Rapid and sensitive detection of gram-negative bacteria using surface-immobilized polymyxin B.

Although detection of gram-negative bacteria (GNB) in body fluids is important for clinical purpose, traditional gram staining and other recently developed methods have inherent limitations in terms of accuracy, sensitivity, and convenience. To overcome the weakness, this study proposed a method detecting GNB based on specific binding of polymyxin B (PMB) to lipopolysaccharides (LPS) of GNB. Fluorescent microscopy demonstrated that surface immobilized PMB using a silane coupling agent was possible to detect fluorescent signal produced by a single Escherichia coli (a model GNB) cell. Furthermore, the signal was selective enough to differentiate between GNB and gram-positive bacteria. The proposed method could detect three cells per ml within one hour, indicating the method was very sensitive and the sensing was rapid. These results suggest that highly multifold PMB binding on each GNB cell occurred, as millions of LPS are present on cell wall of a GNB cell. Importantly, the principle used in this study was realized in a microfluidic chip for a sample containing E. coli cells suspended in porcine plasma, demonstrating its potential application to practical uses. In conclusion, the proposed method was accurate, sensitive, and convenient for detecting GNB, and could be applied clinically.

The Function of Memory CD8+ T Cells in Immunotherapy for Human Diseases.

Memory T (Tm) cells protect against Ags that they have previously contacted with a fast and robust response. Therefore, developing long-lived Tm cells is a prime goal for many vaccines and therapies to treat human diseases. The remarkable characteristics of Tm cells have led scientists and clinicians to devise methods to make Tm cells more useful. Recently, Tm cells have been highlighted for their role in coronavirus disease 2019 vaccines during the ongoing global pandemic. The importance of Tm cells in cancer has been emerging. However, the precise characteristics and functions of Tm cells in these diseases are not completely understood. In this review, we summarize the known characteristics of Tm cells and their implications in the development of vaccines and immunotherapies for human diseases. In addition, we propose to exploit the beneficial characteristics of Tm cells to develop strategies for effective vaccines and overcome the obstacles of immunotherapy.

Atopic dermatitis and risk of gastroesophageal reflux disease: A nationwide population-based study.

BACKGROUND: As atopic dermatitis (AD) has been found to be related to various comorbidities as well as substantial patient burden, questions of a possible relationship between AD and nonallergic diseases beyond allergic diseases have also been raised. OBJECTIVE: The aim of this nationwide matched cohort study was to evaluate whether AD would increase the development of gastroesophageal reflux disease (GERD). METHODS: Patients diagnosed with AD were identified from the National Health Insurance Service-National Sample Cohort (NHIS-NSC) 2.0 database in South Korea from 2002 to 2015. Finally, 9,164 adults with AD (≥20 years old) and age, sex, household income, region of residence, disability, and baseline year-matched 9,164 controls were included in the analysis. Hazard ratio (HR) with 95% confidence interval (CI) for the development of GERD was estimated using a Cox proportional hazard regression model. RESULTS: Overall, 12.3% of the patients in the AD group developed GERD, whereas 10.4% of the individuals in the control group developed GERD. The results of the adjusted model revealed that patients with AD had a significantly increased risk of developing GERD (adjusted HR, 1.15; 95% CI, 1.06-1.26) compared with the matched controls. Increased risk of developing GERD was consistent in subgroup analyses by sex or age groups under 60 years old as well as all the sensitivity analyses performed. CONCLUSIONS: This study suggested that appropriate management should be considered in adults with AD to prevent GERD, because AD was found to be associated with an increased risk of subsequent GERD.

Enhanced delivery of the phototherapeutic nanoparticles via hepatocyte overload.

Phototherapeutic nanoparticles (NPs) were prepared with methylene blue (MB), indocyanine green (ICG), and Solutol through self-assembly. Generation of reactive oxygen species and elevation of temperature were observed that verify the photodynamic/photothermal effects of the NPs. Morphology and size distribution of the NPs were examined by transmittance electron microscopy and dynamic light scattering. The biodistribution of the NPs and their antitumor efficacy were examined using tumor-bearing mice to understand the phototherapeutic effect of the NPs on tumors. To enhance targetability with enhanced therapeutic efficacy, empty NPs (Solutol nanoparticles without MB and ICG) at different concentrations were injected along with the phototherapeutic NPs. Enhanced delivery of the phototherapeutic NPs at the tumor site was examined based on hepatocyte overload.

