Finnish-specific AKT2 gene variant leads to impaired insulin signalling in myotubes.

Finnish-specific gene variant p.P50T/AKT2 (minor allele frequency (MAF) = 1.1%) is associated with insulin resistance and increased predisposition to type 2 diabetes. Here, we have investigated in vitro the impact of the gene variant on glucose metabolism and intracellular signalling in human primary skeletal muscle cells, which were established from 14 male p.P50T/AKT2 variant carriers and 14 controls. Insulin-stimulated glucose uptake and glucose incorporation into glycogen were detected with 2-[1,2-3H]-deoxy-D-glucose and D-[14C]-glucose, respectively, and the rate of glycolysis was measured with a Seahorse XFe96 analyzer. Insulin signalling was investigated with Western blotting. The binding of variant and control AKT2-PH domains to phosphatidylinositol (3,4,5)-trisphosphate (PI(3,4,5)P3) was assayed using PIP StripsTM Membranes. Protein tyrosine kinase and serine-threonine kinase assays were performed using the PamGene® kinome profiling system. Insulin-stimulated glucose uptake and glycogen synthesis in myotubes in vitro were not significantly affected by the genotype. However, the insulin-stimulated glycolytic rate was impaired in variant myotubes. Western blot analysis showed that insulin-stimulated phosphorylation of AKT-Thr308, AS160-Thr642 and GSK3ß-Ser9 was reduced in variant myotubes compared to controls. The binding of variant AKT2-PH domain to PI(3,4,5)P3 was reduced as compared to the control protein. PamGene® kinome profiling revealed multiple differentially phosphorylated kinase substrates, e.g. calmodulin, between the genotypes. Further in silico upstream kinase analysis predicted a large-scale impairment in activities of kinases participating, for example, in intracellular signal transduction, protein translation and cell cycle events. In conclusion, myotubes from p.P50T/AKT2 variant carriers show multiple signalling alterations which may contribute to predisposition to insulin resistance and T2D in the carriers of this signalling variant.

Fear of COVID-19, healthy eating behaviors, and health-related behavior changes as associated with anxiety and depression among medical students: An online survey.

Background: Medical students' health and wellbeing are highly concerned during the COVID-19 pandemic. This study examined the impacts of fear of COVID-19 (FCoV-19S), healthy eating behavior, and health-related behavior changes on anxiety and depression. Methods: We conducted an online survey at 8 medical universities in Vietnam from 7th April to 31st May 2020. Data of 5,765 medical students were collected regarding demographic characteristics, FCoV-19S, health-related behaviors, healthy eating score (HES), anxiety, and depression. Logistic regression analyses were used to explore associations. Results: A lower likelihood of anxiety and depression were found in students with a higher HES score (OR = 0.98; 95%CI = 0.96, 0.99; p = 0.042; OR = 0.98; 95%CI = 0.96, 0.99; p = 0.021), and in those unchanged or more physical activities during the pandemic (OR = 0.54; 95%CI = 0.44, 0.66; p < 0.001; OR = 0.44; 95%CI = 0.37, 0.52; p < 0.001) as compared to those with none/less physical activity, respectively. A higher likelihood of anxiety and depression were reported in students with a higher FCoV-19S score (OR = 1.09; 95%CI = 1.07, 1.12; p < 0.001; OR = 1.06; 95%CI = 1.04, 1.08; p < 0.001), and those smoked unchanged/more during the pandemic (OR = 6.67; 95%CI = 4.71, 9.43; p < 0.001; OR = 6.77; 95%CI = 4.89, 9.38; p < 0.001) as compared to those stopped/less smoke, respectively. In addition, male students had a lower likelihood of anxiety (OR = 0.79; 95%CI = 0.65, 0.98; p = 0.029) compared to female ones. Conclusions: During the pandemic, FCoV-19S and cigarette smoking had adverse impacts on medical students' psychological health. Conversely, staying physically active and having healthy eating behaviors could potentially prevent medical students from anxiety and depressive symptoms.

