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This study investigated the distributions of heavy metals (Cd, Cu, Hg, Ni, Pb, and Zn) in agricultural soils near coal-fired power plants in Taean and Seocheon, South Korea, considering wind direction and distance from the plants. Additionally, pollution assessment for these heavy metals was conducted using the geoaccumulation index (I geo ) and enrichment factor. Results showed that heavy metal concentrations in the studied soil samples were below Korean environmental criteria for agricultural soil (Cd: 4, Cu: 150, Hg: 4, Ni: 100, Pb: 200, and Zn: 300 mg/kg). However, a significant proportion of samples exceeded average levels found in uncontaminated soils. Spatial distribution analysis revealed higher concentrations of Cd and Pb southwest of the Taean plant, influenced by prevailing northeast winds. In Seocheon, soils within 4 km of the plant exhibited elevated levels of Cd and Ni, suggesting coal combustion as a potential contamination source. Pollution assessment indicated that Cd and Pb in soils near both thermal power plants were more enriched by artificial activity compared to agricultural soils in control areas. Sequential extraction results showed that heavy metals in soils within 4 km of the Seocheon plant had higher proportions of exchangeable to organic-associated forms than soils beyond 4 km, indicating a risk of high bioavailability near emission sources. This study highlights the significant impact of coal-fired power plant emissions on soil contamination, emphasizing the need for continuous monitoring and management. Environmental policies should consider wind patterns and proximity to emission sources to effectively mitigate contamination risks., Competing Interests: Declarations. Conflict of interests: The authors have no relevant financial or nonfinancial interests to disclose., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Inside living organisms, proteins are self-assembled into diverse 3D structures optimized for specific functions. This structure-function relationship can be exploited to synthesize functional materials through biotemplating and depositing functional materials onto protein structures. However, conventional biotemplating faces limitations due to the predominantly intracellular existence of proteins and associated challenges in achieving tunability while preserving functionality. In this study, Conversion to Advanced Materials via labeled Biostructures (CamBio), an integrated biotemplating platform that involves labeling target protein structures with antibodies followed by the growth of functional materials, ensuring outstanding nanostructure tunability is proposed. Protein-derived plasmonic nanostructures created by CamBio can serve as precise quantitative tools for assessing target species is demonstrated. The assessment is achieved through highly tunable and efficient surface-enhanced Raman spectroscopy (SERS). CamBio enables the formation of dense nanogap hot spots among metal nanoparticles, templated by diverse fibrous proteins comprising densely repeated monomers. Furthermore, iterative antibody labeling strategies to adjust the antibody density surrounding targets, amplifying the number of nanogaps and consequently improving SERS performance are employed. Finally, cell-patterned substrates and whole meat sections as SERS substrates, confirming their easily accessible, cost-effective, scalable preparation capabilities and dimensional tunability are incorporated., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Lipid droplets (LDs), the essential cytosolic fat storage organelles, have emerged as pivotal regulators of cellular metabolism and are implicated in various diseases. The noninvasive monitoring of LDs necessitates fluorescent probes with precise organelle selectivity and biocompatibility. Addressing this need, we have engineered a probe by strategically modifying the structure of a conventional two-photon-absorbing dipolar dye, acedan. This innovative approach induces nanoaggregate formation in aqueous environments, leading to aggregation-induced fluorescence quenching. Upon cellular uptake via clathrin-mediated endocytosis, the probe selectively illuminates within LDs through a disassembly process, effectively distinguishing LDs from the cytosol with exceptional specificity. This breakthrough enables the high-fidelity imaging of LDs in both cellular and tissue environments. In a pioneering investigation, we probed LDs in a diabetes model induced by streptozotocin, unveiling significantly heightened LD accumulation in cardiac tissues compared to other organs, as evidenced by TP imaging. Furthermore, our exploration of a lipopolysaccharide-mediated cardiomyopathy model revealed an LD accumulation during heart injury. Thus, our developed probe holds immense potential for elucidating LD-associated diseases and advancing related research endeavors.

