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Chemiluminescence immunoassays have been widely employed for diagnosing various diseases. However, because of the extremely low intensity chemiluminescence signals, highly sensitive transducers, such as photomultiplier tubes and image sensors with cooling devices, are required to overcome this drawback. In this study, a hypersensitive photosensor was developed based on cesium lead bromide (CsPbBr3) perovskite quantum dots (QDs) with sufficient high sensitivity for chemiluminescence immunoassays. First, CsPbBr3 QDs with a highly uniform size, that is, 5 nm, were synthesized under thermodynamic control to achieve a high size confinement effect. For the fabrication of the photosensor, MoS2 nanoflakes were used as an electron transfer layer and heat-treated at an optimum temperature. Additionally, a parylene-C film was used as a passivation layer to improve the physical stability and sensitivity of the photosensor. In particular, the trap states on the CsPbBr3 QDs were reduced by the passivation layer, and the sensitivity was increased. Finally, a photosensor based on CsPbBr3 QDs was employed in chemiluminescence immunoassays for the detection of human hepatitis B surface antigen, human immunodeficiency virus antibody, and alpha-fetoprotein (AFP, a cancer biomarker). When compared with the conventionally used equipment, the photosensor was determined to be feasible for application in chemiluminescence immunoassays.

Antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleoprotein (NP) were purified from pig serum through two steps: (1) isolation of anti-NP IgG antibodies using magnetic beads with immobilized human SARS-CoV-2 NP and (2) filtration of anti-spike protein (SP) IgG antibodies using magnetic beads with immobilized human SARS-CoV SP. The enhanced specificity of the purified antibodies to the NP of SARS-CoV-2 was demonstrated using an immunoassay with anti-NP IgG antibodies after the isolation and filtration steps. The binding constants (Kd) of the purified anti-NP IgG antibodies to the NP of SARS-CoV-2 and the SP of SARS-CoV were estimated using a surface plasmon resonance biosensor (SPR). A competitive assay using the two-step purified anti-NP IgG antibodies from pig serum demonstrated (a) the detection of SARS-CoV-2 in viral fluid and (b) the discrimination of SARS-CoV-2 from SARS-CoV, MERS-CoV, and CoV strain 229E in viral fluids.

Oral squamous cell carcinoma (OSCC) frequently invades nearby bone and bone involvement determines the prognosis of patients. Growth factors, stored in the bone matrix and released during bone destruction, are known as key components in the bone-tumor interaction. However, the coordination of growth factor signals and the precise mechanism of bone destruction in oral cancer are still unclear. In the study, we investigated the differential cytokine expression profile of oral cancer cells by TGF-β treatment and the function of altered expression of cytokines on the osteoclast differentiation. We established TGFBR2-knockdown cells using small hairpin RNA. TGF-β was treated to both TGFBR2 expressing and knockdown cells and the culture supernatants were analyzed using a cytokine array kit. We found that the TGF-β inhibited IGFBP3 level and enhanced MMP9 level. We confirmed this regulation of IGFBP3 and MMP9 by TGF-β using ELISA and zymography, respectively. IGFBP3 is known as to modulate the bioavailability of IGF1, which is abundant in the bone microenvironment and regulates osteoclast differentiation. Therefore, we further analyzed the function of IGFBP3 on osteoclastogenesis. Although IGFBP3 increased the viability of murine bone marrow macrophages, the osteoclast differentiation of these cells was blocked by IGFBP3 in a dose-dependent manner. These results revealed a novel pathway for the regulation of osteoclastogenesis by oral cancer cells, which may be a new therapeutic target for osteolysis induced by oral cancer infiltrating into the bone.

One of the key challenges in biology is to understand how individual cells process information and respond to perturbations. However, most of the existing single-cell analysis methods can only provide a glimpse of cell properties at specific time points and are unable to provide cell secretion and protein analysis at single-cell resolution. To address the limits of existing methods and to accelerate discoveries from single-cell studies, we propose and experimentally demonstrate a new sensor based on bound states in the continuum to quantify exosome secretion from a single cell. Our optical sensors demonstrate high-sensitivity refractive index detection. Because of the strong overlap between the medium supporting the mode and the analytes, such an optical cavity has a figure of merit of 677 and sensitivity of 440 nm/RIU. Such results facilitate technological progress for highly conducive optical sensors for different biomedical applications.

Osteoarthritis (OA) is a common joint disease that results from the disintegration of joint cartilage and the underlying bone. Because cartilage and chondrocytes lack the ability to self-regenerate, efforts have been made to utilize stem cells to treat OA. Although various methods have been used to differentiate stem cells into functional chondrocytes, the currently available methods cannot induce stem cells to undergo differentiation into chondrocyte-like cells without inducing characteristics of hypertrophic chondrocytes, which finally lead to cartilage disintegration and calcification. Therefore, an optimized method to differentiate stem cells into chondrocytes that do not display undesired phenotypes is needed. This study focused on differentiating adipose-derived stem cells (ASCs) into functional chondrocytes using a small molecule that regulated the expression of Sox9 as a key factor in cartilage development and then explored its ability to treat OA. We selected ellipticine (ELPC), which induces chondrocyte differentiation of ASCs, using a GFP-Sox9 promoter vector screening system. An in vivo study was performed to confirm the recovery rate of cartilage regeneration with ASC differentiation into chondrocytes by ELPC in a collagenase-induced animal model of OA. Taken together, these data indicate that ellipticine induces ASCs to differentiate into mature chondrocytes without hypertrophic chondrocytes in vitro and in vivo, thus overcoming a problem encountered in previous studies. These results indicate that ELPC is a novel chondrocyte differentiation-inducing drug that shows potential as a cell therapy for OA., Osteoarthritis: Drug-induced stem cell differentiation shows therapeutic promise A novel method of generating healthy cartilage cells from stem cells could repair and regenerate joints and bones damaged by osteoarthritis. The cells that produce and maintain cartilage tissue are known as chondrocytes. Stem cell therapy could potentially re-start cartilage and bone regeneration in patients with osteoarthritis, however scientists have struggled to successfully induce stem cells to differentiate into functioning chondrocytes. A new method developed by Soyeon Lim and Ki-Chul Hwang at the Catholic Kwandong University, Gangneung, South Korea, and co-workers uses a drug called ellipticine to boost the expression of Sox-9, a key protein involved in cartilage development. The researchers found that the ellipticine-based method induced a mature, healthy chondrocyte population in cell cultures and rat models of osteoarthritis. Rats injected with these cells recovered from cartilage damage, suggesting a potential osteoarthritis therapy.