Body mass index, body fat percentage, and visceral fat as mediators in the association between health literacy and hypertension among residents living in rural and suburban areas.

Background: Hypertension is a major cause of death and disability worldwide. Enhancing health literacy (HL) may help to alleviate the risk of hypertension and its burden. However, evidence on the association between HL and hypertension and potential mechanisms remain to be explored. Objectives: This study examined the association between HL and hypertension; and explored whether body mass index (BMI), body fat percentage (PBF), and visceral fat (VF) were mediators of this association in people who resided in rural and suburban areas in Vietnam. Methods: A cross-sectional survey was conducted from 1st July to 31st December 2019, involving 1655 residents and exploring participants' sociodemographic characteristics, HL, health-related behaviors, comorbidities, body composition, and blood pressure (BP). People with systolic BP ≥ 140 mmHg or diastolic BP ≥ 90 mmHg or using antihypertensive medication were classified as having hypertension. Multiple logistic regression and mediation analyses were used to explore associations. Results: The hypertension prevalence was 41.9% (694/1,655). In adjusted models, a higher HL score was associated with a lower hypertension likelihood (OR = 0.96; 95%CI = 0.95-0.97; p < 0.001). Factors associated with a higher odd of hypertension were overweight/obese (OR = 1.69; 95%CI = 1.24-2.29; p = 0.001), high PBF (OR = 2.35; 95%CI = 1.85-2.99; p < 0.001), and high VF (OR = 2.27; 95%CI = 1.63-3.16; p < 0.001). Notably, PBF significantly mediated the association between HL and hypertension (indirect effect, OR = 0.99; 95%CI = 0.98-0.99; p = 0.009; percent mediated = 8.56%). The mediating roles of BMI and VF were not found. Conclusion: The prevalence of hypertension was relatively high. People with better HL were less likely to have hypertension. The association between HL and hypertension was partially explained by PBF. Strategic approaches are required to improve people's HL and body fat which further help to manage hypertension in rural and suburban areas.

Predictors of eHealth Literacy and Its Associations with Preventive Behaviors, Fear of COVID-19, Anxiety, and Depression among Undergraduate Nursing Students: A Cross-Sectional Survey.

Background: The infodemic has been co-existing with the COVID-19 pandemic with an influx of misinformation and conspiracy theories. These affect people's psychological health and adherence to preventive measures. eHealth literacy (eHEALS) may help with alleviating the negative effects of the infodemic. As nursing students are future healthcare professionals, having adequate eHEALS skills is critically important in their clinical practice, safety, and health. This study aimed to (1) explore the eHEALS level and its associated factors, and (2) examine the associations of eHEALS with preventive behaviors, fear of COVID-19 (FCV-19S), anxiety, and depression among nursing students. Methods: We surveyed 1851 nursing students from 7 April to 31 May 2020 from eight universities across Vietnam. Data were collected, including demographic characteristics, eHEALS, adherence to preventive behaviors (handwashing, mask-wearing, physical distancing), FCV-19S, anxiety, and depression. Linear and logistic regression analyses were performed appropriately to examine associations. Results: The mean score of eHEALS was 31.4 ± 4.4. The eHEALS score was significantly higher in males (unstandardized regression coefficient, B, 0.94; 95% confidence interval, 95% CI, 0.15 to 1.73; p = 0.019), and students with a better ability to pay for medication (B, 0.79; 95% CI, 0.39 to 1.19; p < 0.001), as compared to their counterparts. Nursing students with a higher eHEALS score had a higher likelihood of adhering to hand-washing (odds ratio, OR, 1.18; 95% CI, 1.15 to 1.22; p < 0.001), mask-wearing (OR, 1.15; 95% CI, 1.12 to 1.19; p < 0.001), keeping a safe physical distance (OR, 1.20; 95% CI, 1.15 to 1.25; p < 0.001), and had a lower anxiety likelihood (OR, 0.95; 95% CI, 0.92 to 0.99; p = 0.011). Conclusions: Nursing students who were men and with better ability to pay for medication had higher eHEALS scores. Those with higher eHEALS scores had better adherence to preventive measures, and better psychological health. The development of strategies to improve eHEALS of nursing students may contribute to COVID-19 containment and improve their psychological health.

