Korea Research Institute of Bioscience and Biotechnology
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Ultrasensitive stress biomarker detection using polypyrrole nanotube coupled to a field-effect transistor
No full textStress biomarkers such as hormones and neurotransmitters in bodily fluids can indicate an individual's physical and mental state, as well as influence their quality of life and health. Thus, sensitive and rapid detection of stress biomarkers (e.g., cortisol) is important for management of various diseases with harmful symptoms, including post-traumatic stress disorder and depression. Here, we describe rapid and sensitive cortisol detection based on a conducting polymer (CP) nanotube (NT) field-effect transistor (FET) platform. The synthesized polypyrrole (PPy) NT was functionalized with the cortisol antibody immunoglobulin G (IgG) for the sensitive and specific detection of cortisol hormone. The anti-cortisol IgG was covalently attached to a basal plane of PPy NT through an amide bond between the carboxyl group of PPy NT and the amino group of anti-cortisol IgG. The resulting field-effect transistor-type biosensor was utilized to evaluate various cortisol concentrations. Cortisol was sensitively measured to a detection limit of 2.7 x 10-10 M (100 pg/mL), with a dynamic range of 2.7 x 10-10 to 10-7 M; it exhibited rapid responses (<5 s). We believe that our approach can serve as an alternative to time-consuming and labor-intensive health questionnaires; it can also be used for diagnosis of underlying stress-related disorders. ⓒ 2020 by the authors.
Cytotoxic activities and molecular mechanisms of the beauvericin and beauvericin G1 microbial products Against melanoma cells
No full textMelanoma is the most serious type of skin cancer and remains highly drug-resistant. Therefore, the discovery of novel effective agents against melanoma is in high demand. Herein, we investigated the cytotoxic activities in melanoma cells and underlying molecular mechanisms of beauvericin (BEA) and its analogue beauvericin G1 (BEA G1), which are cyclohexadepsipeptides isolated from fungi. BEA and BEA G1 significantly suppressed the growth, clonogenicity, migration, and invasion of A375SM human melanoma cells and promoted caspase-dependent apoptosis through upregulation of death receptors, as well as modulation of pro- and anti-apoptotic Bcl-2 family members. Furthermore, the effects of BEA and BEA G1 were associated with the suppression of multiple molecular targets that play crucial roles in melanoma oncogenesis, including ERK, JNK, p38, NF-κB, STAT3, and MITF. Notably, the cytotoxic efficacy of BEA G1 against A375SM cells was stronger than that of BEA. These findings suggest that BEA and BEA G1 can be further investigated as potent cytotoxic natural compounds for the suppression of melanoma progression. ⓒ 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Exposure of triclosan in porcine oocyte leads to superoxide production and mitochondrial-mediated apoptosis during in vitro maturation
No full textWhile triclosan (TCS) exerts detrimental effects on female reproduction, the effect of TCSderived toxins on porcine oocytes during in vitro maturation (IVM) is unclear. This study investigated the effects of TCS on mitochondrion-derived reactive oxygen species (ROS) production and apoptosis pathways during porcine oocyte maturation. Porcine oocytes were treated with TCS (1, 10, and 100 μM) and triphenylphosphonium chloride (Mito-TEMPO; 0.1 μM), and matured cumulus oocyte complexes (COCs) were stained with orcein, dichlorofluorescein diacetate (DCFDA), and Mito-SOX. Proteins and mRNA levels of factors related to cumulus expansion and mitochondrion-mediated apoptosis and antioxidant enzymes were analyzed by western blotting and reverse-transcription polymerase chain reaction (RT-PCR), respectively. Meiotic maturation and cumulus cell expansion significantly decreased for COCs after TCS treatment along with an increase in mitochondrial superoxide levels at 44 h of IVM. Further, mitochondrion-related antioxidant enzymes and apoptosis markers were significantly elevated in porcine COCs following TCS-mediated oxidative damage. The protective effect of Mito-TEMPO as a specific superoxide scavenger from TCS toxin improved the maturation capacity of porcine COCs. Mito-TEMPO downregulated the mitochondrial apoptosis of TCS-exposed porcine COCs by reducing superoxide level. In conclusion, our data demonstrate that TCS mediates toxicity during porcine oocyte maturation through superoxide production and mitochondrion-mediated apoptosis.
