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Osteocytes are located in the lacunae of fluid-filled bone and communicate with neighboring or distant cells by secreting small extracellular vesicles (sEVs) and growth factors as well as via dendrite–dendrite direct connections. However, the mechanism regulating sEV production in osteocytes is yet to be elucidated. In this study, we investigated sEV production and its underlying mechanism in osteocytes cultured on a three dimensional (3D) scaffold. We employed a perfusion system to apply shear stress stimulation to MLO-Y4 cells cultured on a 3D biphasic calcium phosphate (BCP) scaffold and analyzed sEV production and gene expression using RNA sequencing. We found that the expression of genes associated with sEV biogenesis and the secretory pathway were enhanced by fluid shear stress in MLO-Y4 cells cultured on a 3D BCP scaffold. In particular, fluid shear stress induced the expression of Ank, a pyrophosphate transporter, in 3D-cultured MLO-Y4 cells. The role of Ank in sEV production was further examined. Probenecid, an Ank inhibitor, significantly suppressed shear stress-induced sEV production, whereas Ank cDNA overexpression stimulated it. The inhibition of shear stress-induced sEV production by probenecid was recovered by the exogenous addition of pyrophosphate to MLO-Y4 cells. These findings suggest that shear stress-mediated sEV production in 3D-cultured osteocytes is regulated by extracellular pyrophosphate transported by Ank.

This comprehensive review critically examines the detrimental impacts of endocrine-disrupting chemicals (EDCs) on bone health, with a specific focus on substances such as bisphenol A (BPA), per- and polyfluoroalkyl substances (PFASs), phthalates, and dioxins. These EDCs, by interfering with the endocrine system’s normal functioning, pose a significant risk to bone metabolism, potentially leading to a heightened susceptibility to bone-related disorders and diseases. Notably, BPA has been shown to inhibit the differentiation of osteoblasts and promote the apoptosis of osteoblasts, which results in altered bone turnover status. PFASs, known for their environmental persistence and ability to bioaccumulate in the human body, have been linked to an increased osteoporosis risk. Similarly, phthalates, which are widely used in the production of plastics, have been associated with adverse bone health outcomes, showing an inverse relationship between phthalate exposure and bone mineral density. Dioxins present a more complex picture, with research findings suggesting both potential benefits and adverse effects on bone structure and density, depending on factors such as the timing and level of exposure. This review underscores the urgent need for further research to better understand the specific pathways through which EDCs affect bone health and to develop targeted strategies for mitigating their potentially harmful impacts.

Abstract Proton-activated chloride (PAC) channels, ubiquitously expressed in tissues, regulate intracellular Cl- levels and cell death following acidosis. However, molecular mechanisms and signaling pathways involved in PAC channel modulation are largely unknown. Herein, we determine that phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] of the plasma membrane inner leaflet is essential for the proton activation of PAC channels. PI(4,5)P2 depletion by activating phosphatidylinositol 5-phosphatases or Gq protein-coupled muscarinic receptors substantially inhibits human PAC currents. In excised inside-out patches, PI(4,5)P2 application to the cytoplasmic side increases the currents. Structural simulation reveals that the putative PI(4,5)P2-binding site is localized within the cytosol in resting state but shifts to the cell membrane’s inner surface in an activated state and interacts with inner leaflet PI(4,5)P2. Alanine neutralization of basic residues near the membrane-cytosol interface of the transmembrane helice 2 significantly attenuates PAC currents. Overall, our study uncovers a modulatory mechanism of PAC channel through inner membrane PI(4,5)P2.

Older Korean immigrants are one of the most understudied and marginalized Asian ethnic groups in the United States, despite their rapid population growth. Many older Korean immigrants encounter distinct challenges in assimilating into their new country as first-generation immigrants, including cultural conflict, language barriers, low economic status, and a lack of social support. These issues may be compounded for those who live alone, which is considered a negative factor in their mental and physical health. However, little is known about the correlates and health issues of older Korean immigrants living alone. This study’s objective was to explore correlates and health issues among older Korean immigrants living alone. Based on established scoping review methodology five databases, CINAHL, PubMed, MEDLINE, SocINDEX, and Health Source Nursing/Academic Edition, were used to find relevant studies. Twelve articles were reviewed, and four major themes were identified as correlates and health issues among older Korean immigrants living alone in the United States: depression, changed family relationships, social interactions, and factors on general health and well-being. The findings have significant implications for healthcare professionals for understanding the unique culture, situation, and physical and psychosocial vulnerability of older Korean immigrants living alone.

Abstract SARS-CoV-2 has become a global public health problem. Acute respiratory distress syndrome (ARDS) is the leading cause of death due to the SARS-CoV-2 infection. Pulmonary fibrosis (PF) is a severe and frequently reported COVID-19 sequela. In this study, an in vitro model of ARDS and PF caused by SARS-CoV-2 was established in MH-S, THP-1, and MRC-5 cells using pseudo-SARS-CoV-2 (PSCV). Expression of proinflammatory cytokines (IL-6, IL-1β, and TNF-α) and HIF-1α was increased in PSCV-infected MH-S and THP-1 cells, ARDS model, consistent with other profiling data in SARS-CoV-2-infected patients have been reported. Hypoxia-inducible factor-1 alpha (HIF-1α) siRNA and cobalt chloride were tested using this in vitro model. HIF-1α knockdown reduces inflammation caused by PSCV infection in MH-S and THP-1 cells and lowers elevated levels of CTGF, COLA1, and α-SMA in MRC-5 cells exposed to CPMSCV. Furthermore, apigetrin, a glycoside bioactive dietary flavonoid derived from several plants, including Crataegus pinnatifida, which is reported to be a HIF-1α inhibitor, was tested in this in vitro model. Apigetrin significantly reduced the increased inflammatory cytokine (IL-6, IL-1β, and TNF-α) expression and secretion by PSCV in MH-S and THP-1 cells. Apigetrin inhibited the binding of the SARS-CoV-2 spike protein RBD to the ACE2 protein. An in vitro model of PF induced by SARS-CoV-2 was produced using a conditioned medium of THP-1 and MH-S cells that were PSCV-infected (CMPSCV) into MRC-5 cells. In a PF model, CMPSCV treatment of THP-1 and MH-S cells increased cell growth, migration, and collagen synthesis in MRC-5 cells. In contrast, apigetrin suppressed the increase in cell growth, migration, and collagen synthesis induced by CMPSCV in THP-1 and MH-S MRC-5 cells. Also, compared to control, fibrosis-related proteins (CTGF, COLA1, α-SMA, and HIF-1α) levels were over two-fold higher in CMPSV-treated MRC-5 cells. Apigetrin decreased protein levels in CMPSCV-treated MRC-5 cells. Thus, our data suggest that hypoxia-inducible factor-1 alpha (HIF-1α) might be a novel target for SARS-CoV-2 sequela therapies and apigetrin, representative of HIF-1alpha inhibitor, exerts anti-inflammatory and PF effects in PSCV-treated MH-S, THP-1, and CMPVSC-treated MRC-5 cells. These findings indicate that HIF-1α inhibition and apigetrin would have a potential value in controlling SARS-CoV-2-related diseases.

ABSTRACT Sexual reproduction is crucial for increasing the genetic diversity of populations and providing overwintering structures, such as perithecia and associated tissue, in the destructive plant pathogenic fungus Fusarium graminearum. While mating-type genes serve as master regulators in fungal sexual reproduction, the molecular mechanisms underlying this process remain elusive. Winged-helix DNA-binding proteins are key regulators of embryogenesis and cell differentiation in higher eukaryotes. These proteins are implicated in the morphogenesis and development of several fungal species. However, their involvement in sexual reproduction remains largely unexplored in F. graminearum. Here, we investigated the function of winged-helix DNA-binding proteins in vegetative growth, conidiation, and sexual reproduction, with a specific focus on the FgWING27, which is highly conserved among Fusarium species. Deletion of FgWING27 resulted in an abnormal pattern characterized by a gradual increase in the expression of mating-type genes during sexual development, indicating its crucial role in the stage-specific genetic regulation of MAT genes in the late stages of sexual development. Furthermore, using chromatin immunoprecipitation followed by sequencing analysis, we identified Fg17056 as a downstream gene of Fgwing27, which is essential for sexual reproduction. These findings underscore the significance of winged-helix DNA-binding proteins in fungal development and reproduction in F. graminearum, and highlight the pivotal role of Fgwing27 as a core genetic factor in the intricate genetic regulatory network governing sexual reproduction.IMPORTANCEFusarium graminearum is a devastating plant pathogenic fungus causing significant economic losses due to reduced crop yields. In Fusarium Head Blight epidemics, spores produced through sexual and asexual reproduction serve as inoculum, making it essential to understand the fungal reproduction process. Here, we focus on winged-helix DNA-binding proteins, which have been reported to play crucial roles in cell cycle regulation and differentiation, and address their requirement in the sexual reproduction of F. graminearum. Furthermore, we identified a highly conserved protein in Fusarium as a key factor in self-fertility, along with the discovery of its direct downstream genes. This provides crucial information for constructing the complex genetic regulatory network of sexual reproduction and significantly contribute to further research on sexual reproduction in Fusarium species.