Background/aim We investigated the clinical role of the molecular targets, APEX1 and Jagged-1, and the Apex1 - Jagged-1 cascade in gastric cancer cells. Materials and methods We used 6 human gastric cancer cell lines (SNU-1, SNU-5, SNU-16, NCI-N87, KATO- III and AGS), and demonstrated the chemosensitivity of APEX1 and Jagged-1 through the MTT assay and immunoblotting. Tumor growth was assayed following cisplatin and 5-FU treatment using a xenograft model injected with KATO-III cells. Moreover, gastric tumor samples from 9 patients, divided in 2 groups according to chemotherapy response, were examined by immunocytochemical (IHC) staining, and protein expression levels were scored. Results Following APEX1 knockdown, the MTT assay revealed that the IC50 of cisplatin and 5-FU in AGS cells was decreased approximately 7% and 15%, respectively, however, their decrease in chemoresistant KATO-III cells was decreased by approximately 21% and 67% for cisplatin and 5-FU, respectively. The tumor volume of KATO-III/sicontrol mice treated with cisplatin and 5-FU was affected less, compared with KATO-III/siAPEX1 mice treated with cisplatin and 5-FU. Also, the expression levels of APEX1, Jagged-1 and CD133, assayed by IHC staining, were higher in the chemorefractory group than in the chemoresponsive group. Conclusion Jagged-1-activated signaling by APEX1 plays a role in advanced gastric cancer.

Oral squamous cell carcinoma (OSCC) frequently invades the maxillary or mandibular bone, and this bone invasion is closely associated with poor prognosis and survival. Here, we show that CCL28 functions as a negative regulator of OSCC bone invasion. CCL28 inhibited invasion and epithelial-mesenchymal transition (EMT), and its inhibition of EMT was characterized by induced E-cadherin expression and reduced nuclear localization of β-catenin in OSCC cells with detectable RUNX3 expression levels. CCL28 signaling via CCR10 increased retinoic acid receptor-β (RARβ) expression by reducing the interaction between RARα and HDAC1. In addition, CCL28 reduced RANKL production in OSCC and osteoblastic cells and blocked RANKL-induced osteoclastogenesis in osteoclast precursors. Intraperitoneally administered CCL28 inhibited tumor growth and osteolysis in mouse calvaria and tibia inoculated with OSCC cells. RARβ expression was also increased in tumor tissues. In patients with OSCC, low CCL28, CCR10, and RARβ expression levels were highly correlated with bone invasion. Patients with OSCC who had higher expression of CCL28, CCR10, or RARβ had significantly better overall survival. These findings suggest that CCL28, CCR10, and RARβ are useful markers for the prediction and treatment of OSCC bone invasion. Furthermore, CCL28 upregulation in OSCC cells or CCL28 treatment can be a therapeutic strategy for OSCC bone invasion.

Vascular occlusive disease is a chronic disease with significant morbidity and mortality. Although a variety of therapies and medications have been developed, the likelihood of disease re-emergence is high and this can be life-threatening. Based on a previous screening experiment related to vascular obstructive diseases using 34 types of essential oils, cold-pressed oil (CpO) from Citrus aurantifolia (lime) has been demonstrated to have the best effect for the inhibition of vascular smooth muscle cells (VSMCs) proliferation. The aim of the present study was to evaluate the effect of lime CpO on the pathological changes of VSMCs. To determine this, the effect of lime CpO on VSMC proliferation, a major cause of vascular disease, was investigated. To determine the safe concentration interval for toxicity of CpO during VSMC culture, a dilution of 1x10-5 was determined using Cell Counting Kit-8 assay, which was confirmed to be non-toxic using a lactate dehydrogenase assay. To examine the effect of lime CpO in cellular signaling pathways, changes in phosphorylation of both the PI3K/AKT/mTOR and extracellular signal-regulated MEK/ERK signaling pathways with serum were investigated. Furthermore, lime CpO with FBS also significantly decreased the expression levels of the cell cycle regulators cyclin D1 and proliferating cell nuclear antigen. Additionally, lime CpO with FBS significantly inhibited the sprouting of VSMCs in an ex vivo culture system. These results suggested that lime CpO inhibited the abnormal proliferation of VSMCs and can be developed as a nature-based therapeutic agent for obstructive vascular disease.

Clinical nurses have training needs related to the strategies for infection spread prevention. This study aimed to investigate the levels of importance of and performance in the various areas of care for communicable respiratory infections among clinical nurses and to determine the priority of educational needs. Hospitals in which a baseline survey could be conducted were considered, and nurses working in six hospitals that were designated as COVID-19 care centers in Korea were enrolled. The training needs for the care of patients with communicable respiratory infections were analyzed using Borich’s needs equation, and the locus for focus model. Among participants with prior COVID-19 patient care experience, according to Borich’s equation, the need score was the highest for “initial response to communicable respiratory infection”, followed by “management of aerosol-generating procedures in patients with communicable respiratory infection” and “reporting of patients with communicable respiratory infection and death of patient”. An item rated highly in both Borich’s equation and the locus for focus model among the participants with prior experience was “initial response to communicable respiratory infection”. Among participants without prior relevant experience, according to Borich’s equation, the need score was highest for “management of aerosol-generating procedures in patients with a communicable respiratory infection”, followed by “initial response to communicable respiratory infection” and “reporting of patients with communicable respiratory infection and death of patient”. None of the items were rated high in both Borich’s equation and the locus for focus model among participants without relevant prior experience.