Sex-biased gene expression in antennae of Drosophila suzukii
No full textDrosophila suzukii differs from other members of the genus Drosophila in its host preference and oviposition behavior. The flies are attracted to ripening fruits, and females have a serrated ovipositor enabling eggs to be laid inside the fruit. In addition to its huge economic impact, its unique chemoecological, morphological, and physiological characteristics have garnered considerable research interests. In this study, we analyzed D. suzukii antennal transcriptomes to identify sex-biased genes by comparison of differential gene expressions between male antennae (MA) and female antennae (FA). Among 13,583 total genes of the fly genome, 11,787 genes were expressed in either MA or FA. There are only 132 genes (9 in MA, 7 in FA, and 116 in both, FPKM >1) were expressed in antennae exclusively, and 2,570 genes (9 in MA, 0 in FA, and 2,561 in both) were enriched in antennae containing 185 and 113 sex-biased genes in MA and FA, respectively. Interestingly, many immune-related genes were highly expressed in MA, whereas several chemosensory genes were at high rank in FA. We identified 27 sex-biased chemosensory genes including odorant and gustatory receptors, odorant-binding proteins, chemosensory proteins, ionotropic receptors, and cytochrome P450s, and validated the gene expressions using quantitative real-time PCR. The highly expressed sex-biased genes in antennae are likely involved in the fly specific mating, host-finding behaviors, or sex-specific functions. The molecular results demonstrated here will facilitate to find the unique chemoreception of D. suzukii, as well as on the development of new management strategies for this pest.
New acetohydrazides incorporating 2-oxoindoline and 4-oxoquinazoline: synthesis and evaluation of cytotoxicity and caspase activation activity
No full textIn our search for new small molecules activating procaspase-3, we have designed and synthesized a series of new acetohydrazides incorporating both 2-oxoindoline and 4-oxoquinazoline scaffolds. Biological evaluation showed that a number of these acetohydrazides were comparably or even more cytotoxic against three human cancer cell lines (SW620, colon cancer; PC-3, prostate cancer; NCI？H23, lung cancer) in comparison to PAC-1, a first procaspase-3 activating compound, which was used as a positive control. One of those new compounds, 2-(6-chloro-4-oxoquinazolin-3(4H)-yl)-N′-[(3Z)-5-methyl-2-oxo-1,2-dihydro-3H-indol-3-ylidene]acetohydrazide activated the caspase-3 activity in U937 human lymphoma cells by 5-fold higher than the untreated control. Three of the new compounds significantly induced necrosis and apoptosis in U937 cells.
Single photomask lithography for shape modulation of micropatterns
No full textPhotolithography has been used to fabricate various micropatterns; however, recent efforts have focused on the scaling-down process to reduce the feature sizes of the desired patterns and not redefining existing micropatterns. Here, we describe single-mask photolithography method for the shape modulation of designed patterns with a single-mask that utilizes the intrinsic properties of a monolithic photoresist. Our novel approach can achieve various micropatterns with different open areas by adjusting a few fabrication parameters. To create the modulated micropatterns, the material and physical parameters (e.g., soft-bake temperature and exposure dose) were altered and successfully produced modulated shapes. This approach can produce well-controlled micropatterns with desired shapes over a large area with high throughput. Furthermore, our new approach enables access to numerous varieties of micropatterns and can be expandable to two dimensional (2D) and thre dimensional (3D) multiscale architectures.
Altered mitochondrial functions and morphologies in epithelial cells are associated with pathogenesis of chronic rhinosinusitis with nasal polyps
No full textPurpose: Chronic rhinosinusitis with nasal polyps (CRSwNP) is a complex inflammatory disease of the nasal and paranasal sinus mucosa. The disease is associated with mitochondrial dysfunction, structural changes in the mitochondria, and reactive oxygen species (ROS) generation. This study investigated whether there are functional and morphological changes in the mitochondria in the epithelial cells of nasal polyps (NPs) and Staphylococcus aureus enterotoxin B (SEB)-stimulated nasal epithelial cells.
Methods: In all, 30 patients with CRSwNP and 15 healthy subjects were enrolled. Mitochondrial ROS (mtROS) and changes in mitochondrial functions and structures were investigated in the uncinate tissue (UT) of healthy controls, the UT or NPs of CRSwNP patients, and human nasal epithelial cells with or without SEB stimulation.
Results: Oxidative phosphorylation complexes showed various responses following SEB stimulation in the nasal epithelial cells, and their expressions were significantly higher in the NPs of patients with CRSwNP than in the UT of controls. Generation of mtROS was increased following SEB exposure in nasal epithelial cells and was reduced by pretreatment with MitoTEMPO, which is used as an mtROS scavenger. In the tissues, mtROS was significantly increased in the NPs of CRSwNP patients compared to the UT of controls or CRSwNP patients. The expressions of fusion- and fission-related molecules were also significantly higher in SEB-exposed nasal epithelial cells than in non-exposed cells. In tissues, the expression of fission (fission mediator protein 1)- and fusion (membrane and mitofusin-1, and optic atrophy protein 1)-related molecules was significantly higher in the NPs of CRSwNP patients than in UT of controls or CRSwNP patients. Transmission electron microscopy revealed elongated mitochondria in SEB-exposed nasal epithelial cells and epithelial cells of NPs.