This review article investigates solid organ transplantation-induced osteoporosis, a critical yet often overlooked issue, emphasizing its significance in post-transplant care. The initial sections provide a comprehensive understanding of the prevalence and multifactorial pathogenesis of transplantation osteoporosis, including factors such as deteriorating post-transplantation health, hormonal changes, and the impact of immunosuppressive medications. Furthermore, the review is dedicated to organ-specific considerations in transplantation osteoporosis, with separate analyses for kidney, liver, heart, and lung transplantations. Each section elucidates the unique challenges and management strategies pertinent to transplantation osteoporosis in relation to each organ type, highlighting the necessity of an organ-specific approach to fully understand the diverse manifestations and implications of transplantation osteoporosis. This review underscores the importance of this topic in transplant medicine, aiming to enhance awareness and knowledge among clinicians and researchers. By comprehensively examining transplantation osteoporosis, this study contributes to the development of improved management and care strategies, ultimately leading to improved patient outcomes in this vulnerable group. This detailed review serves as an essential resource for those involved in the complex multidisciplinary care of transplant recipients.

ABSTRACTRegulation of osteoclastogenesis and bone-resorbing activity can be an efficacious strategy for treating bone loss diseases because excessive osteoclastic bone resorption leads to the development of such diseases. Here, we investigated the role of (-)-tubaic acid, a thermal degradation product of rotenone, in osteoclast formation and function in an attempt to identify alternative natural compounds. (-)-Tubaic acid significantly inhibited receptor activator of nuclear factor-κB ligand (RANKL)-mediated osteoclast differentiation at both the early and late stages, suggesting that (-)-tubaic acid affects the commitment and differentiation of osteoclast progenitors as well as the cell-cell fusion of mononuclear osteoclasts. (-)-Tubaic acid attenuated the activation of extracellular signal-regulated kinase (ERK) and expression of nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) and its target genes in response to RANKL. Furthermore, a pit-formation assay revealed that (-)-tubaic acid significantly impaired the bone-resorbing activity of osteoclasts. Our results demonstrated that (-)-tubaic acid exhibits anti-osteoclastogenic and anti-resorptive effects, indicating its therapeutic potential in the management of osteoclast-related bone diseases.

ABSTRACT The mycelium of the plant pathogenic fungus Fusarium graminearum exhibits distinct structures for vegetative growth, asexual sporulation, sexual development, virulence, and chlamydospore formation. These structures are vital for the survival and pathogenicity of the fungus, necessitating precise regulation based on environmental cues. Initially identified in Magnaporthe oryzae, the transcription factor Con7p regulates conidiation and infection-related morphogenesis, but not vegetative growth. We characterized the Con7p ortholog FgCon7, and deletion of FgCON7 resulted in severe defects in conidium production, virulence, sexual development, and vegetative growth. The mycelia of the deletion mutant transformed into chlamydospore-like structures with high chitin level accumulation. Notably, boosting FgABAA expression partially alleviated developmental issues in the FgCON7 deletion mutant. Chromatin immunoprecipitation (ChIP)–quantitative PCR (qPCR) analysis confirmed a direct genetic link between FgABAA and FgCON7. Furthermore, the chitin synthase gene Fg6550 (FGSG_06550) showed significant upregulation in the FgCON7 deletion mutant, and altering FgCON7 expression affected cell wall integrity. Further research will focus on understanding the behavior of the chitin synthase gene and its regulation by FgCon7 in F. graminearum. This study contributes significantly to our understanding of the genetic pathways that regulate hyphal differentiation and conidiation in this plant pathogenic fungus.IMPORTANCEThe ascomycete fungus Fusarium graminearum is the primary cause of head blight disease in wheat and barley, as well as ear and stalk rot in maize. Given the importance of conidia and ascospores in the disease cycle of F. graminearum, precise spatiotemporal regulation of these biological processes is crucial. In this study, we characterized the Magnaporthe oryzae Con7p ortholog and discovered that FgCon7 significantly influences various crucial aspects of fungal development and pathogenicity. Notably, overexpression of FgABAA partially restored developmental defects in the FgCON7 deletion mutant. ChIP-qPCR analysis confirmed a direct genetic link between FgABAA and FgCON7. Furthermore, our research revealed a clear correlation between FgCon7 and chitin accumulation and the expression of chitin synthase genes. These findings offer valuable insights into the genetic mechanisms regulating conidiation and the significance of mycelial differentiation in this plant pathogenic fungus.

Recent studies have highlighted the potential of Mesenchymal Stem Cells (MSCs) as an alternative treatment for Alopecia Areata (AA) due to their immunosuppressive properties. While MSCs have shown promise in cell experiments, their effectiveness in vivo remains uncertain. This study aims to validate local administration of MSC therapy’s efficacy in AA treatment through animal experiments. AA was induced through Interferon-gamma (IFN-γ) administration in mice, and MSC treatment (MSCT)’s effects were assessed visually and through tissue analysis. The MSC-treated group showed more hair regrowth compared to the control (CTL) group. MSCT notably reduced local inflammatory cytokines (JAK1, JAK2, STAT1, STAT3, IFN-γR, IL-1β, IL-16, IL-17α, and IL-18) in AA-induced mice’s skin, but systemic cytokine levels remained unchanged. Furthermore, MSC treatment normalized the expression of Wnt/β-catenin signaling pathway genes (LEF1 and β-catenin) and growth factors (FGF7 and FGF2), which are crucial for hair cycle regulation. This study lays the groundwork for further exploring MSCs as a potential treatment for AA, but more research is needed to fully understand their therapeutic potential.

The α-arrestins form a large family of evolutionally conserved modulators that control diverse signaling pathways, including both G-protein-coupled receptor (GPCR)-mediated and non-GPCR-mediated pathways, across eukaryotes. However, unlike β-arrestins, only a few α-arrestin targets and functions have been characterized. Here, using affinity purification and mass spectrometry, we constructed interactomes for 6 human and 12 Drosophila α-arrestins. The resulting high-confidence interactomes comprised 307 and 467 prey proteins in human and Drosophila, respectively. A comparative analysis of these interactomes predicted not only conserved binding partners, such as motor proteins, proteases, ubiquitin ligases, RNA splicing factors, and GTPase-activating proteins, but also those specific to mammals, such as histone modifiers and the subunits of V-type ATPase. Given the manifestation of the interaction between the human α-arrestin, TXNIP, and the histone-modifying enzymes, including HDAC2, we undertook a global analysis of transcription signals and chromatin structures that were affected by TXNIP knockdown. We found that TXNIP activated targets by blocking HDAC2 recruitment to targets, a result that was validated by chromatin immunoprecipitation assays. Additionally, the interactome for an uncharacterized human α-arrestin ARRDC5 uncovered multiple components in the V-type ATPase, which plays a key role in bone resorption by osteoclasts. Our study presents conserved and species-specific protein–protein interaction maps for α-arrestins, which provide a valuable resource for interrogating their cellular functions for both basic and clinical research.

ABSTRACTThe oxidative stress response is required for plant pathogens to endure host-derived oxidative stress during infection. Previously, we identified the eight transcription factors (TFs) involved in the oxidative stress response in the plant pathogenic fungus Fusarium graminearum and found that of these TFs, the deletion of FgbZIP007 caused hypersensitivity to oxidative stress. However, the underlying mechanisms of Fgbzip007 are not fully understood. Based on chromatin immunoprecipitation followed by sequencing (ChIP-seq) analysis, we found the regulons of Fgbzip007, and further genetic studies demonstrated that Fgbzip007 is a key regulator for sulfur assimilation. The deletion strains of FgbZIP007 and its regulons exhibited low level of glutathione biosynthesis, which led to characterize glutathione biosynthesis. Fgbzip007-mediated sulfur assimilation is required for glutathione biosynthesis, which is essential for oxidative stress resistance and pathogenicity in F. graminearum. Although the reduced resistance of glutathione-deficient mutants against oxidative stress was restored by overexpression of FCA7, encoding a core peroxidase, but not on pathogenicity, suggesting that glutathione in pathogenesis is independent of antioxidant properties. This study characterized the function of genes of glutathione biosynthesis, provides specific insight into how Fgbzip007 regulates pathogenesis in F. graminearum, and establishes a genetic framework for the molecular dissection of a TF Fgbzip007 with the integration of pathogen responses to oxidative stress.IMPORTANCEFusarium graminearum is a destructive fungal pathogen that causes Fusarium head blight (FHB) on a wide range of cereal crops. To control fungal diseases, it is essential to comprehend the pathogenic mechanisms that enable fungi to overcome host defenses during infection. Pathogens require an oxidative stress response to overcome host-derived oxidative stress. Here, we identify the underlying mechanisms of the Fgbzip007-mediated oxidative stress response in F. graminearum. ChIP-seq and subsequent genetic analyses revealed that the role of glutathione in pathogenesis is not dependent on antioxidant functions in F. graminearum. Altogether, this study establishes a comprehensive framework for the Fgbzip007 regulon on pathogenicity and oxidative stress responses, offering a new perspective on the role of glutathione in pathogenicity.

