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Aggregated α-synuclein (α-SYN) proteins, encoded by the SNCA gene, are hallmarks of Lewy body disease (LBD), affecting multiple brain regions. However, the specific mechanisms underlying α-SYN pathology in cortical neurons, crucial for LBD-associated dementia, remain unclear. Here, we recapitulated α-SYN pathologies in human induced pluripotent stem cells (iPSCs)-derived cortical organoids generated from patients with LBD with SNCA gene triplication. Single-cell RNA sequencing, combined with functional and molecular validation, identified synaptic and mitochondrial dysfunction in excitatory neurons exhibiting high expression of the SNCA gene, aligning with observations in the cortex of autopsy-confirmed LBD human brains. Furthermore, we screened 1280 Food and Drug Administration-approved drugs and identified four candidates (entacapone, tolcapone, phenazopyridine hydrochloride, and zalcitabine) that inhibited α-SYN seeding activity in real-time quaking-induced conversion assays with human brains, reduced α-SYN aggregation, and alleviated mitochondrial dysfunction in SNCA triplication organoids and excitatory neurons. Our findings establish human cortical LBD models and suggest potential therapeutic drugs targeting α-SYN aggregation for LBD.

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The maturation of brain microvascular endothelial cells leads to the formation of a tightly sealed monolayer, known as the blood-brain barrier (BBB). The BBB damage is associated with the pathogenesis of age-related neurodegenerative diseases including vascular cognitive impairment and Alzheimer's disease. Growing knowledge in the field of epigenetics can enhance the understanding of molecular profile of the BBB and has great potential for the development of novel therapeutic strategies or targets to repair a disrupted BBB. Histone deacetylases (HDACs) inhibitors are epigenetic regulators that can induce acetylation of histones and induce open chromatin conformation, promoting gene expression by enhancing the binding of DNA with transcription factors. We investigated how HDAC inhibition influences the barrier integrity using immortalized human endothelial cells (HCMEC/D3) and the human induced pluripotent stem cell (iPSC)-derived brain vascular endothelial cells. The endothelial cells were treated with or without a novel compound named W2A-16. W2A-16 not only activates Wnt/ß-catenin signaling but also functions as a class I HDAC inhibitor. We demonstrated that the administration with W2A-16 sustained barrier properties of the monolayer of endothelial cells, as evidenced by increased trans-endothelial electrical resistance (TEER). The BBB-related genes and protein expression were also increased compared with non-treated controls. Analysis of transcript profiles through RNA-sequencing in hCMEC/D3 cells indicated that W2A-16 potentially enhances BBB integrity by influencing genes associated with the regulation of the extracellular microenvironment. These findings collectively propose that the HDAC inhibition by W2A-16 plays a facilitating role in the formation of the BBB. Pharmacological approaches to inhibit HDAC may be a potential therapeutic strategy to boost and/or restore BBB integrity.

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Sinonasal squamous cell carcinoma (SNSCC) is the most common, high-aggressive sinonasal malignancies that have remained relatively stable poor outcomes over the past decade. As a first-line treatment for SNSCC, surgery plus adjuvant radiotherapy is recommended. However, complete surgical resection may not be appropriate due to the proximity of the nasal cavity and sinuses to key structures such as orbit or intracranial. Currently, immune checkpoint inhibitors (ICIs) have been established as one of the first-line therapies for many solid tumors with unresectable stage. However, evidence on the efficacy of ICIs in sinonasal malignancy is scarce and no ICIs are approved for use in SNSCC up to day. In this report, we report a case of a 64-year-old man with SNSCC treated by multi-protocol exploration. The patient achieved pathological complete response (pCR) after receiving two cycles of Docetaxel and cisplatin combined with tislelizumab. To the best of our knowledge, this is the first case of SNSCC treated with tislelizumab that achieved pCR. This case offers real-world evidence that chemotherapy plus immunotherapy is a promising treatment for SNSCC.