Investigation of a Series of 2-(2'-Hydroxyaryl)benzazole Derivatives: Photophysical Properties, Excited-State Intramolecular Proton-Transfer Reactions, and Observation of Long-Lived Triplet Excited States.

Excited state intramolecular proton transfer (ESIPT) has drawn much attention for its important applications in a variety of areas. Here, the steady-state and time-resolved absorption spectroscopic experiments as well as DFT/TD-DFT calculations are employed to study the photophysical properties and photochemical reaction mechanisms of 2-(2'-hydroxyphenyl) benzoxazole (HBO) and selected derivatives (compounds 1-3). Because of their larger π-conjugation framework, compounds 1-3 display red-shifted absorbance but blue-shifted fluorescence compared with HBO. A fast ESIPT process is observed directly for HBO while compound 3 has an enol/keto equilibrium type of ESIPT that exhibits dual emission. Interestingly, only the emission of the enol form is observed for compounds 1 and 2 which suggests that the ESIPT process is strongly inhibited. These results indicate the decoration with electron-withdrawing groups such as thiadiazol and pyrazine on the hydroxyphenyl ring (compounds 1 and 2) apparently suppresses the proton-transfer processes in their excited states. Whereas the ESIPT process is rarely increased for compound 3 that modified with the phenanthrol ring, because the effective conjugation is reduced for compound 3 compared with HBO. The work here provides fundamental insights that may be useful for designing novel ESIPT molecules in the future.

Negative Impact of Fear of COVID-19 on Health-Related Quality of Life Was Modified by Health Literacy, eHealth Literacy, and Digital Healthy Diet Literacy: A Multi-Hospital Survey.

BACKGROUND: The COVID-19 pandemic has been disseminating fear in the community, which has affected people's quality of life, especially those with health problems. Health literacy (HL), eHealth literacy (eHEAL), and digital healthy diet literacy (DDL) may have potential impacts on containing the pandemic and its consequences. This study aimed to examine the association between the fear of COVID-19 scale (FCoV-19S) and the health-related quality of life (HRQoL), and to examine the effect modification by HL, eHEAL, and DDL on this association. METHODS: A cross-sectional study was conducted in 11 hospitals across Vietnam from 7 April to 31 May 2020. Data were collected on 4348 outpatients, including demographic characteristics, HL, eHEAL, DDL, FCoV-19S, and HRQoL. Multiple linear regression and interaction models were used to explore associations. RESULTS: Patients with higher FCoV-19S scores had lower HRQoL scores (unstandardized coefficient, B = -0.78, p < 0.001). HL (B = 0.20, p < 0.001), eHEAL (B = 0.24, p < 0.001), and DDL (B = 0.20, p < 0.001) were positively associated with higher HRQoL scores. The negative impact of FCoV-19S on HRQoL was significantly attenuated by higher eHEAL score groups (from one standard deviation (SD) below the mean, B = -0.93, p < 0.001; to the mean, B = -0.85, p < 0.001; and one SD above the mean, B = -0.77, p < 0.001); and by higher DDL score groups (from one SD below the mean, B = -0.92, p < 0.001; to the mean, B = -0.82, p < 0.001; and one SD above the mean, B = -0.72, p < 0.001). CONCLUSIONS: eHealth literacy and digital healthy diet literacy could help to protect patients' health-related quality of life from the negative impact of the fear of COVID-19 during the pandemic.

Simvastatin profoundly impairs energy metabolism in primary human muscle cells.