Tumor microenvironment is intrinsically hypoxic with abundant hypoxia-inducible factors-1α (HIF-1α), a primary regulator of the cellular response to hypoxia and various stresses imposed on the tumor cells. HIF-1α increases radioresistance and chemoresistance by reducing DNA damage, increasing repair of DNA damage, enhancing glycolysis that increases antioxidant capacity of tumors cells, and promoting angiogenesis. In addition, HIF-1α markedly enhances drug efflux, leading to multidrug resistance. Radiotherapy and certain chemotherapy drugs evoke profound anti-tumor immunity by inducing immunologic cell death that release tumor associated antigens together with numerous pro-immunological factors, leading to priming of cytotoxic CD8+ T cells and enhancing the cytotoxicity of macrophages and NK cells. Radiotherapy and chemotherapy of tumors significantly increase HIF-1α activity in tumor cells. Unfortunately, HIF-1α effectively promotes various immune suppressive pathways including secretion of immune suppressive cytokines, activation of myeloid-derived suppressor cells (MIDSCs), activation of regulatory T cells (Tregs), inhibition of T cells priming and activity, and upregulation of immune checkpoints. Consequently, the anti-tumor immunity elevated by radiotherapy and chemotherapy is counterbalanced or masked by the potent immune suppression promoted by HIF-1α. Effective inhibition of HIF-1α may significantly increase the efficacy of radiotherapy and chemotherapy by increasing radiosensitivity and chemosensitivity of tumor cells and also by upregulating anti-tumor immunity.

Glioblastoma (GBM), the most lethal primary brain cancer, exhibits intratumoral heterogeneity and molecular plasticity, posing challenges for effective treatment. Despite this, the regulatory mechanisms underlying such plasticity, particularly mesenchymal (MES) transition, remain poorly understood. In this study, we elucidate the role of the RNA-binding protein ELAVL2 in regulating aggressive MES transformation in GBM. We found that ELAVL2 is most frequently deleted in GBM compared to other cancers and associated with distinct clinical and molecular features. Transcriptomic analysis revealed that ELAVL2-mediated alterations correspond to specific GBM subtype signatures. Notably, ELAVL2 expression negatively correlated with epithelial-to-mesenchymal transition (EMT)-related genes, and its loss promoted MES process and chemo-resistance in GBM cells, whereas ELAVL2 overexpression exerted the opposite effect. Further investigation via tissue microarray analysis demonstrated that high ELAVL2 protein expression confers a favorable survival outcome in GBM patients. Mechanistically, ELAVL2 was shown to directly bind to the transcripts of EMT-inhibitory molecules, SH3GL3 and DNM3, modulating their mRNA stability, potentially through an m6A-dependent mechanism. In summary, our findings identify ELAVL2 as a critical tumor suppressor and mRNA stabilizer that regulates MES transition in GBM, underscoring its role in transcriptomic plasticity and glioma progression., (© 2024. The Author(s).)

Background: The use of nonintubated video-assisted thoracoscopic surgery (NI-VATS) has been increasingly reported to yield favourable outcomes. However, this technology has not been routinely used because its advantages and safety have not been fully confirmed. The aim of this study was to assess the safety and feasibility of nonintubated spontaneous ventilation (NI-SV) anesthesia compared to intubated mechanical ventilation (I-MV) anesthesia in VATS by evaluating of perioperative complications and practitioners' workloads., Methods: Patients who underwent uniportal VATS were randomly assigned at a 1:1 ratio to receive NI-SV or I-MV anesthesia. The primary outcome was the occurrence of intraoperative airway intervention events, including transient MV, conversion to intubation and repositioning of the double-lumen tube. The secondary outcomes included perioperative complications and modified National Aeronautics and Space Administration Task Load Index (NASA-TLX) scores from anesthesiologists and surgeons., Results: Thirty-five patients in each group were enrolled in the intention-to-treat analysis. The incidence of intraoperative airway intervention events was greater in the NI-SV group than in the I-MV group (12 [34.3%] vs. 3 [8.6%]; OR = 0.180; 95% CI = 0.045-0.710; p = 0.009). No significant difference was found in the postoperative pulmonary complications between the groups (p > 0.05). The median of the anesthesiologists' overall NASA-TLX score was 37.5 (29-52) when administering the NI-SV, which was greater than the 25 (19-34.5) when the I-MV was administered (p < 0.001). The surgeons' overall NASA-TLX score was comparable between the two ventilation strategies (28 [21-38.5] vs. 27 [20.5-38.5], p = 0.814)., Conclusion: The NI-SV anesthesia was feasible for VATS in the selected patients, with a greater incidence of intraoperative airway intervention events than I-MV anesthesia, and with more surgical effort required by anesthesiologists., Trial Registration: Chinese Clinical Trial Registry, ChiCTR2200055427. https://www.chictr.org.cn/showproj.html?proj=147872 was registered on January 09, 2022., (© 2024. The Author(s).)

