene were identified as the metabolites of curcumol by S9 fraction metabolism. Taken together, curcumol metabolites showed the enhanced suppressive effect on colon cancer, suggesting that S9 fraction can be considered as simple, fast, and bio-mimicking platform for the screening of chemical libraries on different chronic diseases.

Cultured meat with enriched organoleptic properties by regulating cell differentiation.

Research on cultured meat has primarily focused on the mass proliferation or differentiation of muscle cells; thus, the food characteristics of cultured meat remain relatively underexplored. As the quality of meat is determined by its organoleptic properties, cultured meat with similar sensory characteristics to animal-derived meat is highly desirable. In this study, we control the organoleptic and nutritional properties of cultured meat by tailoring the 2D differentiation of primary bovine myoblasts and primary bovine adipose-derived mesenchymal stem cells on gelatin/alginate scaffolds with varying stiffness. We assess the effect of muscle and adipose differentiation quality on the sensory properties of cultured meat. Thereafter, we fabricate cultured meat with similar sensory profiles to that of conventional beef by assembling the muscle and adipose constructs composed of highly differentiated cells. We introduce a strategy to produce cultured meat with enriched food characteristics by regulating cell differentiation with scaffold engineering.

Machine learning for predicting hepatitis B or C virus infection in diabetic patients.

Highly prevalent hepatitis B and hepatitis C virus (HBV and HCV) infections have been reported among individuals with diabetes. Given the frequently asymptomatic nature of hepatitis and the challenges associated with screening in some vulnerable populations such as diabetes patients, we conducted an investigation into the performance of various machine learning models for the identification of hepatitis in diabetic patients while also evaluating the significance of features. Analyzing NHANES data from 2013 to 2018, machine learning models were evaluated; random forest (RF), support vector machine (SVM), eXtreme Gradient Boosting (XGBoost), and least absolute shrinkage and selection operator (LASSO) along with stacked ensemble model. We performed hyperparameter tuning to improve the performance of the model, and selected important predictors using the best performance model. LASSO showed the highest predictive performance (AUC-ROC = 0.810) rather than other models. Illicit drug use, poverty, and race were highly ranked as predictive factors for developing hepatitis in diabetes patients. Our study demonstrated that a machine-learning-based model performed optimally in the detection of hepatitis among diabetes patients, achieving high performance. Furthermore, models and predictors evaluated from the current study, we expect, could be supportive information for developing screening or treatment methods for hepatitis care in diabetes patients.

Proximity Labeling Expansion Microscopy (PL-ExM) resolves structure of the interactome.

Elucidating the spatial relationships within the protein interactome is pivotal to understanding the organization and regulation of protein-protein interactions. However, capturing the 3D architecture of the interactome presents a dual challenge: precise interactome labeling and super-resolution imaging. To bridge this gap, we present the Proximity Labeling Expansion Microscopy (PL-ExM). This innovation combines proximity labeling (PL) to spatially biotinylate interacting proteins with expansion microscopy (ExM) to increase imaging resolution by physically enlarging cells. PL-ExM unveils intricate details of the 3D interactome's spatial layout in cells using standard microscopes, including confocal and Airyscan. Multiplexing PL-ExM imaging was achieved by pairing the PL with immunofluorescence staining. These multicolor images directly visualize how interactome structures position specific proteins in the protein-protein interaction network. Furthermore, PL-ExM stands out as an assessment method to gauge the labeling radius and efficiency of different PL techniques. The accuracy of PL-ExM is validated by our proteomic results from PL mass spectrometry. Thus, PL-ExM is an accessible solution for 3D mapping of the interactome structure and an accurate tool to access PL quality.

An amphiphilic material arginine-arginine-bile acid promotes α-synuclein amyloid formation.