The acute demise of stem cells following transplantation significantly compromises the efficacy of stem cell-based cell therapeutics for infarcted hearts. As the stem cells transplanted into the damaged heart are readily exposed to the hostile environment, it can be assumed that the acute death of the transplanted stem cells is also inflicted by the same environmental cues that caused massive death of the host cardiac cells. Pyroptosis, a highly inflammatory form of programmed cell death, has been added to the list of important cell death mechanisms in the damaged heart. However, unlike the well-established cell death mechanisms such as necrosis or apoptosis, the exact role and significance of pyroptosis in the acute death of transplanted stem cells have not been explored in depth. In the present study, we found that M1 macrophages mediate the pyroptosis in the ischemia/reperfusion (I/R) injured hearts and identified miRNA-762 as an important regulator of interleukin 1β production and subsequent pyroptosis. Delivery of exogenous miRNA-762 prior to transplantation significantly increased the post-transplant survival of stem cells and also significantly ameliorated cardiac fibrosis and heart functions following I/R injury. Our data strongly suggest that suppressing pyroptosis can be an effective adjuvant strategy to enhance the efficacy of stem cell-based therapeutics for diseased hearts.

Highly N-doped SiC was presented as an optimal electrode for electrochemical immunoassays with a far higher sensitivity than chemiluminescence detection. As the first step, the electrochemical properties of highly N-doped SiC, such as the double-layer capacitance (Cdl), rate constant for electron transfer (kapp) and ideal polarizable potential range (electrochemical window) were analyzed and compared with those of Au, Pt, and graphite electrodes. The highly N-doped SiC electrode was used for the quantification of oxidized 3,3′,5,5′-tetramethylbenzidine (TMB) which was widely used as chromogenic substrate for commercialized immunoassay kits. In order to enhance the sensitivity for the quantification of the oxidized TMB the chronoamperometry was applied to avoid the background current of i-V measurement. Finally, the chronoamperometry based on the highly N-doped SiC electrode was applied to commercial immunoassay kits for the medical diagnosis of the human immunodeficiency virus (HIV) and the human hepatitis B surface antigen (hHBsAg). The chronoamperometric measurement based on the highly N-doped SiC electrode was proved to detect at far lower limits in comparison with the conventional optical density measurement as well as the chemiluminescence assay based on luminol as a chemiluminescent probe.

Recent studies have shown that several upstream signaling elements of apoptosis and necroptosis are closely associated with acute injury in the heart. In our study, we observed that miR-105 was notably dysregulated in rat hearts with myocardial infarction (MI). Thus, the purpose of this study was to test the hypothesis that miR-105 participates in the regulation of RIP3/p-MLKL- and BNIP3-dependent necroptosis/apoptosis in H9c2 cells and MI rat hearts. Our results show that the RIP3/p-MLKL necroptotic pathway and BNIP3-dependent apoptosis signaling are enhanced in H9c2 cells under hypoxic conditions, whereas, compared with these pathways in the controls, those in miR-105-treated H9c2 cells are suppressed. Mechanistically, we identified miR-105 as the miRNA directly suppressing the expression of RIP3 and BNIP3, two important mediators involved in cell necroptosis and apoptosis. Furthermore, MI rat hearts injected with miR-105 had decreased infarct sizes, indicating that miR-105 is among three miRNAs that function simultaneously to suppress necroptotic/apoptotic cell death pathways and to inhibit MI-induced cardiomyocyte cell death at multiple levels. Taken together, miR-105 may constitute a new therapeutic strategy for cardioprotection in ischemic heart disease. Keywords: apoptosis, ischemic heart, microRNA-105, necroptosis, programmed cell death

Neurodegenerative disease is a brain disorder caused by the loss of structure and function of neurons that lowers the quality of human life. Apart from the limited potential for endogenous regeneration, stem cell-based therapies hold considerable promise for maintaining homeostatic tissue regeneration and enhancing plasticity. Despite many studies, there remains insufficient evidence for stem cell tracing and its correlation with endogenous neural cells in brain tissue with three-dimensional structures. Recent advancements in tissue optical clearing techniques have been developed to overcome the existing shortcomings of cross-sectional tissue analysis in thick and complex tissues. This review focuses on recent progress of stem cell treatments to improve neurodegenerative disease, and introduces tissue optical clearing techniques that can implement a three-dimensional image as a proof of concept. This review provides a more comprehensive understanding of stem cell tracing that will play an important role in evaluating therapeutic efficacy and cellular interrelationship for regeneration in neurodegenerative diseases.

Herein, we present switching-peptides for a one-step immunoassay, without the need for additional antibody treatment or washing steps to detect antigen–antibody interactions. Fluorescently labeled switching-peptides were dissociated from the immobilized antibody soon after the antigens were bound to the binding pockets. In this study, four different parts of the antibody (IgG) frame regions were chemically synthesized, and these peptides were bound to immobilized antibodies as switching-peptides. We presented the design principle of switching-peptides and used Pymol software, based on the changes in thermodynamic parameters, to study the interaction between antibodies and switching-peptides. The binding properties of switching-peptides were analyzed based on Forster resonance energy transfer between switching-peptides as well as between switching-peptides and antibodies (IgGs) isolated from different animals. The binding constants of the four switching-peptides to antibodies were estimated to be in the range of 1.48–3.29 μM. Finally, the feasibility of using switching-peptides for the quantitative one-step immunoassay was demonstrated by human hepatitis B surface antigen (hHBsAg) detection and statistical comparison of the assay results with those of conventional ELISA. The limit of detection for HBsAg was determined to be 56 ng/mL, and the dynamic range was estimated to be 136 ng/mL–33 μg/mL. These results demonstrate the feasibility of the one-step immunoassay for HBsAg.

Patients with oral squamous cell carcinoma (OSCC) bone invasion are surgically treated with bone resection, which results in severe physical and psychological damage. Here, we investigated the potential of fractalkine (CX3CL1), which is regulated by transforming growth factor (TGF-β), as a novel biomarker for correct prediction and early detection of OSCC-associated bone invasion. TGF-β knockdown and treatment with a TGF-β-neutralizing antibody decreased the level of fractalkine in the culture media of HSC-2 and YD10B OSCC cells. Treatment with a fractalkine-neutralizing antibody reduced TGF-β-stimulated invasion by HSC-2 and YD10B cells. Fractalkine treatment increased the viability, invasion, and uPA secretion of both OSCC cell lines. Furthermore, OSCC cell bone invasion was assessed following subcutaneous inoculation of wild-type or TGF-β knockdown OSCC cells in mouse calvaria. TGF-β knockdown prevented erosive bone invasion, reduced the number of osteoclasts at the tumor-bone interface, and downregulated fractalkine expression in mouse tumor tissues. Our results indicate that the production of fractalkine is stimulated by TGF-β and mediates TGF-β-induced cell invasion in several OSCC cell lines showing an erosive pattern of bone invasion. Fractalkine may be a useful predictive marker and therapeutic target for OSCC-induced bone destruction.

