Cellular subpopulations identified using an ensemble average of multiple dielectrophoresis measurements.

While the average value measurement approach can successfully analyze and predict the general behavior and biophysical properties of an isogenic cell population, it fails when significant differences among individual cells are generated in the population by intracellular changes such as the cell cycle, or different cellular responses to certain stimuli. Detecting such single-cell differences in a cell population has remained elusive. Here, we describe an easy-to-implement and generalizable platform that measures the dielectrophoretic cross-over frequency of individual cells by decreasing measurement noise with a stochastic method and computing ensemble average statistics. This platform enables multiple, real-time, label-free detection of individual cells with significant dielectric variations over time within an isogenic cell population. Using a stochastic method in combination with the platform, we distinguished cell subpopulations from a mixture of drug-untreated and -treated isogenic cells. Furthermore, we demonstrate that our platform can identify drug-treated isogenic cells with different recovery rates.

Integrin ß1 regulates the perineural invasion and radioresistance of oral squamous carcinoma cells by modulating cancer cell stemness.

Perineural invasion and radioresistance are the main determinants of treatment outcomes in oral squamous cell carcinoma (OSCC), but the exact mechanism is still unknown. We conducted an in vitro experiment to evaluate the role of integrin ß1 (ITGB1) in the perineural invasion, radioresistance, and tumor aggressiveness of OSCC. Two OSCC cell lines (SCC25, SCC15) and radiation-induced radioresistant OSCC cell lines were used in this study. The expression of ITGB1 was compared between control radiosensitive and radioresistant OSCC cell lines. ITGB1 was inhibited by small hairpin RNA, and then the adhesion to neuronal cells, responsiveness to radiation, and aggressiveness of both OSCC cell lines were evaluated. Expression of ITGB1 and adhesion to neuronal cells were increased in radioresistant OSCC compared with control radiosensitive OSCC, and increased ITGB1 expression was more prominent in cancer stem cell-like cells. When the expression of ITGB1 was inhibited, the adhesion to neuronal cells, resistance to radiation, and invasion and migration of radioresistant OSCC were significantly reduced. Moreover, the expression of cancer stem cell markers and size of spheroid formations were also significantly attenuated by inhibiting ITGB1. These findings suggest that ITGB1 may be a significant contributor to perineural invasion and the maintenance of radioresistance in OSCC cells, and is associated with cancer stem cell-like cells. Furthermore, our results suggest a possible relationship between perineural invasion and radioresistance of OSCC. More detailed research is warranted to evaluate the role of ITGB1 as a novel emerging therapeutic target for radioresistant OSCC.

Attachment to Peers and School: Longitudinal Moderators of the Relation Between Caregiver Psychological Distress and Adolescent Hopelessness.

Research has yet to determine how relationships outside of the family system may buffer negative outcomes associated with hopelessness among racial minority youth. In a sample of Black American youth (N = 512; 49% females) and their parents or caregivers, this study used longitudinal growth models to explore whether youth relationships (attachment to peers and attachment to school) moderated the association between caregiver distress (depressive symptoms and traumatic stress), and youth hopelessness. Adolescents' gender was examined to determine if there were gender differences present in these associations. Four linear growth models showed a significant change in levels of hopelessness over time for youth and a significant positive relation between caregiver distress and youth level of hopelessness. Attachment to peers and attachment to school did not equally moderate the relation between caregiver psychological distress and youth hopelessness. The type of caregiver distress had a differential effect on youth hopelessness in the context of the moderation models and based on gender. The type of caregiver distress had a differential effect on youth hopelessness in the context of the moderation models and based on gender. Implications for the importance of non-familial attachments among Black American youth with distressed parents are discussed.

Association between Tonsillectomy and Cardiovascular Diseases in Adults.

