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Abstract The erythrocyte sedimentation rate (ESR) is a non-specific blood test for determining inflammatory conditions. However, the long measurement time (60 min) to obtain ESR is an obstacle for a prompt evaluation. In this study, to reduce the measurement time of ESR, deep neural networks (DNNs) were applied to the sedimentation tendency of blood samples. DNNs using multilayer perceptron (MLP), long short-term memory (LSTM), and gated recurrent unit (GRU) were assessed and compared to determine a suitable length of time for the input sequence. To avoid overfitting, a stacking ensemble learning was adopted, which combines multiple models by using a meta model. Four meta models were compared: mean, median, least absolute shrinkage and selection operator, and partial least squares regression (PLSR) schemes. From the empirical results, LSTM and GRU models have better prediction than MLP over sequence lengths of 5 to 20 min. The decrease in $$\overline{\mathrm{MAPE} }$$ MAPE ¯ and $$\overline{\mathrm{RMSE} }$$ RMSE ¯ of GRU and LSTM was attenuated after a sequence length of 15 min, so the input sequence length is determined as 15 min. In terms of the meta model, the statistical comparison suggests that GRU combined with PLSR (GRU–PLSR) is the best case. Then, the GRU–PLSR was tested for prediction of ESR data obtained from periodontitis patients to check its applicability to a specific disease. The Bland–Altman plot shows acceptable agreement between measured and predicted ESR values. Based on the results, the GRU–PLSR can predict ESR with improved performance within 15 min and has potential applicability to ESR data with inflammatory and non-inflammatory conditions.

Abstract The formation of droplets or bubbles in a microfluidic system is a significant topic requiring device miniaturization and a small volume of samples. Especially, a two-phase segmented flow can be applied to micro-mixing for chemical reactions and the treatment of heat and mass transfer. In this study, a flow of liquid slugs and bubbles was generated in a 3D-printed chip and controlled by a single pump creating a vacuum at the outlet. The pump and chip device were integrated to form a simple and portable system. The size and flow rate of liquid slugs, obtained through image processing techniques, were analyzed considering several parameters related to hydraulic resistance and pressure drop. In addition, the effect of segmentation on mixing was observed by measuring the intensity change using two different colored inks. The hydraulic resistance of air and liquid flows can be controlled by changing the tube length of air flow and the viscosity of liquid flow. Because the total pressure drop along the channel was produced using a single pump at the outlet of the channel, the size and flow rate of the liquid slugs showed a near linear relation depending on the hydraulic resistances. In contrast, as the total pressure varied with the flow rate of the pump, the size of the liquid slugs showed a nonlinear trend. This indicates that the frequency of the liquid slug formation induced by the squeezed bubble may be affected by several forces during the development of the liquid slugs and bubbles. In addition, each volume of liquid slug segmented by the air is within the range of 10–1 to 2 µL for this microfluidic system. The segmentation contributes to mixing efficiency based on the increased homogeneity factor of liquid. This study provides a new insight to better understand the liquid slug or droplet formation and predict the segmented flow based on the relationship between the resistance, flow rate, and pressure drop.

BackgroundRacial and ethnic minority groups are underrepresented in health research, contributing to persistent health disparities in the United States. Identifying effective recruitment and retention strategies among minority groups and their subpopulations is an important research agenda. Web-based intervention approaches are becoming increasingly popular with the ubiquitous use of the internet. However, it is not completely clear which recruitment and retention strategies have been successful in web-based intervention trials targeting racial and ethnic minorities. ObjectiveThis study aims to describe lessons learned in recruiting and retaining one of the understudied ethnic minority women—Korean Americans—enrolled in a web-based intervention trial and to compare our findings with the strategies reported in relevant published web-based intervention trials. MethodsMultiple sources of data were used to address the objectives of this study, including the study team’s meeting minutes, participant tracking and contact logs, survey reports, and postintervention interviews. In addition, an electronic search involving 2 databases (PubMed and CINAHL) was performed to identify published studies using web-based interventions. Qualitative analysis was then performed to identify common themes addressing recruitment and retention strategies across the trials using web-based intervention modalities. ResultsA total of 9 categories of recruitment and retention strategies emerged: authentic care; accommodation of time, place, and transportation; financial incentives; diversity among the study team; multiple, yet standardized modes of communication; mobilizing existing community relationships with efforts to build trust; prioritizing features of web-based intervention; combined use of web-based and direct recruitment; and self-directed web-based intervention with human support. Although all the studies included in the analysis combined multiple strategies, prioritizing features of web-based intervention or use of human support were particularly relevant for promoting recruitment and retention of racial and ethnic minorities in web-based intervention trials. ConclusionsThe growing prevalence of internet use among racial and ethnic minority populations represents an excellent opportunity to design and deliver intervention programs via the internet. Future research should explore and compare successful recruitment and retention methods among race and ethnic groups for web-based interventions. Trial RegistrationClinicalTrials.gov NCT03726619; https://clinicaltrials.gov/ct2/show/NCT03726619.

Abstract Background Despite the growing demand for antimicrobial peptides (AMPs) for clinical use as an alternative approach against antibiotic-resistant bacteria, the manufacture of AMPs relies on expensive, small-scale chemical methods. The small ubiquitin-related modifier (SUMO) tag is industrially practical for increasing the yield of recombinant proteins by increasing solubility and preventing degradation in expression systems. Results A new vector system, pKSEC1, was designed to produce AMPs, which can work in prokaryotic systems such as Escherichia coli and plant chloroplasts. 6xHis was tagged to SUMO for purification of SUMO-fused AMPs. Abaecin, a 34-aa-long antimicrobial peptide from honeybees, was expressed in a fusion form to 6xHis-SUMO in a new vector system to evaluate the prokaryotic expression platform of the antimicrobial peptides. The fusion sequences were codon-optimized in three different combinations and expressed in E. coli. The combination of the native SUMO sequence with codon-optimized abaecin showed the highest expression level among the three combinations, and most of the expressed fusion proteins were detected in soluble fractions. Cleavage of the SUMO tag by sumoase produced a 29-aa-long abaecin derivative with a C-terminal deletion. However, this abaecin derivative still retained the binding sequence for its target protein, DnaK. Antibacterial activity of the 29-aa long abaecin was tested against Bacillus subtilis alone or in combination with cecropin B. The combined treatment of the abaecin derivative and cecropin B showed bacteriolytic activity 2 to 3 times greater than that of abaecin alone. Conclusions Using a SUMO-tag with an appropriate codon-optimization strategy could be an approach for the production of antimicrobial peptides in E.coli without affecting the viability of the host cell.