Summary: Peptic ulcer disease caused by environmental factors increases the risk of developing gastric cancer (GC), one of the most common and deadly cancers in the world. However, the mechanisms underlying this association remain unclear. A major type of GC uniquely undergoes spasmolytic polypeptide-expressing metaplasia (SPEM) followed by intestinal metaplasia. Notably, intestinal-type GC patients with high levels of YAP signaling exhibit a lower survival rate and poor prognosis. YAP overexpression in gastric cells induces atrophy, metaplasia, and hyperproliferation, while its deletion in a Notch-activated gastric adenoma model suppresses them. By defining the YAP targetome genome-wide, we demonstrate that YAP binds to active chromatin elements of SPEM-related genes, which correlates with the activation of their expression in both metaplasia and ulcers. Single-cell analysis combined with our YAP signature reveals that YAP signaling is activated during SPEM, demonstrating YAP as a central regulator of SPEM in gastric neoplasia and regeneration.

Prostate cancer is frequently diagnosed and the leading cause of death in men worldwide. Prostate-specific antigen (PSA) blood tests and biopsies are the primary methods for diagnosing prostate cancer; however, their accuracy is less than 50%. Therefore, there is a need to develop diagnostic tests that minimize patient discomfort during examination and adequate biomarkers that are more accurate, sensitive, and specific for the detection of prostate cancer. This study investigated the application of metabolomics to identify biomarkers in prostate cancer biofluids. In addition, changes in prostate cancer metabolite levels induced by chrysin, a natural anticancer compound, were evaluated and compared with those in non-treated prostate cancer cells. Gas chromatography-mass spectrometry (GC-MS)-based metabolomic profiling was performed to investigate the differences in metabolic alterations among prostate cancer, normal prostate, and chrysin-treated prostate cancer cells. Pairwise comparisons of the extracellular fluid metabolomes were performed using principal component analysis (PCA), partial least squares–discriminant analysis (PLS-DA), and Student’s t-test. The results revealed significantly different patterns among the metabolite groups, including alcohols, amino acids, carboxylic acids, organic acids, sugars, and urea. The RWPE-1- and chrysin-treated PC-3 (PC-3 Chr) cell groups showed similar tendencies for 23 metabolites, while the groups showed significant differences from the PC-3 group. Most amino acids showed higher concentrations in PC-3 cells than in the normal cell line RWPE-1 cells and PC-3 Chr cells. Our results revealed that GC-MS might be an effective diagnostic tool to detect prostate cancer and contribute to finding new tumor markers for prostate cancer as the basis for new ideas.

ABSTRACT In plant-pathogen interactions, oxidative bursts are crucial for plants to defend themselves against pathogen infections. Rapid production and accumulation of reactive oxygen species kill pathogens directly and cause local cell death, preventing pathogens from spreading to adjacent cells. Meanwhile, the pathogens have developed several mechanisms to tolerate oxidative stress and successfully colonize plant tissues. In this study, we investigated the mechanisms responsible for resistance to oxidative stress by analyzing the transcriptomes of six oxidative stress-sensitive strains of the plant pathogenic fungus Fusarium graminearum. Weighted gene co-expression network analysis identified several pathways related to oxidative stress responses, including the DNA repair system, autophagy, and ubiquitin-mediated proteolysis. We also identified hub genes with high intramodular connectivity in key modules and generated deletion or conditional suppression mutants. Phenotypic characterization of those mutants showed that the deletion of FgHGG4, FgHGG10, and FgHGG13 caused sensitivity to oxidative stress, and further investigation on those genes revealed that transcriptional elongation and DNA damage responses play roles in oxidative stress response and pathogenicity. The suppression of FgHGL7 also led to hypersensitivity to oxidative stress, and we demonstrated that FgHGL7 plays a crucial role in heme biosynthesis and is essential for peroxidase activity. This study increases the understanding of the adaptive mechanisms to cope with oxidative stress in plant pathogenic fungi. IMPORTANCE Fungal pathogens have evolved various mechanisms to overcome host-derived stresses for successful infection. Oxidative stress is a representative defense system induced by the host plant, and fungi have complex response systems to cope with it. Fusarium graminearum is one of the devastating plant pathogenic fungi, and understanding its pathosystem is crucial for disease control. In this study, we investigated adaptive mechanisms for coping with oxidative stress at the transcriptome level using oxidative stress-sensitive strains. In addition, by introducing genetic modification technique such as CRISPR-Cas9 and the conditional gene expression system, we identified pathways/genes required for resistance to oxidative stress and also for virulence. Overall, this study advances the understanding of the oxidative stress response and related mechanisms in plant pathogenic fungi.

Background: Human immunodeficiency virus-1 (HIV-1) that binds to the coreceptor CCR5 (R5 viruses) can evolve into viruses that bind to the coreceptor CXCR4 (X4 viruses), with high viral replication rates governing this coreceptor switch. Korean Red Ginseng (KRG) treatment of HIV-1 infected patients has been found to slow the depletion of CD4+ T cells. This study assessed whether the KRG-associated slow depletion of CD4+ T cells was associated with coreceptor switching. Methods: This study included 146 HIV-1-infected patients naïve to antiretroviral therapy (ART) and seven patients receiving ART. A total of 540 blood samples were obtained from these patients over 122 ± 129 months. Their env genes were amplified by nested PCR or RT-PCR and subjected to direct sequencing. Tropism was determined with a 10% false positive rate (FPR) cutoff. Results: Of the 146 patients naïve to ART, 102 were KRG-naïve, and 44 had been treated with KRG. Evaluation of initial samples showed that coreceptor switch had occurred in 19 patients, later occurring in 38 additional patients. There was a significant correlation between the amount of KRG and FPR. Based on initial samples, the R5 maintenance period was extended 2.35-fold, with the coreceptor switch being delayed 2.42-fold in KRG-treated compared with KRG-naïve patients. The coreceptor switch occurred in 85% of a homogeneous cohort. The proportion of patients who maintained R5 for ≥10 years was significantly higher in long-term slow progressors than in typical progressors. Conclusion: KRG therapy extends R5 maintenance period by increasing FPR, thereby slowing the coreceptor switch.

Background: The number of primary human immunodeficiency virus (HIV)-1 non-B subtype infections (non-B) and that of reports regarding the differences in the pathogenesis of subtype B and non-B infections are increasing. However, to the best of our knowledge, there have been no reports on gross deletion in the nef gene (gΔnef) in non-B infections. Methods: To determine whether there is a difference in the change in CD4+ T cells after treatment with Korean Red Ginseng (KRG) between patients with subtype B and non-B infections, we retrospectively analyzed and compared the annual decrease in CD4+ T cells (AD) and the proportion of gΔnef in 77 patients who were followed for more than 10 years in the absence of combination antiretroviral therapy. Results: Overall, AD was significantly faster in patients with non-B infections than in those with subtype B infections. Survival analysis showed that the survival probability was significantly higher in subtype B than in non B-infected patients. These differences mainly resulted from significant differences in the amount of KRG and age. In the patients treated with KRG, there was a significant correlation between the amount of KRG and the AD in both subtypes. Interestingly, there was a significant correlation between the amount of KRG and the proportion of gΔnef in patients infected with subtype B, but not in those infected with non-B. The same phenomenon was observed when the KRG dose was adjusted. Conclusion: Our results suggest that non-B may be biologically more stable than subtype B.

RNA lariat debranching enzyme Dbr1 is required for intron turnover in the fungal plant pathogen Fusarium graminearum , and accumulation of lariat RNA affects its development and pathogenesis.