OBJECTIVES: Simvastatin use is associated with muscular side effects, and increased risk for type 2 diabetes (T2D). In clinical use, simvastatin is administered in inactive lipophilic lactone-form, which is then converted to active acid-form in the body. Here, we have investigated if lactone- and acid-form simvastatin differentially affect glucose metabolism and mitochondrial respiration in primary human skeletal muscle cells. METHODS: Muscle cells were exposed separately to lactone- and acid-form simvastatin for 48 h. After pre-exposure, glucose uptake and glycogen synthesis were measured using radioactive tracers; insulin signalling was detected with Western blotting; and glycolysis, mitochondrial oxygen consumption and ATP production were measured with Seahorse XFe96 analyzer. RESULTS: Lactone-form simvastatin increased glucose uptake and glycogen synthesis, whereas acid-form simvastatin did not affect glucose uptake and decreased glycogen synthesis. Phosphorylation of insulin signalling targets Akt substrate 160 kDa (AS160) and glycogen synthase kinase 3ß (GSK3ß) was upregulated with lactone-, but not with acid-form simvastatin. Exposure to both forms of simvastatin led to a decrease in glycolysis and glycolytic capacity, as well as to a decrease in mitochondrial respiration and ATP production. CONCLUSIONS: These data suggest that lactone- and acid-forms of simvastatin exhibit differential effects on non-oxidative glucose metabolism as lactone-form increases and acid-form impairs glucose storage into glycogen, suggesting impaired insulin sensitivity in response to acid-form simvastatin. Both forms profoundly impair oxidative glucose metabolism and energy production in human skeletal muscle cells. These effects may contribute to muscular side effects and risk for T2D observed with simvastatin use.

Correlations between Excessive Body Mass Index, Body Perception, Physical Activity, and Respiratory Functions among Youths in an Urban Setting of Vietnam.

Young adults are susceptible to overweight and obesity and their adverse outcomes. However, limited studies have been conducted to understand this health problem in Vietnamese youths. This study was conducted to examine the rate of overweight and obesity, as well as the relationship between this condition on body perception, physical activity, and respiratory function among young adults in Hanoi, Vietnam. We performed a cross-sectional survey with 367 students aged 18-25 years at the Hanoi University of Pharmacy from June 2017 to June 2018. The result showed that the rate of overweight and obesity in our sample was 16.6%. There were 55.7% of overweight/obese students having the misperception of their body image. Results of multivariate regression analysis showed that overweight/obesity increased nearly three times (OR = 2.8, 95% CI: 1.5-5.0) the ability to be active. Overweight/obese students with physical activity had a better respiratory function. To conclude, this study underlined the associations between overweight and obesity with physical activity, body image, and respiratory function in our young sample. Further longitudinal studies should be warranted to examine their causal relationships.

Using mobile multimedia platforms in teaching dental diagnosis.

OBJECTIVE: Mobile Multimedia Platforms (MMPs) are prolific tools that can be used by individuals and corporations to share content. However, few studies have shown the effectiveness of MMPs as educational tools. Through this study, we aimed to evaluate the effectiveness of MMPs in improving basic dental diagnostic skills. In addition, we captured student feedback on the use of MMPs in a dental curriculum. METHOD: In this voluntary interventional study on 89 senior dental students, we created pilot learning modules through an MMP called Instagram Stories to teach dental diagnosis. We evaluated the efficacy of the modules through diagnostic tests that were given to dental students who were close to graduating. RESULTS: The students showed a significant increase in diagnostic test scores from 49% to 73% (p < 0.05) after the use of an MMP. Furthermore, the students' feedback on the MMP indicated that most students found it easy and enjoyable to use. CONCLUSION: Our study data show that MMPs may be used to improve training in basic dental diagnostic skills and can serve as an adjunct teaching tool. Moreover, MMP modules can potentially enrich professional education in developing countries where access to educational resources is limited.

Fear of COVID-19 Scale-Associations of Its Scores with Health Literacy and Health-Related Behaviors among Medical Students.

