Munc13 activates the Munc18-1/syntaxin-1 complex and enables Munc18-1 to prime SNARE assembly.

Priming of synaptic vesicles involves Munc13-catalyzed transition of the Munc18-1/syntaxin-1 complex to the SNARE complex in the presence of SNAP-25 and synaptobrevin-2; Munc13 drives opening of syntaxin-1 via the MUN domain while Munc18-1 primes SNARE assembly via domain 3a. However, the underlying mechanism remains unclear. In this study, we have identified a number of residues in domain 3a of Munc18-1 that are crucial for Munc13 and Munc18-1 actions in SNARE complex assembly and synaptic vesicle priming. Our results showed that two residues (Q301/K308) at the side of domain 3a mediate the interaction between the Munc18-1/syntaxin-1 complex and the MUN domain. This interaction enables the MUN domain to drive the opening of syntaxin-1 linker region, thereby leading to the extension of domain 3a and promoting synaptobrevin-2 binding. In addition, we identified two residues (K332/K333) at the bottom of domain 3a that mediate the interaction between Munc18-1 and the SNARE motif of syntaxin-1. This interaction ensures Munc18-1 to persistently associate with syntaxin-1 during the conformational change of syntaxin-1 from closed to open, which reinforces the role of Munc18-1 in templating SNARE assembly. Taken together, our data suggest a mechanism by which Munc13 activates the Munc18-1/syntaxin-1 complex and enables Munc18-1 to prime SNARE assembly.

Cytoplasmic tail of transmembrane dscam controls antibacterial responses by regulating cell proliferation-related genes in hemocytes of Chinese mitten crab (Eriocheir sinensis).

In arthropods, the involvement of Dscam (Down syndrome cell adhesion molecule) in innate immunity has been extensively demonstrated. Its cytoplasmic tail contains multiple conserved functional sites, which indicates its involvement in different intracellular signaling pathways. In this study, we focused on the role of the cytoplasmic tail of Dscam in the Chinese mitten crab (Eriocheir sinensis) immune defense. In the group with cytoplasmic tail knockdown (the site was located on constant exons 37 and 38), 3885 differentially expressed genes (DEGs) were identified. The DEGs were enriched in small molecule binding, protein-containing complex binding, and immunity-related pathways. The expression of selected genes were validated using quantitative real-time reverse transcription PCR. We identified key Cell cycle, Janus kinase (JAK)-signal transducer, activator of transcription (STAT) and mitogen-activated protein kinase (MAPK) signaling pathway genes, the results indicated that the cytoplasmic tail of Dscam controls antibacterial responses by regulating cell proliferation-related genes in hemocytes.

Identification of residues critical for the extension of Munc18-1 domain 3a.

BACKGROUND: Neurotransmitter release depends on the fusion of synaptic vesicles with the presynaptic membrane and is mainly mediated by SNARE complex assembly. During the transition of Munc18-1/Syntaxin-1 to the SNARE complex, the opening of the Syntaxin-1 linker region catalyzed by Munc13-1 leads to the extension of the domain 3a hinge loop, which enables domain 3a to bind SNARE motifs in Synaptobrevin-2 and Syntaxin-1 and template the SNARE complex assembly. However, the exact mechanism of domain 3a extension remains elusive. RESULTS: Here, we characterized residues on the domain 3a hinge loop that are crucial for the extension of domain 3a by using biophysical and biochemical approaches and electrophysiological recordings. We showed that the mutation of residues T323/M324/R325 disrupted Munc13-1-mediated SNARE complex assembly and membrane fusion starting from Munc18-1/Syntaxin-1 in vitro and caused severe defects in the synaptic exocytosis of mouse cortex neurons in vivo. Moreover, the mutation had no effect on the binding of Synaptobrevin-2 to isolated Munc18-1 or the conformational change of the Syntaxin-1 linker region catalyzed by the Munc13-1 MUN domain. However, the extension of the domain 3a hinge loop in Munc18-1/Syntaxin-1 was completely disrupted by the mutation, leading to the failure of Synaptobrevin-2 binding to Munc18-1/Syntaxin-1. CONCLUSIONS: Together with previous results, our data further support the model that the template function of Munc18-1 in SNARE complex assembly requires the extension of domain 3a, and particular residues in the domain 3a hinge loop are crucial for the autoinhibitory release of domain 3a after the MUN domain opens the Syntaxin-1 linker region.