The present study translated the Vocal Tract Discomfort Scale (VTDS) into Korean (K-VTDS) and evaluated its reliability and validity.This was a prospective study.The VTDS was first translated into Korean and validated. One hundred and fifty-nine patients with voice disorders were divided into three different diagnostic groups (functional, structural, and neurologic voice disorder) and 131 vocally healthy adults were also included. All participants completed the K-VTDS and the Korean version of the Voice Handicap Index (K-VHI) and Korean versions of the Voice-Related Quality of Life (K-VRQOL). The internal consistency of the K-VTDS was analyzed through Cronbach's α coefficient. The VTDS score differences related to the diagnostic groups were assessed with t test and analysis of variance. We assessed the correlation between the K-VTDS, the K-VHI, and the K-VRQOL using Pearson's correlation analysis.High internal consistency and the test-retest reliability of the K-VTDS were found. The voice disorder group had significantly higher K-VTDS scores for the subscales and total scores than those in the healthy group (P0.001). The K-VTDS scores for the subscale of frequency and total scores were highest in the functional voice disorder group. We found a significant difference in frequency and total score of the K-VTDS between the functional voice disorder group and the structural voice disorder group (P0.05). We observed a strong positive correlation among the scores for the subscales of frequency and severity, and total scores in the K-VTDS. The K-VTDS showed moderate correlation with the K-VHI and the K-VRQOL.The K-VTDS is a reliable and valid instrument for voice assessment for voice disorders in Korean-speaking patients.

Background/Aims: Bone metastasis of cancer cells decreases patient survival and quality of life. Hybridization via the covalent coupling of two bioactive natural products is a useful strategy for developing more potent anticancer agents by enhancing their bioavailability and avoiding drug resistance. Methods: The in vivo activities of artemisinin-daumone hybrid 15 (ARTD) were estimated in cancer cell-inoculated mice and ovariectomized mice. The viability, migration, and invasion of cancer cells were measured via MTT, wound-healing, and transwell invasion assays. ARTD-regulated transcription factors were detected with an RT2 profiler PCR array kit and Western blotting. Osteoclastogenesis and osteoclast activity were detected with tartrate-resistant acid phosphatase staining, a pit formation assay, gelatin zymography, and a cathepsin K ELISA assay. Results: ARTD blocked cancer-associated osteolysis more potently than artemisinin in mice with intratibially inoculated breast cancer or lung cancer cells. ARTD inhibited the viability, migration, and invasion of breast and lung cancer cells in the absence or presence of transforming growth factor-β1. ARTD treatment induced the expression of tumor suppressive activating transcription factor 3 and inhibited oncogenic E2F transcription factor 1 expression at the mRNA and protein levels. ARTD inhibited receptor activator of nuclear factor kappa-B ligand-induced osteoclast formation and bone resorbing activity by reducing the secreted levels of matrix metalloproteinase-9 and cathepsin K. Furthermore, ARTD prevented estrogen deficiency-induced bone loss in ovariectomized mice. Conclusion: ARTD may be a promising candidate for inhibiting cancer-induced bone destruction. The application of ARTD may be extended to patients with chemotherapy-induced ovarian failure or postmenopausal osteoporosis.

BACKGROUND: Although low-intensity pulsed ultrasound has been reported to be potential cartilage regeneration, there still unresolved treatment due to cartilage fibrosis and degeneration by a lack of rapid and high-efficiency treatment. The purpose of this study was to investigate the effect of a combination therapy of focused acoustic force and stem cells at site for fast and efficient healing on cartilage regeneration. METHODS: Using a rat articular cartilage defects model, one million adipose tissue-derived stem cells (ASCs) were injected into the defect site, and low-intensity focused ultrasound (LOFUS) in the range of 100–600 mV was used for 20 min/day for 2 weeks. All experimental groups were sacrificed after 4 weeks in total. The gross appearance score and hematoxylin and eosin (H&E), Alcian blue, and Safranin O staining were used for measuring the chondrogenic potential. The cartilage characteristics were observed, and type II collagen, Sox 9, aggrecan, and type X collagen were stained with immunofluorescence. The results of the comprehensive analysis were calculated using the Mankin scoring method. RESULTS: The gross appearance scores of regenerated cartilage and chondrocyte-like cells in H&E images were higher in LOFUS-treated groups compared to those in negative control or ASC-treated groups. Safranin O and Alcian blue staining demonstrated that the 100 and 300 mV LOFUS groups showed greater synthesis of glycosaminoglycan and proteoglycan. The ASC + LOFUS 300 mV group showed positive regulation of type II collagen, Sox 9 and aggrecan and negative regulation of type X collagen, which indicated the occurrence of cartilage regeneration based on the Mankin score result. CONCLUSION: The combination therapy, which involved treatment with ASC and 300 mV LOFUS, quickly and effectively reduced articular cartilage defects.