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Chemosensory protein (CSP) genes significantly enriched in the female antennae are potential molecular candidates for mediating female oviposition behaviors. In this study, we presented the interaction mechanisms of a female-antenna-biased PxutCSP19 in Papilio xuthus to 47 host volatiles, four biopesticides and 24 synthetic insecticides. Using a bioinformatics-based homology search, 22 genes orthologous to PxutCSP19 were identified from 22 other Papilio butterflies with high sequence identities to each other (73.20~98.72%). Multiple alignment analyses revealed a particularly extended N-terminus of Papilio CSP19s (an average of 154 residues) compared to insects' typical CSPs (approximately 120 residues). The expression profiles indicated that PxutCSP19 was significantly enriched in the female antennae, with a 31.81-fold difference relative to the male antennae. In ligand-binding assays, PxutCSP19 could strongly bind six host odorants with high affinities, ranging from dissociation constant (Ki) values of 20.44 ± 0.64 µM to 22.71 ± 0.73 µM. Notably, this protein was tuned to a monoterpenoid alcohol, linalool, which generally existed in the Rutaceae plants and elicited electrophysiological and behavioral activities of the swallowtail butterfly. On the other hand, PxutCSP19 was also capable of binding eight insecticides with stronger binding abilities (Ki < 12 µM) compared to host odorants. When an extended N-terminal region of PxutCSP19 was truncated into two different proteins, they did not significantly affect the binding of PxutCSP19 to ligands with high affinities, suggesting that this extended N-terminal sequences were not involved in the specificity of ligand recognition. Altogether, our study sheds light on the putative roles of PxutCSP19 enriched in the female antennae of P. xuthus in the perception of host volatiles and the sequestering of insecticides, and it complements the knowledge of butterfly CSPs in olfaction and insecticide resistance.

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Hyperlipidemia is a prevalent chronic metabolic disease that severely affects human health. Currently, commonly used clinical therapeutic drugs are prone to drug dependence and toxic side effects. Dietary intervention for treating chronic metabolic diseases has received widespread attention. Rosa sterilis is a characteristic fruit tree in China whose fruits are rich in flavonoids, which have been shown to have a therapeutic effect on hyperlipidemia; however, their exact molecular mechanism of action remains unclear. Therefore, this study aimed to investigate the therapeutic effects of R. sterilis total flavonoid extract (RS) on hyperlipidemia and its possible mechanisms. A hyperlipidemic zebrafish model was established using egg yolk powder and then treated with RS to observe changes in the integral optical density in the tail vessels. Network pharmacology and molecular docking were used to investigate the potential mechanism of action of RS for the treatment of hyperlipidemia. The results showed that RS exhibited favorable hypolipidemic effects on zebrafish in the concentration range of 3.0-30.0 µg/mL in a dose-dependent manner. Topological and molecular docking analyses identified HSP90AA1, PPARA, and MMP9 as key targets for hypolipidemic effects, which were exerted mainly through lipolytic regulation of adipocytes and lipids; pathway analysis revealed enrichment in atherosclerosis, chemical carcinogenic-receptor activation pathways in cancers, and proteoglycans in prostate cancer and other cancers. Mover, chinensinaphthol possessed higher content and better target binding ability, which suggested that chinensinaphthol might be an important component of RS with hypolipidemic active function. These findings provide a direction for further research on RS interventions for the treatment of hyperlipidemia.

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BACKGROUND/PURPOSE: This study compared the clinical efficacy and safety of laparoscopic versus open resection for hilar cholangiocarcinoma (HCCA) and analyzed potential prognostic factors. METHODS: The study included patients who underwent HCCA resection at our center from March 2012 to February 2022. Perioperative complications and postoperative prognosis were compared between the laparoscopic surgery (LS) and open surgery (OS) groups. RESULTS: After screening 313 HCCA patients, 68 patients were eligible for the study in the LS group (n = 40) and OS group (n = 28). Kaplan-Meier survival curve analysis revealed that overall survival > 2 years and 3-year disease-free survival (DFS) were more common in the LS than OS group, but the rate of 2-year DFS was lower in the LS group than OS group. Cox multivariate regression analysis revealed age (< 65 years), radical resection, and postoperative adjuvant therapy were associated with reduced risk of death (hazard ratio [HR] = 0.380, 95% confidence interval [CI] = 0.150-0.940, P = 0.036; HR = 0.080, 95% CI = 0.010-0.710, P = 0.024 and HR = 0.380, 95% CI = 0.150-0.960, P = 0.040), whereas preoperative biliary drainage was an independent factor associated with increased risk of death (HR = 2.810, 95% CI = 1.130-6.950, P = 0.026). Perineuronal invasion was identified as an independent risk factor affecting DFS (HR = 5.180, 95% CI = 1.170-22.960, P = 0.030). CONCLUSIONS: Compared with OS, laparoscopic HCCA resection does not significantly differ in terms of clinical efficacy. Age (<65 years), radical resection, and postoperative adjuvant therapy reduce the risk of death, and preoperative biliary drainage increases the risk of death.

