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Purpose: This study aims to explore the potential mechanisms of esculin in the treatment of renal cell carcinoma (RCC). Methods: We employed network pharmacology to predict the potential mechanisms and targets of esculin in RCC. Molecular docking techniques were then employed to validate the predicted targets. Additionally, a series of in vitro experiments were conducted to verify the anticancer effects of esculin on RCC cells, including the CCK-8 assay, EdU assay, wound healing assay, apoptosis assay, and Western blot. Results: Network pharmacology and molecular docking results identified GAPDH, TNF, GSK3B, CCND1, MCL1, IL2, and CDK2 as core targets. GO and KEGG analyses suggested that esculin may influence apoptotic processes and target the PI3K/Akt pathway in RCC. Furthermore, the CCK-8 assay demonstrated that esculin inhibited RCC cell viability. Microscopic observations revealed that following esculin treatment, there was an increase in cell crumpling, a reduction in cell density, and an accumulation of floating dead cells. Additionally, with increasing esculin concentrations, the proportion of EdU-positive cells decreased, the wound closure ratio decreased, the proportion of PI-positive cells increased, the expression levels of BAX and cleaved-caspase-3 proteins increased, and the expression level of Bcl2 protein decreased. These findings suggested that esculin inhibits the proliferation and migration of RCC cells while promoting apoptosis. Moreover, esculin was found to target GAPDH and inhibit the PI3K/Akt pathway. Conclusions: This study is the first to elucidate the therapeutic effects of esculin on RCC cells. The results provide evidence supporting the clinical application of esculin and introduce a promising new candidate for RCC treatment.

Abstract Introduction The optimal first-line immunotherapy regimen for advanced non-squamous non-small cell lung cancer (NS-NSCLC) patients with programmed cell death ligand 1 (PD-L1) expression ≥ 50% remains unclear. Our aim is to determine the most effective treatment regimen through a network meta-analysis (NMA) comparing these treatments. Methods A systematic search was performed in PubMed, Cochrane Library, Web of Science, and Embase databases, and a Bayesian network meta-analysis was conducted. To ensure transparency, the study was registered in the International Prospective Register of Systematic Reviews (CRD42022349712). Results The analysis included 11 randomized controlled trials (RCTs) with 2037 patients and 12 immunotherapy combinations. ICI-ICI, ICI alone, and chemotherapy-ICI showed significant advantages over chemotherapy in terms of overall survival (OS) and progression-free survival (PFS). Pembrolizumab plus chemotherapy showed the best OS results compared to chemotherapy. Tislelizumab plus chemotherapy and sintilimab plus chemotherapy provided the best PFS results. Conclusions For NS-NSCLC patients with PD-L1 ≥ 50%, pembrolizumab plus chemotherapy, tislelizumab plus chemotherapy, and sintilimab plus chemotherapy are recommended as good treatment options based on the results of this Network meta-analysis (NMA).

Abstract Background Although spatial organization of compartments and topologically associating domains at large scale is relatively well studied, the spatial organization of regulatory elements at fine scale is poorly understood in plants. Results Here we perform high-resolution chromatin interaction analysis using paired-end tag sequencing approach. We map chromatin interactions tethered with RNA polymerase II and associated with heterochromatic, transcriptionally active, and Polycomb-repressive histone modifications in Arabidopsis. Analysis of the regulatory repertoire shows that distal active cis-regulatory elements are linked to their target genes through long-range chromatin interactions with increased expression of the target genes, while poised cis-regulatory elements are linked to their target genes through long-range chromatin interactions with depressed expression of the target genes. Furthermore, we demonstrate that transcription factor MYC2 is critical for chromatin spatial organization, and propose that MYC2 occupancy and MYC2-mediated chromatin interactions coordinately facilitate transcription within the framework of 3D chromatin architecture. Analysis of functionally related gene-defined chromatin connectivity networks reveals that genes implicated in flowering-time control are functionally compartmentalized into separate subdomains via their spatial activity in the leaf or shoot apical meristem, linking active mark- or Polycomb-repressive mark-associated chromatin conformation to coordinated gene expression. Conclusion The results reveal that the regulation of gene transcription in Arabidopsis is not only by linear juxtaposition, but also by long-range chromatin interactions. Our study uncovers the fine scale genome organization of Arabidopsis and the potential roles of such organization in orchestrating transcription and development.