Is Omalizumab Related to Ear and Labyrinth Disorders? A Disproportionality Analysis Based on a Global Pharmacovigilance Database.

INTRODUCTION: Asthma is a chronic disease, characterized by reversible airway obstruction, hypersensitivity reactions, and inflammation. Oral corticosteroids are an important treatment option for patients with severe or steroid-resistant asthma. Biologics for asthma are recommended in patients with severe asthma, owing to their steroid-sparing effect as well as their ability to reduce the severity and aggravation of uncontrolled asthma. Most clinical trials of omalizumab in patients with asthma have suggested its tolerability and safety. However, some studies reported eosinophilic comorbidities in the ear, nose, and throat during omalizumab treatment, particularly eosinophilic otitis media. This study examined the relationship between ear disorders and omalizumab compared with that of other biologics for asthma using a large real-world database. MATERIALS AND METHODS: Individual case safety reports from the Uppsala Monitoring Centre Vigibase of biologics for asthma (omalizumab, mepolizumab, reslizumab, benralizumab, and dupilumab) up to 29 December 2019, were used. A disproportionality analysis was performed using the proportional reporting ratio (PRR), reporting odds ratio (ROR), and information components (IC). A hierarchy analysis used the Medical Dictionary for Regulatory Activities Terminology. A tree map was generated using R studio version 4.2. RESULTS: In 32,618 omalizumab reports, 714 adverse events (AEs) were detected as signals. Among the 714 signals, seventeen AEs were detected as signals of omalizumab-related ear and labyrinth disorders in 394 reports. Only three AEs (ear pain, ear disorder, and ear discomfort) were detected from mepolizumab. No signal was detected from reslizumab, benralizumab, and dupilumab. CONCLUSIONS: Careful monitoring of ear disorders is recommended when omalizumab treatment is started, with decreased oral corticosteroid use in patients with severe asthma. Further studies are necessary to confirm the omalizumab-related signals.

The extrinsic factors important to the homeostasis of allergen-specific memory CD4 T cells.

Memory T cells, which are generated after the primary immune response to cognate antigens, possess unique features compared to naïve or effector T cells. These memory T cells are maintained for a long period of time and robustly reactivate in lymphoid or peripheral tissues where they re-encounter antigens. Environments surrounding memory T cells are importantly involved in the process of the maintenance and reactivation of these T cells. Although memory T cells are generally believed to be formed in response to acute infections, the pathogenesis and persistence of chronic inflammatory diseases, including allergic diseases, are also related to the effector functions of memory CD4 T cells. Thus, the factors involved in the homeostasis of allergen-specific memory CD4 T cells need to be understood to surmount these diseases. Here, we review the characteristics of allergen-specific memory CD4 T cells in allergic diseases and the importance of extrinsic factors for the homeostasis and reactivation of these T cells in the view of mediating persistence, recurrence, and aggravation of allergic diseases. Overall, this review provides a better understanding of memory CD4 T cells to devise effective therapeutic strategies for refractory chronic inflammatory diseases.

Neural-Cadherin Influences the Homing of Terminally Differentiated Memory CD8 T Cells to the Lymph Nodes and Bone Marrow.