The coronavirus disease 2019 (COVID-19) pandemic causes fear, as its immediate consequences for the public have produced unprecedented challenges for the education and healthcare systems. We aimed to validate the fear of COVID-19 scale (FCoV-19S) and examine the association of its scores with health literacy and health-related behaviors among medical students. A cross-sectional study was conducted from 7 to 29 April 2020 on 5423 students at eight universities across Vietnam, including five universities in the North, one university in the Center, two universities in the South. An online survey questionnaire was used to collect data on participants' characteristics, health literacy, fear of COVID-19 using the FCoV-19S, and health-related behaviors. The results showed that seven items of the FCoV-19S strongly loaded on one component, explained 62.15% of the variance, with good item-scale convergent validity and high internal consistency (Cronbach's alpha = 0.90). Higher health literacy was associated with lower FCoV-19S scores (coefficient, B, -0.06; 95% confidence interval, 95%CI, -0.08, -0.04; p < 0.001). Older age or last academic years, being men, and being able to pay for medication were associated with lower FCoV-19S scores. Students with higher FCoV-19S scores more likely kept smoking (odds ratio, OR, 1.11; 95% CI, 1.08, 1.14; p < 0.001) or drinking alcohol (OR, 1.04; 95% CI, 1.02, 1.06; p < 0.001) at an unchanged or higher level during the pandemic, as compared to students with lower FCoV-19S scores. In conclusion, the FCoV-19S is valid and reliable in screening for fear of COVID-19. Health literacy was found to protect medical students from fear. Smoking and drinking appeared to have a negative impact on fear of COVID-19. Strategic public health approaches are required to reduce fear and promote healthy lifestyles during the pandemic.

Deprotection Strategies for Thioimidates during Fmoc Solid-Phase Peptide Synthesis: A Safe Route to Thioamides.

Thioamides are important biophysical probes of peptide folding but are prone to α-C epimerization during Fmoc solid-phase peptide synthesis. The stereochemical integrity of thioamide-containing peptides can be dramatically improved by protecting the thioamide as a thioimidate during synthesis. A drawback of this approach, however, is that once synthesis of the peptide is complete, regeneration of the thioamide requires the toxic, corrosive, and flammable gas H2S. This work examines several approaches to supplant H2S as a deprotection reagent in favor of a safer and more convenient alternative. Ultimately, a new application of the 4-azidobenzyl protecting group to thioamides was found to provide the most suitable means of both protection of α-C stereochemistry and conversion back to thioamide.

Community-wide Screening for Tuberculosis in a High-Prevalence Setting.

BACKGROUND: The World Health Organization has set ambitious targets for the global elimination of tuberculosis. However, these targets will not be achieved at the current rate of progress. METHODS: We performed a cluster-randomized, controlled trial in Ca Mau Province, Vietnam, to evaluate the effectiveness of active community-wide screening, as compared with standard passive case detection alone, for reducing the prevalence of tuberculosis. Persons 15 years of age or older who resided in 60 intervention clusters (subcommunes) were screened for pulmonary tuberculosis, regardless of symptoms, annually for 3 years, beginning in 2014, by means of rapid nucleic acid amplification testing of spontaneously expectorated sputum samples. Active screening was not performed in the 60 control clusters in the first 3 years. The primary outcome, measured in the fourth year, was the prevalence of microbiologically confirmed pulmonary tuberculosis among persons 15 years of age or older. The secondary outcome was the prevalence of tuberculosis infection, as assessed by an interferon gamma release assay in the fourth year, among children born in 2012. RESULTS: In the fourth-year prevalence survey, we tested 42,150 participants in the intervention group and 41,680 participants in the control group. A total of 53 participants in the intervention group (126 per 100,000 population) and 94 participants in the control group (226 per 100,000) had pulmonary tuberculosis, as confirmed by a positive nucleic acid amplification test for Mycobacterium tuberculosis (prevalence ratio, 0.56; 95% confidence interval [CI], 0.40 to 0.78; P<0.001). The prevalence of tuberculosis infection in children born in 2012 was 3.3% in the intervention group and 2.6% in the control group (prevalence ratio, 1.29; 95% CI, 0.70 to 2.36; P = 0.42). CONCLUSIONS: Three years of community-wide screening in persons 15 years of age or older who resided in Ca Mau Province, Vietnam, resulted in a lower prevalence of pulmonary tuberculosis in the fourth year than standard passive case detection alone. (Funded by the Australian National Health and Medical Research Council; ACT3 Australian New Zealand Clinical Trials Registry number, ACTRN12614000372684.).