This study aimed to evaluate the association between tonsillectomy and cardiovascular diseases (CVDs) in the Korean adult population. Using data from the 2002-2015 Korean National Health Insurance Service-Health Screening Cohort, a total of 1082 participants aged 40 years or older who had undergone tonsillectomy were matched with 4328 control individuals for age, sex, income, and region of residence. We evaluated the incidence of CVDs in both the tonsillectomy and control groups and calculated the hazard ratios (HRs) of stroke, ischemic heart disease (IHD), and heart failure (HF) for participants who underwent tonsillectomy using a stratified Cox proportional hazard model. The incidence rates of stroke (81.3 vs. 46.6 per 10,000 person-years) and IHD (112.3 vs. 64.9 per 10,000 person-years) were significantly higher in patients who had undergone tonsillectomy than in the control group. After adjustment, the tonsillectomy group exhibited a 1.78-fold and 1.60-fold higher occurrence of stroke (CI = 1.32-2.42, p < 0.001) and IHD (CI = 1.24-2.08, p < 0.001), respectively, compared to the control group. However, there was no significant difference in the incidence rate of tonsillectomy and control groups (11.1 vs. 6.1 per 10,000 person-years). The HR of HF did not differ significantly between the tonsillectomy and control groups in the adjusted model (p = 0.513). We identified a significant relationship between a history of tonsillectomy and occurrence of stroke/IHD in the Korean adult population.

On-Chip Single-Cell Bioelectrical Analysis for Identification of Cell Electrical Phenotyping in Response to Sequential Electric Signal Modulation.

In recent years, an interesting biomarker called membrane breakdown voltage has been examined using artificial planar lipid bilayers. Even though they have great potential to identify cell electrical phenotyping for distinguishing similar cell lines or cells under different physiological conditions, the biomarker has not been evaluated in the context of living cell electrical phenotyping. Herein, we present a single-cell analysis platform to continuously measure the electric response in a large number of cells in parallel using electric frequency and voltage variables. Using this platform, we measured the direction of cell displacement and transparent cell image alteration as electric polarization of the cell responds to signal modulation, extracting the dielectrophoretic crossover frequency and membrane breakdown voltage for each cell, and utilizing the measurement results in the same spatiotemporal environment. We developed paired parameters using the dielectrophoretic crossover frequency and membrane breakdown voltage for each cell and evaluated the paired parameter efficiency concerning the identification of two different breast cancer cells and cell drug response. Moreover, we showed that the platform was able to identify cell electrical phenotyping, which was generated by subtle changes in cholesterol depletion-induced cell membrane integrity disruption when the paired parameter was used. Our platform introduced in this paper is extremely useful for facilitating more accurate and efficient evaluation of cell electrical phenotyping in a variety of applications, such as cell biology and drug discovery.

Patterns of patients with polypharmacy in adult population from Korea.

Polypharmacy and its rising global prevalence is a growing public health burden. Using a large representative nationwide Korean cohort (N = 761,145), we conducted a retrospective cross-sectional study aiming to identify subpopulations of patients with polypharmacy and characterize their unique patterns through cluster analysis. Patients aged ≥ 30 years who were prescribed at least one medication between 2014 and 2018 were included in our study. Six clusters were identified: cluster 1 mostly included patients who were hospitalized for a long time (4.3 ± 5.3 days); cluster 2 consisted of patients with disabilities (100.0%) and had the highest mean number of prescription drugs (7.7 ± 2.8 medications); cluster 3 was a group of low-income patients (99.9%); cluster 4 was a group of high-income patients (80.2%) who frequently (46.4 ± 25.9 days) visited hospitals/clinics (7.3 ± 2.7 places); cluster 5 was mostly elderly (74.9 ± 9.8 years) females (80.3%); and cluster 6 comprised mostly middle-aged (56.4 ± 1.5 years) males (88.6%) (all P < 0.001). Patients in clusters 1-5 had more prescribed medications and outpatient visit days than those in cluster 6 (all P < 0.001). Given limited health care resources, individuals with any of the identified phenotypes may be preferential candidates for participation in intervention programs for optimal medication use.

Ultra-sensitive dielectrophoretic surface charge multiplex detection inside a micro-dielectrophoretic device.