Memory T (TM) cells play an important role in the long-term defense against pathogen reinvasion. However, it is still unclear how these cells receive the crucial signals necessary for their longevity and homeostatic turnover. To understand how TM cells receive these signals, we infected mice with lymphocytic choriomeningitis virus (LCMV) and examined the expression sites of neural cadherin (N-cadherin) by immunofluorescence microscopy. We found that N-cadherin was expressed in the surroundings of the white pulps of the spleen and medulla of lymph nodes (LNs). Moreover, TM cells expressing high levels of killer cell lectin-like receptor G1 (KLRG1), a ligand of N-cadherin, were co-localized with N-cadherin+ cells in the spleen but not in LNs. We then blocked N-cadherin in vivo to investigate whether it regulates the formation or function of TM cells. The numbers of CD127hiCD62Lhi TM cells in the spleen of memory P14 chimeric mice declined when N-cadherin was blocked during the contraction phase, without functional impairment of these cells. In addition, when CD127loKLRG1hi TM cells were adoptively transferred into anti-N-cadherin-treated mice compared with control mice, the number of these cells was reduced in the bone marrow and LNs, without functional loss. Taken together, our results suggest that N-cadherin participates in the development of CD127hiCD62Lhi TM cells and homing of CD127loKLRG1hi TM cells to lymphoid organs.

Synthesis and Biochemical Evaluation of Baicalein Prodrugs.

Baicalein (5,6,7-trihydroxy-2-phenyl-4H-1-benzopyran-4-one), a flavonoid analog from Scutellaria baicalensis, possesses several pharmacological activities including antioxidant, antiproliferative, and anti-inflammatory activities. We previously reported that baicalein inhibits the thymic stromal lymphopoietin (TSLP)/TSLP receptor (TSLPR) signaling pathways and can be used as an active ingredient in the treatment of asthma and atopic dermatitis. However, baicalein is rapidly metabolized to baicalin and baicalein-6-O-glucuronide in vivo, which limits its preclinical and clinical use. In this study, we designed, synthesized, and evaluated baicalein prodrugs that protect the OH group at the 7-position of the A ring in baicalein with the amino acid carbamate functional group. Comprehensive in vitro and in vivo studies identified compound 2 as a baicalein prodrug candidate that improved the plasma exposure of baicalein in mouse animal studies. Our results demonstrated that this prodrug approach could be further adopted to discover oral baicalein prodrugs.

Ocular surface disorders associated with the use of dupilumab based on WHO VigiBase.

Dupilumab is a dual inhibitor of interleukin-4 and interleukin-13 and is mainly used to treat moderate-to-severe atopic dermatitis. Post-marketing safety data related to dupilumab have been accumulated, and it has been found that ocular surface diseases are closely associated with dupilumab treatment. The aim of this study was to detect dupilumab-related signals and to determine the safety characteristics of dupilumab with respect to eye disorders using real-world big data. Data on dupilumab use until December 29, 2019 were collected. The data were mined by calculating three indices: proportional reporting ratios, reporting odds ratios, and information components. The detected signals were classified using the primary system organ class in MedDRA terminology. Among 21,161,249 reports for all drugs, 20,548 reports were recorded for dupilumab. A total of 246 signals in the preferred terms were detected for dupilumab. Among the 246 positive signals obtained, 61 signals were related to eye disorders, which accounted for the largest percentage (24.8%), and 38 signals were anatomically related to the ocular surface. Dupilumab may cause extensive eye disorders; however, the underlying mechanisms and risk factors remain unclear. Our findings may facilitate broad safety screening of dupilumab-related eye disorders using real-world big data.

Effect of Human or Mouse IL-7 on the Homeostasis of Porcine T Lymphocytes.

Due to the inconsistent fluctuation of blood supply for transfusion, much attention has been paid to the development of artificial blood using other animals. Although mini-pigs are candidate animals, contamination of mini-pig T cells in artificial blood may cause a major safety concern. Therefore, it is important to analyze the cross-reactivity of IL-7, the major survival factor for T lymphocytes, between human, mouse, and mini-pig. Thus, we compared the protein sequences of IL-7 and found that porcine IL-7 was evolutionarily different from human IL-7. We also observed that when porcine T cells were cultured with either human or mouse IL-7, these cells did not increase the survival or proliferation compared to negative controls. These results suggest that porcine T cells do not recognize human or mouse IL-7 as their survival factor.

The Roles of CCR7 for the Homing of Memory CD8+ T Cells into Their Survival Niches.