Amyloid generation plays essential roles in various human diseases, biological functions, and nanotechnology. However, developing efficient chemical and biological candidates for regulating amyloid fibrillation remains difficult because information on the molecular actions of modulators is insufficient. Thus, studies are needed to understand how the intermolecular physicochemical properties of the synthesised molecules and amyloid precursors influence amyloidogenesis. In this study, we synthesised a novel amphiphilic sub-nanosized material, arginine-arginine (RR)-bile acid (BA), by conjugating positively charged RR to hydrophobic BA. The effects of RR-BA on amyloid formation were investigated on α-synuclein (αSN) in Parkinson's disease and on K18 and amyloid-ß (1-42) (Aß42) in Alzheimer's disease. RR-BA showed no appreciable effect on the kinetics of K18 and Aß42 amyloid fibrillation because of their weak and non-specific interactions. However, RR-BA specifically bound to αSN with moderate binding affinity through electrostatic interactions between the positively charged RR and the negatively charged cluster in the C-terminus of αSN. In addition, hydrophobic BA in the αSN-RR-BA complex transiently condensed αSN for primary nucleation, thereby accelerating αSN amyloid fibrillation. We propose an electrostatic binding and hydrophobic condensation model of RR-BA-driven amyloid formation of αSN, which will contribute to the rational design and development of molecules for controlling amyloid aggregation in diverse fields.

Zwitterionic Inhaler with Synergistic Therapeutics for Reprogramming of M2 Macrophage to Pro-Inflammatory Phenotype.

Myriad lung diseases are life threatening and macrophages play a key role in both physiological and pathological processes. Macrophages have each pro-/anti-inflammatory phenotype, and each lung disease can be aggravated by over-polarized macrophage. Therefore, development of a method capable of mediating the macrophage phenotype is one of the solutions for lung disease treatment. For mediating the phenotype of macrophages, the pulmonary delivery system (PDS) is widely used due to its advantages, such as high efficiency and accessibility of the lungs. However, it has a low drug delivery efficiency ironically because of the perfect lung defense system consisting of the mucus layer and airway macrophages. In this study, zwitterion-functionalized poly(lactide-co-glycolide) (PLGA) inhalable microparticles (ZwPG) are synthesized to increase the efficiency of the PDS. The thin layer of zwitterions formed on PLGA surface has high nebulizing stability and show high anti-mucus adhesion and evasion of macrophages. As a reprogramming agent for macrophages, ZwPG containing dexamethasone (Dex) and pirfenidone (Pir) are treated to over-polarized M2 macrophages. As a result, a synergistic effect of Dex/Pir induces reprogramming of M2 macrophage to pro-inflammatory phenotypes.

Case report: Generalized lymphatic anomaly of multiple abdominal organs in a young dog.

A 9-month-old, female Pomeranian dog presented with vomiting and lethargy. Ultrasonography revealed multilobulated anechoic round shape structures at the ovarian and uterine locations. Through computed tomography scan, an extensive non-contrast multilobulated fluid-filled mass suspected of originating from the walls of the ovary, uterus, urinary bladder and rectum was observed. Ovariohysterectomy and urinary bladder biopsy were performed. Histopathological examination revealed numerous cystic lesions lined by plump cuboidal cells believed to be of epithelial origin. Immunohistochemical staining showed that the cyst-like lesions lining cells were strongly positive for lymphatic vessel endothelial hyaluronan receptor 1. Based on these results, lesions were identified as generalized lymphatic anomaly (GLA), in which lymphangiomas develop in multiple organs. After 6 months follow-up, the size of the cysts remaining in the region of the bladder did not undergo much change. GLA should be included in the differential diagnosis when multiple cystic lesions are interspersed in multiple organs.

Development of a Peptide-Based Nano-Sized Cathepsin B Inhibitor for Anticancer Therapy.