Label-free dielectrophoretic force-based surface charge detection has shown great potential for highly sensitive and selective sensing of metal ions and small biomolecules. However, this method suffers from a complex calibration process and measurement signal interference in simultaneous multi-analyte detection, thus creating difficulties in multiplex detection. We have developed a method to overcome these issues based on the optical discrimination of the dielectrophoretic behaviors of multiple microparticle probes considering the surface charge difference before and after self-assembling conjugation. In this report, we demonstrate and characterize this dielectrophoretic force-based surface charge detection method with particle probes functionalized by various biomolecules. This technique achieved an attomolar limit of detection (LOD) for Hg2+ in distilled water and a femtomolar LOD in drinking water using DNA aptamer-functionalized particle probes. More importantly, using two different DNA aptamer-functionalized particle probes for Hg2+ and Ag+, label-free dielectrophoretic multiplex detection of these species in drinking water with a femtomolar and a nanomolar LOD was achieved for the first time.

A Novel Device for Blood Drainage after Le Fort I Osteotomy: Maxillary Sinus Ventilation Drainage (MSVD).

The purpose of this study is to present a novel maxillary sinus ventilation drainage (MSVD) device which facilitates blood drainage and nasal breathing after Le Fort I osteotomy. One hundred patients who underwent bimaxillary orthognathic surgery from January 2016 to June 2016 at the Department of Oral and Maxillofacial Surgery, Chung-Ang University Hospital were retrospectively selected and divided into two groups. MSVD was applied in 50 patients, who were allocated to the MSVD group, while the remaining 50 patients, in whom MSVD was not applied, were allocated to the non-MSVD group. All patients underwent a cone-beam computed tomography (CBCT) scan before and 2 days after surgery. CBCT was used to analyze middle meatus patency and the percentage of hematoma volume per entire maxillary sinus volume. Statistical comparisons between the two groups were performed using the Chi-squared and Mann-Whitney U tests to investigate the clinical effectiveness of MSVD. The MSVD group showed significantly higher maintenance ratio of the middle meatus patency and a higher percentage of maxillary sinus air volume (p < 0.05) than the non-MSVD group. MSVD facilitated nasal breathing after Le Fort I osteotomy by reducing hematoma inside the maxillary sinus and promoting middle meatal patency.

Tonsillectomy in Adults over 40 Years of Age Does Not Increase the Risk of Pneumonia: A Three-Year Longitudinal Follow-Up Study.

To evaluate the effect of tonsillectomy on the subsequent risk of pneumonia in an adult population, a longitudinal follow-up case control study was conducted using a national health screening cohort dataset between 2003 and 2012. A total of 1005 tonsillectomy participants were 1:4 matched with 4020 control participants for age, sex, income, and region of residence. The number of pneumonia diagnoses were counted from the index date (ID) to the date after the first-year (post-ID 1y), second-year (post-ID 2y), and third-year (post-ID 3y) periods. Simple linear regression and multiple linear regression were conducted to calculate estimated values (EVs) and 95% confidence intervals for each post-ID pneumonia and compared between the two groups. Subgroup analyses were performed according to age, sex, and the number of pneumonia cases during the year prior to the ID (pre-ID 1y). In the simple linear regression model, post-ID pneumonia did not show a significant correlation with tonsillectomy (post-ID 1y: EV = 0.003; post-ID 2y: EV = 0.007; post-ID 3y: EV = 0.013; all p > 0.05). In the multiple regression model, post-ID pneumonia also did not show a significant correlation with tonsillectomy (post-ID 1y: EV = 0.001; post-ID 2y: EV = 0.006; post-ID 3y: EV = 0.011; all p > 0.05). In the subgroup analyses, tonsillectomy did not show a significant correlation with post-ID pneumonia in either the simple linear regression or multiple linear regression models (all p > 0.05). Tonsillectomy performed in the adult population did not show any effect in increasing the incidence of pneumonia during the first three postoperative years.

Role of Primary Care and Challenges for Public-Private Cooperation during the Coronavirus Disease 2019 Pandemic: An Expert Delphi Study in South Korea.