In studying blood flow in the vessels, the characteristics of non-Newtonian fluid are important, considering the role of viscosity in rheology. Stenosis, which is an abnormal narrowing of the vessel, has an influence on flow behavior. Therefore, analysis of blood flow in stenosed vessels is essential. However, most of them exist as simulation outcomes. In this study, non-Newtonian fluid was observed in stenosed microchannels under the pulsatile flow condition. A polydimethylsiloxane channel with 60% stenosis was fabricated by combining an optic fiber and a petri dish, resembling a mold. Three types of samples were prepared by changing the concentrations of xanthan gum, which induces a shear thinning effect (phosphate buffered saline (PBS) solution as the Newtonian fluid and two non-Newtonian fluids mimicking normal blood and highly viscous blood analog). The viscosity of the samples was measured using a Y-shaped microfluidic viscometer. Thereafter, velocity profiles were analyzed under the pulsatile flow condition using the micro-particle image velocimetry (PIV) method. For the Newtonian fluid, the streamline was skewed more to the wall of the channel. The velocity profile of the non-Newtonian fluid was generally blunter than that of the Newtonian fluid. A highly oscillating wall shear stress (WSS) during the pulsatile phase may be attributed to such a bluntness of flow under the same wall shear rate condition with the Newtonian fluid. In addition, a highly viscous flow contributes to the variation in the WSS after passing through the stenosed structures. A similar tendency was observed in simulation results. Such a variation in the WSS was associated with plaque instability or rupture and damage of the tissue layer. These results, related to the influence on the damage to the endothelium or stenotic lesion, may help clinicians understand relevant mechanisms.

Bimaxillary orthognathic surgery is widely used to treat skeletal class III malocclusion. Changes in jaw position affect the shape of surrounding soft tissues. We used computational fluid dynamics (CFD) simulation to observe changes in airways observed in a patient who underwent bimaxillary orthognathic surgery. For CFD simulation, we performed cone beam computed tomography (CBCT) preoperatively (T0), 3 days postoperatively (T1), and 7 months postoperatively (T2). The values of velocity, pressure drop (ΔP), and wall shear stress all increased 7 months after surgery (Vmax 7.038 m/s to 12.054 m/s, ΔP −7.723 Pa to −53.739 Pa, WSSmax 4.214 Pa to 14.323 Pa). Locations where the velocity and pressure gradients are large included the velopharynx, oropharynx, and epiglottis, with narrow cross-sectional areas. Wall shear stress was also observed at these locations. The velopharynx, oropharynx, and epiglottis are structures most vulnerable to morphological changes, that is, they can easily become obstructed.

In order to realize high-performance thermoelectric materials, a way to obtain small grain size is necessary for intensification of the phonon scattering. Here, we use a melt-spinning-spark plasma sintering process for making p-type Bi0.36Sb1.64Te3 thermoelectric materials and evaluate the relation between the process conditions and thermoelectric performance. We vary the Cu wheel rotation speed from 1000 rpm (~13 ms−1) to 4000 rpm (~52 ms−1) during the melt spinning process to change the cooling rate, allowing us to control the characteristic size of nanostructure in melt-spun Bi0.36Sb1.64Te3 ribbons. The higher wheel rotation speed decreases the size of nanostructure, but the grain sizes of sintered pellets are inversely proportional to the nanostructure size after the same sintering condition. As a result, the ZT values of the bulks fabricated from 1000–3000 rpm melt-spun ribbons are comparable each other, while the ZT value of the bulk from the 4000 rpm melt-spun ribbons is rather lower due to reduction of grain boundary phonon scattering. In this work, we can conclude that the smaller nanostructure in the melt spinning process does not always guarantee high-performance thermoelectric bulks, and an adequate following sintering process must be included.

BACKGROUND: Pandemic influenza poses a serious threat to global health and the world economy. While vaccines are currently under development, passive immunization could offer an alternative strategy to prevent and treat influenza virus infection. Attempts to develop monoclonal antibodies (mAbs) have been made. However, passive immunization based on mAbs may require a cocktail of mAbs with broader specificity in order to provide full protection since mAbs are generally specific for single epitopes. Chicken immunoglobulins (IgY) found in egg yolk have been used mainly for treatment of infectious diseases of the gastrointestinal tract. Because the recent epidemic of highly pathogenic avian influenza virus (HPAIV) strain H5N1 has resulted in serious economic losses to the poultry industry, many countries including Vietnam have introduced mass vaccination of poultry with H5N1 virus vaccines. We reasoned that IgY from consumable eggs available in supermarkets in Vietnam could provide protection against infections with HPAIV H5N1. METHODS AND FINDINGS: We found that H5N1-specific IgY that are prepared from eggs available in supermarkets in Vietnam by a rapid and simple water dilution method cross-protect against infections with HPAIV H5N1 and related H5N2 strains in mice. When administered intranasally before or after lethal infection, the IgY prevent the infection or significantly reduce viral replication resulting in complete recovery from the disease, respectively. We further generated H1N1 virus-specific IgY by immunization of hens with inactivated H1N1 A/PR/8/34 as a model virus for the current pandemic H1N1/09 and found that such H1N1-specific IgY protect mice from lethal influenza virus infection. CONCLUSIONS: The findings suggest that readily available H5N1-specific IgY offer an enormous source of valuable biological material to combat a potential H5N1 pandemic. In addition, our study provides a proof-of-concept for the approach using virus-specific IgY as affordable, safe, and effective alternative for the control of influenza outbreaks, including the current H1N1 pandemic.