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Centromere protein L (CENPL) is involved in the mitotic process of eukaryotic cells and the development of various types of cancer. However, its role in hepatocellular carcinoma (HCC) remains unclear. This study aimed to investigate the expression and clinical significance of CENPL in HCC, and explore its involvement in regulating HCC cell proliferation, apoptosis, cell cycle, and glycolysis both in vivo and in vitro. CENPL expression was analyzed in HCC and normal liver tissues using The Cancer Genome Atlas, Gene Expression Omnibus mining, real-time quantitative polymerase chain reaction, and immunohistochemistry. Functional assays were used to assess the role of CENPL in HCC cell proliferation, apoptosis, cell cycle, and glycolysis. The potential pathways underlying the regulatory effects of CENPL, as well as the expression of mitogen-activated protein kinase (MAPK) signaling pathway-related molecules and markers of proliferation and glycolysis were investigated. CENPL was significantly upregulated in HCC tissue and associated with multiple clinicopathological features and poor patient prognosis. Univariate and multivariate analyses demonstrated that CENPL may serve as an independent prognostic factor for HCC. Upregulation of CENPL in HCC regulated tumor proliferation and glycolytic processes. Mechanistic studies revealed that differentially expressed genes between the CENPL-overexpressing and control groups were mainly concentrated in the MAPK signaling pathway. Pathway inhibition analysis indicated that CENPL activated the MEK1/2-ERK1/2 signaling pathway to promote proliferation and glycolysis in HCC cells. This study elucidated the role of CENPL in regulating cell proliferation, apoptosis, cell cycle, and glycolysis in HCC. CENPL may represent a therapeutic target and prognostic biomarker for HCC.

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Rosa roxburghii and Rosa sterilis, two species belonging to the Rosaceae family, are widespread in the southwest of China. These species have gained recognition for their remarkable abundance of ascorbate in their fresh fruits, making them an ideal vitamin C resource. In this study, we generated two high-quality chromosome-scale genome assemblies for R. roxburghii and R. sterilis, with genome sizes of 504 and 981.2 Mb, respectively. Notably, we present a haplotype-resolved, chromosome-scale assembly for diploid R. sterilis. Our results indicated that R. sterilis originated from the hybridization of R. roxburghii and R. longicuspis. Genome analysis revealed the absence of recent whole-genome duplications in both species and identified a series of duplicated genes that possibly contributing to the accumulation of flavonoids. We identified two genes in the ascorbate synthesis pathway, GGP and GalLDH, that show signs of positive selection, along with high expression levels of GDP-d-mannose 3', 5'-epimerase (GME) and GDP-l-galactose phosphorylase (GGP) during fruit development. Furthermore, through co-expression network analysis, we identified key hub genes (MYB5 and bZIP) that likely regulate genes in the ascorbate synthesis pathway, promoting ascorbate biosynthesis. Additionally, we observed the expansion of terpene synthase genes in these two species and tissue expression patterns, suggesting their involvement in terpenoid biosynthesis. Our research provides valuable insights into genome evolution and the molecular basis of the high concentration of ascorbate in these two Rosa species.