Understanding how ecological communities assemble is an urgent research priority. In this study, we used a community ecology approach to examine how ecological and evolutionary processes shape biodiversity patterns of plant pathogenic fungi, Fusarium graminearum and F. asiaticum. High-throughput screening revealed that the isolates had a wide range of phenotypic variation in stress tolerance traits. Net Relatedness Index (NRI) and Nearest Taxon Index (NTI) values were computed based on stress-tolerant distance matrices. Certain local regions exhibited positive values of NRI and NTI, indicating phenotypic clustering within the fungal communities. Competition assays of the pooled strains were conducted to investigate the cause of clustering. During stress conditions and wheat colonization, only a few strains dominated the fungal communities, resulting in reduced diversity. Overall, our findings support the modern coexistence theory that abiotic stress and competition lead to phenotypic similarities among coexisting organisms by excluding large, low-competitive clades. We suggest that agricultural environments and competition for host infection lead to locally clustered communities of plant pathogenic fungi in the field.

ABSTRACT Lipases, which catalyze the hydrolysis of long-chain triglycerides, diglycerides, and monoglycerides into free fatty acids and glycerol, participate in various biological pathways in fungi. In this study, we examined the biological functions and regulatory mechanisms of fungal lipases via two approaches. First, we performed a systemic functional characterization of 86 putative lipase-encoding genes in the plant-pathogenic fungus Fusarium graminearum. The phenotypes were assayed for vegetative growth, asexual and sexual reproduction, stress responses, pathogenicity, mycotoxin production, and lipase activity. Most mutants were normal in the assessed phenotypes, implying overlapping roles for lipases in F. graminearum. In particular, FgLip1 and Fgl1 were revealed as core extracellular lipases in F. graminearum. Second, we examined the lipase activity of previously constructed transcription factor (TF) mutants of F. graminearum and identified three TFs and one histone acetyltransferase that significantly affect lipase activity. The relative transcript levels of FgLIP1 and FGL1 were markedly reduced or enhanced in these TF mutants. Among them, Gzzc258 was identified as a key lipase regulator that is also involved in the induction of lipase activity during sexual reproduction. To our knowledge, this study is the first comprehensive functional analysis of fungal lipases and provides significant insights into the genetic and regulatory mechanisms underlying lipases in fungi. IMPORTANCE Fusarium graminearum is an economically important plant-pathogenic fungus that causes Fusarium head blight (FHB) on wheat and barley. Here, we constructed a gene knockout mutant library of 86 putative lipase-encoding genes and established a comprehensive phenotypic database of the mutants. Among them, we found that FgLip1 and Fgl1 act as core extracellular lipases in this pathogen. Moreover, several putative transcription factors (TFs) that regulate the lipase activities in F. graminearum were identified. The disruption mutants of F. graminearum-lipase regulatory TFs all showed defects in sexual reproduction, which implies a strong relationship between sexual development and lipase activity in this fungus. These findings provide valuable insights into the genetic mechanisms regulating lipase activity as well as its importance to the developmental stages of this plant-pathogenic fungus.

Background: Gyejibokryeong-hwan (GBH), a herbal mixture that is widely used for climacteric syndrome, is studied for its efficacy; however, no study evaluated the GBH indication, which is a blood-stasis pattern based on traditional Chinese medicine theory. Methods: This was a randomized, double-blinded, placebo-controlled clinical pilot trial. Fifty subjects with climacteric syndrome were recruited and randomly assigned to GBH group or placebo group. Subjects were administered GBH or placebo granules for 4 weeks followed by 4 weeks of observation period. For the primary outcome, the Menopause Rating Scale (MRS) was evaluated. For the secondary outcomes, quality of life, degrees of abdominal resistance and tenderness, blood-stasis pattern questionnaire and degree of upward movement of Qi were evaluated. Results: After 4-week intervention, the mean change of total MRS score significantly decreased in the GBH group compared to the placebo group (p = 0.037). The quality of life related to physical health (p = 0.008) and blood-stasis pattern (p = 0.018) significantly improved in the GBH group but not in the placebo group. Conclusion: Our findings provide evidence of the feasibility of recruiting subjects with GBH indications and show that GBH may have clinical efficacy for the treatment of menopausal symptoms, especially urogenital symptoms, without any significant adverse events. Trial registration: This trial was registered at Clinical Research Information Service (CRIS identifier: KCT0002040).

Psoriasis is a chronic inflammatory skin disease characterized by epidermal hyperproliferation, aberrant differentiation of keratinocytes, and dysregulated immune responses. WW domain-containing oxidoreductase (WWOX) is a non-classical tumor suppressor gene that regulates multiple cellular processes, including proliferation, apoptosis, and migration. This study aimed to explore the possible role of WWOX in the pathogenesis of psoriasis. Immunohistochemical analysis showed that the expression of WWOX was increased in epidermal keratinocytes of both human psoriatic lesions and imiquimod-induced mice psoriatic model. Immortalized human epidermal keratinocytes were transduced with a recombinant adenovirus expressing microRNA specific for WWOX to downregulate its expression. Inflammatory responses were detected using Western blotting, real-time quantitative reverse transcription polymerase chain reaction (PCR), and enzyme-linked immunosorbent assay. In human epidermal keratinocytes, WWOX knockdown reduced nuclear factor-kappa B signaling and levels of proinflammatory cytokines induced by polyinosinic: polycytidylic acid [(poly(I:C)] in vitro. Furthermore, calcium chelator and protein kinase C (PKC) inhibitors significantly reduced poly(I:C)-induced inflammatory reactions. WWOX plays a role in the inflammatory reaction of epidermal keratinocytes by regulating calcium and PKC signaling. Targeting WWOX could be a novel therapeutic approach for psoriasis in the future.

Boxwood is a representative ornamental shrub that is widely used in landscaping horticulture. After pruning, damaged leaves or stems of boxwoods are unavoidably vulnerable to infection by various plant pathogens. Several boxwood diseases caused by fungi, such as Volutella blight and Macrophoma leaf spot, have been reported worldwide including Republic of Korea. In this study, we isolated and identified fungal pathogens of boxwood diseases that occurred in Korea and characterized their morphological and taxonomic characteristics. Boxwood samples showing blight symptoms were collected in Seoul, Republic of Korea, and the putative fungal pathogens Pseudonectria buxi, P. foliicola, and Neofusicoccum buxi were successfully identified. Investigation of the morphological features of the field isolates, including mycelial growth and conidial morphology, and phylogenetic analysis of multiple DNA barcode loci revealed that there were some morphological and genetic variations among isolates, but all of the analyzed isolates were closely related to the corresponding reference strains. We also found that P. foliicola strains were more virulent than P. buxi, and the N. buxi strains isolated in this study were weak pathogens or saprophytes. The results of our study will contribute to the development of control strategies for boxwood diseases caused by fungi and accelerate research on the complex ecology of boxwood diseases.

Plant pathogenic fungi cause severe yield losses and mycotoxin contamination in crops. The precise and rapid detection of fungal pathogens is essential for effective disease management. Sequencing universal DNA barcodes has become the standard method for the diagnosis of fungal diseases, as well as for identification and phylogenetic analysis. A major bottleneck in obtaining DNA sequence data from many samples was the laborious and time-consuming process of sample preparation for genomic DNA. Here, we describe a direct PCR approach that bypasses the DNA extraction steps to streamline the molecular identification of fungal species. Using a direct PCR approach, we successfully sequenced the nuclear ribosomal internal transcribed spacer (ITS) region for the representatives of major fungal lineages. To demonstrate the usefulness of this approach, we performed a phylogenetic analysis of the Fusarium fujikuroi species complex, which causes bakanae (“foolish seedling”) disease of rice and mycotoxin contamination. A total of 28 candidate strains were isolated from rice seeds in the Republic of Korea, and the identity of the isolates was determined using the DNA sequence of both ITS and translation elongation factor 1-α regions. In addition, 17 F. fujikuroi isolates were examined for fumonisin (FB) production in rice medium using an enzyme-linked immunosorbent assay. Phylogenetic and toxigenic analyses showed that the F. fujikuroi strains could be distinguished into two groups: FB producers (B14-type) and non-producers (B20-type). These results will accelerate the molecular identification of fungal pathogens and facilitate the effective management of fungal diseases.

Abstract Background and aims Despite global vaccination efforts, the number of confirmed cases of coronavirus disease 2019 (COVID‐19) remains high. To overcome the crisis precipitated by the ongoing pandemic, characteristic studies such as virus diagnosis, isolation, and genome analysis of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) are necessary. Herein, we report the isolation and molecular characterization of SARS‐CoV‐2 from the saliva of patients who had tested positive for COVID‐19 at Proving Ground in Taean County, Republic of Korea, in 2020. Methods We analyzed the whole‐genome sequence of SARS‐CoV‐2 isolated from the saliva samples of patients through next‐generation sequencing. We also successfully isolated SARS‐CoV‐2 from the saliva samples of two patients by using cell culture, which was used to study the cytopathic effects and viral replication in Vero E6 cells. Results Whole‐genome sequences of the isolates, SARS‐CoV‐2 ADD‐2 and ADD‐4, obtained from saliva were identical, and phylogenetic analysis using Bayesian inference methods showed SARS‐CoV‐2 GH clade (B.1.497) genome‐specific clustering. Typical coronavirus‐like particles, with diameters of 70–120 nm, were observed in the SARS‐CoV‐2 infected Vero E6 cells using transmission electron microscopy. Conclusion In conclusion, this report provides insights into the molecular diagnosis, isolation, genetic characteristics, and diversity of SARS‐CoV‐2 isolated from the saliva of patients. Further studies are needed to explore and monitor the evolution and characteristics of SARS‐CoV‐2 variants.