PURPOSE: The aim of this study was to explore the role of primary care facilities and their support measures in response to coronavirus disease 2019 (COVID-19) and to identify challenges to achieving public-private cooperation in South Korea. MATERIALS AND METHODS: Twenty-four leading experts were selected and recruited to participate in this iterative web-based Delphi study. An open-ended questionnaire was administered to collect the expert panel's views in the first round. In the second round, the panel was asked to rate on a 5-point Likert scale their agreement with individual items gleaned from qualitative content analysis of views expressed in the first round. The participants were offered the opportunity to reevaluate and correct their initial responses in subsequent rounds. Responses in the second and following rounds were analyzed using quantitative descriptive statistics. RESULTS: The first and second rounds were completed by 54.2% (n=13/24) and 58.3% (n=14/24) of the selected panel, respectively, while 10 out of these 14 participants completed the third round. The panel cited in-person essential medical services, telehealth for fever/respiratory symptoms, surveillance for influenzae-like illness, and minimization of spread to staff as important and appropriate roles of primary care, which are urgent and feasible during a pandemic. Regarding conditions/support for these roles, the panel indicated that institutional support and funding for separate areas, workforce, and telehealth, along with public-private collaborative governance, are urgent, but not feasible. CONCLUSION: This study provides guidance on strategies for continuing the required roles of primary care and highlights a need to strengthen public-private partnerships during pandemic events in Korea.

Programmable Aggregation of Artificial Cells with DNA Signals.

Cell aggregation is a complex behavior that is closely related to the viability, differentiation, and migration of cells. An effort to create synthetic analogs could lead to considerable advances in cell physiology and biophysics. Rendering and modulating such a dynamic artificial cell system require mechanisms for receiving, transducing, and transmitting intercellular signals, yet effective tools are limited at present. Here we construct synthetic cells from engineered lipids and show their programmable aggregation behaviors using DNA oligonucleotides as signaling molecules. The artificial cells have transmembrane channels made of DNA origami that are used to recognize and process intercellular signals. We demonstrate that multiple small vesicles aggregate onto a giant vesicle after a transduction of external DNA signals by an intracellular enzyme and that the small vesicles dissociate when receiving "release" signals. This work provides new possibilities for building synthetic protocells capable of chemical communication and coordination.

Assessment of Additional MRI-Detected Breast Lesions Using the Quantitative Analysis of Contrast-Enhanced Ultrasound Scans and Its Comparability with Dynamic Contrast-Enhanced MRI Findings of the Breast.

Purpose: To assess the diagnostic performance of contrast-enhanced ultrasound (CEUS) for additional MR-detected enhancing lesions and to determine whether or not kinetic pattern results comparable to dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) of the breast can be obtained using the quantitative analysis of CEUS. Materials and Methods: In this single-center prospective study, a total of 71 additional MR-detected breast lesions were included. CEUS examination was performed, and lesions were categorized according to the Breast Imaging-Reporting and Data System (BI-RADS). The sensitivity, specificity, and diagnostic accuracy of CEUS were calculated by comparing the BI-RADS category to the final pathology results. The degree of agreement between CEUS and DCE-MRI kinetic patterns was evaluated using weighted kappa. Results: On CEUS, 46 lesions were assigned as BI-RADS category 4B, 4C, or 5, while 25 lesions category 3 or 4A. The diagnostic performance of CEUS for enhancing lesions on DCE-MRI was excellent, with 84.9% sensitivity, 94.4% specificity, and 97.8% positive predictive value. A total of 57/71 (80%) lesions had correlating kinetic patterns and showed good agreement (weighted kappa = 0.66) between CEUS and DCE-MRI. Benign lesions showed excellent agreement (weighted kappa = 0.84), and invasive ductal carcinoma (IDC) showed good agreement (weighted kappa = 0.69). Conclusion: The diagnostic performance of CEUS for additional MR-detected breast lesions was excellent. Accurate kinetic pattern assessment, fairly comparable to DCE-MRI, can be obtained for benign and IDC lesions using CEUS.

Variable Membrane Dielectric Polarization Characteristic in Individual Live Cells.

Investigation of the dielectric properties of cell membranes plays an important role in understanding the biological activities that sustain cellular life and realize cellular functionalities. Herein, the variable dielectric polarization characteristics of cell membranes are reported. In controlling the dielectric polarization of a cell using dielectrophoresis force spectroscopy, different cellular crossover frequencies were observed by modulating both the direction and sweep rate of the frequency. The crossover frequencies were used for the extraction of the variable capacitance, which is involved in the dielectric polarization across the cell membranes. In addition, this variable phenomenon was investigated by examining cells whose membranes were cholesterol-depleted with methyl-ß-cyclodextrin, which verified a strong correlation between the variable dielectric polarization characteristics and membrane composition changes. This study presented the dielectric polarization properties in live cells' membranes that can be modified by the regulation of external stimuli and provided a powerful platform to explore cellular membrane dielectric polarization.