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Covalent organic frameworks (COFs) represent an important class of crystalline porous materials with designable structures and functions. The interconnected organic monomers, featuring pre-designed symmetries and connectivities, dictate the structures of COFs, endowing them with high thermal and chemical stability, large surface area, and tunable micropores. Furthermore, by utilizing pre-functionalization or post-synthetic functionalization strategies, COFs can acquire multifunctionalities, leading to their versatile applications in gas separation/storage, catalysis, and optoelectronic devices. Our review provides a comprehensive account of the latest advancements in the principles, methods, and techniques for structural design and determination of COFs. These cutting-edge approaches enable the rational design and precise elucidation of COF structures, addressing fundamental physicochemical challenges associated with host-guest interactions, topological transformations, network interpenetration, and defect-mediated catalysis.

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The orientation of the oligophagous cone-feeding moth Dioryctria abietella (Lepidoptera: Pyralidae) to host plants primarily relies on olfactory-related proteins, particularly those candidates highly expressed in antennae. Here, through a combination of expression profile, ligand-binding assay, molecular docking and site-directed mutagenesis strategies, we characterized the chemosensory protein (CSP) gene family in D. abietella. Quantitative real-time PCR (qPCR) analyses revealed the detectable expression of all 22 DabiCSPs in the antennae, of which seven genes were significantly enriched in this tissue. In addition, the majority of the genes (19/22 relatives) had the expression in at least one reproductive tissue. In the interactions of four antenna-dominant DabiCSPs and different chemical classes, DabiCSP1 was broadly tuned to 27 plant-derived odors, three man-made insecticides and one herbicide with high affinities (Ki < 6.60 µM). By contrast, three other DabiCSPs (DabiCSP4, CSP6 and CSP17) exhibited a narrow odor binding spectrum, in response to six compounds for each protein. Our mutation analyses combined with molecular docking simulations and binding assays further identified four key residues (Tyr25, Thr26, Ile65 and Val69) in the interactions of DabiCSP1 and ligands, of which binding abilities of this protein to 12, 15, 16 and three compounds were significantly decreased compared to the wildtype protein, respectively. Our study reveals different odor binding spectra of four DabiCSPs enriched in antennae and identifies key residues responsible for the binding of DabiCSP1 and potentially active compounds for the control of this pest.

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Volatile organic compounds (VOCs) and flavor characteristics of Rosa roxburghii Tratt. (RR) and Rosa sterilis (RS) were analyzed using headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS). The flavor network was constructed by combining relative odor activity values (ROAVs), and the signature differential flavor components were screened using orthogonal partial least squares discriminant analysis (OPLS-DA) and random forest (RF). The results showed that 61 VOCs were detected in both RR and RS: 48 in RR, and 26 in RS. There were six key flavor components (ROAVs ≥ 1) in RR, namely nonanal, ethyl butanoate, ethyl hexanoate, (3Z)-3-hexen-1-yl acetate, ethyl caprylate, and styrene, among which ethyl butanoate had the highest contribution, whereas there were eight key flavor components (ROAVs ≥ 1) in RS, namely 2-nonanol, (E)-2-hexenal, nonanal, methyl salicylate, ß-ocimene, caryophyllene, α-ionone, and styrene, among which nonanal contributed the most to RS. The flavor of RR is primarily fruity, sweet, green banana, and waxy, while the flavor of RS is primarily sweet and floral. In addition, OPLS-DA and RF suggested that (E)-2-hexenal, ethyl caprylate, ß-ocimene, and ethyl butanoate could be the signature differential flavor components for distinguishing between RR and RS. In this study, the differences in VOCs between RR and RS were analyzed to provide a basis for further development and utilization.

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A high-efficiency drug screening method is urgently needed due to the expanding number of potential targets and the extremely long time required to assess them. To date, high throughput and high content have not been successfully combined in image-based drug screening, which is the main obstacle to improve the efficiency. Here, we establish a high-throughput and high-content drug screening method by preparing a superhydrophobic microwell array plate (SMAP) and combining it with protein-retention expansion microscopy (proExM). Primarily, we described a flexible method to prepare the SMAP based on photolithography. Cells were cultured in the SMAP and treated with different drugs using a microcolumn-microwell sandwiching technology. After drug treatment, proExM was applied to realize super-resolution imaging. As a demonstration, a 7 × 7 image array of microtubules was successfully collected within 3 h with 68 nm resolution using this method. Qualitative and quantitative analyses of microtubule and mitochondria morphological changes after drug treatment suggested that more details were revealed after applying proExM, demonstrating the successful combination of high throughput and high content.