Enterovirus infections induce infectious diseases in young children, such as hand, foot, and mouth disease which is characterized by highly contagious rashes or blisters around the hands, feet, buttocks, and mouth. This predominantly arises from enterovirus A71 or coxsackievirus A16 infections and in severe cases, they can lead to encephalitis, paralysis, pulmonary edema, or even fatality, representing a global health threat. Due to the absence of effective therapeutic strategies for these infections, various experimental animal models are being investigated for the development of vaccines. During the early stages of research on enterovirus infections, non-human primate infections exhibited symptoms like those in humans, leading to their utilization as model animals. However, due to economic and ethical considerations, their current usage is limited. While enterovirus infections do not readily occur in mice, an infection model with mouse-adapted strain in neonatal mice has been employed. Cellular receptors have been identified in human cells, and genetically modified mice expressing these receptors have been used. Most recently, the utilization of Mongolian gerbil model is actively being considered and should be pursued for further animal model development. So, herein, we provide a summarized overview of the current portfolio of available enterovirus infection models, emphasizing their respective advantages and limitations. [ABSTRACT FROM AUTHOR]

Periodontitis and diabetes mellitus (DM) have a bidirectional relationship. Periodontitis is initiated by dysbiosis of oral microorganisms, and in particular, the characteristics of the microorganisms that have penetrated the tissue are directly related to the disease; therefore, we investigated the effect of DM on intragingival microbial profiling of patients with periodontitis. A total of 39 subjects were recruited and divided into three groups in this case control study as follows: healthy (NA, 10), periodontitis only (PD, 18), and periodontitis with DM (PD_DM, 11). Gingival tissue was collected, DNA was extracted, and whole-genome sequencing was performed. PD and PD_DM showed different characteristics from NA in diversity and composition of the microbial community; however, no difference was found between the PD nad PD_DM. PD_DM showed discriminatory characteristics for PD in the network analysis. PD showed a network structure in which six species were connected, including three red complex species, and PD_DM's network was more closely connected and expanded, with six additional species added to the PD network. Although DM did not significantly affect a- and b-diversity or abundance of phyla and genera of microbiota that invaded the gingival tissue of patients with periodontitis, DM will affect the progression of periodontitis by strengthening the bacterial network in the gingival tissue. [ABSTRACT FROM AUTHOR]

Since the widespread introduction of oral and live attenuated rotavirus vaccines around the world in 2009, the impacts of disease burden and the effects of disease reduction in developing countries have been proven. However, in low and middle-income countries, the vaccine efficacy is somewhat lower than in developed countries due to differences in nutritional conditions, microbial environments of individuals, and other factors. In addition, as oral, live vaccines have been found to be associated with rare but serious side effects, the development of a next-generation vaccine with safety, improved effectiveness, and ease of storage is currently underway. New vaccine strain developed by the Centers for Disease Control and Prevention in the United States are undergoing preclinical testing of efficacy, antigen dose, and administration route in the form of a heat-treated inactive vaccine, and a recombinant protein-based trivalent subunit vaccine developed by the Program for Appropriate Technology in Health is undergoing clinical trial in phase III. Several research groups are also developing non-replicating protein-based rotavirus vaccines using virus-like particles and nanoparticles. This review provides a brief overview of the development status and technology of parenteral, non-live rotavirus vaccines worldwide. [ABSTRACT FROM AUTHOR]

Purpose: Virus-like particles (VLPs) are being developed as a promising vaccine platform and therapeutic delivery. Various strategies for effectively constructing VLPs have been studied, but relatively few studies have been done on various factors affecting storage. In this study, we investigated the antigenic changes of VLPs in an acidic or basic pH environment using influenza VLPs as an experimental model. Materials and Methods: Influenza VLPs containing hemagglutination and M1 proteins were generated and their antigenicity and protective immunity in vitro and in vivo were evaluated after exposure to acidic (pH 4 and 5) or basic (pH 9 and 10) pH buffers. Results: VLP exposed to basic pH showed similar levels of antigenicity to those stored in neutral pH, while antigenicity of VLP exposed to acidic pH was found to be significantly reduced compared to those expose neutral or basic pH. All groups of mice responded effectively to low concentrations of virus infections; however, VLP vaccine groups exposed to acid pH were found not to induce sufficient protective immune responses when a high concentration of influenza virus infection. Conclusion: In order for VLP to be used as a more powerful vaccine platform, it should be developed in a strategic way to respond well to external changes such as acidic pH conditions. [ABSTRACT FROM AUTHOR]

Purpose: In this study, we investigated whether the antigenic changes of the virus-like particles (VLPs) are affected by the temperature during storage. Materials and Methods: After exposing the recombinant influenza VLPs to various temperatures for a period, antigenic changes were examined through in vitro hemagglutination receptor binding assay and in vivo mouse experiments. Results: Influenza VLPs were exposed at three different temperatures of low, middle, and high on a thermo-hygrostat. High temperature exposed influenza VLPs were showed significantly reduced HA activity and immunogenicity after mouse single immunization over time compared low and middle. When the VLPs exposed to the high temperature were inoculated once in the mice, it was found that the immunogenicity was significantly reduced compared to the VLPs exposed to the low temperature. However, these differences were almost neglected when mice were inoculated twice even with VLPs exposed to high temperatures. Conclusion: This study suggests that similar protective effects can be expected by controlling the number of vaccination and storage conditions, although the antigenic change in the VLP vaccines occurred when exposed to high temperature. [ABSTRACT FROM AUTHOR]

Polyphenols including catechins from green tea (Camellia sinensis) have been reported to have anti-infective activities against a broad spectrum of viruses and other pathogens. During the last two decades, antiviral activities of catechins with different modes of action have been demonstrated on diverse families of viruses, such as human immunodeficiency virus, Herpes simplex virus, influenza virus, hepatitis B and C virus. In this study, we focused on the antiviral properties of catechins and their derivatives against viral hepatitis which have become a key public health issue due to their serious impact on human health with liver diseases. [ABSTRACT FROM AUTHOR]

Influenza virus M2 extracellular domain (M2e) has been a target for developing cross-protective vaccines. However, the efficacy and immune correlates of M2e vaccination are poorly understood in the different host genetic backgrounds in comparison with influenza vaccines. We previously reported the cross-protective efficacy of virus-like particle (M2e5x VLP) vaccines containing heterologous tandem M2e repeats (M2e5x) derived from human, swine, and avian influenza viruses. In this study to gain better understanding of cross-protective influenza vaccines, we compared immunogenicity and efficacy of M2e5x VLP, H5 hemagglutinin VLP (HA VLP), and inactivated H3N2 virus (H3N2i) in wild-type strains of BALB/c and C57BL/6 mice, and CD4 and CD8 knockout (KO) mice. M2e5x VLP was superior to HA VLP in conferring cross-protection whereas H3N2i inactivated virus vaccine provided high efficacy of homologous protection. After M2e5x VLP vaccination and challenge, BALB/c mice induced higher IgG responses, lower lung viral loads, and less body weight loss when compared with those in C57BL/6 mice. M2e5x VLP but not H3N2i immune mice after primary challenges developed strong immunity against a secondary heterosubtypic virus as a model of future pandemics. M2e5x VLP and HA VLP vaccines were able to raise IgG isotypes in CD4 KO mice. T cells were found to contribute to cross-protection by playing a role in reducing lung viral loads. In conclusion, M2e5x VLP vaccination induced better cross-protection than HA VLP, and its efficacy varied depending on the genetic backgrounds of mice, supporting the important roles of T cells. [ABSTRACT FROM AUTHOR]

