Bacillus genus produces several secondary metabolites with biocontrol ability against various phytopathogens. Bacillus velezensis AK-0 (AK-0), an antagonistic strain isolated from Korean ginseng ...rhizospheric soil, was found to exhibit antagonistic activity against several phytopathogens. To further display the genetic mechanism of the biocontrol traits of AK-0, we report the complete genome sequence of AK-0 and compared it with complete genome sequences of closely related strains. We report the biocontrol activity of AK-0 against apple bitter rot caused by Colletotrichum gloeosporioides, which could lead to commercialization of this strain as a microbial biopesticide in Korea. To retain its biocontrol efficacy for a longer period, AK-0 has been formulated with ingredients for commercialization, named AK-0 product formulation (AK-0PF). AK-0PF played a role in the suppression of the mycelial growth of the fungicide-resistant pathogen C. gloeosporioides YCHH4 at a greater level than the non-treated control. Moreover, AK-0PF exhibited greater disease suppression of bitter rot in matured under field conditions. Here, we report the complete genome sequence of the AK-0 strain, which has a 3,969,429 bp circular chromosome with 3808 genes and a G+C content of 46.5%. The genome sequence of AK-0 provides a greater understanding of the Bacillus species, which displays biocontrol activity via secondary metabolites. The genome has eight potential secondary metabolite biosynthetic clusters, among which, ituD and bacD genes were expressed at a greater level than other genes. This work provides a better understanding of the strain AK-0, as an effective biocontrol agent (BCA) against phytopathogens, including bitter rot in apple.
Background and Objectives
Botulinum neurotoxin (BoNT) is a substance used to treat chronic sialorrhea, muscle dystonia, and is used in cosmetic applications. Measuring the potency of BoNT is crucial ...because it acts even with a small amount. However, the current methods for measuring the potency of BoNT involve using two‐dimensional neuroblastoma cell line‐based methods. In this study, we aimed to develop a new method to measure the potency of BoNT using a three‐dimensional organoid culture system.
Materials and Method
We established the optimal conditions for coculturing N2a neuronal cells with murine salivary gland organoids (SGOs). After determining the appropriate chemical concentrations, we treated the SGOs cocultured with N2a cells with BoNT type A (BoNT/A). We confirmed the expression of salivary gland‐related genes and proteins using real‐time polymerase chain reaction (PCR) and immunofluorescence staining.
Results
The SGOs cocultured with N2a cells showed that the dendrites or axons of neuronal cells were in contact with the outermost layer of the SGOs. When we applied acetylcholine and neostigmine to the coculture systems, the mRNA expression of Aqp5 and Bhlha15, associated with salivary gland secretory cells, increased. However, this effect was reversed when BoNT/A was applied, as confirmed through real‐time PCR.
Conclusion
We found that the coculture system of SGOs and N2a neuronal cells can potentially serve as a potency testing platform for BoNT.
Level of Evidence
NA Laryngoscope, 134:2697–2704, 2024
This study introduces a salivary gland organoid‐N2a coculture system enabling to evaluate the potency of botulinum neurotoxin type A.
Previously, we showed that exposure of human normal and cancer cells to a 6 mT, 60 Hz gradient electromagnetic field (EMF) induced genotoxicity. Here, we investigated the cellular effects of a ...uniform EMF. Single or repetitive exposure to a 6 mT, 60 Hz uniform EMF neither induced DNA damage nor affected cell viability in HeLa and primary IMR-90 fibroblasts. However, continuous exposure of these cells to an EMF promoted cell proliferation. Cell viability increased 24.4% for HeLa and 15.2% for IMR-90 cells after a total 168 h exposure by subculture. This increase in cell proliferation was directly correlated with EMF strength and exposure time. When further incubated without EMF, cell proliferation slowed down to that of unexposed cells, suggesting that the proliferative effect is reversible. The expression of cell cycle markers increased in cells continuously exposed to an EMF as expected, but the distribution of cells in each stage of the cell cycle did not change. Notably, intracellular reactive oxygen species levels decreased and phosphorylation of Akt and Erk1/2 increased in cells exposed to an EMF, suggesting that reduced levels of intracellular reactive oxygen species play a role in increased proliferation. These results demonstrate that EMF uniformity at an extremely low frequency (ELF) is an important factor in the cellular effects of ELF-EMF.