As a potent and selective allosteric inhibitor of MEK, TAK-733 has been shown to exert anti-cancer effects for a wide range of cancers both in vitro and in vivo. However, its effects on inhibiting growth have never been investigated in the cardiovascular system, where regulation of abnormal vascular smooth muscle cell growth in neointimal hyperplasia is an important area of focus. Angiotensin II was used to mimic inflammatory neointimal hyperplasia in an in vitro environment, and balloon catheter-induced injury with an infusion of angiotensin II was used to generate an in vivo rat restenosis model under inflammatory conditions. TAK-733 exerted anti-proliferative and anti-migratory effects on human vascular smooth muscle cells. These multiple effects of TAK-733 were evaluated using various assays, such as cell cycle analysis and wound healing. Interestingly, TAK-733 did not induce apoptosis in smooth muscle cells but only reduced the proliferation rate; additionally, it did not affect EC viability. TAK-733 also exhibited anti-inflammatory activity, as observed by attenuated monocyte adhesion to smooth muscle cells via inhibition of ICAM1 and VCAM1 overexpression. The in vivo study demonstrated that neointimal hyperplasia after balloon injury and angiotensin II stimulation was suppressed by TAK-733, and downregulation of the inflammatory signal and enhanced re-endothelialization were observed. TAK-733 may have therapeutic potential for treating neointimal hyperplasia by attenuating smooth muscle cell proliferation, migration, and inflammation. Thus, TAK-733 could be a promising drug candidate for treating patients with restenosis., Atherosclerosis: Potential therapy based on prospective cancer drug Drug undergoing trials as a cancer treatment shows promise in tackling the narrowing of blood vessels in other diseases. The thickening of arterial walls and consequent restriction of blood flow, typical of conditions like atherosclerosis, increases the risk of heart attacks and strokes. Soyeon Lim and Ki-Chul Hwang at the Catholic Kwandong University, South Korea, and co-workers have investigated the efficacy of a prospective cancer drug called TAK-733 against atherosclerosis both in cell culture and in trials on rat models of the disease. The team hypothesized that TAK-733’s anti-cancer effects could prove useful in treating other conditions. They demonstrated that TAK-733 successfully blocked the proliferation and migration of vascular smooth muscle cells, the overexpression of which is a dominant factor in the development of atherosclerosis.

Introduction Previous occasional successes after delayed replantation suggest that the presence of viable cells may not be the only factor for successful periodontal regeneration in delayed replantation. Various other factors such as proteins or the extracellular matrix (ECM) may play a role in this process. The purpose of this study was to characterize changes in the proteome components of periodontal ligament (PDL) tissue after hypothermic preservation of the tooth. Methods Extracted teeth were divided into 4 groups: immediate sampling, sampling after 1 week of preservation at 4°C, sampling after 2 weeks of preservation at 4°C, and sampling after 1 week of dry storage at room temperature as a negative control. PDL tissues were collected from the root and stored immediately in liquid nitrogen. The tissues were subjected to 2-dimensional gel electrophoresis, and spot selection was executed. Selected spots that maintained the protein volume were then processed with matrix-assisted laser desorption and ionization time-of-flight mass spectrometry to identify the nature of the proteins. Results Thirty-five selected spots were analyzed. Sixteen spots were identified as vimentin, and 3 spots were type VI collagen. The size of the 16 vimentin spots decreased gradually with increasing storage time, from 0 to 2 weeks, and decreased rapidly after dry storage. However, only the dry storage group differed significantly from the immediately sampled group. Conclusions Vimentin was the most prominent protein followed by type VI collagen in volumetrically maintained protein spots. Although these proteins are known to be closely related with ECM integrity, the role of these proteins in delayed replantation is beyond the scope of this study. Further studies are needed to elucidate the possible role of these proteins for periodontal healing of delayed replantation.

Oral squamous cell carcinoma (OSCC) frequently invades mandibular bone, and outcomes for treatment with surgical resection are typically poor, ultimately resulting in death. Holarrhena antidysenterica L. (Apocynaceae), distributed throughout Sri Lanka and India, has been used as a folk remedy to treat various diseases. Treatment with methanol extract of H. antidysenterica bark (HABE) inhibited cell viability and BrdU incorporation and induced apoptotic cell death in Ca9-22 gingival and HSC-3 tongue SCC cells. Flow cytometric analysis indicated that HABE treatment preferentially induces apoptotic cell death via increasing the sub-G1 peak in Ca9-22 cells and cell cycle arrest at the G1 phase in HSC-3 cells. HABE treatment in the presence of zVAD-fmk, a pan-caspase inhibitor, rescued cell viabilities in both OSCC cell lines. The ratio of Bax to Bcl-2 increased with reductions in the Bcl-2 protein expression, and the activation of caspase 3 and subsequent cleavage of PARP was detected in HABE-treated Ca9-22 and HSC-3 cells. Furthermore, HABE treatment at noncytotoxic concentrations inhibited osteoclast formation in RANKL-stimulated bone marrow macrophages. Taken together, HABE possesses the inhibitory activity on the growth of OSCC cells and antiosteoclastogenic activity. Therefore, HABE may be a promising alternative and complementary agent for preventing and treating OSCC.

// Junhee Park 1, 2 , Hyun-Jeong Kim 2 , Ki Rim Kim 3 , Sun Kyoung Lee 2 , Hyungkeun Kim 2, 4 , Kwang-Kyun Park 1, 2, 4 , Won-Yoon Chung 1, 2, 4 1 Department of Dentistry, Graduate School, Yonsei University, Seoul 120-749, Republic of Korea 2 Department of Oral Biology, Oral Cancer Research Institute, and BK21 PLUS Project, Yonsei University College of Dentistry, Seoul 120-752, Republic of Korea 3 Department of Dental Hygiene, Kyungpook National University, Sangju 742-711, Korea 4 Department of Applied Life Sciences, Graduate School, Yonsei University, Seoul 120-749, Republic of Korea Correspondence to: Won-Yoon Chung, email: wychung@yuhs.ac Kwang-Kyun Park, email: biochelab@yuhs.ac Keywords: oral squamous cell carcinoma, bone invasion, runt-related transcription factor 3, transforming growth factor-β Received: October 08, 2015 Accepted: December 15, 2016 Published: December 21, 2016 ABSTRACT High recurrence and lower survival rates in patients with oral squamous cell carcinoma (OSCC) are associated with its bone invasion. We identified the oncogenic role of RUNX3 during bone invasion by OSCC. Tumor growth and the generation of osteolytic lesions were significantly inhibited in mice that were subcutaneously inoculated with RUNX3-knockdown human OSCC cells. RUNX3 knockdown enhanced TGF-β-induced growth arrest and inhibited OSCC cell migration and invasion in the absence or presence of transforming growth factor-β (TGF-β), a major growth factor abundant in the bone microenvironment. RUNX3 knockdown induced cell cycle arrest at the G1 and G2 phases and promoted G2 arrest by TGF-β in Ca9.22 OSCC cells. RUNX3 knockdown also inhibited both the basal and TGF-β-induced epithelial-to-mesenchymal transition by increasing E-cadherin expression and suppressing the nuclear translocation of β-catenin. In addition, the expression and TGF-β-mediated induction of parathyroid hormone-related protein (PTHrP), one of key osteolytic factors, was blocked in RUNX3-knockdown OSCC cells. Furthermore, treating human osteoblastic cells with conditioned medium derived from RUNX3-knockdown OSCC cells reduced the receptor activator of nuclear factor-kappaB ligand (RANKL)/osteoprotegerin ratio compared with treatment with conditioned medium from RUNX3-expressing cells. These findings indicate that RUNX3 expression in OSCC cells contributes to their bone invasion and the resulting osteolysis by inducing their malignant behaviors and production of osteolytic factors. RUNX3 alone or in combination with TGF-β and PTHrP may be a useful predictive biomarker and therapeutic target for bone invasion by oral cancer.