Purpose: Nitrocellulose membrane–based filtration system (NCFS) is widely used for protein concentration. In this study, we applied NCFS for production of virus-like particle (VLP) as a vaccine candidate and evaluated yield property and immunogenicity. Materials and Methods: Influenza VLPs were generated by baculovirus-insect cell protein expression system. NCFS and sucrose gradient ultracentrifugation were used for purification of VLP. Immunogenicity of VLP was evaluated by animal experiment. Results: Influenza VLPs expressing hemagglutinin (HA) and neuraminidase proteins derived from highly pathogenic influenza virus (H5N8) were effectively produced and purified by NCFS. HA activity of VLP which correlated with antigenicity was well conserved during multiple purification steps. This NCFS based purified VLPs induced influenza virus–specific antibody responses. Conclusion: Our results indicate that the influenza VLP vaccine could be prepared by NCFS without loss of immunogenicity and elicit antigen-specific immune responses. [ABSTRACT FROM AUTHOR]

Since the start of the industrial revolution, human activities have caused a rapid increase in atmospheric CO2 concentrations, which have in turn been cited as the cause of a variety climate changes such as global warming and ocean acidification. Various approaches have been proposed to reduce atmospheric CO2 concentrations. The ''Martin (or Iron) Hypothesis'' suggests that ocean iron fertilization (OIF) should be an efficient method for stimulating the biological pump in iron-limited high nutrient-low chlorophyll regions. To test the Martin hypothesis, a total 13 OIF experiments have been performed since 1990 in the Southern Ocean (7 times), in the subarctic Pacific (3 times), in the equatorial Pacific (twice), and in the subtropical Atlantic (once). These OIF field experiments demonstrated that primary production could be significantly increased after artificial iron addition. However, export production efficiency revealed by the OIF experiments was unexpectedly low compared to production from natural processes in all, except one of the experiments (i.e., the Southern Ocean European Iron Fertilization Experiment, EIFEX). These results, including side effects such as N2O production and hypoxia development, have been scientifically debated amongst those who support and oppose OIF experimentation. In the context of increasing global and political concerns associated with climate change, it is valuable to examine the validity and usefulness of the OIF. We provide a general overview of the OIF experiments conducted over the last 25 years (past), a discussion of OIF considerations including possible side effects (present), and an introduction to the OIF experiment plan currently being designed by Korean oceanographers (future). [ABSTRACT FROM AUTHOR]

The article discusses a study that examined the use of cone-beam computerized tomography (CBCT) for the morphologic analysis of the edentulous posterior mandible to determine the safety and utility of mandibular body bone (MBB) graft as an autogenous bone graft material. It describes the use of a three-dimensional (3D) imaging program for the analysis, as well as the results of buccal cortical bone thickness measurements. It also suggests a method for improving the surgical procedure's safety.

Background: This cross-sectional study sought to identify factors associated with antenatal depression in pregnant Korean females, including sociodemographic parameters, social support, social conflict, and bipolarity. Methods: Eighty-four pregnant women were recruited to complete questionnaires on sociodemographic factors, obstetric history, depressive symptoms, and bipolarity. Depressive symptoms were assessed using the Korean version of the Edinburgh Postnatal Depression Scale. Bipolarity was assessed using the Korean version of the Mood Disorder Questionnaire. Results: Nineteen participants (22.6%) had positive Mood Disorder Questionnaire scores, suggesting the presence of bipolarity, and were significantly more likely to score high on the Edinburgh Postnatal Depression Scale. Antenatal depression was associated with bad marital communication and marital dissatisfaction. Conclusion: These results suggest that spousal interactions play a significant role in antenatal depression, and pregnant women with bipolarity may be more depressed than those without bipolarity. [ABSTRACT FROM AUTHOR]

Purpose: Neuraminidase (NA) of influenza virus contains stalk region that shows a great deal of variability in both amino acid sequence and length. In this paper, we investigated generation of recombinant influenza viruses that had hepatitis B virus (HBV) B cell epitopes in the NA stalk region as a dual vaccine candidate. Materials and Methods: We used the WSH-HK reassortant helper virus for rescue of recombinant influenza virus containing HBV epitopes and reverse genetic protocol based on the use of micrococcal nuclease-treated virus cores for reconstitution of ribonucleoproteins. Results: We successfully generated a chimeric influenza viruses which contained 22 amino acid peptides in the stalk region derived from the surface and pre-surface protein HBV. The growth kinetics of the recombinant viruses was investigated after infection of Madin-Darby canine kidney (MDCK) and Madin-Darby bovine kidney (MDBK) cells and the rIV-BVPreS virus showed higher titer than other viruses in MDCK cells. We also confirmed the presence of HBV epitopes in the chimeric viruses by enzyme-linked immunosorbent assay (ELISA) using anti-HBV polyclonal antibody. When the ratio of recombinant virus verse wild type virus was calculated by ELISA, recombinant viruses exhibited 2 fold higher values than the wild type virus. Conclusion: These results suggest that chimeric influenza virus which contained foreign anti-gens can be used as dual vaccine against both HBV and influenza viruses. [ABSTRACT FROM AUTHOR]

Ginseng polysaccharide has been known to have multiple immunomodulatory effects. In this study, we investigated whether Panax ginseng polysaccharide (GP) would have a preventive effect on influenza infection. Administration of mice with GP prior to infection was found to confer a survival benefit against infection with H1N1 (A/PR/8/34) and H3N2 (A/ Philippines/82) influenza viruses. Mice infected with the 2009 H1N1 virus suspended in GP solution showed moderately enhanced survival rates and lower levels of lung viral titers and the inflammatory cytokine (IL-6). Daily treatment of vaccinated mice with GP improved their survival against heterosubtypic lethal challenge. This study demonstrates the first evidence that GP can be used as a remedy against influenza viral infection. [ABSTRACT FROM AUTHOR]