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BACKGROUND: The apolipoprotein E (APOE) gene is the strongest genetic risk factor for Alzheimer's disease (AD); however, how it modulates brain homeostasis is not clear. The apoE protein is a major lipid carrier in the brain transporting lipids such as cholesterol among different brain cell types. METHODS: We generated three-dimensional (3-D) cerebral organoids from human parental iPSC lines and its isogenic APOE-deficient (APOE-/-) iPSC line. To elucidate the cell-type-specific effects of APOE deficiency in the cerebral organoids, we performed scRNA-seq in the parental and APOE-/- cerebral organoids at Day 90. RESULTS: We show that APOE deficiency in human iPSC-derived cerebral organoids impacts brain lipid homeostasis by modulating multiple cellular and molecular pathways. Molecular profiling through single-cell RNA sequencing revealed that APOE deficiency leads to changes in cellular composition of isogenic cerebral organoids likely by modulating the eukaryotic initiation factor 2 (EIF2) signaling pathway as these events were alleviated by the treatment of an integrated stress response inhibitor (ISRIB). APOE deletion also leads to activation of the Wnt/ß-catenin signaling pathway with concomitant decrease of secreted frizzled-related protein 1 (SFRP1) expression in glia cells. Importantly, the critical role of apoE in cell-type-specific lipid homeostasis was observed upon APOE deletion in cerebral organoids with a specific upregulation of cholesterol biosynthesis in excitatory neurons and excessive lipid accumulation in astrocytes. Relevant to human AD, APOE4 cerebral organoids show altered neurogenesis and cholesterol metabolism compared to those with APOE3. CONCLUSIONS: Our work demonstrates critical roles of apoE in brain homeostasis and offers critical insights into the APOE4-related pathogenic mechanisms.

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The prevalence of Alzheimer's disease is greater in women, but the underlying mechanisms remain to be elucidated. We herein demonstrated that α-secretase ADAM10 was downregulated and ADAM10 inhibitor sFRP1 was upregulated in 5xFAD mice. While there were no sex effects on ADAM10 protein and sFRP1 mRNA levels, female 5xFAD and age-matched non-transgenic mice exhibited higher levels of sFRP1 protein than corresponding male mice. Importantly, female 5xFAD mice accumulated more Aß than males, and sFRP1 protein levels were positively associated with Aß42 levels in 5xFAD mice. Our study suggests that sFRP1 is associated with amyloid pathology in a sex-dependent manner.

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BACKGROUND: Alzheimer's disease (AD) is a progressive and age-associated neurodegenerative disorder that affects women disproportionally. However, the underlying mechanisms are poorly characterized. Moreover, while the interplay between sex and ApoE genotype in AD has been investigated, multi-omics studies to understand this interaction are limited. Therefore, we applied systems biology approaches to investigate sex-specific molecular networks of AD. METHODS: We integrated large-scale human postmortem brain transcriptomic data of AD from two cohorts (MSBB and ROSMAP) via multiscale network analysis and identified key drivers with sexually dimorphic expression patterns and/or different responses to APOE genotypes between sexes. The expression patterns and functional relevance of the top sex-specific network driver of AD were further investigated using postmortem human brain samples and gene perturbation experiments in AD mouse models. RESULTS: Gene expression changes in AD versus control were identified for each sex. Gene co-expression networks were constructed for each sex to identify AD-associated co-expressed gene modules shared by males and females or specific to each sex. Key network regulators were further identified as potential drivers of sex differences in AD development. LRP10 was identified as a top driver of the sex differences in AD pathogenesis and manifestation. Changes of LRP10 expression at the mRNA and protein levels were further validated in human AD brain samples. Gene perturbation experiments in EFAD mouse models demonstrated that LRP10 differentially affected cognitive function and AD pathology in sex- and APOE genotype-specific manners. A comprehensive mapping of brain cells in LRP10 over-expressed (OE) female E4FAD mice suggested neurons and microglia as the most affected cell populations. The female-specific targets of LRP10 identified from the single cell RNA-sequencing (scRNA-seq) data of the LRP10 OE E4FAD mouse brains were significantly enriched in the LRP10-centered subnetworks in female AD subjects, validating LRP10 as a key network regulator of AD in females. Eight LRP10 binding partners were identified by the yeast two-hybrid system screening, and LRP10 over-expression reduced the association of LRP10 with one binding partner CD34. CONCLUSIONS: These findings provide insights into key mechanisms mediating sex differences in AD pathogenesis and will facilitate the development of sex- and APOE genotype-specific therapies for AD.