Demonstration of Baird's rule complementarity in the singlet state with implications for excited-state intramolecular proton transfer.

The aromatic character of an arene is proposed to switch from aromatic in the ground state (S0) to antiaromatic in the S1 and T1 excited states. This behavior is known as Baird's rule and has been invoked to explain excited-state properties, primarily in the triplet state, whereas rationalization of antiaromaticity in the singlet state is less developed. This work demonstrates the first application of Baird's rule to rationalize previously unexplained experimental behavior of the singlet state process known as excited-state intramolecular proton transfer (ESIPT). Further, by analyzing the variations in isotropic magnetic shielding around the base arenes (benzene and naphthalene) of ESIPT fluorophores in the S0 and S1 electronic states, different shielding distributions indicate a complementarity to Baird's rule: greater aromaticity in S0 leads to greater antiaromaticity in S1 and vice versa. These findings have immediate application in the design of functional ESIPT fluorophores and, more generally, for photochemical reactions that are driven by the relief of antiaromaticity in the excited state. Notably, a tenet of traditional chromophore design states that expansion of conjugation generally leads to a red-shift in absorbance and emission wavelengths. The results of this study show that ESIPT fluorophores run contrary to those conventional design principles and this behavior can only be rationalized by considering Baird's rule.

Open-Resonance-Assisted Hydrogen Bonds and Competing Quasiaromaticity.

The delocalization of electron density upon tautomerization of a proton across a conjugated bridge can alter the strength of hydrogen bonds. This effect has been dubbed resonance-assisted hydrogen bonding (RAHB) and plays a major role in the energetics of the tautomeric equilibrium. The goal of this work was to investigate the role that π-delocalization plays in the stability of RAHBs by engaging other isomerization processes. Similarly, acid-base chemistry has received little experimental attention in studies of RAHB, and we address the role that acid-base effects play in the tautomeric equilibrium. We find that π-delocalization and the disruption of adjacent aromatic rings is the dominant effect in determining the stability of a RAHB.

Palmitate and oleate exert differential effects on insulin signalling and glucose uptake in human skeletal muscle cells.

Saturated fatty acids are implicated in the development of insulin resistance, whereas unsaturated fatty acids may have a protective effect on metabolism. We tested in primary human myotubes if insulin resistance induced by saturated fatty acid palmitate can be ameliorated by concomitant exposure to unsaturated fatty acid oleate. Primary human myotubes were pretreated with palmitate, oleate or their combination for 12 h. Glucose uptake was determined by intracellular accumulation of [3H]-2-deoxy-d-glucose, insulin signalling and activation of endoplasmic reticulum (ER) stress by Western blotting, and mitochondrial reactive oxygen species (ROS) production by fluorescent dye MitoSOX. Exposure of primary human myotubes to palmitate impaired insulin-stimulated Akt-Ser473, AS160 and GSK-3ß phosphorylation, induced ER stress signalling target PERK and stress kinase JNK 54 kDa isoform. These effects were virtually abolished by concomitant exposure of palmitate-treated myotubes to oleate. However, an exposure to palmitate, oleate or their combination reduced insulin-stimulated glucose uptake. This was associated with increased mitochondrial ROS production in palmitate-treated myotubes co-incubated with oleate, and was alleviated by antioxidants MitoTempo and Tempol. Thus, metabolic and intracellular signalling events diverge in myotubes treated with palmitate and oleate. Exposure of human myotubes to excess fatty acids increases ROS production and induces insulin resistance.

Characterization of bispecific T-cell Engager (BiTE) antibodies with a high-capacity T-cell dependent cellular cytotoxicity (TDCC) assay.