Numerous cathepsin B inhibitors have been developed and are under investigation as potential cancer treatments. They have been evaluated for their ability to inhibit cathepsin B activity and reduce tumor growth. However, they have shown critical limitations, including low anticancer efficacy and high toxicity, due to their low selectivity and delivery problems. In this study, we developed a novel peptide and drug conjugate (PDC)-based cathepsin B inhibitor using cathepsin-B-specific peptide (RR) and bile acid (BA). Interestingly, this RR and BA conjugate (RR-BA) was able to self-assemble in an aqueous solution, and as a result, it formed stable nanoparticles. The nano-sized RR-BA conjugate showed significant cathepsin B inhibitory effects and anticancer effects against mouse colorectal cancer (CT26) cells. Its therapeutic effect and low toxicity were also confirmed in CT26 tumor-bearing mice after intravenous injection. Therefore, based on these results, the RR-BA conjugate could be developed as an effective anticancer drug candidate for inhibiting cathepsin B in anticancer therapy.

High protein-containing new food by cell powder meat.

Demand for a new protein source to replace meat is increasing to solve various issues such as limited resources and food shortages. Diverse protein sources are being developed, but alternative proteins such as plants or insects need to improve people's perceptions and organoleptic properties. Therefore, cell-based meat research is intensively conducted, and most studies are aimed at scale-up and cost-down via the research of scaffolds and culture media. Here, we proposed a new food by cell powder meat (CPM), which has a high protein content and a meaty flavor. The powder was manufactured 76% more cost-effectively with less serum than the conventional culture medium and without 3D scaffold. Due to its comprehensive characteristics, the potential applicability of CPM in the cell-based meat industry could be expected.

Fructose malabsorption in ChREBP-deficient mice disrupts the small intestine immune microenvironment and leads to diarrhea-dominant bowel habit changes.

BACKGROUND: The mechanism by which incompletely absorbed fructose causes gastrointestinal symptoms is not fully understood. In this study, we investigated the immunological mechanisms of bowel habit changes associated with fructose malabsorption by examining Chrebp-knockout mice exhibiting defective fructose absorption. METHODS: Mice were fed a high-fructose diet (HFrD), and stool parameters were monitored. The gene expression in the small intestine was analyzed by RNA sequencing. Intestinal immune responses were assessed. The microbiota composition was determined by 16S rRNA profiling. Antibiotics were used to assess the relevance of microbes for HFrD-induced bowel habit changes. RESULTS: Chrebp-knockout (KO) mice fed HFrD showed diarrhea. Small-intestine samples from HFrD-fed Chrebp-KO mice revealed differentially expressed genes involved in the immune pathways, including IgA production. The number of IgA-producing cells in the small intestine decreased in HFrD-fed Chrebp-KO mice. These mice showed signs of increased intestinal permeability. Chrebp-KO mice fed a control diet showed intestinal bacterial imbalance, which the HFrD exaggerated. Bacterial reduction improved diarrhea-associated stool parameters and restored the decreased IgA synthesis induced in HFrD-fed Chrebp-KO mice. CONCLUSIONS: The collective data indicate that gut microbiome imbalance and disrupting homeostatic intestinal immune responses account for the development of gastrointestinal symptoms induced by fructose malabsorption.

Inhibition of ANO1 by Cis- and Trans-Resveratrol and Their Anticancer Activity in Human Prostate Cancer PC-3 Cells.

Anoctamin1 (ANO1), a calcium-activated chloride channel, is involved in the proliferation, migration, and invasion of various cancer cells including head and neck squamous cell carcinoma, lung cancer, and prostate cancer. Inhibition of ANO1 activity or downregulation of ANO1 expression in these cancer cells is known to exhibit anticancer effects. Resveratrol, a natural polyphenol abundant in wines, grapes, berries, soybeans, and peanuts, shows a wide variety of biological effects including anti-inflammatory, antioxidant, and anticancer activities. In this study, we investigated the effects of two stereoisomers of resveratrol on ANO1 activity and found that cis- and trans-resveratrol inhibited ANO1 activity with different potencies. Cis- and trans-resveratrol inhibited ANO1 channel activity with IC50 values of 10.6 and 102 µM, respectively, and had no significant effect on intracellular calcium signaling at 10 and 100 µM, respectively. In addition, cis-resveratrol downregulated mRNA and protein expression levels of ANO1 more potently than trans-resveratrol in PC-3 prostate cancer cells. Cis- and trans-resveratrol significantly reduced cell proliferation and cell migration in an ANO1-dependent manner, and both resveratrol isomers strongly increased caspase-3 activity, PARP cleavage, and apoptotic sub-G1 phase ratio in PC-3 cells. These results revealed that cis-resveratrol is a potent inhibitor of ANO1 and exhibits ANO1-dependent anticancer activity against human metastatic prostate cancer PC-3 cells.