A novel automatic segmentation and tracking method to measure cellular dielectrophoretic mobility from individual cell trajectories for high throughput assay.

BACKGROUND AND OBJECTIVE: The dielectrophoresis (DEP) technique is increasingly being recognised as a potentially valuable tool for non-contact manipulation of numerous cells as well as for biological single cell analysis with non-invasive characterisation of a cell's electrical properties. Several studies have attempted to track multiple cells to characterise their cellular DEP mobility. However, they encountered difficulties in simultaneously tracking the movement of a large number of individual cells in a bright-field image sequence because of interference from the background electrode pattern. Consequently, this present study aims to develop an automatic system for imaging-based characterisation of cellular DEP mobility, which enables the simultaneous tracking of several hundred of cells inside a microfluidic device. METHODS: The proposed method for segmentation and tracking of cells consists of two main stages: pre-processing and particle centre localisation. In the pre-processing stage, background subtraction and contrast enhancement were performed to distinguish the cell region from the background image. In the particle centre localisation stage, the unmarked cell was automatically detected via graph-cut algorithm-based K-means clustering. RESULTS: Our algorithm enabled segmentation and tracking of numerous Michigan Cancer Foundation-7 (MCF-7) cell trajectories while the DEP force was oscillated between positive and negative. The cell tracking accuracy and cell count capability was at least 90% of the total number of cells with the newly developed algorithm. In addition, the cross-over frequency was measured by analysing the segmented and tracked trajectory data of the cellular movements caused by the positive and negative DEP force. The measured cross-over frequency was compared with previous results. The multi-cellular movements investigation based on the measured cross-over frequency was repeated until the viability of cells was unchanged in the same environment as in a microfluidic device. The results were statistically consistent, indicating that the developed algorithm was reliable for the investigation of DEP cellular mobility. CONCLUSION: This study developed a powerful platform to simultaneously measure the DEP-induced trajectories of numerous cells, and to investigate in a robust, efficient, and accurate manner the DEP properties at both the single cell and cell ensemble level.

ERK1/2-Nanog signaling pathway enhances CD44(+) cancer stem-like cell phenotypes and epithelial-to-mesenchymal transition in head and neck squamous cell carcinomas.

Head and neck squamous cell carcinomas (HNSCCs) harbor a subset of cells that are CD44(+) and present with malignancy and radiotherapy resistance. As a key regulator of self-renewal, Nanog expression not only determines cell fate in pluripotent cells but also mediates tumorigenesis in cancer cells; thus, we examined the role of Nanog in CD44(+) HNSCC. Three HNSCC cell lines, tumor xenografts, and patient tumors were examined. Nanog levels were significantly higher in CD44(+) HNSCC spheroids than in CD44(-) spheroids, and further increased when grown as spheroids to enrich for CSCs. CD44(+) spheroids showed a 3.4-7.5-fold increase in migration and invasion compared with CD44(-) spheroids and were resistant to radiation therapy, which was reversed by inhibiting Nanog. Nanog knockdown also decreased spheroid formation by 66.5-68.8%. Moreover, a phosphokinase array identified upregulated ERK1/2 signaling in CD44(+) HNSCC cells compared with that in CD44(-) cells. ERK1/2 signaling was found to regulate Nanog expression, aiding tumor progression, metastasis, and radiotherapy resistance. In xenograft models, the combination of radiation and Nanog or ERK1/2 inhibition inhibited tumor growth by 75.6% and 79.1%, respectively. In lung metastasis models, CD44(+) cells injected into the tail vein of mice led to significantly more lung metastases and higher Nanog expression level compared with that by ERK1/2-knockdown CD44(+) cells. Finally, in tumor tissues, CD44 and Nanog expression levels were correlated with tumorigenesis in HNSCC patients. Thus, targeting Nanog and the ERK1/2 signaling pathway may prevent or reverse CSC phenotypes and epithelial-mesenchymal transition that drive tumor progression, metastasis, and radiotherapy resistance in HNSCC.

Can Osteopenia Induce Residual Dizziness After Treatment of Benign Paroxysmal Positional Vertigo?