Tonsillectomy is the most common type of surgical procedure performed in preschool children. Due to short period of hospitalization, mothers are expected to manage their children's care at home. However, they are rarely provided with sufficient information about postoperative management. This study aims to determine the effectiveness of providing caregivers with information on tonsillectomy care by smartphone text messaging in increasing their mothers' knowledge, reducing the anxiety, and improving the sick-role behavior of pediatric tonsillectomy patients.A sample of 61 pediatric patients and their mothers was recruited. Participants were randomly assigned into either the experimental group (n = 27) or the control group (n = 34). The control group was given information about the tonsillectomy by conventional textual and verbal means, whereas the experimental group received the same information in the form of 10 text messages during the period from hospitalization to their first follow-up visits.Results of mixed design, two-way analysis of variance indicated significant interaction effects between time points and groups for mothers' knowledge (F = 4.26, p = 0.043) and children's anxiety (F = 3.32; p = 0.037). Thus, the results do support the effectiveness of tonsillectomy education using smartphone text messaging in increasing mothers' knowledge and reducing children's anxiety.These results can be applied to preoperative and postoperative interventions for children not only for tonsillectomy but also for many other operations. The development of various educational programs using smartphone text messaging for postoperative patient management would also be valuable.

Background/aim Colorectal cancer (CRC) is one of the most common in the world and its prevalence is rapidly increasing. Jagged-1-activated Notch signaling by apurinic/apyrimidinic endodeoxyribonuclease 1 (APEX1) promotes CRC, and high expression of Jagged-1 is associated with poor prognosis. However, its clinical implication is unknown. The aim of this study was to investigate the clinical role of Jagged-1-activated Notch signaling by APEX1. Materials and methods The 5-dimethylthiazol-2-yl) 2, 5-diphenyltetrazolium bromide (MTT) assay was used to evaluate the anti-cancer efficacy of 5-fluorouracil (5-FU), oxaliplatin, and irinotecan. Tissue from CRC patients was analyzed to assess the clinical specificity of Jagged-1 activated by APEX1. Results The half-maximal inhibitory concentration (IC50) in cells co-expressing APEX1 and Jagged-1 cells was higher than that in cells expressing only APEX1. These results indicated that the simultaneous expression of APEX1 and Jagged-1 might be associated with chemoresistance toward 5-FU, oxaliplatin, and irinotecan. Analysis of tissue from CRC patients revealed that high expression of Jagged-1 was associated with a statistically significantly low response to chemotherapy. Conclusion Overexpression of Jagged-1 by APEX1 might serve as a predictor of response to chemotherapy and of poor prognosis, and moreover may be a therapeutic target for chemotherapy of advanced CRC.

ABO genotyping is a molecular diagnostic technique important for transfusion and transplantation in medicine, and human identification in forensic science. Because ABO genotyping are labor intensive and time consuming, the genotyping cannot be firstly used to resolve the serological ABO discrepancy in blood bank. For rapid one-step ABO genotyping, we developed direct, real-time, allele-specific polymerase chain reaction (PCR), and melting curve analysis (DRAM assay) without DNA preparation. In DRAM assay, we used a special PCR buffer for direct PCR, a rapid RBC lysis buffer, white blood cells as template without DNA preparation, allele-specific primers for discriminating three ABO alleles (261G/del, 796C/A, and 803G/C), and melting curve analysis as a detection method. There was 100% concordance among the results of ABO genotyping by the DRAM assay, serologic typing, PCR–RFLP and PCR-direct sequencing of 96 venous blood samples. We were able to reduce the number of manual steps to three and the hands-on time to 12 min, compared to seven steps and approximately 40 min for conventional ABO genotyping using allele-specific PCR with purified DNA and agarose gel electrophoresis. We have established and validated the DRAM assay for rapid and reliable one-step ABO genotyping in a closed system. The DRAM assay with an appropriate number of allele-specific primers could help in resolving ABO discrepancies and should be valuable in clinical laboratory and blood bank.

Previous studies have reported that Hedyotis diffusa Willdenow extract shows various biological activities on cerebropathia, such as neuroprotection and short-term memory enhancement. However, there has been a lack of studies on the inhibitory activity on neurodegenerative diseases such as Alzheimer&rsquo, s disease (AD) through enzyme assays of H. diffusa. Therefore, H. diffusa extract and fractions were evaluated for their inhibitory effects through assays of enzymes related to AD, including acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), and &beta, site amyloid precursor protein cleaving enzyme 1 (BACE1), and on the formation of advanced glycation end-product (AGE). In this study, ten bioactive compounds, including nine iridoid glycosides 1&ndash, 9 and one flavonol glycoside 10, were isolated from the ethyl acetate and n-butanol fractions of H. diffusa using a bioassay-guided approach. Compound 10 was the strongest inhibitor of cholinesterase, BACE1, and the formation of AGEs of all isolated compounds, while compound 5 had the lowest inhibitory activity. Compounds 3, 6, and 9 exhibited better inhibitory activity than other compounds on AChE, and two pairs of diastereomeric iridoid glycoside structures (compounds 4, 8, and 6, 7) showed higher inhibitory activity than others on BChE. In the BACE1 inhibitory assay, compounds 1&ndash, 3 were good inhibitors, and compound 10 showed higher inhibitory activity than quercetin, the positive control. Moreover, compounds 1 and 3 were stronger inhibitors of the formation of AGE than aminoguanidine (AMG), the positive control. In conclusion, this study is significant since it demonstrated that the potential inhibitory activity of H. diffusa on enzymes related to AD and showed the potential use for further study as a natural medicine for AD treatment on the basis of the bioactive components isolated from H. diffusa.