Technological Breakthroughs in Chip Fabrication, Transfer, and Color Conversion for High-Performance Micro-LED Displays.

The implementation of high-efficiency and high-resolution displays has been the focus of considerable research interest. Recently, micro light-emitting diodes (micro-LEDs), which are inorganic light-emitting diodes of size <100 µm2 , have emerged as a promising display technology owing to their superior features and advantages over other displays like liquid crystal displays and organic light-emitting diodes. Although many companies have introduced micro-LED displays since 2012, obstacles to mass production still exist. Three major challenges, i.e., low quantum efficiency, time-consuming transfer, and complex color conversion, have been overcome with technological breakthroughs to realize cost-effective micro-LED displays. In the review, methods for improving the degraded quantum efficiency of GaN-based micro-LEDs induced by the size effect are examined, including wet chemical treatment, passivation layer adoption, LED structure design, and growing LEDs in self-passivated structures. Novel transfer technologies, including pick-up transfer and self-assembly methods, for developing large-area micro-LED displays with high yield and reliability are discussed in depth. Quantum dots as color conversion materials for high color purity, and deposition methods such as electrohydrodynamic jet printing or contact printing on micro-LEDs are also addressed. This review presents current status and critical challenges of micro-LED technology and promising technical breakthroughs for commercialization of high-performance displays.

Synthesis and Biological Evaluation of a Fused Structure of Indolizine and Pyrrolo[1,2-c]pyrimidine: Identification of Its Potent Anticancer Activity against Liver Cancer Cells.

A highly efficient approach to a new indolizine scaffold fused with pyrrolo[1,2-c]pyrimidine was achieved via one-pot three-component coupling followed by an oxidative cyclization reaction. The simple two-step sequence allowed rapid access to various tetracyclic compounds from commercially available starting materials with the formation of five new bonds. Here, we observed the effects of these compounds on cell viability in HepG2, H1299, HT29, AGS, and A549 cancer cell lines. Interestingly, this fused scaffold had more potent anticancer activity in hepatocellular carcinoma HepG2 and Huh7 cells than other cancer cells. In particular, 5r strongly decreased cell viability in HepG2 and Huh7 cells with an IC50 value of 0.22 ± 0.08 and 0.10 ± 0.11 µM, respectively, but had a very weak inhibitory effect on the cell viability of other cancer cell lines. In addition, 5r significantly inhibited cell migration and induced apoptosis in HepG2 and Huh7 cells via the activation of caspase-3 and cleavage of PARP in a dose-dependent manner. Notably, the co-treatment of 5r with gemcitabine resulted in the significant additional inhibition of cell viability in HepG2 and Huh7 cells. Our results suggest that 5r could be used to develop new chemotype anticancer agents against liver cancers.

Label-Retention Expansion Microscopy (LR-ExM) Enables Super-Resolution Imaging and High-Efficiency Labeling.

Expansion microscopy (ExM) is a sample preparation technique that can be combined with most light microscopy methods to increase the resolution. After embedding cells or tissues in swellable hydrogel, samples can be physically expanded three-to sixteen-fold (linear dimension) compared to the original size. Therefore, the effective resolution of any microscope is increased by the expansion factor. A major limitation of the previously introduced ExM is reduced fluorescence after polymerization and the digestion procedure. To overcome this limitation, label-retention expansion microscopy (LR-ExM) has been developed, which prevents signal loss and greatly enhances labeling efficiency using a set of novel trifunctional anchors. This technique allows one to achieve higher resolution when investigating cellular or subcellular structures at a nanometric scale with minimal fluorescent signal loss. LR-ExM can be used not only for immunofluorescence labeling, but also with self-labeling protein tags, such as SNAP- and CLIP-tags, thus achieving higher labeling efficiency. This work presents the procedure and troubleshooting for this immunostaining-based approach, as well as discussion of self-labeling tagging approaches of LR-ExM as an alternative.