OBJECTIVE: This study aimed to determine whether there is an association between osteopenia and residual dizziness after successful treatment of benign paroxysmal positional vertigo (BPPV). METHODS: In all, 62 patients with canalolithiasis-type BPPV were included in the study. Patients were divided into two groups according to the presence of residual dizziness after resolution of BPPV. Univariate and multivariate analyses were performed to determine the factors associated with residual dizziness. Patients were analyzed based on age, sex, affected semicircular canal, affected side, BPPV duration, and presence of hypertension, diabetes mellitus, hyperlipidemia, and osteopenia. RESULTS: In univariate analysis, BPPV duration and osteopenia showed a relatively significant association (p < 0.20) with the development of residual dizziness. On subsequent multivariate analysis using these factors, osteopenia remained a statistically significant factor in association with residual dizziness (p = 0.012, odds ratio, 9.916). CONCLUSION: Osteopenia is associated with the development of residual dizziness. BPPV patients with osteopenia more frequently suffer from residual dizziness after successful treatment of BPPV than those without osteopenia.

Cytoplasmic HMGB1 and HMGB1-Beclin1 complex are increased in radioresistant oral squamous cell carcinoma.

Cytoplasmic high mobility group box 1 (HMGB1) is an autophagy regulator, and autophagy is important in the radioresistance of various solid cancers. We evaluated the degree of autophagy and cytoplasmic HMGB1 in radioresistant oral squamous cell carcinoma (SCC) by culturing the SCC15 and quasiliquid layer 1 (QLL1) SCC cell lines that originate from cancer of the oral tongue and a metastatic lymph node, respectively, and then delivered radiation to induce radioresistance to cells. We then compared the degree of autophagy between non-irradiated control and radioresistant cancer cells using a western blot assay. We also compared the total and cytoplasmic concentrations of HMGB1 between the non-irradiated control and radioresistant cancer cells by western blot assay, and extracellular concentrations of HMGB1 with an enzyme-linked immunosorbent assay (ELISA). Formation of an HMGB1-Beclin1 complex was evaluated by immunofluorescence and co-immunoprecipitation assays. Autophagy increased in the radioresistant SCC15 cells (compared with non-irradiated control SCC15 cells) but not in the radioresistant QLL1 cells. The total amount of HMGB1 expression within cells did not differ; however, the degree of cytoplasmic HMGB1 expression was higher in radioresistant SCC15 cells than in non-irradiated control SCC15 cells. The HMGB1-Beclin1 complex, which is a main regulator of autophagy, was also increased in radioresistant SCC15 cells compared with non-irradiated control SCC15 cells. Autophagy flux and cytoplasmic HMGB1-Beclin1 increased after the acquisition of radioresistance in oral SCC.

Clinical utility of cervical vestibular-evoked myogenic potentials in predicting residual dizziness after benign paroxysmal positional vertigo.

OBJECTIVES: In the present study, the value of cervical vestibular-evoked myogenic potential (cVEMP) as a predictive factor for residual dizziness after recovery of benign paroxysmal positional vertigo (BPPV) was evaluated. METHODS: The present study included 65 patients who had BPPV and underwent cVEMP testing. Patients were divided into two groups depending on the presence or absence of residual dizziness after recovery of BPPV. Univariate and multivariate analyses were performed to determine the factors associated with residual dizziness using age, gender, affected semicircular canal, affected side, BPPV duration, and cVEMP parameters. RESULTS: In univariate analysis, cVEMP-modified interaural amplitude difference (IAD) ratio and p13 latency showed a relatively significant association (p < 0.20) with residual dizziness. Based on multivariate analysis, increased cVEMP-modified interaural amplitude difference (IAD) ratio at the affected side (≥25%; p = 0.018, OR 6.623) remained as an associated factor. CONCLUSIONS: Increased cVEMP-modified IAD ratio at the affected side is associated with residual dizziness. BPPV patients with increased cVEMP-modified IAD ratio at the affected side are more likely to have residual dizziness after recovery of BPPV. SIGNIFICANCE: cVEMP testing could be used for the prediction of residual dizziness. An increased cVEMP-modified IAD ratio at the affected side may be used as a predictor of residual dizziness.