Background: Current influenza vaccines based on the hemagglutinin protein are strain specific and do not provide good protection against drifted viruses or emergence of new pandemic strains. An influenza vaccine that can confer cross-protection against antigenically different influenza A strains is highly desirable for improving public health. Methodology/Principal Findings: To develop a cross protective vaccine, we generated influenza virus-like particles containing the highly conserved M2 protein in a membrane-anchored form (M2 VLPs), and investigated their immunogenicity and breadth of cross protection. Immunization of mice with M2 VLPs induced anti-M2 antibodies binding to virions of various strains, M2 specific T cell responses, and conferred long-lasting cross protection against heterologous and heterosubtypic influenza viruses. M2 immune sera were found to play an important role in providing cross protection against heterosubtypic virus and an antigenically distinct 2009 pandemic H1N1 virus, and depletion of dendritic and macrophage cells abolished this cross protection, providing new insight into cross-protective immune mechanisms. Conclusions/Significance: These results suggest that presenting M2 on VLPs in a membrane-anchored form is a promising approach for developing broadly cross protective influenza vaccines. [ABSTRACT FROM AUTHOR]

There is a significant gap in our fundamental understanding of early morphological and migratory changes in human Langerhans cells (LCs) in response to vaccine stimulation. As the vast majority of LCs studies are conducted in small animal models, substantial interspecies variation in skin architecture and immunity must be considered when extrapolating the results to humans. This study aims to determine whether excised human skin, maintained viable in organ culture, provides a useful human model for measuring and understanding early immune response to intradermally delivered vaccine candidates. Excised human breast skin was maintained viable in air-liquid-interface organ culture. This model was used for the first time to show morphological changes in human LCs stimulated with influenza virus-like particle (VLP) vaccines delivered via intradermal injection. Immunohistochemistry of epidermal sheets and skin sections showed that LCs in VLP treated skin lost their typical dendritic morphology. The cells were more dispersed throughout the epidermis, often in close proximity to the basement membrane, and appeared vertically elongated. Our data provides for increased understanding of the complex morphological, spatial and temporal changes that occur to permit LC migration through the densely packed keratinocytes of the epidermis following exposure to vaccine. Significantly, the data not only supports previous animal data but also provides new and essential evidence of host response to this vaccination strategy in the real human skin environment. [ABSTRACT FROM AUTHOR]

Background: Recurrent outbreaks of highly pathogenic H5N1 avian influenza virus pose a threat of eventually causing a pandemic. Early vaccination of the population would be the single most effective measure for the control of an emerging influenza pandemic. Methodology/Principal Findings: Influenza virus-like particles (VLPs) produced in insect cell-culture substrates do not depend on the availability of fertile eggs for vaccine manufacturing. We produced VLPs containing influenza A/Viet Nam1203/04 (H5N1) hemagglutinin, neuraminidase, and matrix proteins, and investigated their preclinical immunogenicity and protective efficacy. Mice immunized intranasally with H5N1 VLPs developed high levels of H5N1 specific antibodies and were 100% protected against a high dose of homologous H5N1 virus infection at 30 weeks after immunization. Protection is likely to be correlated with humoral and cellular immunologic memory at systemic and mucosal sites as evidenced by rapid anamnestic responses to re-stimulation with viral antigen in vivo and in vitro. Conclusions/Significance: These results provide support for clinical evaluation of H5N1 VLP vaccination as a public health intervention to mitigate a possible pandemic of H5N1 influenza. [ABSTRACT FROM AUTHOR]

Besides well-known health benefits, green tea catechins exert antimicrobial and antiviral activities against a variety of infectious agents. Although the detailed mechanism of the antimicrobial activity of tea catechins remains to be explored, the broad-spectrum activity of catechins may involve common target(s), such as the cell membrane, in addition to specific targets for each pathogen. This extends to antiviral activities, where many pronounced activities were reported for enveloped viruses. Yet, the effectiveness of tea catechins as antimicrobials is compromised by relative chemical instability and poor bioavailability. Whether tea catechins will emerge as a viable option as alternative medicine or as a synergistic combination therapy with pre-existing antivirals or antibiotics must therefore depend on a method of delivery that ensures its stability and bioavailability. However, green tea may provide an option for mitigating the health and economic burdens associated with emerging and re-emerging infectious diseases, especially considering the paucity of effective control measures. Considering the zoonotic nature of newly arising infectious diseases, the dual use of green tea components in both humans and livestock may reduce animal-human transmission, which would complement the current management of infectious diseases. [ABSTRACT FROM AUTHOR]

Abstract Type 2 diabetes mellitus (T2DM) and periodontitis (PD) have intricated connections as chronic inflammatory diseases. While the immune response is a key factor that accounts for their association, the underlying mechanisms remain unclear. To gain a deeper understanding of the connection, we conducted research using a multiomics approach. We generated whole genome and methylation profiling array data from the periodontium of PD patients with DM (PDDM) and without DM to confirm genetic and epigenetic changes. Independent bulk and single‐cell RNA sequencing data were employed to verify the expression levels of hypo‐methylated genes. We observed a gradual rise in C>T base substitutions and hypomethylation in PD and PDDM patients compared with healthy participants. Furthermore, specific genetic and epigenetic alterations were prominently associated with the Fc‐gamma receptor‐mediated phagocytosis pathway. The upregulation of these genes was confirmed in both the periodontal tissues of PD patients and the pancreatic tissues of T2DM patients. Through single‐cell RNA analysis of peripheral blood mononuclear cells, substantial upregulation of Fc‐gamma receptors and related genes was particularly identified in monocytes. Our findings suggest that targeting the Fc‐gamma signaling pathway in monocytes holds promise as a potential treatment strategy for managing systemic complications associated with diabetes.