RCEAU-Net: Cascade Multi-Scale Convolution and Attention-Mechanism-Based Network for Laser Beam Target Image Segmentation with Complex Background in Coal Mine.

Accurate and reliable pose estimation of boom-type roadheaders is the key to the forming quality of the tunneling face in coal mines, which is of great importance to improve tunneling efficiency and ensure the safety of coal mine production. The multi-laser-beam target-based visual localization method is an effective way to realize accurate and reliable pose estimation of a roadheader body. However, the complex background interference in coal mines brings great challenges to the stable and accurate segmentation and extraction of laser beam features, which has become the main problem faced by the long-distance visual positioning method of underground equipment. In this paper, a semantic segmentation network for underground laser beams in coal mines, RCEAU-Net, is proposed based on U-Net. The network introduces residual connections in the convolution of the encoder and decoder parts, which effectively fuses the underlying feature information and improves the gradient circulation performance of the network. At the same time, by introducing cascade multi-scale convolution in the skipping connection section, which compensates for the lack of contextual semantic information in U-Net and improves the segmentation effect of the network model on tiny laser beams at long distance. Finally, the introduction of an efficient multi-scale attention module with cross-spatial learning in the encoder enhances the feature extraction capability of the network. Furthermore, the laser beam target dataset (LBTD) is constructed based on laser beam target images collected from several coal mines, and the proposed RCEAU-Net model is then tested and verified. The experimental results show that, compared with the original U-Net, RCEAU-Net can ensure the real-time performance of laser beam segmentation while increasing the Accuracy by 0.19%, Precision by 2.53%, Recall by 22.01%, and Intersection and Union Ratio by 8.48%, which can meet the requirements of multi-laser-beam feature segmentation and extraction under complex backgrounds in coal mines, so as to further ensure the accuracy and stability of long-distance visual positioning for boom-type roadheaders and ensure the safe production in the working face.

Study on the Skincare Effects of Red Rice Fermented by Aspergillus oryzae In Vitro.

Red rice, a variety of pigmented grain, serves dual purposes as both a food and medicinal resource. In recent years, we have witnessed an increasing interest in the dermatological benefits of fermented rice extracts, particularly their whitening and hydrating effects. However, data on the skincare advantages derived from fermenting red rice with Aspergillus oryzae remain sparse. This study utilized red rice as a substrate for fermentation by Aspergillus oryzae, producing a substance known as red rice Aspergillus oryzae fermentation (RRFA). We conducted a preliminary analysis of RRFA's composition followed by an evaluation of its skincare potential through various in vitro tests. Our objective was to develop a safe and highly effective skincare component for potential cosmetic applications. RRFA's constituents were assessed using high-performance liquid chromatography (HPLC), Kjeldahl nitrogen determination, the phenol-sulfuric acid method, and enzyme-linked immunosorbent assay (ELISA). We employed human dermal fibroblasts (FB) to assess RRFA's anti-aging and antioxidative properties, immortalized keratinocytes (HaCaT cells) and 3D epidermal models to examine its moisturizing and reparative capabilities, and human primary melanocytes (MCs) to study its effects on skin lightening. Our findings revealed that RRFA encompasses several bioactive compounds beneficial for skin health. RRFA can significantly promote the proliferation of FB cells. And it markedly enhances the mRNA expression of ECM-related anti-aging genes and reduces reactive oxygen species production. Furthermore, RRFA significantly boosts the expression of Aquaporin 3 (AQP3), Filaggrin (FLG), and Hyaluronan Synthase 1 (HAS1) mRNA, alongside elevating moisture levels in a 3D epidermal model. Increases were also observed in the mRNA expression of Claudin 1 (CLDN1), Involucrin (IVL), and Zonula Occludens-1 (ZO-1) in keratinocytes. Additionally, RRFA demonstrated an inhibitory effect on melanin synthesis. Collectively, RRFA contains diverse ingredients which are beneficial for skin health and showcases multifaceted skincare effects in terms of anti-aging, antioxidant, moisturizing, repairing, and whitening capabilities in vitro, highlighting its potential for future cosmetic applications.

Constructing a Myxobacterial Natural Product Database to Facilitate NMR-Based Metabolomics Bioprospecting of Myxobacteria.