Salivary glands that produce and secrete saliva, which is essential for lubrication, digestion, immunity, and oral homeostasis, consist of diverse cells. The long-term maintenance of diverse salivary ...gland cells in organoids remains problematic. Here, we establish long-term murine and human salivary gland organoid cultures. Murine and human salivary gland organoids express gland-specific genes and proteins of acinar, myoepithelial, and duct cells, and exhibit gland functions when stimulated with neurotransmitters. Furthermore, human salivary gland organoids are established from isolated basal or luminal cells, retaining their characteristics. Single-cell RNA sequencing also indicates that human salivary gland organoids contain heterogeneous cell types and replicate glandular diversity. Our protocol also enables the generation of tumoroid cultures from benign and malignant salivary gland tumor types, in which tumor-specific gene signatures are well-conserved. In this study, we provide an experimental platform for the exploration of precision medicine in the era of tissue regeneration and anticancer treatment.
Smoking cessation aids in restoring lung function. However, whether long-term cessation can fully restore lung function has not been studied thoroughly, especially in Asian countries. This study ...aimed to evaluate the association between smoking cessation status and obstructive spirometry pattern among Koreans aged 40-79 years. In total, 6298 men and 8088 women aged 40-79 years from the Korea National Health and Nutrition Examination Survey (2015-2019) were analyzed for smoking cessation status, including the duration after quitting. Current-smokers showed a higher likelihood of having an obstructive spirometry pattern than never-smokers among both men (odds ratio OR: 3.15, 95% confidence interval CI: 2.32-4.29) and women (OR: 2.60, 95% CI: 1.59-4.23). In men, the effect tended to decrease with longer duration after cessation, but male ex-smokers who had quit smoking ≥ 20 years ago still showed a higher likelihood of having an obstructive spirometry pattern than male never-smokers (OR: 1.40, 95% CI: 1.05-1.89). In female ex-smokers, there was no significant association with the obstructive spirometry pattern, compared to that in female never-smokers. This study emphasizes the benefits of smoking cessation, possibility of long-lasting harm to lung function due to tobacco smoking, and importance of smoking prevention.
In the present study, a novel polar-solvent vapor annealing (PSVA) was used to induce a significant structural rearrangement in poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS) ...films in order to improve their electrical conductivity and work function. The effects of polar-solvent vapor annealing on PEDOT:PSS were systematically compared with those of a conventional solvent additive method (SAM) and investigated in detail by analyzing the changes in conductivity, morphology, top and bottom surface composition, conformational PEDOT chains, and work function. The results confirmed that PSVA induces significant phase separation between excess PSS and PEDOT chains and a spontaneous formation of a highly enriched PSS layer on the top surface of the PEDOT:PSS polymer blend, which in turn leads to better 3-dimensional connections between the conducting PEDOT chains and higher work function. The resultant PSVA-treated PEDOT:PSS anode films exhibited a significantly enhanced conductivity of up to 1057 S cm–1 and a tunable high work function of up to 5.35 eV. The PSVA-treated PEDOT:PSS films were employed as transparent anodes in polymer light-emitting diodes (PLEDs) and polymer solar cells (PSCs). The cell performances of organic optoelectronic devices with the PSVA-treated PEDOT:PSS anodes were further improved due to the significant vertical phase separation and the self-organized PSS top surface in PSVA-treated PEDOT:PSS films, which can increase the anode conductivity and work function and allow the direct formation of a functional buffer layer between the active layer and the polymeric electrode. The results of the present study will allow better use and understanding of polymeric-blend materials and will further advance the realization of high-performance indium tin oxide (ITO)-free organic electronics.
Three-dimensional spheroid culture enhances cell-to-cell interactions among stem cells and promotes the expression of stem cell properties; however, subsequent retrieval and delivery of these cells ...remain a challenge. We fabricated a thermoresponsive fiber-based microwell scaffold by combining electrospinning and hydrogel micropatterning. The resultant scaffold appeared to facilitate the formation of cellular spheroids of uniform size and enabled the expression of more stem cell-secreting growth factor genes (EGF, IGF-1, FGF1, FGF2, and HGF), pluripotent stem cell-related genes (SOX2 and NANOG), and adult epithelial stem cell-related genes (LGR4, LGR5, and LGR6) than salivary gland stem cells in a monolayer culture (SGSCmonolayer). The spheroids could be retrieved efficiently by decreasing temperature. SGSC-derived spheroid (SGSCspheroid) cells were then implanted into the submandibular glands of mice at 2 weeks after fractionated X-ray irradiation at a dose of 7.5 Gy/day. At 16 weeks post-irradiation, restoration of salivary function was detected only in SGSCspheroid-implanted mice. The production of submandibular acini specific mucin increased in SGSCspheroid-implanted mice, compared with PBS control. More MIST1+ mature acinar cells were preserved in the SGSCspheroid-implanted group than in the PBS control group. Intriguingly, SGSCspheroid-implanted mice exhibited greater amelioration of tissue damage and preservation of KRT7+ terminally differentiated luminal ductal cells than SGSCmonolayer-implanted mice. The SGSCspheroid-implanted mice also showed less DNA damage and apoptotic cell death than the SGSCmonolayer-implanted mice at 2 weeks post-implantation. Additionally, a significant increase in Ki67+AQP5+ proliferative acinar cells was noted only in SGSCspheroid-implanted mice. Our results suggest that a thermoresponsive fiber-based scaffold could be of use to facilitate the production of function-enhanced SGSCspheroid cells and their subsequent retrieval and delivery to damaged salivary glands to alleviate radiation-induced apoptotic cell death and promote salivary gland regeneration.