Visual evaluation of the surface anchoring energies in a nematic liquid crystal (LC) cell is characterized by the direction of the convection roll pattern that appears in the low-frequency conduction regime. The convection roll pattern in a twisted nematic LC (TNLC) cell is oriented perpendicular to the midplane LC director dominating the direction of convection flow, and its direction is determined by the relative surface anchoring energy between two surface boundaries. Thus the direction of the roll pattern generated at the TNLC cell with asymmetric LC alignment layers can provide information on the surface anchoring energies at the two boundaries. We demonstrate a method for determining the two anchoring energies through a measured midplane LC director applied to the Ericksen-Leslie equation. [ABSTRACT FROM AUTHOR]

Abstract Background Type 2 diabetes mellitus (DM) is a complex metabolic disorder that causes various complications, including periodontitis (PD). Although a bidirectional relationship has been reported between DM and PD, their immunological relationship remains poorly understood. Therefore, this study aimed to compare the immune response in patients with PD alone and in those with both PD and DM (PDDM) to expand our knowledge of the complicated connection between PD and DM. Methods Peripheral blood mononuclear cells were collected from 11 healthy controls, 10 patients with PD without DM, and six patients with PDDM, followed by analysis using single‐cell RNA sequencing. The differences among groups were then compared based on intracellular and intercellular perspectives. Results Compared to the healthy state, classical monocytes exhibited the highest degree of transcriptional change, with elevated levels of pro‐inflammatory cytokines in both PD and PDDM. DM diminished the effector function of CD8+ T and natural killer (NK) cells as well as completely modified the differentiation direction of these cells. Interestingly, a prominent pathway, RESISTIN, which is known to increase insulin resistance and susceptibility to diabetes, was found to be activated under both PD and PDDM conditions. In particular, CAP1+ classical monocytes from patients with PD and PDDM showed elevated nuclear factor kappa B‐inducing kinase activity. Conclusions Overall, this study elucidates how the presence of DM contributes to the deterioration of T/NK cell immunity and the immunological basis connecting PD to DM.

Abstract Background The purpose of this study was to measure the time of the conventional surgical planning (CSP) and virtual surgical planning (VSP) in orthognathic surgery and to compare them in terms of cost. Material and method This is a retrospective study of the patients who underwent orthognathic surgery at the OOOOO University Dental Hospital from December 2017 to August 2018. All the patients were analyzed through both CSP and VSP, and all the surgical stents were fabricated through manual and 3-dimensional (3D) printing. The predictor variables were the planning method (CSP vs. VSP) and the surgery type (group I: Le Fort I osteotomy+bilateral sagittal split osteotomy [LFI+BSSO] or group II: only bilateral sagittal split osteotomy [BSSO]), and the outcomes were the time and cost. The results were analyzed using paired t test. Results Thirty patients (12 females, 18 males) met the inclusion criteria, and 17 patients were excluded from the study due to missing or incomplete data. There were 20 group I patients (LFI+BSSO regardless of genioplasty) and 10 group II patients (BSSO regardless of genioplasty). The average time of CSP for group I was 385±7.8 min, and that for group II was 195±8.33 min. The time reduction rate of VSP compared with CSP was 62.8% in group I and 41.5% in group II. On the other hand, there was no statistically significant cost reduction. Conclusions The time investment in VSP in this study was significantly smaller than that in CSP, and the difference was greater in group I than in group II.

Abstract Background Stability of the grafted bone volume is one of the important factors to the success of alveolar bone grafts. For this, platelet-rich plasma (PRP) or fibrin sealant is mixed with the bone graft material. Bio-Oss® is a protein-free bovine mineral commonly used in bone graft procedures. The grafting particles are commonly combined with a standard fibrin sealant (Tisseel®) to fabricate a plastic implantable product. The purpose of this experiment was to evaluate the efficacy of fibrin sealant (Tisseel®) in bone regeneration performance in a rabbit maxillary sinus model. Methods A total of five 3.5 kg weight New Zealand white rabbits were used for the study. After elevating the sinus membrane in both maxillary sinus cavities, Bio-Oss® mixed with normal saline (group 1) was filled into the right side, and Tisseel® mixed Bio-Oss® (group 2) was inserted into the other side. The bone mineral density and bone volume were analyzed with microscopic computed tomography (micro-CT) and histomorphometric 12 weeks after application. Results Histologically, new bone formation rate was 14.8%, and grafted bone rate was 70.5% in group 1. In group 2, they were 18.5% and 60.4%, respectively. According to micro-CT analysis, bone mineral density (mg/cm3, BMD) was 2.5% larger in group 1. Conclusions The findings from this study suggest that, although the difference in the bone formation between group 1 and group 2 appears to be insignificant, group 2 had an advantage in using smaller amount of bone substances to achieve the reliable bone formation.

The format of influenza virus-like particles (VLPs) as a nonreplicating particulate vaccine candidate is a promising alternative to conventional egg-based vaccines. In this study, we have investigated the detailed kinetics of immune responses and protective efficacy after a single intranasal immunization with different doses of VLPs alone or in the presence of an Escherichia coli mutant heat-labile enterotoxin [mLT(R192G)] or cholera toxin subunit B as adjuvants. Analysis of immune responses showed differential kinetics in a VLP antigen dose-dependent manner and dynamic changes in the ratios of antibody immunoglobulin G isotypes over the time course. Protection against lethal challenge was observed with a single immunization with influenza VLPs even without adjuvant. The addition of adjuvant showed significant antigen-sparing effects with improved protective efficacy. The protective immune responses, efficacies of protection, and antigen-sparing effects were significantly improved by a second immunization as determined by the levels of neutralizing antibodies, morbidity postchallenge, lung viral titers, and inflammatory cytokines. Our results are informative for a better understanding of the protective immunity induced by a single dose or two doses of influenza VLPs, which is dependent on antigen dosage and the presence of adjuvant, and will provide insights into designing effective vaccines based on VLPs. [ABSTRACT FROM AUTHOR]

Abstract Background The purpose of this study was to review the clinical features of oromaxillofacial infections in patients presenting to a hospital emergency ward, to identify the key factors affecting the requirement for hospitalization, and the potential risk factors predisposing to a prolonged length of hospital stay. Methods A retrospective medical record review of the 598 patients treated for oromaxillofacial infection from 2013 to 2017 at the oral and maxillofacial surgery department, Yangsan Pusan National University Hospital, was conducted. The following information was collected from each patient: sex, age, past medical history, site of infection, etiology, admission or outpatient care, level of C-reactive protein (mg/dL), fascial spaces involved, treatment method, and duration of hospitalization. Chi-squared tests were used to identify risk factors, which were further analyzed using multivariable logistic regression. Results A total of 606 patients were eligible for inclusion in the study, of which eight were excluded due to having incomplete charts; thus, 598 patients were included: 55% were male, mean patient age was 47.1 ± 19.9 years, and 12.9% of patients were diabetic. Furthermore, 71.2% of patients had infection originating in the mandible; the most common tooth of origin was lower posterior, and 29.8% of patients were hospitalized. Risk factors for hospital admission were elderly patients with concurrent disease, elevated C-reactive protein level, and multiple-space infection in the oromaxillofacial area. The duration of hospitalization was correlated with both diabetes and age. Conclusions The requirement for hospital admission is determined by the severity of the infection; even severe infections, once treated with appropriate surgery, have no relation to the length of hospital stay. The important risk factors for increased duration of hospitalization are diabetes mellitus and older age. The understanding of risk factors associated with a prolonged hospital stay during the treatment of oromaxillofacial infection will aid in treatment planning as well as highlight the importance of adequate diabetes control in patients at risk of such infection.