The Bispecific T-cell Engager (BiTE) antibody modality is a clinically validated immunotherapeutic approach for targeting tumors. Using T-cell dependent cellular cytotoxicity (TDCC) assays, we measure the percentage of specific cytotoxicity induced when a BiTE molecule engages T-cells, redirects T-cell mediated cytolysis, and ultimately kills target cells. We establish a novel luminescence-based TDCC assay quantified by measuring cell viability via constitutive expression of luciferase. The luciferase-based TDCC assay performance is valid and comparable to an adenosine triphosphate (ATP)-based detection method. We demonstrate that the luciferase-based TDCC assay is an efficient homogeneous assay format that is amenable to both suspension and adherent target cells. The luciferase-based TDCC assay eliminates the need for plate-washing protocols, allowing for higher-throughput screening of BiTE antibodies and better data quality. Assay capacity is also improved by performing serial dilutions of BiTE antibodies in 384-well format with an automated liquid handler. We describe here a robust, homogeneous TDCC assay platform with capacity for in vitro assessment of BiTE antibody potency and efficacy using multiple tumor cell lines and T-cell donors.

Limited density of an antigen presented by RMA-S cells requires B7-1/CD28 signaling to enhance T-cell immunity at the effector phase.

The association of B7-1/CD28 between antigen presenting cells (APCs) and T-cells provides a second signal to proliferate and activate T-cell immunity at the induction phase. Many reports indicate that tumor cells transfected with B7-1 induced augmented antitumor immunity at the induction phase by mimicking APC function; however, the function of B7-1 on antitumor immunity at the effector phase is unknown. Here, we report direct evidence of enhanced T-cell antitumor immunity at the effector phase by the B7-1 molecule. Our experiments in vivo and in vitro indicated that reactivity of antigen-specific monoclonal and polyclonal T-cell effectors against a Lass5 epitope presented by RMA-S cells is increased when the cells expressed B7-1. Use of either anti-B7-1 or anti-CD28 antibodies to block the B7-1/CD28 association reduced reactivity of the T effectors against B7-1 positive RMA-S cells. Transfection of Lass5 cDNA into or pulse of Lass5 peptide onto B7-1 positive RMA-S cells overcomes the requirement of the B7-1/CD28 signal for T effector response. To our knowledge, the data offers, for the first time, strong evidence that supports the requirement of B7-1/CD28 secondary signal at the effector phase of antitumor T-cell immunity being dependent on the density of an antigenic peptide.

Studying cancer stem cell dynamics on PDMS surfaces for microfluidics device design.

This systematic study clarified a few interfacial aspects of cancer cell phenotypes on polydimethylsiloxane (PDMS) substrates and indicated that the cell phenotypic equilibrium greatly responds to cell-to-surface interactions. We demonstrated that coatings of fibronectin, bovine serum albumin (BSA), or collagen with or without oxygen-plasma treatments of the PDMS surfaces dramatically impacted the phenotypic equilibrium of breast cancer stem cells, while the variations of the PDMS elastic stiffness had much less such effects. Our results showed that the surface coatings of collagen and fibronectin on PDMS maintained breast cancer cell phenotypes to be nearly identical to the cultures on commercial polystyrene Petri dishes. The surface coating of BSA provided a weak cell-substrate adhesion that stimulated the increase in stem-cell-like subpopulation. Our observations may potentially guide surface modification approaches to obtain specific cell phenotypes.

Microfluidics separation reveals the stem-cell-like deformability of tumor-initiating cells.

Here we report a microfluidics method to enrich physically deformable cells by mechanical manipulation through artificial microbarriers. Driven by hydrodynamic forces, flexible cells or cells with high metastatic propensity change shape to pass through the microbarriers and exit the separation device, whereas stiff cells remain trapped. We demonstrate the separation of (i) a mixture of two breast cancer cell types (MDA-MB-436 and MCF-7) with distinct deformabilities and metastatic potentials, and (ii) a heterogeneous breast cancer cell line (SUM149), into enriched flexible and stiff subpopulations. We show that the flexible phenotype is associated with overexpression of multiple genes involved in cancer cell motility and metastasis, and greater mammosphere formation efficiency. Our observations support the relationship between tumor-initiating capacity and cell deformability, and demonstrate that tumor-initiating cells are less differentiated in terms of cell biomechanics.