Myxobacteria are fascinating prokaryotes featuring a potent capacity for producing a wealth of bioactive molecules with intricate chemical topology as well as intriguing enzymology, and thus it is critical to developing an efficient pipeline for bioprospecting. Herein, we construct the database MyxoDB, the first public compendium solely dedicated to myxobacteria, which enabled us to provide an overview of the structural diversity and taxonomic distribution of known myxobacterial natural products. Moreover, we demonstrated that the cutting-edge NMR-based metabolomics was effective to differentiate the biosynthetic priority of myxobacteria, whereby MyxoDB could greatly streamline the dereplication of multifarious known compounds and accordingly speed up the discovery of new compounds. This led to the rapid identification of a class of linear di-lipopeptides (archangimins) and a rare rearranged sterol (corasterol) that were endowed with unique chemical architectures and/or biosynthetic enzymology. We also showcased that NMR-based metabolomics, MyxoDB, and genomics can also work concertedly to accelerate the targeted discovery of a polyketidic compound pyxipyrrolone C. All in all, this study sets the stage for the discovery of many more novel natural products from underexplored myxobacterial resources.

Efficient framework of solving time-gated reflection matrix for imaging through turbid medium.

Imaging through turbid medium is a long pursuit in many research fields, such as biomedicine, astronomy and automatic vehicle, in which the reflection matrix-based method is a promising solution. However, the epi-detection geometry suffers from round-trip distortion and it is challenging to isolate the input and output aberrations in non-ideal cases due to system imperfections and measurement noises. Here, we present an efficient framework based on single scattering accumulation together with phase unwrapping that can accurately separate input and output aberrations from the noise-affected reflection matrix. We propose to only correct the output aberration while suppressing the input aberration by incoherent averaging. The proposed method is faster in convergence and more robust against noise, avoiding precise and tedious system adjustments. In both simulations and experiments, we demonstrate the diffraction-limited resolution capability under optical thickness beyond 10 scattering mean free paths, showing the potential of applications in neuroscience and dermatology.

Discovery and Biosynthesis of Glycosylated Cycloheximide from a Millipede-Associated Actinomycete.

Chemical redundancy of microbial natural products (NPs) underscores the importance to exploit new resources of microorganisms. Insect-associated microbes are prolific but largely underexplored sources of diverse NPs. Herein, we discovered the new compound α-l-rhamnosyl-actiphenol (1) from a millipede-associated Streptomyces sp. ML6, which is the first glycosylated cycloheximide-class natural product. Interestingly, bioinformatics analysis of the ML6 genome revealed that the biosynthesis of 1 involves a cooperation between two gene clusters (chx and rml) located distantly on the genome of ML6. We also carried out in vitro enzymatic glycosylation of cycloheximide using an exotic promiscuous glycosyltransferase BsGT-1, which resulted in the production of an additional cycloheximide glycoside cycloheximide 7-O-ß-d-glucoside (5). Although the antifungal and cytotoxic activities of the new compounds 1 and 5 were attenuated relative to those of cycloheximide, our work not only enriches the chemical repertoire of the cycloheximide family but also provides new insights into the structure-activity relationship optimization and ecological roles of cycloheximide.

C-type lectin receptors Dectin-3 and Dectin-2 form a heterodimeric pattern-recognition receptor for host defense against fungal infection.

C-type lectin receptors (CLRs) play critical roles as pattern-recognition receptors (PRRs) for sensing Candida albicans infection, which can be life-threatening for immunocompromised individuals. Here we have shown that Dectin-3 (also called CLECSF8, MCL, or Clec4d), a previously uncharacterized CLR, recognized α-mannans on the surfaces of C. albicans hyphae and induced NF-κB activation. Mice with either blockade or genetically deleted Dectin-3 were highly susceptible to C. albicans infection. Dectin-3 constantly formed heterodimers with Dectin-2, a well-characterized CLR, for recognizing C. albicans hyphae. Compared to their respective homodimers, Dectin-3 and Dectin-2 heterodimers bound α-mannans more effectively, leading to potent inflammatory responses against fungal infections. Together, our study demonstrates that Dectin-3 forms a heterodimeric PRR with Dectin-2 for sensing fungal infection and suggests that different CLRs may form different hetero- and homodimers, which provide different sensitivity and diversity for host cells to detect various microbial infections.

Comparison of Rural Childhood BMI Percentiles: Prevalence and Trends in a Midwest County, 2008-2016.