Memory CD8+ T cells in the immune system are responsible for the removal of external Ags for a long period of time to protect against re-infection. Naïve to memory CD8+ T cell differentiation and memory CD8+ T cell maintenance require many different factors including local environmental factors. Thus, it has been suggested that the migration of memory CD8+ T cells into specific microenvironments alters their longevity and functions. In this review, we have summarized the subsets of memory CD8+ T cells based on their migratory capacities and described the niche hypothesis for their survival. In addition, the basic roles of CCR7 in conjunction with the migration of memory CD8+ T cells and recent understandings of their survival niches have been introduced. Finally, the applications of altering CCR7 signaling have been discussed.

Author Correction: Structure-Activity Relationships of Baicalein and its Analogs as Novel TSLP Inhibitors.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Tumor microenvironment dictates regulatory T cell phenotype: Upregulated immune checkpoints reinforce suppressive function.

BACKGROUND: Regulatory T (Treg) cells have an immunosuppressive function in cancer, but the underlying mechanism of immunosuppression in the tumor microenvironment (TME) is unclear. METHODS: We compared the phenotypes of T cell subsets, including Treg cells, obtained from peripheral blood, malignant effusion, and tumors of 103 cancer patients. Our primary focus was on the expression of immune checkpoint (IC)-molecules, such as programmed death (PD)-1, T-cell immunoglobulin and mucin-domain containing (TIM)-3, T cell Ig and ITIM domain (TIGIT), and cytotoxic T lymphocyte antigen (CTLA)-4, on Treg cells in paired lymphocytes from blood, peritumoral tissue, and tumors of 12 patients with lung cancer. To identify the immunosuppressive mechanisms acting on tumor-infiltrating Treg cells, we conducted immunosuppressive functional assays in a mouse model. RESULTS: CD8+, CD4+ T cells, and Treg cells exhibited a gradual upregulation of IC-molecules the closer they were to the tumor. Interestingly, PD-1 expression was more prominent in Treg cells than in conventional T (Tconv) cells. In lung cancer patients, higher levels of IC-molecules were expressed on Treg cells than on Tconv cells, and Treg cells were also more enriched in the tumor than in the peri-tumor and blood. In a mouse lung cancer model, IC-molecules were also preferentially upregulated on Treg cells, compared to Tconv cells. PD-1 showed the greatest increase on most cell types, especially Treg cells, and this increase occurred gradually over time after the cells entered the TME. PD-1 high-expressing tumor-infiltrating Treg cells displayed potent suppressive activity, which could be partially inhibited with a blocking anti-PD-1 antibody. CONCLUSIONS: We demonstrate that the TME confers a suppressive function on Treg cells by upregulating IC-molecule expression. Targeting IC-molecules, including PD-1, on Treg cells may be effective for cancer treatment.

Structure-Activity Relationships of Baicalein and its Analogs as Novel TSLP Inhibitors.

Thymic stromal lymphopoietin (TSLP) plays an important role in the differentiation and proliferation of Th2 cells, resulting in eosinophilic inflammation and numerous allergic diseases. Baicalein (1), a major component of Scutellaria baicalensis, was found to be the first small molecule to block TSLP signaling pathways. It inhibited effectively eosinophil infiltration in house dust mite-induced and ovalbumin-challenged mouse models. Structure-activity relationship studies identified compound 11a, a biphenyl flavanone analog, as a novel human TSLP inhibitor for the discovery and development of new anti-allergic drugs.

Mediators of the homeostasis and effector functions of memory Th2 cells as novel drug targets in intractable chronic allergic diseases.

Allergic patients have life-long chronic inflammatory diseases with repeated relapses and exacerbations. Currently used allergy therapeutics have some limitations, which warrants a search for novel drug targets for allergy treatment. The studies on conventional allergic disease therapeutics have been focused on the pathology of allergy involving effector type 2 helper T cells (Th2). However, it has been suggested that allergen-specific memory Th2 cells are developed after the initial allergen exposure, which may play a critical role in the allergic relapses. Here, we discuss the contribution of memory Th2 cells to allergic diseases and the microenvironmental factors for chronic allergic disease persistence. Since most allergy drugs are prescribed to suppress symptoms of the diseases, targeting the different types of cells or factors contributing to allergic diseases persistence may cure the disease.

Pluronics: Intelligent building units for targeted cancer therapy and molecular imaging.