Tailoring a Gelatin/Agar Matrix for the Synergistic Effect with Cells to Produce High-Quality Cultured Meat.

Edible scaffolds are needed in cultured meat to mimic meat's three-dimensional structure by organizing cells and replenishing the insufficient meat mass of cells alone. However, there is still a large gap between slaughtered meat and cells developed into tissues using scaffolds. This is mainly due to the difference in size, texture, flavor, and taste. In this study, we develop a coating matrix to modify the surface of textured vegetable protein (TVP), a vegetable cell support, to produce cultured meat having slaughtered meat's essential characteristics. We optimized the fish gelatin/agar matrix's microstructure by controlling the ratio of the two biopolymers, stably introducing a cell adhesive environment on the TVP. By coating the optimized gelatin/agar matrix on the TVP's surface using an easy and fast dipping method, hybrid cultured meat composed of animal cells and plant protein was produced. As the cells proliferated, their synergistic effect permitted the cultured meat's texture, flavor, and taste to reach a level comparable to that of slaughtered meat. The TVP-based cultured meat prepared with the present technology has been recreated as high-quality cultured meat by satisfying five challenging factors: cells, texture, cost, mass, and flavor.

Ultralight and Ultrathin Electrospun Membranes with Enhanced Air Permeability for Chemical and Biological Protection.

With the growing interest in chemical and biological warfare agents (CWAs/BWAs), the focus has shifted toward aerosol protection using protective clothing. However, compared to air-permeable membranes, those with water vapor permeability have been investigated more extensively. Filtering membranes without air permeability have limited practical usage in personal protective suits and masks. In this study, polyacrylonitrile membranes with tightly attached activated carbon and doped copper(II) oxide were prepared via electrospinning. The nanofibers with uniformly controlled diameters and smooth morphologies enable water/air breathability and protection against aerosol (100 nm polystyrene nanobeads similar to SARS-CoV-2) penetration. The uniformly distributed and tightly attached activated carbon and doped copper(II) oxide particles enhance the sorptive performance of the membranes by blocking gaseous CWAs, including soman, nerve chemical agents, and BWAs. Such dual-purpose membranes can be implemented in protective equipment owing to their high performance and easy processing.

Amelioration of SARS-CoV-2 infection by ANO6 phospholipid scramblase inhibition.

As an enveloped virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) delivers its viral genome into host cells via fusion of the viral and cell membranes. Here, we show that ANO6/TMEM16F-mediated cell surface exposure of phosphatidylserine is critical for SARS-CoV-2 entry and that ANO6-selective inhibitors are effective against SARS-CoV-2 infections. Application of the SARS-CoV-2 Spike pseudotyped virus (SARS2-PsV) evokes a cytosolic Ca2+ elevation and ANO6-dependent phosphatidylserine externalization in ACE2/TMPRSS2-positive mammalian cells. A high-throughput screening of drug-like chemical libraries identifies three different structural classes of chemicals showing ANO6 inhibitory effects. Among them, A6-001 displays the highest potency and ANO6 selectivity and it inhibits the single-round infection of SARS2-PsV in ACE2/TMPRSS2-positive HEK 293T cells. More importantly, A6-001 strongly inhibits authentic SARS-CoV-2-induced phosphatidylserine scrambling and SARS-CoV-2 viral replications in Vero, Calu-3, and primarily cultured human nasal epithelial cells. These results provide mechanistic insights into the viral entry process and offer a potential target for pharmacological intervention to protect against coronavirus disease 2019 (COVID-19).