The number of children who are obese and overweight continues as a public health challenge despite decades of research. The purpose of this article is to describe trends in body mass index (BMI) percentile data collected from 11- to 14-year-old school children in 2008-2009 and 2015-2016 in rural Wisconsin. The BMI percentiles from 1,347 students were compared using time, gender, age, and school (public vs. parochial) as predictors. The trend over time indicated a decrease in students of healthy weight and an increase in those overweight or obese. Also noted was a significantly higher proportion of children who were overweight or obese in parochial compared to public schools. Discussed are the observed trends, community-wide initiatives implemented, as well as how schools can employ a more comprehensive approach to childhood obesity that first ensures community readiness and involves school, home, and community.

Effect of germination potential on storage lipids and transcriptome changes in premature developing seeds of oilseed rape (Brassica napus L.).

KEY MESSAGE: We provided a gene pool of moderate size for selecting or manipulating the candidate genes that favour the acquisition of seed dormancy, shedding light on the elevation of seed oil content in oilseed rape by blocking lipid degradation in developing seeds. In oilseed rape, the association between the germination potential of premature seeds and the final level of seed lipids, and the underlying mechanism, is elusive. Here, we investigated phenotypic differences in the germination percentage of premature seeds in a collection of oilseed rape cultivars. We compared the dynamic lipid accumulation between the deep-, moderate- and non-dormant genotypes and compared the transcriptomes of the seeds at 40 days after pollination between multiple pairs of deep- and non-dormant genotypes. We identified a wide range of differences in germination percentage of premature seeds and the association between the germination potential and the change of fatty acid content at late stage of seed maturation. The comparisons of transcriptomes between deep- and non-dormant seeds revealed the genetic basis for the dormant difference, e.g. the different expression levels of the genes involved in gibberellic and abscisic acid biosynthesis and/or signalling, fatty acid metabolic pathways, and the structure of seed cell wall. We provided a gene pool of moderate size for selecting or manipulating the candidate genes that favour the acquisition of seed dormancy, shedding light on the elevation of seed oil content in oilseed rape by blocking lipid degradation in developing seeds.

Unified Classification of Bacterial Colonies on Different Agar Media Based on Hyperspectral Imaging and Machine Learning.

A universal method by considering different types of culture media can enable convenient classification of bacterial species. The study combined hyperspectral technology and versatile chemometric algorithms to achieve the rapid and non-destructive classification of three kinds of bacterial colonies (Escherichia coli, Staphylococcus aureus and Salmonella) cultured on three kinds of agar media (Luria-Bertani agar (LA), plate count agar (PA) and tryptone soy agar (TSA)). Based on the extracted spectral data, partial least squares discriminant analysis (PLS-DA) and support vector machine (SVM) were employed to established classification models. The parameters of SVM models were optimized by comparing genetic algorithm (GA), particle swarm optimization (PSO) and grasshopper optimization algorithm (GOA). The best classification model was GOA-SVM, where the overall correct classification rates (OCCRs) for calibration and prediction of the full-wavelength GOA-SVM model were 99.45% and 98.82%, respectively, and the Kappa coefficient for prediction was 0.98. For further investigation, the CARS, SPA and GA wavelength selection methods were used to establish GOA-SVM simplified model, where CARS-GOA-SVM was optimal in model accuracy and stability with the corresponding OCCRs for calibration and prediction and the Kappa coefficients of 99.45%, 98.73% and 0.98, respectively. The above results demonstrated that it was feasible to classify bacterial colonies on different agar media and the unified model provided a continent and accurate way for bacterial classification.

Four New Diterpene Glucosides from Perovskia atriplicifolia.

Four new diterpene glucosides, namely perovskiaditerpenosides A - D (1 - 4), were isolated from the BuOH extract of Perovskia atriplicifolia. Their structures were well elucidated by chemical methods and comprehensive spectroscopic analyses including MS, IR, and NMR (1D and 2D). The newly isolated compounds were screened for their cytotoxic activity against HepG2, NB4, HeLa, K562, MCF7, PC3, and HL60. The obtained results indicated that the new compounds possessed considerable cytotoxic activity.

C-type lectin receptor dectin-3 mediates trehalose 6,6'-dimycolate (TDM)-induced Mincle expression through CARD9/Bcl10/MALT1-dependent nuclear factor (NF)-κB activation.