Biological systems are composed of hierarchical structures made of a large number of proteins. These structures are highly sophisticated and challenging to replicate using artificial synthesis methods. To exploit these structures in materials science, biotemplating is used to achieve biocomposites that accurately mimic biological structures and impart functionality of inorganic materials, including electrical conductivity. However, the biological scaffolds used in previous studies are limited to stereotypical and simple morphologies with little synthetic diversity because of a lack of control over their morphologies. This study proposes that the specific protein assemblies within the cell-derived extracellular matrix (ECM), whose morphological features are widely tailorable, can be employed as versatile biotemplates. In a typical procedure, a fibrillar assembly of fibronectin-a constituent protein of the ECM-is metalized through an antibody-guided biotemplating approach. Specifically, the antibody-bearing nanogold is attached to the fibronectin through antibody-antigen interactions, and then metals are grown on the nanogold acting as a seed. The biomimetic structure can be adapted for hydrogen production and sensing after improving its electrical conductivity through thermal sintering or additional metal growth. This study demonstrates that cell-derived ECM can be an attractive option for addressing the diversity limitation of a conventional biotemplate., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Targeted protein degradation (TPD) provides unique advantages over gene knockdown in that it can induce selective degradation of disease-associated proteins attributed to pathological mutations or aberrant post-translational modifications (PTMs). Herein, we report a protein degrader, PRZ-18002, that selectively binds to an active form of p38 MAPK. PRZ-18002 induces degradation of phosphorylated p38 MAPK (p-p38) and a phosphomimetic mutant of p38 MAPK in a proteasome-dependent manner. Given that the activation of p38 MAPK plays pivotal roles in the pathophysiology of Alzheimer's disease (AD), selective degradation of p-p38 may provide an attractive therapeutic option for the treatment of AD. In the 5xFAD transgenic mice model of AD, intranasal treatment of PRZ-18002 reduces p-p38 levels and alleviates microglia activation and amyloid beta (Aβ) deposition, leading to subsequent improvement of spatial learning and memory. Collectively, our findings suggest that PRZ-18002 ameliorates AD pathophysiology via selective degradation of p-p38, highlighting a novel therapeutic TPD modality that targets a specific PTM to induce selective degradation of neurodegenerative disease-associated protein., Competing Interests: The authors declare the following competing financial interest(s): Kyung Hee University and Prazer Therapeutics, Inc. have filed patent applications (PCT/KR2021/002361) based on the results of this study. The remaining authors declare no competing interests., (© 2023 The Authors. Published by American Chemical Society.)

In biological studies and diagnoses, brightfield (BF), fluorescence, and electron microscopy (EM) are used to image biomolecules inside cells. When compared, their relative advantages and disadvantages are obvious. BF microscopy is the most accessible of the three, but its resolution is limited to a few microns. EM provides a nanoscale resolution, but sample preparation is time-consuming. In this study, we present a new imaging technique, which we termed decoration microscopy (DecoM), and quantitative investigations to address the aforementioned issues in EM and BF microscopy. For molecular-specific EM imaging, DecoM labels proteins inside cells using antibodies bearing 1.4 nm gold nanoparticles (AuNPs) and grows silver layers on the AuNPs' surfaces. The cells are then dried without buffer exchange and imaged using scanning electron microscopy (SEM). Structures labeled with silver-grown AuNPs are clearly visible on SEM, even they are covered with lipid membranes. Using stochastic optical reconstruction microscopy, we show that the drying process causes negligible distortion of structures and that less structural deformation could be achieved through simple buffer exchange to hexamethyldisilazane. Using DecoM, we visualize the nanoscale alterations in microtubules by microtubule-severing proteins that cannot be observed with diffraction-limited fluorescence microscopy. We then combine DecoM with expansion microscopy to enable sub-micron resolution BF microscopy imaging. We first show that silver-grown AuNPs strongly absorb white light, and the structures labeled with them are clearly visible on BF microscopy. We then show that the application of AuNPs and silver development must follow expansion to visualize the labeled proteins clearly with sub-micron resolution., Competing Interests: The authors declare no competing financial interest., (This journal is © The Royal Society of Chemistry.)