Low-frequency driving of a liquid crystal display (LCD) panel to minimise power consumption has drawn much attention recently. In case an LCD panel is driven by a low-frequency fringe field, image flickering phenomenon is detected by the naked human eye when the sign of the applied electric field is reversed. We investigated the effect of electrode spacing on the image flickering phenomenon induced by the flexoelectric effect in a fringe-field-switching (FFS) liquid crystal cell. We found that the image flicker in an FFS cell under low-frequency driving can be eliminated in a simple manner by changing the electrode spacing.

We previously published a liquid crystal alignment method using an ultraviolet (UV)-curable polymer for the near-zero pretilt angle. We have shown that the pretilt angle can be lowered by curing a reactive mesogen in the alignment layer while applying a vertical electric field. In this study, we analyzed the mechanism behind these observations. Using X-ray photoelectron spectroscopy, we confirmed that the C 1s and O 1s peaks show neither a wavelength shift nor an intensity change by UV curing, regardless of application of an electric field. We also confirmed the characteristics of the flattened surface after UV curing under an applied electric field, using atomic force microscopy. We found by atomic force microscopy that the decrease in pretilt angle is related to the flattened surface in a homogeneously aligned LC cell fabricated with the field-induced UV-alignment method.

Objective Hearing loss is a major side effect of cisplatin chemotherapy. Although cell death in cisplatin-induced ototoxicity is primarily caused by apoptosis, the exact mechanism behind the ototoxic effects of cisplatin is not fully understood. Autophagy is generally known as a pro-survival mechanism that protects cells under starvation or stress conditions. However, recent research has reported that autophagy plays a functional role in cell death also. This study aimed to investigate the role of autophagy in cisplatin-induced ototoxicity in an auditory cell line. Methods Cultured HEI-OC1 cells were exposed to 30 μM cisplatin for 48 h, and cell viability was tested using MTT assays. To evaluate whether autophagy serves to cell death after cisplatin exposure, western blotting and immunofluorescence staining for LC3-II were performed. Markers of two autophagy-related pathways, mTOR and class III PI3K, were also investigated. Results The formation of the autophagic protein LC3-II in response to 30 μM cisplatin increased with time. The early upregulation of autophagy exerted cytoprotective activity via the class III PI3K pathway. But later increase in autophagy induced cell death by suppressing the mTOR pathway. Conclusion Our results prove that autophagy could induce cell death during cisplatin-induced ototoxicity, and modulating the autophagic pathway might be another strategy against cisplatin-induced ototoxicity.

Objective Remodeling of alveolar bone is controlled by osteoclast-mediated bone resorption and osteoblast-induced bone formation. LPS of Porphyromonas gingivalis, a major causative agent of periodontitis, produces proinflammatory cytokines in host immune cells, which thereby triggers osteoclastogenesis and leads to alveolar bone resorption. We investigated the anti-periodontitis potential of Platycarya strobilacea leaf extract (PLE), which is used as a traditional medicine in Asian countries. Design TNF-α levels in cell culture media were measured using a commercially available enzyme-linked immunosorbent assay kit. Osteoclast differentiation was observed by tartrate-resistant acid phosphatase staining, and the expression levels of osteoclastogenic genes were measured by quantitative real-time PCR. Bone-resorbing activity was confirmed by the resorption pit formation, gelatin zymographic, and the cathepsin K activity assays. Osteogenic differentiation was confirmed with an ALP activity assay and alizarin red S staining. Results PLE treatment inhibited the production of TNF-α in P. gingivalis LPS-stimulated RAW264.7 macrophages. In bone marrow-derived macrophages serving as osteoclast precursors, PLE treatment blocked RANKL-induced osteoclastogenesis and gene expression levels of the osteoclastogenic transcription factor NFATc1, DC-STAMP for osteoclast fusion, and cathepsin K for osteoclast activity. In addition, PLE treatment reduced the formation of resorption pits and the secretion of MMP 9 and cathepsin K from the differentiated osteoclasts. Furthermore, PLE treatment induced osteogenesis by increasing ALP activity and calcium content in preosteoblastic cells. Conclusion PLE inhibits P. gingivalis LPS-induced TNF-α production and bone resorption and induces bone formation. PLE may be a beneficial agent to promote oral health by inhibiting periodontitis-induced alveolar bone loss.

In this work, the gram-negative bacterium Escherichia coli strain BL21(DE3) (with lipopolysaccharide (LPS) in its outer membrane) and its modified ClearColi™ strain (lacking LPS) were used for the separation of anti-LPS antibodies from human serum by the following steps: (1) binding of the serum proteins to BL21(DE3); (2) dissociation of the bound proteins (including anti-LPS antibodies) from BL21(DE3) with acid; (3) filtering of the dissociated proteins using ClearColi to remove unwanted proteins; and (4) separation of the antibody fraction by protein-A column chromatography. The binding properties of the separated antibodies were analyzed by fluorescence-activated cell sorting to confirm their selective binding to LPS on the outer membrane of BL21(DE3), and by thermophoretic immunoassay to estimate their dissociation constant. The in vitro applicability of the separated anti-LPS antibodies was demonstrated through a fluorescence assay of BL21(DE3), after immobilizing the antibodies onto a modified microplate surface. The electrochemical detection of BL21(DE3) was also achieved after immobilizing the anti-LPS antibodies onto a gold electrode.