Tumor-targeting antibody (Ab)-fused cytokines, referred to as immunocytokines, are designed to increase antitumor efficacy and reduce toxicity through the tumor-directed delivery of cytokines. However, the poor localization and intratumoral penetration of immunocytokines, especially in solid tumors, pose a challenge to effectively stimulate antitumor immune cells to kill tumor cells within the tumor microenvironment. Here, we investigated the influence of the tumor antigen-binding kinetics of a murine interleukin 12 (mIL12)-based immunocytokine on tumor localization and diffusive intratumoral penetration, and hence the consequent antitumor activity, by activating effector T cells in immunocompetent mice bearing syngeneic colon tumors. Based on tumor-associated antigen HER2-specific Ab Herceptin (HCT)-fused mIL12 carrying one molecule of mIL12 (HCT-mono-mIL12 immunocytokine), we generated a panel of HCT-mono-mIL12 variants with different affinities (KD) mainly varying in their dissociation rates (koff) for HER2. Systemic administration of HCT-mono-mIL12 required an anti-HER2 affinity above a threshold (KD = 130 nM) for selective localization and antitumor activity to HER2-expressing tumors versus HER2-negative tumors. However, the high affinity (KD = 0.54 or 46 nM) due to the slow koff from HER2 antigen limited the depth of intratumoral penetration of HCT-mono-mIL12 and the consequent tumor infiltration of T cells, resulting in inferior antitumor activity compared with that of HCT-mono-mIL12 with moderate affinity of (KD = 130 nM) and a faster koff. The extent of intratumoral penetration of HCT-mono-mIL12 variants was strongly correlated with their tumor infiltration and intratumoral activation of CD4+ and CD8+ T cells to kill tumor cells. Collectively, our results demonstrate that when developing antitumor immunocytokines, tumor antigen-binding kinetics and affinity of the Ab moiety should be optimized to achieve maximal antitumor efficacy.

BackgroundWhole-genome sequencing plays a critical role in the genomic epidemiology intended to improve understanding the spread of emerging viruses. Dabie bandavirus, causing severe fever with thrombocytopenia syndrome (SFTS), is a zoonotic tick-borne virus that poses a significant public health threat. We aimed to evaluate a novel amplicon-based nanopore sequencing tool to obtain whole-genome sequences of Dabie bandavirus, also known as SFTS virus (SFTSV), and investigate the molecular prevalence in wild ticks, Republic of Korea (ROK).Principal findingsA total of 6,593 ticks were collected from Gyeonggi and Gangwon Provinces, ROK in 2019 and 2020. Quantitative polymerase chain reaction revealed the presence of SFSTV RNA in three Haemaphysalis longicornis ticks. Two SFTSV strains were isolated from H. longicornis captured from Pocheon and Cheorwon. Multiplex polymerase chain reaction-based nanopore sequencing provided nearly full-length tripartite genome sequences of SFTSV within one hour running. Phylogenetic and reassortment analyses were performed to infer evolutionary relationships among SFTSVs. Phylogenetic analysis grouped SFTSV Hl19-31-4 and Hl19-31-13 from Pocheon with sub-genotype B-1 in all segments. SFTSV Hl20-8 was found to be a genomic organization compatible with B-1 (for L segment) and B-2 (for M and S segments) sub-genotypes, indicating a natural reassortment between sub-genotypes.Conclusion/significanceAmplicon-based next-generation sequencing is a robust tool for whole-genome sequencing of SFTSV using the nanopore platform. The molecular prevalence and geographical distribution of SFTSV enhanced the phylogeographic map at high resolution for sophisticated prevention of emerging SFTS in endemic areas. Our findings provide important insights into the rapid whole-genome sequencing and genetic diversity for the genome-based diagnosis of SFTSV in the endemic outbreak.

Acne vulgaris is a common skin disease characterized by increased sebum production, inflammation, and Cutibacterium acnes (CA: formerly Propionibacterium acnes) hyperproliferation in pilosebaceous follicles. This study evaluated the efficacy of FRO, a formula composed of fermented Rhus verniciflua Stokes and Orostachys japonicus, against acne pathogenesis via antimicrobial assessment and an in vitro analysis. Stimulated model cells treated with hormones, CA, or lipopolysaccharide (LPS) were designed based on the characteristics of acne pathogenesis, including inflammation and sebum hypersecretion. High-performance liquid chromatography, disc diffusion, MTS, and western blotting assays were used to examine potential anti-acne effects. FRO was determined to contain phenolics such as gallic acid, fisetin, quercetin, and kaempferol. FRO exerted antimicrobial activity against CA and inhibited reactive oxygen species production that was otherwise increased by LPS or CA in HaCaT cells. Additionally, FRO exerted anti-inflammatory effects by inhibiting iNOS, TNF-α, IL-6, p-STAT-3, and p-NF-κB, which were previously upregulated by LPS or CA in THP-1 and HaCaT cells. FRO inhibited lipogenesis induced by steroid hormones and CA by decreasing FAS and SREBP-1 levels in sebocytes. Additionally, FRO down-regulated the androgen receptor, 5α-reductase, SREBP-1, and FAS levels, which were upregulated by steroid hormone in LNCaP cells. Taken together, our findings suggest that FRO alleviates acne by inhibiting the growth of CA, inflammation, and excess sebum and could be used for functional cosmetics or acne treatments.

Several studies have reported the pathogenic role of Malassezia in atopic dermatitis (AD); the significance of Malassezia’s influence on AD needs to be further investigated. Dupilumab, a monoclonal antibody to anti-Interleukin (IL) 4Rα, and ruxolitinib, a Janus kinase (JAK)1/2 inhibitor, are the first approved biologics and inhibitors widely used for AD treatment. In this study, we aimed to investigate how Malassezia Restricta (M. restricta) affects the skin barrier and inflammation in AD and interacts with the AD therapeutic agents ruxolitinib and anti-IL4Rα. To induce an in vitro AD model, a reconstructed human epidermis (RHE) was treated with IL-4 and IL-13. M. restricta was inoculated on the surface of RHE, and anti-IL4Rα or ruxolitinib was supplemented to model treated AD lesions. Histological and molecular analyses were performed. Skin barrier and ceramide-related molecules were downregulated by M. restricta and reverted by anti-IL4Rα and ruxolitinib. Antimicrobial peptides, VEGF, Th2-related, and JAK/STAT pathway molecules were upregulated by M. restricta and suppressed by anti-IL4Rα and ruxolitinib. These findings show that M. restricta aggravated skin barrier function and Th2 inflammation and decreased the efficacy of anti-IL4Rα and ruxolitinib.

Background: We have reported that internal deletions in the nef, gag, and pol genes in HIV-1–infected patients are induced in those treated with Korean Red Ginseng (KRG). KRG delays the development of resistance mutations to antiretroviral drugs. Methods: The vif-vpr genes over 26 years in 20 hemophiliacs infected with HIV-1 from a single source were sequenced to investigate whether vif-vpr genes were affected by KRG and KRG plus highly active antiretroviral therapy (ART) (hereafter called GCT) and compared the results with our previous data. Results: A significantly higher number of in-frame small deletions were found in the vif-vpr genes of KRG-treated patients than at the baseline, in control patients, and in ART-alone patients (p

Background: Cancer-related cognitive impairment (CRCI) is a significant problem for cancer patients, as the number of cancer survivors experiencing cognitive impairments is increasing in the absence of standard treatment. There have been attempts to improve the cognitive function of patients with cancer using acupuncture; however, no studies have been conducted using electroacupuncture. Thus, we designed a preliminary study to investigate the feasibility of a clinical trial using electroacupuncture in CRCI patients. Methods: We conducted a single-arm, pilot, clinical trial to investigate the feasibility of a study protocol for further large-scale clinical trials of electroacupuncture in CRCI patients. All participants were treated with electroacupuncture twice a week for 30 minutes at a time, for 8 weeks on acupoints GV20, GV24, EX-HN1, and GB20, HT7, PC6, and KI3. Both subjective and objective outcomes of cognitive function, quality of life (QoL), and psychological factors were measured in all participants at baseline, week 4, 8, and 12. For safety assessment, vital signs, laboratory examinations, and adverse events (AEs) were observed throughout the trial. Results: A total of 12 participants were enrolled at Daejeon and Dunsan Korean Medicine Hospital of Daejeon University from 21 April 2017 to 31 January 2018. After 8 weeks of treatment, electroacupuncture significantly improved both subjective and objective cognitive outcomes, including the perceived cognitive impairments scale of the Functional Assessment of Cancer Therapy-Cognitive Function, QoL scale of the European Organization for Research and Treatment of Cancer Quality of Life Core Questionnaire-C30, Korean version of Montreal Cognitive Assessment, Boston Naming Test, Seoul Verbal Learning Test, and Rey Complex Figure Test. During the entire trial period, 19 AEs were observed, with no serious AEs. Additionally, it was found that all feasibility outcomes, including recruitment, completion, and adherence rates, achieved successful results as the ratio exceeded 0.8. Conclusion: Our study results revealed that electroacupuncture improved cognitive complaints in cancer patients, and we expect electroacupuncture to be a safe and effective management therapy for CRCI patients. These feasibility trial results will be used as preliminary data for future randomized controlled clinical trials. Trial registration number: Korean Clinical Trial Registry (KCT0002168).