Background: Bentonite, a montmorillonite clay, has been used as a classical remedy strategy for a long time. Recently, bentonite has been used as a raw material in cosmetic products because of its antibacterial and antioxidant properties. However, the therapeutic effect of bentonite on burn injuries has not yet been identified., Objectives: The aim of this study was to explore the therapeutic effect of a novel bentonite complex, which was developed for medical use, on burn wounds and the anti-inflammatory function of bentonite clay in the complex in vitro., Methods: A novel bentonite complex and bentonite clay were prepared for each in vivo and in vitro assay (C&L Biotech, Seoul, Korea). Burn wounds were induced on the dorsal skin of two Yucatan minipigs, and effects of tissue regeneration and anti-inflammation were assessed by histological and gene expression analysis., Results: The bentonite complex improved skin regeneration in burn wounds by inducing collagen synthesis, cell proliferation and angiogenesis in vivo. Furthermore, expression of inflammatory cytokines was significantly inhibited by treatment of the bentonite complex. In vitro study for identification of anti-inflammatory effect showed that bentonite clay significantly regulated COX-2 signalling in both HacaT keratinocytes and 3D4/2 macrophage cell lines., Conclusions: This study identified the therapeutic effect of a novel bentonite complex in burn wounds by inducing skin regeneration and bentonite-mediated anti-inflammatory responses., (© 2022 The Authors. Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Discoidin domain receptor 2 (DDR2) belongs to the receptor tyrosine kinase (RTK) family, other RTKs have been reported to regulate phagocytic function of Sertoli cells (SCs), yet little is known about the function of DDR2 in Sertoli cells. In the present study, we aim to explore the function and mechanism of ectopic discoidin domain receptor 2 (DDR2) expression in Sertoli cells of Sertoli cell-only syndrome (SCOS) testes. We found that discoidin domain receptor 2 (DDR2) was absent in Sertoli cells of normal testis but was expressed in Sertoli cells of SCOS testes. This Sertoli cell DDR2 expression was induced by impaired androgen receptor (AR) signaling, but was inhibited by increased AR signaling from testosterone administration. The Sertoli cell DDR2 expression led to an increase in phagocytosis through up-regulation of Scavenger receptor class B member 1 (SR-BI) levels. However, loss of DDR2 by knock-out or knock-down weakened the phagocytotic capacity of Sertoli cells. Furthermore, the expression of DDR2 in Sertoli cells activated matrix metallopeptidase 9 (MMP-9) to consume abnormal collagen increase in seminiferous tubules which was responsible for the block of testosterone transportation and AR loss and to compensate for the impaired blood-testis-barrier (BTB). Our data suggest that the AR/DDR2 cascade may serve as a negative feedback mechanism to help compensate for the homeostasis of seminiferous epithelium in SCOS testis., Competing Interests: None., (AJTR Copyright © 2022.)

Biological systems consist of hierarchical protein structures, each of which has unique 3D geometries optimized for specific functions. In the past decades, the growth of inorganic materials on specific proteins has attracted considerable attention. However, the use of specific proteins as templates has only been demonstrated in relatively simple organisms, such as viruses, limiting the range of structures that can be used as scaffolds. This study proposes a method for synthesizing metallic structures that resemble the 3D assemblies of specific proteins in mammalian cells and animal tissues. Using 1.4 nm nanogold-conjugated antibodies, specific proteins within cells and ex vivo tissues are labeled, and then the nanogold acts as nucleation sites for growth of metal particles. As proof of concept, various metal particles are grown using microtubules in cells as templates. The metal-containing cells are applied as catalysts and show catalytic stability in liquid-phase reactions due to the rigid support provided by the microtubules. Finally, this method is used to produce metal structures that replicate the specific protein assemblies of neurons in the mouse brain or the extracellular matrices in the mouse kidney and heart. This new biotemplating approach can facilitate the conversion of specific protein structures into metallic forms in ex vivo multicellular organisms., (© 2022 Wiley-VCH GmbH.)