Conclusions: Production of mtROS, disrupted mitochondrial function, and structural changes in nasal epithelial cells might be involved in the pathogenesis of CRSwNP.
Integrin-mediated adhesions in regulation of cellular senescence
No full textBioinformatic and functional data link integrin-mediated cell adhesion to cellular senescence; however, the significance of and molecular mechanisms behind these connections are unknown. We now report that the focal adhesion？localized βPAK-interacting exchange factor (βPIX)？G protein？coupled receptor kinase interacting protein (GIT) complex controls cellular senescence in vitro and in vivo. βPIX and GIT levels decline with age. βPIX knockdown induces cellular senescence, which was prevented by reexpression. Loss of βPIX induced calpain cleavage of the endocytic adapter amphiphysin 1 to suppress clathrin-mediated endocytosis (CME); direct competition of GIT1/2 for the calpain-binding site on paxillin mediates this effect. Decreased CME and thus integrin endocytosis induced abnormal integrin signaling, with elevated reactive oxygen species production. Blocking integrin signaling inhibited senescence in human fibroblasts and mouse lungs in vivo. These results reveal a central role for integrin signaling in cellular senescence, potentially identifying a new therapeutic direction.
Demonstration of arithmetic calculations by DNA tile-based algorithmic self-assembly
No full textOwing to its high information density, energy efficiency, and massive parallelism, DNA computing has undergone several advances and made significant contributions to nanotechnology. Notably, arithmetic calculations implemented by multiple logic gates such as adders and subtractors have received much attention because of their well-established logic algorithms and feasibility of experimental implementation. Although small molecules have been used to implement these computations, a DNA tile-based calculator has been rarely addressed owing to complexity of rule design and experimental challenges for direct verification. Here, we construct a DNA-based calculator with three types of building blocks (propagator, connector, and solution tiles) to perform addition and subtraction operations through algorithmic self-assembly. An atomic force microscope is used to verify the solutions. Our method provides a potential platform for the construction of various types of DNA algorithmic crystals (such as flip-flops, encoders, and multiplexers) by embedding multiple logic gate operations in the DNA base sequences.
GATA3 induces the upregulation of UCP-1 by directly binding to PGC-1α during adipose tissue browning
No full textObjective: Obesity is recognized as the cause of multiple metabolic diseases and is rapidly increasing worldwide. As obesity is due to an imbalance in energy homeostasis, the promotion of energy consumption through browning of white adipose tissue (WAT) has emerged as a promising therapeutic strategy to counter the obesity epidemic. However, the molecular mechanisms of the browning process are not well understood. In this study, we investigated the effects of the GATA family of transcription factors on the browning process.
Methods: We used qPCR to analyze the expression of GATA family members during WAT browning. In order to investigate the function of GATA3 in the browning process, we used the lentivirus system for the ectopic expression and knockdown of GATA3. Western blot and real-time qPCR analyses revealed the regulation of thermogenic genes upon ectopic expression and knockdown of GATA3. Luciferase reporter assays, co-immunoprecipitation, and chromatin immunoprecipitation were performed to demonstrate that GATA3 interacts with proliferator-activated receptor-γ co-activator-1α (PGC-1α) to regulate the promoter activity of uncoupling protein-1 (UCP-1). Enhanced energy expenditure by GATA3 was confirmed using oxygen consumption assays, and the mitochondrial content was assessed using MitoTracker. Furthermore, we examined the in vivo effects of lentiviral GATA3 overexpression and knockdown in inguinal adipose tissue of mice.
Results: Gata3 expression levels were significantly elevated in the inguinal adipose tissue of mice exposed to cold conditions. Ectopic expression of GATA3 enhanced the expression of UCP-1 and thermogenic genes upon treatment with norepinephrine whereas GATA3 knockdown had the opposite effect. Luciferase reporter assays using the UCP-1 promoter region showed that UCP-1 expression was increased in a dose-dependent manner by GATA3 regardless of norepinephrine treatment. GATA3 was found to directly bind to the promoter region of UCP-1. Furthermore, our results indicated that GATA3 interacts with the transcriptional coactivator PGC-1α to increase the expression of UCP-1. Taken together, we demonstrate that GATA3 has an important role in enhancing energy expenditure by increasing the expression of thermogenic genes both in vitro and in vivo.
Conclusion: GATA3 may represent a promising target for the prevention and treatment of obesity by regulating thermogenic capacity.