Previous studies indicate that both Dectin-3 (also called MCL or Clec4d) and Mincle (also called Clec4e), two C-type lectin receptors, can recognize trehalose 6,6'-dimycolate (TDM), a cell wall component from mycobacteria, and induce potent innate immune responses. Interestingly, stimulation of Dectin-3 by TDM can also induce Mincle expression, which may enhance the host innate immune system to sense Mycobacterium infection. However, the mechanism by which Dectin-3 induces Mincle expression is not fully defined. Here, we show that TDM-induced Mincle expression is dependent on Dectin-3-mediated NF-κB, but not nuclear factor of activated T-cells (NFAT), activation, and Dectin-3 induces NF-κB activation through the CARD9-BCL10-MALT1 complex. We found that bone marrow-derived macrophages from Dectin-3-deficient mice were severely defective in the induction of Mincle expression in response to TDM stimulation. This defect is correlated with the failure of TDM-induced NF-κB activation in Dectin-3-deficient bone marrow-derived macrophages. Consistently, inhibition of NF-κB, but not NFAT, impaired TDM-induced Mincle expression, whereas NF-κB, but not NFAT, binds to the Mincle promoter. Dectin-3-mediated NF-κB activation is dependent on the CARD9-Bcl10-MALT1 complex. Finally, mice deficient for Dectin-3 or CARD9 produced much less proinflammatory cytokines and keyhole limpet hemocyanin (KLH)-specific antibodies after immunization with an adjuvant containing TDM. Overall, this study provides the mechanism by which Dectin-3 induces Mincle expression in response to Mycobacterium infection, which will have significant impact to improve adjuvant and design vaccine for antimicrobial infection.

Asymmetric annulation of 3-alkynylacrylaldehydes with styrene-type olefins by synergetic relay catalysis from AgOAc and chiral phosphoric acid.

Asymmetric annulation of 3-alkynylacrylaldehydes with 2-hydroxystyrenes has been studied and achieved by synergetic catalysis from AgOAc and chiral phosphoric acid, providing the corresponding multiring products with decent total yields and moderate to excellent enantioselectivities (up to 95% ee). The developed mild multibond-formation cascade reaction involves in situ generation of pyrylium intermediate by Ag(I)-catalyzed alkyne/carbonyl cycloisomerization, counteranion-directed asymmetric oxa [4 + 2]-cycloaddition, and intramolecular nucleophilic substitution.

Administrative monopoly regulation and environmental innovation: An analysis based on China's asymmetric market structure.

To better understand the role of the competitive environment for firm innovation, this study takes China's Fair Competition Review System as an example to explore the impact of administrative monopoly regulation on environmental innovation. By utilizing China's market Structure, the study devises a quasi-natural experiment. It employs the DID method to assess the causal relationship between administrative monopoly regulation and the firm's environmental innovation. The findings indicate that implementing the Fair Competition Review System induces a marked reduction in environmental innovation by firms in the administrative monopoly sector, suggesting a substantial inhibition effect of the policy. Further research shows that this inhibition effect primarily manifests in the short term. Breaking administrative monopoly and optimizing the market environment wield the potential to increase firm environmental innovation in the long term. Evidence from the mechanism analysis highlights the critical roles played by factors such as industry competition, industry environmental uncertainty, R&D investments, financial constraints, and government subsidies in driving this process.

The effect of Dendrobium officinale extract in inhibiting the senescence of H2O2-induced fibroblasts based on the senescence-associated secretory phenotype.

BACKGROUND: Dendrobium officinale is widely used for a long time in China, with effect of antioxidation, antitumor, enhancing immunity and so on. In recent years, Dendrobium officinale has been gradually used in cosmetics due to its powerful beauty effects. AIMS: Based on senescence-associated secretory phenotype (SASP), we studied the antiaging effect of Dendrobium officinale extract (DOE) on skin. METHODS: The senescent model of human skin fibroblasts was established by the induction of H2O2, and the content of SASP factors was tested after the treatment of DOE, such as interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1) and matrix metalloproteinase-1 (MMP-1). RESULTS: It was found that after the treatment with different concentrations of DOE, the contents of IL-6, MCP-1 and MMP-1 all decreased in different degrees. CONCLUSIONS: It indicated that DOE could inhibit the secretion of SASP factors and was a promising natural antiaging agent.