Bacterial wilt caused by Ralstonia solanacearum is a fatal disease that affects the production of tomatoes and many other crops worldwide. As an effective strategy to manage bacterial wilt, biological control agents using plant growth-promoting rhizobacteria (PGPR) are being developed. In this study, we screened 2,3-butanediol (BDO)-producing PGPR to control tomato bacterial wilt and investigated the action mechanism of the disease control agent. Of the 943 strains isolated from soil, Klebsiella pneumoniae strain JCK-2201 produced the highest concentration of 2,3-BDO. The culture broth of K. pneumoniae JCK-2201 did not show any direct activity on R. solanacearum in vitro , but a 100-fold dilution effectively controlled tomato bacterial wilt with a control value of 77% in vivo . Fermentation utilizing K. pneumoniae JCK-2201 was optimized to produce 48 g/L of meso-2,3-BDO, which is 50% of the sucrose conversion efficiency. In addition, the control efficacy and mechanism of meso-2,3-BDO produced by JCK-2201 in tomato bacterial wilt were determined by comparative analysis with Bacillus licheniformis DSM13 producing meso-2,3-BDO and B. licheniformis DSM13 ΔalsS that did not produce 2,3-BDO, as the step of converting pyruvate to α-acetolactate was omitted. Tomato seedlings treated with the K. pneumoniae JCK-2201 (500-fold dilution) and B. licheniformis DSM13 (100-fold dilution) culture broth produced meso-2,3-BDO that significantly reduced R. solanacearum -induced disease severity with control values of 55% and 63%, respectively. The formulated meso-2,3-BDO 9% soluble concentrate (SL; 1,000-fold dilution) showed 87% control against tomato bacterial wilt in the field condition. Klebsiella pneumoniae JCK-2201 and B. licheniformis DSM13 treatment induced the expression of plant defense marker genes, such as LePR1 , LePR2 , LePR5 , LePR3 , and PI-II , in the salicylic acid and jasmonic acid signaling pathways at 4 days after inoculation. These results show that 2,3-BDO-producing bacteria and 2,3-BDO are potential biological control agents that act through induction of resistance for controlling tomato bacterial wilt., Competing Interests: CS and HS were employed by the GS Caltex Corporation of South Korea. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kim, Park, Song, Song and Kim.)

High-dose hypofractionated radiation such as SABR (stereotactic ablative radiotherapy) evokes an anti-tumor immune response by promoting a series of immune-stimulating processes, including the release of tumor-specific antigens from damaged tumor cells and the final effector phase of immune-mediated lysis of target tumor cells. High-dose hypofractionated radiation also causes vascular damage in tumors, thereby increasing tumor hypoxia and upregulation of hypoxia-inducible factors HIF-1α and HIF-2α, the master transcription factors for the cellular response to hypoxia. HIF-1α and HIF-2α are critical factors in the upregulation of immune suppression and are the master regulators of immune evasion of tumors. Consequently, SABR-induced increase in anti-tumor immunity is counterbalanced by the increase in immune suppression mediated by HIFα. Inhibition of HIF-1α with small molecules such as metformin downregulates immunosuppressive pathways, including the expression of immune checkpoints, and it improves or restores the anti-tumor immunity stimulated by irradiation. Combinations of HIFα inhibitors, particularly HIF-1α inhibitors, with immune checkpoint blocking antibodies may represent a novel approach to boost the overall anti-tumor immune profile in patients and thus enhance outcomes after SABR.

Turfgrass, the most widely grown ornamental crop, is severely affected by fungal pathogens including Sclerotinia homoeocarpa, Rhizoctonia solani, and Magnaporthe poae. At present, turfgrass fungal disease management predominantly relies on synthetic fungicide treatments. However, the extensive application of fungicides to the soil increases residual detection frequency, raising concerns for the environment and human health. The bacterial volatile compound, 2,3-butanediol (BDO), was found to induce plant resistance. In this study, we evaluated the disease control efficacy of a combination of stereoisomers of 2,3-BDO and commercial fungicides against turfgrass fungal diseases in both growth room and fields. In the growth room experiment, the combination of 0.9% 2R,3R-BDO (levo) soluble liquid (SL) formulation and 9% 2R,3S-BDO (meso) SL with half concentration of fungicides significantly increased the disease control efficacy against dollar spot and summer patch disease when compared to the half concentration of fungicide alone. In field experiments, the disease control efficiency of levo 0.9% and meso 9% SL, in combination with a fungicide, was confirmed against dollar spot and large patch disease. Additionally, the induction of defense-related genes involved in the salicylic acid and jasmonic acid/ethylene signaling pathways and reactive oxygen species detoxification-related genes under Clarireedia sp. infection was confirmed with levo 0.9% and meso 9% SL treatment in creeping bentgrass. Our findings suggest that 2,3-BDO isomer formulations can be combined with chemical fungicides as a new integrated tool to control Clarireedia sp. infection in turfgrass, thereby reducing the use of chemical fungicides.