Abstract Background In the general population, the trabecular bone score (TBS) represents the bone microarchitecture and predicts fracture risk independent of bone mineral density (BMD). A few studies reported that TBS is significantly reduced in dialysis patients. Chronic kidney disease-mineral and bone disorder (CKD-MBD) are accompanied by increased fracture risk, cardiovascular morbidity, and mortality. We investigated whether TBS is associated with comorbidity related to CKD-MBD or frailty in hemodialysis patients. Methods In this prospective observational study, TBS was obtained using the TBS iNsight software program (Med-Imaps) with BMD dual energy x-ray absorptiometry (DXA) images (L1–L4) from prevalent hemodialysis patients. A Tilburg frailty indicator was used to evaluate frailty, and hand grip strength and bio-impedance (InBody) were measured. A patient-generated subjective global assessment (PG-SGA) was used for nutritional assessment. The history of cardiovascular events (CVE) and demographic, clinical, laboratory, and biomarker data were collated. We then followed up patients for the occurrence of CKD-MBD related complications. Results We enrolled 57 patients in total. The mean age was 56.8 ± 15.9 years (50.9% female). Prevalence of Diabetes mellitus (DM) was 40.4% and CVE was 36.8%. Mean TBS was 1.44 ± 0.10. TBS significantly reduced in the CVE group (1.38 ± 0.08 vs. 1.48 ± 0.10, p

Abstract The CCAAT sequence is a ubiquitous cis-element of eukaryotic promoters, and genes containing CCAAT sequences have been shown to be activated by the CCAAT-binding transcription factor complex in several eukaryotic model organisms. In general, CCAAT-binding transcription factors form heterodimers or heterotrimeric complexes that bind to CCAAT sequences within the promoters of target genes and regulate various cellular processes. To date, except Hap complex, CCAAT-binding complex has been rarely reported in fungi. In this study, we characterized two CCAAT-binding transcription factors (Fct1 and Fct2) in the plant pathogenic fungus Fusarium graminearum. Previously, FCT1 and FCT2 were shown to be related to DNA damage response among eight CCAAT-binding transcription factors in F. graminearum. We demonstrate that the nuclear CCAAT-binding complex of F. graminearum has important functions in various fungal developmental processes, not just DNA damage response but virulence and mycotoxin production. Moreover, the results of biochemical and genetic analyses revealed that Fct1 and Fct2 may form a complex and play distinct roles among the eight CCAAT-binding transcription factors encoded by F. graminearum. To the best of our knowledge, the results of this study represent a substantial advancement in our understanding of the molecular mechanisms underlying the functions of CCAAT-binding factors in eukaryotes.

Genome editing using the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) system has greatly facilitated the genetic analysis of fungal pathogens. The head blight fungus, Fusarium graminearum, causes destructive losses of economically important cereal crops. The recent development of the CRISPR-Cas9 system for use with F. graminearum has enabled more efficient genome editing. In this study, we described a CRISPR-Cas9-based genome-editing tool for the direct delivery of preassembled Cas9 ribonucleoproteins (RNPs) into the protoplasts of F. graminearum. The use of RNPs significantly increased both the number of transformants and percentage of transformants in which the target gene was successfully replaced with a selectable marker. We showed that a single double-strand DNA break mediated by the Cas9 ribonucleoprotein was sufficient for gene deletion. In addition, short-homology recombination required only 50 base pair regions flanking the target gene. The high efficiency of Cas9 RNPs enables large-scale functional analysis, the identification of essential genes, and gene deletion that is difficult with conventional methods. We expect that our approach will accelerate genetic studies of F. graminearum.

Oral mucositis (OM) is a debilitating adverse event in patients undergoing treatment for cancer. This study aimed to evaluate the therapeutic effect of a novel handheld photobiomodulation therapy (PBMT) device on chemoradiation therapy (CRT)-induced OM in patients with head and neck cancer. Head and neck cancer patients undergoing CRT who developed moderate-to-severe OM during treatment were enrolled. After PBMT and at 2 and 4 weeks after PBMT, the mean value of OM grade decreased significantly from 2.63 to 2.13, 1.31, and 0.75, respectively (p < 0.05, p < 0.001, and p < 0.001). Moreover, we observed significant improvement in health-related quality of life (HRQoL) after PBMT compared to baseline through a validated questionnaire; EORTC QLQ-C30. In the present study, the use of this PBMT device in the management of CRT-induced OM in patients with head and neck cancer was generally well tolerated and resulted in the improvement of OM. However, evidence supporting its use remains lacking owing to limitations such as the small number of participants and lack of a control group. Therefore, further mechanistic studies and large-scale randomized controlled trials are needed to confirm the effectiveness of PBMT in the treatment of CRT-induced OM, as shown in our results.

Objectives: Emerging studies found the potential effects of acupuncture for treating chronic pain and mental disorders, namely, depressive and anxiety disorders. Acupuncture is widely used for treating culture-related anger syndrome, Hwa-byung. This pilot trial aimed to investigate the feasibility of a clinical trial testing acupuncture for the psychosomatic symptoms of Hwa-byung.Methods: A total of 26 patients with Hwa-byung planned to be randomly assigned to the acupuncture or sham acupuncture groups. About 10 treatment sessions were applied over 4 weeks. The 100-mm visual analog scale was used to measure the six major Hwa-byung symptoms: stuffiness in the chest, heat sensations, pushing-up in the chest, feeling a mass in the throat, feelings of unfairness, and hard feelings. The criteria for assessing the success of this pilot trial were defined as improvement in three or more of the six Hwa-byung symptoms after treatment, with an effect size >0.2.Results: A total of 15 patients were finally included and randomly assigned to the acupuncture group (n = 7) or the sham acupuncture group (n = 8). After 10 treatment sessions, the Cohen's d effect sizes for acupuncture compared to sham acupuncture were >0.2 for each one of the six major Hwa-byung symptoms, which met our a priori criteria for success. Also, the effect size for the somatic symptoms of “stuffiness in the chest” was 0.81 (95% CI −0.40, 2.20), referring to a large effect size.Conclusions: Our results suggest that acupuncture treatment would be regarded as an acceptable intervention for a full-scale study of psychosomatic symptoms in patients with Hwa-byung.Trial Registration:cris.nih.go.kr, identifier: KCT0001732.

Alopecia areata (AA) is an autoimmune condition related to the collapse of the immune privilege of hair follicles. Certain AA populations present severe clinical manifestations, such as total scalp hair or body hair loss and a treatment refractory property. The aim of this study was to assess the effects of allogenic human mesenchymal stem cells (hMSCs) from healthy donors on the peripheral blood mononuclear cells (PBMCs) of severe AA patients, with a focus on the change in the cell fraction of Th1, Th17, and Treg cells and immunomodulatory functions. PBMCs of 10 AA patients and eight healthy controls were collected. Levels of Th17, Th1, and Treg subsets were determined via flow cytometry at baseline, activation status, and after co-culturing with hMSCs. All participants were severe AA patients with SALT > 50 and with a long disease duration. While the baseline Th1 and Treg levels of AA patients were comparable to those of healthy controls, their Th17 levels were significantly lower than those of the controls. When stimulated, the levels of CD4+IFN-γ+ T cells of the AA patients rose sharply compared to the baseline, which was not the case in those of healthy controls. The cell fraction of CD4+Foxp3+ regulatory T cells also abruptly increased in AA patients only. Co-culturing with allogenic hMSCs in activated AA PBMCs slightly suppressed the activation levels of CD4+INF-γ+ T cells, whereas it significantly induced the differentiation of CD4+Foxp3+ regulatory T cells. However, these changes were not prominent in the PBMCs of health controls. To examine the pathomechanisms, PBMCs of healthy donors were treated with IFN-γ to induce AA-like environment and then treated with allogenic grants and compared with ruxolitinib as a positive treatment control. hMSC treatment was shown to significantly inhibit the mRNA levels of proinflammatory cytokines, such as IFN-γ, TNF-α, IL-1α, IL-2R, IL-15, and IL-18, and chemokines, such as CCR7 and CCR10, in IFN-treated PBMCs. Interestingly, hMSCs suppressed the activation of JAK/STAT signaling by IFN in PBMCs with an effect that was comparable to that of ruxolitinib. Furthermore, the hMSC treatment showed stronger efficacy in inducing Foxp3, IL-10, and TGF-β mRNA transcription than ruxolitinib in IFN-treated PBMCs. This study suggests that allogenic hMSC treatments have therapeutic potential to induce immune tolerance and anti-inflammatory effects in severe AA patients.