Abstract Background The purpose of this study was to measure the time of the conventional surgical planning (CSP) and virtual surgical planning (VSP) in orthognathic surgery and to compare them in terms of cost. Material and method This is a retrospective study of the patients who underwent orthognathic surgery at the Pusan National University Dental Hospital from December 2017 to August 2018. All the patients were analyzed through both CSP and VSP, and all the surgical stents were fabricated through manual and three-dimensional (3D) printing. The predictor variables were the planning method (CSP vs. VSP) and the surgery type (group I: Le Fort I osteotomy + bilateral sagittal split osteotomy [LFI+BSSO] or group II: only bilateral sagittal split osteotomy [BSSO]), and the outcomes were the time and cost. The results were analyzed using the paired t test. Results Thirty patients (12 females, 18 males) met the inclusion criteria, and 17 patients were excluded from the study due to missing or incomplete data. There were 20 group I patients (LFI+BSSO regardless of genioplasty) and 10 group II patients (BSSO regardless of genioplasty). The average time of CSP for group I was 385 ± 7.8 min, and that for group II was 195 ± 8.33 min. The time reduction rate of VSP compared with CSP was 62.8% in group I and 41.5% in group II. On the other hand, there was no statistically significant cost reduction. Conclusions The time investment in VSP in this study was significantly smaller than that in CSP, and the difference was greater in group I than in group II.

Abstract Background Free fibular flap is one of the most useful methods in the hard tissue reconstruction of the maxilla-mandible. Free fibular flap presents some advantages in which the reconstruction of both soft and hard tissues can be done at the same time. It also provides a safe and successful bone graft for the reconstruction, along with a low rate of complications. Despite these advantages and the rarity of a postoperative complication, particularly in oral and maxillofacial surgery procedures, a prolonged operation might exhibit some complications related with rhabdomyolysis. We experienced the rare event of rhabdomyolysis after oral cancer surgery. Case presentation In this article, we report the case of a patient who developed rhabdomyolysis after undergoing free fibular flap surgery. Conclusions Despite the advantages of the free fibular flap operation, clinicians must be aware of the risk of complications because there are multiple factors that could result in rhabdomyolysis, such as duration of operation, position of the subject, and pre-existing conditions of diabetes and hypertension. Once the diagnosis of rhabdomyolysis is confirmed, a prompt treatment plan should be made and applied as soon as possible. This will increase the chance of a full recovery for the patient who is exhibiting symptoms of rhabdomyolysis.

Low-level laser therapy (LLLT) is recognized as an effective medical tool for the treatment of various conditions requiring tissue repair, pain relief, inflammation treatment, and restoration of tissue dysfunction, and its development and research are growing rapidly. However, studies that analyze molecular biology by applying LLLT to osteoclasts are still insufficient to understand the mechanism. In order for LLLT to be suggested as an appropriate treatment method for the treatment of various bone diseases, it is necessary to elucidate the effect and mechanism of LLLT on osteoclast differentiation. In this study, we investigated the effect of LLLT on osteoclast differentiation using murine macrophage (RAW 264.7) cells by means of a Ga-As-Al laser (λ = 810, 80 mW). Our results indicate that LLLT did not induce cytotoxicity in RAW 264.7 cells. When LLLT was applied for 15 s to osteoclasts exposed to RANKL, the expression of NF-κB, ERK, p38, and c-Fos, which are associated with expression of NFATc1, was increased. The RT-PCR results also demonstrated significantly increased expression of osteoclast-specific genes, including NFATc1, TRAP, the calcitonin receptor, and cathepsin K, compared with the control. Taken together, we concluded that low-level laser irradiation induces osteoclastogenesis by enhancing the expression of NF-κB, MAPKs (ERK, p38), c-Fos, and NFATc1 in RAW 264.7 cells. These findings indicate that low-level laser irradiation could be considered a potential treatment option in various metabolic bone diseases that require osteoclastic activity and bone formation.

The complement pathway is involved in eliminating antigen immune complexes. However, the role of the C3 complement system remains largely unknown in influenza virus M2 extracellular (M2e) domain or hemagglutinin (HA) vaccine-mediated protection after vaccination. Using a C3 knockout (C3 KO) mouse model, we found that complement protein C3 was required for effective induction of immune responses to vaccination with M2e-based or HA-based vaccines, which include isotype class-switched antibodies and effector CD4 and CD8 T cell responses. C3 KO mice after active immunization with cross-protective nonneutralizing M2e-based vaccine were not protected against influenza virus, although low levels of M2e-specific antibodies were protective after passive coadministration with virus in wild-type mice. In contrast, C3 KO mice that were immunized with strain-specific neutralizing HA-based vaccine were protected against homologous virus challenge despite lower levels of HA antibody responses. C3 KO mice showed impaired maintenance of innate immune cells and a defect in innate immune responses upon exposure to antigens. The findings in this study suggest that C3 is required for effective induction of humoral and cellular adaptive immune responses as well as protective immunity after nonneutralizing influenza M2e vaccination. [ABSTRACT FROM AUTHOR]

Traditionally, chemical agents such as formalin (FA) and β-propiolactone (BPL) have long been used for the preparation of inactivated vaccines or toxoids. It has been shown that FA extensively modifies vaccine antigens and thus affects immunogenicity profiles, sometimes compromising the protective efficacy of the vaccines or even exacerbating the disease upon infection. In this study, we show that natural catechins from green tea extracts (GT) can be used as an inactivating agent to prepare inactivated viral vaccines. GT treatment resulted in complete and irreversible inactivation of influenza virus as well as dengue virus. In contrast to FA that reacted extensively with multiple amino acids including lysine, a major anchor residue for epitope binding to MHC molecules, GT catechin epigallocatechin-3-gallate (EGCG) crosslinked primarily with cysteine residues and thus preserved the major epitopes of the influenza hemagglutinin. In a mouse model, vaccination with GT-inactivated influenza virus (GTi virus) elicited higher levels of viral neutralizing antibodies than FA-inactivated virus (FAi virus). The vaccination completely protected the mice from a lethal challenge and restricted the challenge viral replication in the lungs. Of note, the quality of antibody responses of GTi virus was superior to that with FAi virus, in terms of the magnitude of antibody titer, cross-reactivity to hetero-subtypes of influenza viruses, and the avidity to viral antigens. As the first report of using non-toxic natural compounds for the preparation of inactivated viral vaccines, the present results could be translated into a clinically relevant vaccine platform with improved efficacy, safety, productivity, and public acceptance.