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Subcritical water extraction was used to extract bioactive phenolic compounds from Vaccinium dunalianum Wight leaves. The optimal extraction conditions were determined as an extraction temperature of 150 °C, an extraction time of 40 min, and a liquid-solid ratio of 35 : 1 mL/g. The total phenolic content reached 21.35 mg gallic acid /g, which was 16 % higher than that by hot water extraction. The subcritical water extraction extract exhibited strong scavenging activity of DPPH free radical and ABTS+ free radical, as well as significant tyrosinase inhibitory activity. The study suggests that subcritical water extraction can alter the composition of the extracts, leading to the production of various phenolic compounds, effective antioxidants, and tyrosinase inhibitors from Vaccinium dulciana Wight leaves. These findings confirm the potential of Vaccinium dunalianum Wight as a natural antioxidant molecule source for the medicine and food industries, and for the therapy of skin pigmentation disorders.

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Wnt and R-spondin (Rspo) proteins are two major types of endogenous Wnt/ß-catenin signaling agonists. While Wnt/ß-catenin signaling is greatly diminished in Alzheimer's disease (AD), it remains to be elucidated whether the inhibition of this pathway is associated with dysregulation of Wnt and Rspo proteins. By analyzing temporal cortex RNA-seq data of the human postmortem brain samples, we found that WNT1 and RRPO2 were significantly downregulated in human AD brains. In addition, the expression of Wnt acyltransferase porcupine (PORCN), which is essential for Wnt maturation and secretion, was greatly deceased in these human AD brains. Interestingly, the lowest levels of WNT1, PORCN, and RSPO2 expression were found in human AD brains carrying two copies of APOE4 allele, the strongest genetic risk factor of late-onset AD. Importantly, there were positive correlations among the levels of WNT1, PORCN, and RSPO2 expression in human AD brains. Supporting observations in humans, Wnt1, PORCN, and Rspo2 were downregulated and Wnt/ß-catenin signaling was diminished in the 5xFAD amyloid model mice. In human APOE-targeted replacement mice, downregulation of WNT1, PORCN, and RSPO2 expression was positively associated with aging and APOE4 genotype. Finally, WNT1 and PORCN expression and Wnt/ß-catenin signaling were inhibited in human APOE4 iPSC-derived astrocytes when compared to the isogenic APOE3 iPSC-derived astrocytes. Altogether, our findings suggest that the dysregulations of Wnt1, PORCN, and Rspo2 could be coordinated together to diminish Wnt/ß-catenin signaling in aging- and APOE4-dependent manners in the AD brain.

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Aqueous film-forming foam (AFFF) in firefighting foam can effectively extinguish oil fire. However, AFFF will be phased out due to the presence of persistent organic pollutants such as perfluorooctane sulfonates. It is necessary to explore environment-friendly foam extinguishing agent. Silicone surfactant is a kind of environmentally friendly surfactants with high surface activity, which can be used as a substitute for fluorocarbon surfactant. In this study, silicone surfactant and cationic surfactant were combined to adsorb nanosilica particles to form a multiphase system. The foaming property and foam stability of the multiphase system were tested by stirring method. The foam suppression effect on fuel vapor, fire extinguishing, and burn-back performance was tested by self-designed foam suppression device and foam generating device of porous glass with recyclable liquid supply. The experimental results show that foam stability is enhanced and the foaming property is slightly decreased after the addition of nanoparticles. When the component of three-phase foam reaches the optimum, its suppression effect on fuel vapor is better than that of two-phase foam. The results of fire extinguishing and burn-back performance test showed that the fire extinguishing effect of three-phase foam was slightly worse than that of AFFF, but it has extremely strong burn-back performance.