Hypoxia, a typical feature of locally advanced solid tumors including prostate cancer, is a critical contributor to tumor progression and causes resistance to therapy. In this study, we investigated the effects of chrysin on tumor progression in hypoxic PC-3 cells. Chrysin exerted a significant inhibitory effect on 3D cell growth under normoxic and hypoxic conditions. It also decreased the hypoxia-induced vasculogenic mimicry and attenuated the expression of HIF-1α and VE-cadherin. Chrysin inhibited HIF-1α accumulation in a concentration- and time-dependent manner in hypoxic PC-3 cells, while also suppressing the expression of HIF-1α by inhibiting SPHK-1 in both CoCl2 and hypoxic PC-3 cells. At high concentrations of chrysin, there was a greater increase in apoptosis in the hypoxic cells compared to that in normoxic cells, which was accompanied by sub-G1 phase arrest. Chrysin-induced apoptosis inhibited VEGF and Bcl-2 and induced the cleavage of PARP and caspase-3. SPHK-1 knockdown induced apoptosis and inhibited epithelial–mesenchymal transition. Consistent with the in vitro data, 50 mg/kg of chrysin suppressed the tumor growth of PC-3 xenografts by 80.4% compared to that in the untreated control group. The immunohistochemistry of tumor tissues revealed decreased Ki-67, HIF-1α, and VEGF expression in the chrysin-treated group compared to an untreated control. Western blotting data for tumor tissues showed that chrysin treatment decreased SPHK-1, HIF-1α, and PARP expression while inducing caspase-3 cleavage. Overall, our findings suggest that chrysin exerts anti-tumor activity by inhibiting SPHK-1/HIF-1α signaling and thus represents a potent chemotherapeutic agent for hypoxia, which promotes cancer progression and is related to poor prognoses in prostate cancer patients.

Seoul virus (SEOV), an etiological agent for hemorrhagic fever with renal syndrome, poses a significant public health threat worldwide. This study evaluated the feasibility of a mobile Biomeme platform for facilitating rapid decision making of SEOV infection. A total of 27 Rattus norvegicus were collected from Seoul Metropolitan City and Gangwon Province in Republic of Korea (ROK), during 2016–2020. The serological and molecular prevalence of SEOV was 5/27 (18.5%) and 2/27 (7.4%), respectively. SEOV RNA was detected in multiple tissues of rodents using the Biomeme device, with differences in Ct values ranging from 0.6 to 2.1 cycles compared to a laboratory benchtop system. Using amplicon-based next-generation sequencing, whole-genome sequences of SEOV were acquired from lung tissues of Rn18-1 and Rn19-5 collected in Gangwon Province. Phylogenetic analysis showed a phylogeographical diversity of rat-borne orthohantavirus collected in Gangwon Province. We report a novel isolate of SEOV Rn19-5 from Gangwon Province. Our findings demonstrated that the Biomeme system can be applied for the molecular diagnosis of SEOV comparably to the laboratory-based platform. Whole-genome sequencing of SEOV revealed the phylogeographical diversity of orthohantavirus in the ROK. This study provides important insights into the field-deployable diagnostic assays and genetic diversity of orthohantaviruses for the rapid response to hantaviral outbreaks in the ROK.

Background: We have reported that defective nef and gag genes are induced in HIV-1–infected patients treated with Korean Red Ginseng (KRG). Methods: To investigate whether KRG treatment and highly active antiretroviral therapy (HAART) affect genetic defects in the pol gene, we amplified and sequenced a partial pol gene (p-pol) containing the integrase portion (1.2 kb) by nested PCR with sequential peripheral blood mononuclear cells over 20 years and compared it with those patients at baseline, in control patients, those taking ginseng-based combination therapy (GCT; KRG plus combinational antiretroviral therapy) and HAART alone. We also compared our findings to look for the full-length pol gene (pol) (3.0-kb) Results: Twenty-patients infected with subtype B were treated with KRG for 116 ± 58 months in the absence of HAART. Internal deletion in the pol gene (Δpol) was significantly higher in the KRG group (11.9%) than in the control group and at baseline; its detection was significantly inhibited during GCT as much as during HAART. In addition, the Δpol in p-pol significantly depended on the duration of KRG treatment. In pol, the proportion of Δpol was significantly higher in the KRG group (38.7%) than in the control group, and it was significantly inhibited during GCT and HAART. In contrast, the proportion of stop codon appeared not to be affected by KRG treatment. The PCR success rate was significantly decreased with longer GCT. Conclusion: The proportion of Δpol depends on template size as well as KRG treatment. HAART decreases the detection of Δpol. Keywords: AIDS, HIV-1, Korean Red Ginseng, Internal deletion, pol

Background: To date, no study has described disease progression in Asian patients infected with HIV-1 subtype D. Methods: To determine whether the disease progression differs in patients infected with subtypes D and B prior to starting combination antiretroviral therapy, the annual decline (AD) in CD4+ T cell counts over 96 ± 59 months was retrospectively analyzed in 163 patients and compared in subtypes D and B based on the nef gene. Results: CD4+ T cell AD was significantly higher in the six subtype D–infected patients than in the 157 subtype B–infected patients irrespective of Korean Red Ginseng (KRG) treatment (p 2-fold higher risk of death and a 2.9-fold greater rate of progression than subtype B, regardless of KRG treatment. Keywords: AIDS, Disease progression, HIV-1 subtype D, Korean Red Ginseng, nef gene

Background Lung cancer is a leading cause of morbidity and mortality worldwide, and the incidence continues to rise. Although many prognostic factors have been identified, the clinical characteristics and outcomes in Korean lung cancer patients are not well defined. Methods Of the 23,254 new lung cancer cases registered at the Korea Central Cancer Registry in 2013, total 489 patients from 19 hospitals were abstracted by the Korean Central Cancer Registry. The clinical data retrospectively analyzed, patients were followed up until December 2015. Results The median age was 69 years (interquartile range, 60–74 years); 65.4% were male and 62.1% were ever-smokers. Cough was the most common initial symptom (33.5%); 13.1% of patients were asymptomatic. While squamous cell carcinoma was the most common subtype in male patients (37.2%), adenocarcinoma was the most frequent histological type in all patients (48.7%) and females (76.3%). The majority of patients received treatment (76.5%), which included surgery, radiation therapy, and chemotherapy. Older age (hazard ratio [HR], 1.037), lower body mass index (HR, 0.904), ever-smoker (HR, 2.003), small cell lung cancer (HR, 1.627), and distant metastasis (HR, 3.990) were independent predictors of mortality. Patients without symptoms (HR, 0.387) and without treatment (HR, 0.364) were associated with a favorable outcome in multivariate Cox analysis. Conclusion Lung cancer in Korea occurs predominantly in elderly patients, with adenocarcinoma being the most frequent subtype. The prognosis was poorer in ever-smokers and older, malnourished, and untreated patients with advanced lung cancer.

Successful spinal surgery demands high levels of concentration and cooperation from participating health care workers. The intraoperative stress levels and concentration levels of surgeons have been studied previously; however, those of nurses are rarely studied. Therefore, the purpose of this study is to understand the stresses affecting surgical nurses by their participating role during spinal surgery. A total of 160 surgical stress records were obtained during 40 surgeries, including electroencephalography (EEG) signals and heart rate variability (HRV) from three orthopedic spinal surgeons and six nurses; concentration, tension level and physical stress were analyzed. Levels of both concentration and tension were significantly higher in circulating nurses during all surgical stages (p < 0.05). Both beats per minute and low frequency/high frequency ratios, which reflect physical stress, were higher in scrub nurses (p < 0.05). As the surgical experience of scrub nurses increased, the key parameters related to stress tended to decrease (p < 0.01). These results will contribute to understanding the pattern of intraoperative stress of surgical nurses, and therefore help in enhancing the teamwork of the surgical team for optimal outcomes.