Enveloped virus vaccines can be damaged by high osmotic strength solutions, such as those used to protect the vaccine antigen during drying, which contain high concentrations of sugars. We therefore studied shrinkage and activity loss of whole inactivated influenza virus in hyperosmotic solutions and used those findings to improve vaccine coating of microneedle patches for influenza vaccination. Using stopped-flow light scattering analysis, we found that the virus underwent an initial shrinkage on the order of 10% by volume within 5 s upon exposure to a hyperosmotic stress difference of 217 milliosmolarity. During this shrinkage, the virus envelope had very low osmotic water permeability (1 - 6×10-4 cm s-1) and high Arrhenius activation energy (Ea = 15.0 kcal mol-1), indicating that the water molecules diffused through the viral lipid membranes. After a quasi-stable state of approximately 20 s to 2 min, depending on the species and hypertonic osmotic strength difference of disaccharides, there was a second phase of viral shrinkage. At the highest osmotic strengths, this led to an undulating light scattering profile that appeared to be related to perturbation of the viral envelope resulting in loss of virus activity, as determined by in vitro hemagglutination measurements and in vivo immunogenicity studies in mice. Addition of carboxymethyl cellulose effectively prevented vaccine activity loss in vitro and in vivo, believed to be due to increasing the viscosity of concentrated sugar solution and thereby reducing osmotic stress during coating of microneedles. These results suggest that hyperosmotic solutions can cause biphasic shrinkage of whole inactivated influenza virus which can damage vaccine activity at high osmotic strength and that addition of a viscosity enhancer to the vaccine coating solution can prevent osmotically driven damage and thereby enable preparation of stable microneedle coating formulations for vaccination.

Major histocompatibility complex class II-deficient (MHC-II KO; Aß-/-) mice were used to assess the roles of MHC-II molecules in inducing protective immune responses to vaccination. After vaccination with influenza A/PR8 virus-like particle (VLP) vaccine, in vivo and in vitro vaccine antigen-specific IgG isotype antibodies were not detected in MHC-II KO mice, which is quite different from CD4 T cell-deficient mice that induced vaccine-specific IgG antibodies. The deficiency in MHC-II did not significantly affect the induction of antigen-specific IgM antibody in sera. MHC-II KO mice that were vaccinated with influenza VLP, whole inactivated influenza virus, or live attenuated influenza virus vaccines were not protected against lethal infection with influenza A/PR8 virus. Adoptive transfer of fractionated spleen cells from wild-type mice to MHC-II KO mice indicated that CD43+ cell populations with MHC-II contributed more significantly to producing vaccine-specific IgG antibodies than CD43- B220+ conventional B cell or CD4 T cell populations, as well as conferring protection against lethal infection. Bone marrowderived dendritic cells from MHC-II KO mice showed a significant defect in producing interleukin-6 and tumor necrosis factor alpha cytokines. Thus, results indicate that MHC-II molecules play multiple roles in inducing protective immunity to influenza vaccination. [ABSTRACT FROM AUTHOR]

There is a significant gap in our fundamental understanding of early morphological and migratory changes in human Langerhans cells (LCs) in response to vaccine stimulation. As the vast majority of LCs studies are conducted in small animal models, substantial interspecies variation in skin architecture and immunity must be considered when extrapolating the results to humans. This study aims to determine whether excised human skin, maintained viable in organ culture, provides a useful human model for measuring and understanding early immune response to intradermally delivered vaccine candidates. Excised human breast skin was maintained viable in air-liquid-interface organ culture. This model was used for the first time to show morphological changes in human LCs stimulated with influenza virus-like particle (VLP) vaccines delivered via intradermal injection. Immunohistochemistry of epidermal sheets and skin sections showed that LCs in VLP treated skin lost their typical dendritic morphology. The cells were more dispersed throughout the epidermis, often in close proximity to the basement membrane, and appeared vertically elongated. Our data provides for increased understanding of the complex morphological, spatial and temporal changes that occur to permit LC migration through the densely packed keratinocytes of the epidermis following exposure to vaccine. Significantly, the data not only supports previous animal data but also provides new and essential evidence of host response to this vaccination strategy in the real human skin environment.

Influenza virus-like particles (VLPs) are a promising cell culture-based vaccine, and the skin is considered an attractive immunization site. In this study, we examined the immunogenicity and protective efficacy of influenza VLPs (H1N1 A/PR/8/34) after skin vaccination using vaccine dried on solid microneedle arrays. Coating of microneedles with influenza VLPs using an unstabilized formulation was found to decrease hemagglutinin (HA) activity, whereas inclusion of trehalose disaccharide preserved the HA activity of influenza VLP vaccines after microneedles were coated. Microneedle vaccination of mice in the skin with a single dose of stabilized influenza VLPs induced 100% protection against challenge infection with a high lethal dose. In contrast, unstabilized influenza VLPs, as well as intramuscularly injected vaccines, provided inferior immunity and only partial protection (=40%). The stabilized microneedle vaccination group showed IgG2a levels that were 1 order of magnitude higher than those of other groups and had the lowest lung viral titers after challenge. Also, levels of recall immune responses, including hemagglutination inhibition titers, neutralizing antibodies, and antibody-secreting plasma cells, were significantly higher after skin vaccination with stabilized formulations. Therefore, our results indicate that HA stabilization, combined with vaccination via the skin using a vaccine formulated as a solid microneedle patch, confers protection superior to that with intramuscular injection and enables potential dose-sparing effects which are reflected by pronounced increases in rapid recall immune responses against influenza virus. [ABSTRACT FROM AUTHOR]