Chitinase-3-like protein 1 (CHI3L1) is a secreted glycoprotein that mediates inflammation, macrophage polarization, apoptosis, and carcinogenesis. The expression of CHI3L1 is strongly upregulated by ...various inflammatory and immunological diseases, including several cancers, Alzheimer's disease, and atherosclerosis. Several studies have shown that CHI3L1 can be considered as a marker of disease diagnosis, prognosis, disease activity, and severity. In addition, the proinflammatory action of CHI3L1 may be mediated via responses to various proinflammatory cytokines, including tumor necrosis factor-α, interleukin-1β, interleukin-6, and interferon-γ. Therefore, CHI3L1 may contribute to a vast array of inflammatory diseases. However, its pathophysiological and pharmacological roles in the development of inflammatory diseases remain unclear. In this article, we review recent findings regarding the roles of CHI3L1 in the development of inflammatory diseases and suggest therapeutic approaches that target CHI3L1.
Korean isolates of the Mycobacterium chelonae-Mycobacterium abscessus group, which had been isolated from two different hospitals in South Korea, were identified by PCR restriction analysis (PRA) and ...comparative sequence analysis of 16S rRNA genes, rpoB, and hsp65 to evaluate the proportion of four closely related species (M. chelonae, M. abscessus, M. massiliense, and M. bolletii). Of the 144 rapidly growing mycobacterial strains tested, 127 strains (88.2%) belonged to the M. chelonae-M. abscessus group. In this group, M. chelonae, M. abscessus, M. massiliense, and M. bolletii accounted for 0.8% (n = 1), 51.2% (n = 65), 46.5% (n = 59), and 1.6% (n = 2), respectively. Two isolates which showed discordant results, M. massiliense by rpoB sequence analysis and M. abscessus by hsp65 sequence analysis, were finally identified as M. massiliense based on the additional analysis of sodA and the 16S-23S internal transcribed spacer. M. abscessus group I isolates previously identified by hsp65 PRA were all found to be M. abscessus, whereas group II isolates were further identified as M. massiliense or M. bolletii by sequencing of rpoB and hsp65. Smooth, rough, or mixed colonies of both M. abscessus and M. massiliense isolates were observed. M. massiliense strains that were highly resistant to clarithromycin had a point mutation at the adenine at position 2058 (A₂₀₅₈) or 2059 (A₂₀₅₉) in the peptidyltransferase region of the 23S rRNA gene.
Stem cells produce nanosized particles known as extracellular vesicles (SC-EVs), which therapeutically affect stem cells. EVs are more abundantly produced, exhibit better stability, and possess lower ...immune rejection rates than stem cells. However, the traditional methods of isolating EVs, such as ultracentrifugation, possess limitations that require a complex process and consume more time. Moreover, it is difficult to isolate specific EVs that have target surface proteins that affect regenerative effects. To address these limitations, a new dual-mode horseshoe-shaped orifice micromixer (DM-HOMM) chip that can bind antibody-conjugated micromagnetic beads and SC-EVs and sequentially elute specific SC-EVs on the beads using an eluent was developed. For effective elution from the microbead-SC-EV complex, four types of eluents were used to control pH and ionic strength between antibodies and surface proteins in EVs. In addition, we investigated the reusability of antibody-conjugated micromagnetic beads. The beads indicated identical binding efficiencies between the antibodies and specific SC-EVs for three repeated cycles using the dual-mode chip. CD63
+
EVs collected by the chip exhibited higher cell viability and regeneration effects than untreated and total EVs. This SC-EVs’ isolation method possesses the potential for targeted therapeutic applications and enhanced regenerative effects.
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