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The influence of son of sevenless homolog 1 (SOS1) on invasion and metastasis of hepatocellular carcinoma (HCC) cells was investigated. HCC cells were transfected with siRNA and lentivirus to achieve SOS1 knock down/overexpression and changes in RNA and protein levels analyzed by q-PCR and Western blotting (WB). Transwell assay was utilized to assess variations in cell invasion and migration in vitro and by a lung metastasis model of liver cancer in vivo. High expression of SOS1 was observed in most human liver cancers, which indicated a worse prognosis. SOS1 knockout in HepG2 cells significantly decreased cell invasion and migration. SOS1 knockout also reduced the number of metastatic foci in a lung metastasis model of HCC established in nude mice. SOS1 knockout inhibited the epithelial-mesenchymal transition (EMT) in HepG2 cells as well as the PI3K/AKT/mTOR pathway. Overexpression of SOS1 in Huh7 cells had the opposite effect. To conclude, SOS1 may induce the EMT by the activation of the PI3K/AKT/mTOR pathway, thereby enhancing invasion, migration and metastasis of HCC cells. These findings may expose SOS1 as a new HCC therapeutic target.

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BACKGROUND: The aggregation and spread of α-synuclein (α-Syn) protein and related neuronal toxicity are the key pathological features of Parkinson's disease (PD) and Lewy body dementia (LBD). Studies have shown that pathological species of α-Syn and tau can spread in a prion-like manner between neurons, although these two proteins have distinct pathological roles and contribute to different neurodegenerative diseases. It is reported that the low-density lipoprotein receptor-related protein 1 (LRP1) regulates the spread of tau proteins; however, the molecular regulatory mechanisms of α-Syn uptake and spread, and whether it is also regulated by LRP1, remain poorly understood. METHODS: We established LRP1 knockout (LRP1-KO) human induced pluripotent stem cells (iPSCs) isogenic lines using a CRISPR/Cas9 strategy and generated iPSC-derived neurons (iPSNs) to test the role of LRP1 in α-Syn uptake. We treated the iPSNs with fluorescently labeled α-Syn protein and measured the internalization of α-Syn using flow cytometry. Three forms of α-Syn species were tested: monomers, oligomers, and pre-formed fibrils (PFFs). To examine whether the lysine residues of α-Syn are involved in LRP1-mediated uptake, we capped the amines of lysines on α-Syn with sulfo-NHS acetate and then measured the internalization. We also tested whether the N-terminus of α-Syn is critical for LRP1-mediated internalization. Lastly, we investigated the role of Lrp1 in regulating α-Syn spread with a neuronal Lrp1 conditional knockout (Lrp1-nKO) mouse model. We generated adeno-associated viruses (AAVs) that allowed for distinguishing the α-Syn expression versus spread and injected them into the hippocampus of six-month-old Lrp1-nKO mice and the littermate wild type (WT) controls. The spread of α-Syn was evaluated three months after the injection. RESULTS: We found that the uptake of both monomeric and oligomeric α-Syn was significantly reduced in iPSNs with LRP1-KO compared with the WT controls. The uptake of α-Syn PFFs was also inhibited in LRP1-KO iPSNs, albeit to a much lesser extent compared to α-Syn monomers and oligomers. The blocking of lysine residues on α-Syn effectively decreased the uptake of α-Syn in iPSNs and the N-terminus of α-Syn was critical for LRP1-mediated α-Syn uptake. Finally, in the Lrp1-nKO mice, the spread of α-Syn was significantly reduced compared with the WT littermates. CONCLUSIONS: We identified LRP1 as a key regulator of α-Syn neuronal uptake, as well as an important mediator of α-Syn spread in the brain. This study provides new knowledge on the physiological and pathological role of LRP1 in α-Syn trafficking and pathology, offering insight for the treatment of synucleinopathies.

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