Introduction: Taiwan has several hepatitis C virus (HCV) hyper-endemic areas. We aimed to evaluate the effectiveness and safety of a collaborative HCV care system with an outreach decentralized strategy among the resource-constrained rural/remote areas of Taiwan. Methods: The pilot study was conducted in four high HCV-endemic townships in the rural/remote areas of Taoyuan, Alishan, Zhuoxi and Xiulin. Registered residents who worked or lived in the four areas and were aged 30–75 years were invited to participate in this program. Multidisciplinary HCV care teams provided outreach decentralized services of anti-HCV screening, link-to-diagnosis, and link-to-treatment with direct-acting antiviral agents (DAA). The primary end-point was sustained virological response (SVR). Results: Of 8291 registered residents who were invited as the target population, 7807 (94.2%) subjects received anti-HCV screening, with the average anti-HCV prevalence rate of 14.2% (1108/7807) (range among four areas: 11.8%–16.7%). The rate of link-to-diagnosis was 94.4% (1046/1108) of anti-HCV-positive subjects (range: 90.9%–100%) with an average HCV-viremic rate of 55.1% (576/1046) (range: 50.0%–64.3%). The link-to-treat rate was 94.4% (544/576) in HCV-viremic subjects (range from 92.7% to 97.2%). Overall, 523 (96.1%) patients achieved an SVR (range: 94.7%–97.6%). Eventually, the overall effectiveness was 80.7% (range: 74.6%–93.1%). The presence of hepatocellular carcinoma at baseline was the only factor associated with DAA failure. The DAA regimens were well-tolerated. Conclusion: The outreach decentralized community-based care system with DAA therapy was highly effective and safe in the achievement of HCV micro-elimination in the resource-constrained rural and remote regions, which could help us to tackle the disparity.

Abstract Somatic activating MAP2K1 mutations in endothelial cells (ECs) cause extracranial arteriovenous malformation (AVM). We previously reported the generation of a mouse line allowing inducible expression of constitutively active MAP2K1 (p.K57N) from the Rosa locus (R26 GT-Map2k1-GFP/+) and showed, using Tg-Cdh5CreER, that EC expression of mutant MAP2K1 is sufficient for the development of vascular malformations in the brain, ear, and intestines. To gain further insight into the mechanism by which mutant MAP2K1 drives AVM development, we induced MAP2K1 (p.K57N) expression in ECs of postnatal-day-1 pups (P1) and investigated the changes in gene expression in P9 brain ECs by RNA-seq. We found that over-expression of MAP2K1 altered the transcript abundance of > 1600 genes. Several genes had > 20-fold changes between MAP2K1 expressing and wild-type ECs; the highest were Col15a1 (39-fold) and Itgb3 (24-fold). Increased expression of COL15A1 in R26 GT-Map2k1-GFP/+; Tg-Cdh5CreER +/− brain ECs was validated by immunostaining. Ontology showed that differentially expressed genes were involved in processes important for vasculogenesis (e.g., cell migration, adhesion, extracellular matrix organization, tube formation, angiogenesis). Understanding how these genes and pathways contribute to AVM formation will help identify targets for therapeutic intervention.

Purpose: Medicinal and recreational cannabis use has grown exponentially, however, its effect on testicular function and spermatogenesis remains uncertain. The aim of this study was to evaluate the association between cannabis use and semen parameters in a cohort of Asian-American men with unknown fertility. Materials and Methods: Asian men were recruited to complete an online survey and submit a semen sample. Semen analysis, demographic data, lifestyle factors, and cannabis use habits were collected. Linear and logistic regression analyses were used to determine. Results: Among the 112 men included in this study, 51 used cannabis at least once in their lifetime, 30 men used cannabis at least once in the last 12 months, and 26 men used cannabis at least once in the last 30 days. Adjusted linear regression analyses identified an association between cannabis use in the previous 30 days and worse sperm morphology (β: -0.45, p=0.025) and sperm motility (β: -1.64, p=0.016). However, when stratifying by subfertile semen quality (i.e., WHO criteria), no association was identified between semen quality and cannabis use. Lower sperm morphology and motility are partially associated with recent cannabis use, while all other semen parameters are not. Conclusions: We did not observe any consistent associations between cannabis use on any semen parameters in Asian- American men. Further studies within the field are needed to explore racial and ethnic differences in semen quality and lifestyle factors.