Pinus koraiensis is a valuable plant source of functional health foods and medicinal materials. Hypertriglyceridemia affects about 15–20% of adults and is related to stroke, metabolic syndromes, cardiovascular diseases, and diabetes mellitus. Dietary fructose, a risk factor for developing hypertriglyceridemia, significantly increases postprandial triglyceride (TG) levels and aggravates non-alcoholic fatty liver disease. In this study, we aimed to analyze the effect of ethanol extract from P. koraiensis needles (EPK) on fructose (Fr)-induced cell culture and animal models, respectively. Our team determined the bioactivity, such as anti-cancer, anti-obesity, anti-diabetic, and anti-hyperlipidemic functions, of P. koraiensis needle extract. The EPK markedly reduced TG levels in the liver and serum and enhanced TG excretion through feces in high-fructose-fed rats. Furthermore, the EPK inhibited de novo lipogenesis and its markers—carbohydrate response element-binding protein (ChREBP), sterol regulatory element-binding protein 1 (SREBP-1), fatty acid synthase (FAS), 3-Hydroxy-3-Methylglutaryl-CoA Reductase (HMGCR), and tumor necrosis factor-alpha (TNF-α), a pro-inflammatory marker. Consistent with the results of the in vivo experiment, the EPK decreased SREBP-1, ChREBP, HMGCR, FAS, TNF-α, and iNOS expression levels, resulting in slower lipid accumulation and lower TG levels in Fr-induced HepG2 cells. These findings suggest that EPK mitigates hypertriglyceridemia and hepatic TG accumulation by inhibiting de novo lipogenic and pro-inflammatory factors.

Apigetrin is a flavonoid glycoside phytochemical that is derived from various herbs and exhibits several beneficial biological activities, including anti-oxidant, anti-inflammatory, anti-obesity, and anti-cancer effects. In the present study, we elucidated the anti-cancer effect and targeting mechanism of apigetrin in LNCaP and PC-3 cells through various experiments, including cell viability by CELLOMAXTM Viability Assay kit, cell migration by scratch wound assays, and 2D-and 3D- cell growth assay. Apigetrin inhibited the viability, migration, proliferation, and growth of cells in long-term 2D- and 3D- cultures cell growth. A high dose of apigetrin induced apoptosis, as evidenced by increased cleavage of poly ADP-ribose polymerase (PARP) and caspase-3 (c-cas3) in both LNCaP and PC-3 cells. Furthermore, apigetrin inhibited AR, PSA, HIF-1α, and VEGF expression in LNCaP and PC-3 cells. Apigetrin also suppressed the hypoxia-induced HIF-1α expression in these cells. Furthermore, apigetrin reduced hypoxia-induced VEGF secretion in the culture medium and inhibited hypoxia-induced tube formation of HUVECs. Silencing of AKT revealed that the anti-cancer activity of apigetrin is mediated via AKT. Thus, our data suggest that apigetrin exerts anti-cancer effects by inhibiting AKT, a central key of HIF-1α and AR signaling, in early-and late-stage prostate cancer cells.

Bone is a highly dynamic tissue that is continuously remodeled to attain and maintain optimal bone integrity, mass, and strength [...]

Patients with severe eosinophilic asthma (SEA; characterized by persistent eosinophilia in blood and airway tissues) experience frequent asthma exacerbations with poor clinical outcomes. Interleukin 5 (IL-5) and IL-5 receptor alpha subunit (IL-5α) play key roles in eosinophilia maintenance, and relevant therapeutic strategies include the development of antibodies (Abs) against IL-5 or IL-5α to control eosinophilia. Benralizumab, an anti–IL-5α Ab that depletes eosinophils mainly via Ab-dependent cell-mediated cytotoxicity and through blockage of IL-5 function on eosinophils, has been clinically approved for patients with SEA. Here, we report engineering of a new humanized anti–IL-5Rα Ab with potent biological activity. We first raised murine Abs against human IL-5Rα, humanized a leading murine Ab, and then further engineered the humanized Abs to enhance their affinity for IL-5Rα using the yeast surface display technology. The finally engineered version of the Ab, 5R65.7, with affinity (KD ≈ 4.64 nM) stronger than that of a clinically relevant benralizumab analogue (KD ≈ 26.8 nM) showed improved neutralizing activity toward IL-5–dependent cell proliferation in a reporter cell system. Domain level Ab epitope mapping revealed that 5R65.7 recognizes membrane-proximal domain 3 of IL-5Rα, distinct from domain I epitope of the benralizumab analogue. In ex vivo assays with peripheral eosinophils from patients with SEA and healthy donors, 5R65.7 manifested more potent biological activities than the benralizumab analogue did, including inhibition of IL-5–dependent proliferation of eosinophils and induction of eosinophil apoptosis through autologous natural-killer-cell–mediated Ab-dependent cell-mediated cytotoxicity. Our study provides a potent anti–IL-5Rα Ab, 5R65.7, which is worthy of further testing in preclinical and clinical trials against SEA as a potential alternative to the current therapeutic arsenal.

Megakaryocytes are large hematopoietic cells present in the bone marrow cavity, comprising less than 0.1% of all bone marrow cells. Despite their small number, megakaryocytes play important roles in blood coagulation, inflammatory responses, and platelet production. However, little is known about changes in gene expression during megakaryocyte maturation. Here we identified the genes whose expression was changed during K562 leukemia cell differentiation into megakaryocytes using an Affymetrix GeneChip microarray to determine the multifunctionality of megakaryocytes. K562 cells were differentiated into mature megakaryocytes by treatment for 7 days with phorbol 12-myristate 13-acetate, and a microarray was performed using RNA obtained from both types of cells. The expression of 44,629 genes was compared between K562 cells and mature megakaryocytes, and 954 differentially expressed genes (DEGs) were selected based on a p-value < 0.05 and a fold change >2. The DEGs was further functionally classified using five major megakaryocyte function-associated clusters—inflammatory response, angiogenesis, cell migration, extracellular matrix, and secretion. Furthermore, interaction analysis based on the STRING database was used to generate interactions between the proteins translated from the DEGs. This study provides information on the bioinformatics of the DEGs in mature megakaryocytes after K562 cell differentiation.

Localized surface plasmon resonance (LSPR) is a powerful platform for detecting biomolecules including proteins, nucleotides, and vesicles. Here, we report a colloidal gold (Au) nanoparticle-based assay that enhances the LSPR signal of nanoimprinted Au strips. The binding of the colloidal Au nanoparticle on the Au strip causes a red-shift of the LSPR extinction peak, enabling the detection of interleukin-10 (IL-10) cytokine. For LSPR sensor fabrication, we employed a roll-to-roll nanoimprinting process to create nanograting structures on polyethylene terephthalate (PET) film. By the angled deposition of Au on the PET film, we demonstrated a double-bent Au structure with a strong LSPR extinction peak at ~760 nm. Using the Au LSPR sensor, we developed an enzyme-linked immunosorbent assay (ELISA) protocol by forming a sandwich structure of IL-10 capture antibody/IL-10/IL-10 detection antibody. To enhance the LSPR signal, we introduced colloidal Au nanocube (AuNC) to be cross-linked with IL-10 detection antibody for immunogold assay. Using IL-10 as a model protein, we successfully achieved nanomolar sensitivity. We confirmed that the shift of the extinction peak was improved by 450% due to plasmon coupling between AuNC and Au strip. We expect that the AuNC-assisted LSPR sensor platform can be utilized as a diagnostic tool by providing convenient and fast detection of the LSPR signal.

No effective therapeutic strategies have been developed against food allergies. Immunomodulation during early infant period could prevent the development of food allergies. We investigated the preventive effects of human hematopoietic mesenchymal stem cells (hHMSCs) in mice with ovalbumin (OVA)-induced food allergy. BALB/c mice with OVA-induced food allergy were divided into 3 groups, and each group was treated with hHMSCs or hHMSC culture medium (hHMSC-CM) or saline. Ear thickness, allergy score, rectal temperature, and diarrhea occurrence were checked. Total IgE, OVA-specific IgE, and mucosal mast cell protease-1 (mMCP-1) were measured by ELISA. Other allergic parameters were analyzed using histology specimens, RT-PCR, and flow cytometry. Treatment with hHMSCs or hHMSC-CM significantly suppressed the frequency of anaphylactic response and rectal temperature decline, reduced diarrhea, total IgE, OVA-specific IgE, and mMCP-1. While the treatment decreased the level of Th2 cytokines, it enhanced IL-10 and TGF-β1 mRNA. Exposure to hHMSC or hHMSC-CM did not generate regulatory T cells, but reduced mast cells. The immunomodulatory effect on the Th2 cytokines was greater in hHMSC-CM than in hHMSCs. hHMSC treatment may be a promising preventive intervention against food allergy. Further studies are needed to elucidate the key substances released from hHMSC to induce immune tolerance.