Pluronics are triblock copolymers, in which two hydrophilic poly (ethylene oxide) (PEO) blocks are connected via a hydrophobic poly propylene oxide (PPO) block. Because of their low molecular weight and high content of PEO, Pluronics have demonstrated the micellization phenomenon, which is dependent on temperature and/or concentration. With an understanding of micellization phenomenon in more detail, information on the morphology, micelle core radius, aggregation behavior with critical micelle concentration (CMC) and critical micelle temperature (CMT) and so on has been revealed. Based on this acquired information, various studies have been performed for biomedical applications such as drug delivery systems, tissue regeneration scaffolders, and biosurfactants. This review discusses the delivery of small molecules and macromolecules using Pluronic-based NPs and their composites.

Decreased Expression of Sphingosine-1-Phosphate Receptor 1 in the Blood Leukocyte of Rheumatoid Arthritis Patients.

Sphingosine-1-phosphate (S1P) plays an important role in trafficking leukocytes and developing immune disorders including autoimmunity. In the synovium of rheumatoid arthritis (RA) patients, increased expression of S1P was reported, and the interaction between S1P and S1P receptor 1 (S1P1) has been suggested to regulate the expression of inflammatory genes and the proliferation of synovial cells. In this study, we investigated the level of S1P1 mRNA expression in the blood leukocytes of RA patients. In contrast to the previous reports, the expression level of this gene was not correlated to their clinical scores, disease durations and ages. However, S1P1 was transcribed at a significantly lower level in the circulating leukocytes of RA patients when compared to age-, and sex-matched healthy controls. Since these data may suggest the participation of S1P1, further studies are needed to determine the role of this receptor in the pathogenesis of RA.

Surveillance of adverse drug events associated with etanercept prescribed for juvenile idiopathic arthritis in a single center up to 9-years: A retrospective observational study.

The introduction of biologic agents opened a new era of treatment of juvenile idiopathic arthritis (JIA) over the past decade. From clinical experience, it appears that biological agents are well tolerated overall, and serious adverse events are rare. However, such clinical studies have not been conducted in Korea. Therefore, we examined the safety profile of JIA patients with biologics in a single center in Korea. All JIA outpatients treated from April 2004 to June 2013 were enrolled and retrospectively reviewed. Pharmacy-based surveillance of adverse drug events (ADEs) was identified by recording the patient's symptoms in the medical record and suspected ADEs were additionally explored by screening laboratory test values and observing changes in medication orders. Finally, 83 patients were enrolled and experienced 109 ADEs in 52 patients. Most ADEs (99.1%) were mild to moderate in severity assessment. The total follow-up time was 328 patient-treatment years and the overall rate of ADEs was 0.33 per patient-years for etanercept. Infection including upper respiratory tract was the most common ADE and concomitant corticosteroids contributed to the risk of infections. If the dose of prednisolone increases 0.34 mg/kg/day, the probability of developing infections increases 3.29 times. Also, all 11 patients who stopped etanercept with injection site reactions were receiving a single use prefilled syringe. In our study, etanercept appears well tolerated and safe. Children affected by JIA should be carefully monitoring so as to limit the risk of ADEs during etanercept as much as possible. To gain further knowledge about risk profiles, national collaboration for the accumulation of long-term data should be encouraged in Korea.

Eukaryotic DNA damage responses: Homologous recombination factors and ubiquitin modification.

To prevent genomic instability disorders, cells have developed a DNA damage response. The response involves various proteins that sense damaged DNA, transduce damage signals, and effect DNA repair. In addition, ubiquitin modifications modulate the signaling pathway depending on cellular context. Among various types of DNA damage, double-stranded breaks are highly toxic to genomic integrity. Homologous recombination (HR) repair is an essential mechanism that fixes DNA damage because of its high level of accuracy. Although factors in the repair pathway are well established, pinpointing the exact mechanisms of repair and devising therapeutic applications requires more studies. Moreover, essential functions of ubiquitin modification in the DNA damage signaling pathway have emerged. In this review, to explore the eukaryotic DNA damage response, we will mention the functions of main factors in the HR repair pathway and ubiquitin modification.