Among abiotic stresses in plants, drought and chilling stresses reduce the supply of moisture to plant tissues, inhibit photosynthesis, and severely reduce plant growth and yield. Thus, the application of water stress-tolerant agents can be a useful strategy to maintain plant growth under abiotic stresses. This study assessed the effect of exogenous bio-based 2,3-butanediol (BDO) application on drought and chilling response in tomato and turfgrass, and expression levels of several plant signaling pathway-related gene transcripts. Bio-based 2,3-BDOs were formulated to levo-2,3-BDO 0.9% soluble concentrate (levo 0.9% SL) and meso-2,3-BDO 9% SL (meso 9% SL). Under drought and chilling stress conditions, the application of levo 0.9% SL in creeping bentgrass and meso 9% SL in tomato plants significantly reduced the deleterious effects of abiotic stresses. Interestingly, pretreatment with levo-2,3-BDO in creeping bentgrass and meso-2,3-BDO in tomato plants enhanced JA and SA signaling pathway-related gene transcript expression levels in different ways. In addition, all tomato plants treated with acibenzolar- S -methyl (as a positive control) withered completely under chilling stress, whereas 2,3-BDO-treated tomato plants exhibited excellent cold tolerance. According to our findings, bio-based 2,3-BDO isomers as sustainable water stress-tolerant agents, levo- and meso-2,3-BDOs, could enhance tolerance to drought and/or chilling stresses in various plants through somewhat different molecular activities without any side effects.

We present a new type of stretchable dichroic film in which Au and Ag alloy nanoparticles (NPs) are dispersed in polydimethylsiloxane (PDMS). The alloy NPs are synthesized with different atomic compositions and sizes to modulate their plasmonic resonance frequencies and absorption and scattering cross sections. The PDMS dichroic film in which 100 nm alloy NPs with a Au/Ag ratio of 7:3 are dispersed shows exotic optical properties under tensile strain. When 40% tensile strain is applied, the film exhibits a strain-sensitive transmission and strain-insensitive reflection behavior in which the transmittance is increased up to 2.6 times, whereas the reflectance remains unchanged. Moreover, we demonstrate a stretchable anticounterfeiting film and a flexible dichroic sculpture fabricated with the PDMS composite. This work demonstrates a new type of plasmonic application that has great potential in various applications, such as special-purpose optical films, security patterns, and smart windows.

A hot filament chemical vapor deposition (HFCVD) method was adopted to deposit diamond films at deposition pressures ranging from 2-6 kPa. The effects of deposition pressure on the deposition rate, phase structure, and microstructure of diamond films were investigated. The surface morphology, grain size, micro-structure, and growth rate of the diamond films were analyzed using scanning electron microscopy, X-ray diffraction (XRD), and Raman spectrometry. The experimental results showed that granules on the surface exhibited increasingly compact structure with increasing deposition pressure. The diamond films deposited at various pressures have good compactness, and the particles on the film surfaces are arranged in an ordered manner. All films exhibited orientation along the (111) plane, which was the significant characteristic XRD peak of each diamond film. The (111) peak intensity was the strongest for the film prepared at 2 kPa deposition pressure. Overall, the deposition rate and grain size decreased with increasing deposition pressure, provided other deposition conditions remained unchanged. However, the densification of the microstructure and the nucleation density increased with increasing deposition pressure. Secondary nucleation became more pronounced as deposition pressure increased, and grain size decreased as nucleation density increased.

NAD(P)H quinone oxidoreductase-1 (NQO1), a protective enzyme against cellular oxidative stress, is expressed abnormally high in solid tumors and thus recognized as a cancer biomarker. To develop a fluorescent NQO1 probe with practicality, we investigated benzo-rosol fluorophores linked with a known self-immolative quinone substrate. Four probe candidates exhibited ratiometric sensing behavior toward the enzyme, satisfying our orbital mismatch stratagem proposed before, under dual-excitation and dual-emission conditions that alleviate the spectral overlap issue commonly observed with the ratiometric probes based on intramolecular charge-transfer change. Among the candidates, two ester-linked compounds exhibited hydrolytic instability to water or an esterase, discouraging us to develop such ester-linked probes. One ether-linked, hydrolytically stable probe provided brighter cellular fluorescence than the other and thus was applied to ratiometric imaging of NQO1 in cells and tissues. We found that the enzyme activity levels are much different in organ tissues: stomach (56), kidney (22), colon (9.8), testis (7.8), bladder (5.6), lung (1.2), and muscle (1.0). Furthermore, a markedly high enzyme level (14.6-fold) was observed in a xenograft tumor tissue compared with that in a normal tissue, which suggests that such an NQO1 probe is promising for cancer diagnosis and for studying the enzyme-associated biology.