Synergistic effect of Panax ginseng, Polygonatum cyrtonema, Epiphyllum oxypetalum, Nelumbo nucifera and Osmanthus fragrans extracts on skin aging regulation. From in silico predictions to in vitro outcome.

Intrinsic and extrinsic aging affect the health of human skin. Extracellular matrix protein degradation, DNA damage and oxidative stress are known to disturb skin architecture and skin homeostasis leading to skin aging. Traditional Chinese Medicine (TCM) delivers a large amount of knowledge regarding the phytotherapeutic power of diverse plants. Panax ginseng, Polygonatum cyrtonema, Epiphyllum oxypetalum, Nelumbo nucifera and Osmanthus fragrans are five plants used in TCM for their protective effect. In this study, several combinations of these TCM plants were explored: first, an in silico analysis was performed to predict their potential to target biological activities in the skin and then, some predictions were verified with in vitro studies to underline the synergistic effect of plant extracts. The results showed a stronger anti-aging activity for the combination with the five plants compared to the combination with Panax ginseng, Polygonatum cyrtonema, Epiphyllum oxypetalum and, compared to Panax ginseng alone.

A new species of the genus Achalinus (Squamata, Xenodermatidae) from southwest Hunan Province, China.

A new species of xenodermid snake, Achalinusnanshanensis H. Li, L.-Q. Zhu, Z.-Q. Zhang & X.-Y. Mo, sp. nov., is described based on three specimens collected from Nanshan National Park and Tongdao County of southwest Hunan Province. This new species is genetically distinct amongst its congeners with the mitochondrial COI uncorrected p-distance ranging from 4.4% (in A.yangdatongi) to 17.7% (in A.meiguensis). In addition, this new species can be distinguished from its congeners by a combination of the following morphological characters: (1) dorsal scales with 23 or 25 rows throughout and strongly keeled; (2) tail relatively longer so that TaL/ToL = 0.215-0.248; (3) length of suture between internasals significantly longer than that between prefrontals, LSBI/LSBP = 1.66-1.84; (4) single loreal scale present; (5) SPL 6 in number, with the fourth and fifth contacting eye; (6) IFL 6 in number, with the first three touching the first pair of chin shields; (7) TMP is 2-2-4/2-2(3)-4, with the anterior pair elongated and in contact with the eye; (8) ventrals 2 + 147-158; (9) subcaudals 64-77, unpaired; (10) dorsal body brownish black, with a bright yellow neck collar extending to the head and abdomen in the occipital region. The recognition of the new species increases the number of described Achalinus species to 28, of which 21 are found in China.

Alpha5 nicotine acetylcholine receptor subunit promotes intrahepatic cholangiocarcinoma metastasis.

Neurotransmitter-initiated signaling pathway were reported to play an important role in regulating the malignant phenotype of tumor cells. Cancer cells could exhibit a "neural addiction" property and build up local nerve networks to achieve an enhanced neurotransmitter-initiated signaling through nerve growth factor-mediated axonogenesis. Targeting the dysregulated nervous systems might represent a novel strategy for cancer treatment. However, whether intrahepatic cholangiocarcinoma (ICC) could build its own nerve networks and the role of neurotransmitters in the progression ICC remains largely unknown. Immunofluorescence staining and Enzyme-linked immunosorbent assay suggested that ICC cells and the infiltrated nerves could generate a tumor microenvironment rich in acetylcholine that promotes ICC metastasis by inducing epithelial-mesenchymal transition (EMT). Acetylcholine promoted ICC metastasis through interacting with its receptor, alpha 5 nicotine acetylcholine receptor subunits (CHRNA5). Furthermore, acetylcholine/CHRNA5 axis activated GSK3ß/ß-catenin signaling pathway partially through the influx of Ca2+-mediated activation of Ca/calmodulin-dependent protein kinases (CAMKII). In addition, acetylcholine signaling activation also expanded nerve infiltration through increasing the expression of Brain-Derived Neurotrophic Factor (BDNF), which formed a feedforward acetylcholine-BDNF axis to promote ICC progression. KN93, a small-molecule inhibitor of CAMKII, significantly inhibited the migration and enhanced the sensitivity to gemcitabine of ICC cells. Above all, Acetylcholine/CHRNA5 axis increased the expression of ß-catenin to promote the metastasis and resistance to gemcitabine of ICC via CAMKII/GSK3ß signaling, and the CAMKII inhibitor KN93 may be an effective therapeutic strategy for combating ICC metastasis.