Background Adipose tissue-derived mesenchymal stem cells (AT-MSCs) offer potential as a therapeutic option for chronic discogenic low back pain (LBP) because of their immunomodulatory functions and capacity for cartilage differentiation. The goal of this study was to assess the safety and tolerability of a single intradiscal implantation of combined AT-MSCs and hyaluronic acid (HA) derivative in patients with chronic discogenic LBP. Methods We performed a single-arm phase I clinical trial with a 12-month follow-up and enrolled 10 eligible chronic LBP patients. Chronic LBP had lasted for more than 3 months with a minimum intensity of 4/10 on a visual analogue scale (VAS) and disability level ≥ 30% on the Oswestry Disability Index (ODI). The 10 patients underwent a single intradiscal injection of combined HA derivative and AT-MSCs at a dose of 2 × 107 cells/disc (n = 5) or 4 × 107 cells/disc (n = 5). Safety and treatment outcomes were evaluated by assessing VAS, ODI, Short Form-36 (SF-36), and imaging (lumbar spine X-ray imaging and MRI) at regular intervals over 1 year. Results No patients were lost at any point during the 1-year clinical study. We observed no procedure or stem cell-related adverse events or serious adverse events during the 1-year follow-up period. VAS, ODI, and SF-36 scores significantly improved in both groups receiving both low (cases 2, 4, and 5) and high (cases 7, 8, and 9) cell doses, and did not differ significantly between the two groups. Among six patients who achieved significant improvement in VAS, ODI, and SF-36, three patients (cases 4, 8, and 9) were determined to have increased water content based on an increased apparent diffusion coefficient on diffusion MRI. Conclusions Combined implantation of AT-MSCs and HA derivative in chronic discogenic LBP is safe and tolerable. However, the efficacy of combined AT-MSCs and HA should be investigated in a randomized controlled trial in a larger population. Trial registration ClinicalTrials.gov NCT02338271 . Registered 7 January 2015.

Non-small cell lung cancer (NSCLC) frequently metastasizes to bone, which is associated with significant morbidity and a dismal prognosis. RUNX3 functions as a tumour suppressor in lung cancer and loss of expression occurs more frequently in invasive lung adenocarcinoma than in pre-invasive lesions. Here, we show that RUNX3 and RUNX3-regulated chemokines are linked to NSCLC-mediated bone resorption. Notably, the receptor activator of nuclear factor-κB ligand (RANKL)/osteoprotegerin (OPG) ratio, an index of osteoclastogenic stimulation, was significantly increased in human osteoblastic cells treated with conditioned media derived from RUNX3-knockdown NSCLC cells. We aimed to identify RUNX3-regulated factors that modify the osteoblastic RANKL/OPG ratio and found that RUNX3 knockdown led to CCL5 up-regulation and down-regulation of CCL19 and CXCL11 in NSCLC cells. Tumour size was noticeably increased and more severe osteolytic lesions were induced in the calvaria and tibiae of mice that received RUNX3-knockdown cells. In response to RUNX3 knockdown, serum and tissue levels of CCL5 increased, whereas CCL19 and CXCL11 decreased. Furthermore, CCL5 increased the proliferation, migration, and invasion of lung cancer cells in a dose-dependent manner; however, CCL19 and CXCL11 did not show any significant effects. The RANKL/OPG ratio in osteoblastic cells was increased by CCL5 but reduced by CCL19 and CXCL11. CCL5 promoted osteoclast differentiation, but CCL19 and CXCL11 reduced osteoclastogenesis in RANKL-treated bone marrow macrophages. These findings suggest that RUNX3 and related chemokines are useful markers for the prediction and/or treatment of NSCLC-induced bone destruction.

Under distinct pathological heart conditions, the expression of a single miRNA can display completely opposite patterns. However, the mechanism underlying the bidirectional regulation of a single miRNA and the clinical implications of this regulation remain largely unknown. To address this issue, we examined the regulation of miR-1, one of the most abundant miRNAs in the heart, during cardiac hypertrophy and ischemia/reperfusion (I/R). Our data indicated that different magnitudes and chronicities of ROS levels in cardiomyocytes resulted in differential expression of miR-1, subsequently altering the expression of myocardin. In animal models, the administration of a miR-1 mimic attenuated cardiac hypertrophy by suppressing the transverse aortic constriction-induced increase in myocardin expression, whereas the administration of anti-miR-1 ameliorated I/R-induced cardiac apoptosis and deterioration of heart function. Our findings indicated that a pathologic stimulus such as ROS can bidirectionally alter the expression of miRNA to contribute to the development of pathological conditions exhibiting distinct phenotypes and that the meticulous adjustment of the pathological miRNA levels is required to improve clinical outcomes.

Dynamin-related protein-1 (Drp1) plays a critical role in mitochondrial fission which allows cell proliferation and Mdivi-1, a specific small molecule Drp1 inhibitor, is revealed to attenuate proliferation. However, few molecular mechanisms-related to Drp1 under stimulus for restenosis or atherosclerosis have been investigated in vascular smooth muscle cells (vSMCs). Therefore, we hypothesized that Drp1 inhibition can prevent vascular restenosis and investigated its regulatory mechanism. Angiotensin II (Ang II) or hydrogen peroxide (H2 O2 )-induced proliferation and migration in SMCs were attenuated by down-regulation of Drp1 Ser 616 phosphorylation, which was demonstrated by in vitro assays for migration and proliferation. Excessive amounts of ROS production and changes in mitochondrial membrane potential were prevented by Drp1 inhibition under Ang II and H2 O2 . Under the Ang II stimulation, activated Drp1 interacted with PKCδ and then activated MEK1/2-ERK1/2 signaling cascade and MMP2, but not MMP9. Furthermore, in ex vivo aortic ring assay, inhibition of the Drp1 had significant anti-proliferative and -migration effects for vSMCs. A formation of vascular neointima in response to a rat carotid artery balloon injury was prevented by Drp1 inhibition, which shows a beneficial effect of Drp1 regulation in the pathologic vascular condition. Drp1-mediated SMC proliferation and migration can be prevented by mitochondrial division inhibitor (Mdivi-1) in in vitro, ex vivo and in vivo, and these results suggest the possibility that Drp1 can be a new therapeutic target for restenosis or atherosclerosis.

In case an LCD panel is driven by a low-frequency fringe field, image flickering phenomenon is detected by the naked human eye. We investigated the dependence on the electrode spacing of the image flickering phenomenon induced by the flexoelectric effect in an FFS LCD panel. We found that the image flicker in an FFS LCD panel under low-frequency driving can be eliminated in a simple manner by optimizing the electrode spacing.