Rainfall frequency analysis, an essential work for water resources management, is often conducted by using the annual maximum rainfall series. For rainfall stations with short record lengths and outliers presence, the use of annual maximum series for rainfall frequency analysis may yield design rainfall estimates of higher uncertainties. Moreover, for regions with cyclostationary climate patterns, the annual maximum rainfalls may be caused by different prevalent storm types, which differ in terms of their occurrence frequency and storm rainfall characteristics. In this study, we propose a novel event-maximum-rainfall-based mixture distribution modeling approach for rainfall frequency analysis. By considering the event-maximum rainfalls of individual storm events, the sample size for parameter estimation increases, and the uncertainty of design rainfall estimates reduces. Mixture distribution modeling enables a thorough investigation of the contributing probabilities of different storm types to the annual maximum rainfall. Through rigorous stochastic simulation, we demonstrated the superiority of the proposed approach over the conventional annual maximum rainfall approach. The proposed approach was applied to four representative rainfall stations in Taiwan, and the results revealed that the proposed approach is more robust than the conventional annual maximum rainfall approach. The results provide insights into the contributions of individual storm types to the annual maximum rainfall.

BackgroundEndovascular treatment of severe intracranial atherosclerotic stenosis (ICAS) using coronary drug-eluting stents (DESs) significantly reduces the risk of in-stent restenosis (ISR) and stroke recurrence. However, there are few reports regarding the treatment of ICAS with intracranial dedicated DES. Herein, we present our experience with the feasibility, safety, and medium-term follow-up outcomes of a novel intracranial DES, named NOVA stent, in patients with symptomatic severe ICAS (≥70%).MethodsFrom December 2021 to May 2022, patients with symptomatic severe ICAS who underwent implantation of the NOVA stent in our institution were retrospectively analyzed for procedural results, perioperative complications, imaging and clinical follow-up outcomes.ResultsTwenty-four patients, 16 (66.7%) with anterior circulation lesions and 8 (33.3%) with posterior circulation lesions, were enrolled. All patients with intracranial ICA (n = 6), middle cerebral artery (n = 10), basilar artery (n = 3), intracranial vertebral artery (n = 3), and the vertebrobasilar junction (n = 2) stenosis were treated successfully using NOVA stents. The severity of stenosis ranged from 75 to 96% (mean 85.9%) before treatment and this was reduced to 0 to 20% (mean 8.6%) immediately after stent placement. Symptomatic distal embolism occurred in one case; however, there were no other perioperative complications. The mean follow-up duration was 12.2 ± 1.06 months. No symptomatic ischemic events occurred during follow-up. Follow-up cerebral angiography was performed in 22 of 24 patients (91.7%), and significant ISR occurred in one patient (4.2%).ConclusionOur results demonstrate that implantation of the novel intracranial DES NOVA in severe ICAS is feasible, safe, and effective in selected cases, reducing the incidence of ISR, and showing excellent midterm clinical outcomes, providing a promising option for ICAS treatment.

Objectives: Lung adenocarcinomas have different prognoses depending on their histological growth patterns. Micropapillary growth within lung adenocarcinoma, particularly metastasis, is related to dismal prognostic outcome. Metastasis accounts for a major factor leading to mortality among lung cancer patients. Understanding the mechanisms underlying early stage metastasis can help develop novel treatments for improving patient survival. Methods: Here, quantitative mass spectrometry was conducted for comparing protein expression profiles among various histological subtypes, including adenocarcinoma in situ, minimally invasive adenocarcinoma, and invasive adenocarcinoma (including acinar and micropapillary [MIP] types). To determine the mechanism of MIP-associated metastasis, we identified a protein that was highly expressed in MIP. The expression of the selected highly expressed MIP protein was verified via immunohistochemical (IHC) analysis and its function was validated by an in vitro migration assay. Results: Proteomic data revealed that low-density lipoprotein receptor-related protein–associated protein 1 (LRPAP1) was highly expressed in MIP group, which was confirmed by IHC. The co-expressed proteins in this study, PSMD1 and HSP90AB1, have been reported to be highly expressed in different cancers and play an essential role in metastasis. We observed that LRPAP1 promoted lung cancer progression, including metastasis, invasion and proliferation in vitro and in vivo. Conclusion: LRPAP1 is necessary for MIP-associated metastasis and is the candidate novel anti-metastasis therapeutic target.

Abstract Background Resistance to immune checkpoint inhibitor (ICI) therapy narrows the efficacy of cancer immunotherapy. Although 4-1BB is a promising drug target as a costimulatory molecule of immune cells, no 4-1BB agonist has been given clinical approval because of severe liver toxicity or limited efficacy. Therefore, a safe and efficient immunostimulatory molecule is urgently needed for cancer immunotherapy. Methods HK010 was generated by antibody engineering, and the Fab/antigen complex structure was analyzed using crystallography. The affinity and activity of HK010 were detected by multiple in vitro bioassays, including enzyme-linked immunosorbent assay (ELISA), surface plasmon resonance (SPR), flow cytometry, and luciferase-reporter assays. Humanized mice bearing human PD-L1-expressing MC38 (MC38/hPDL1) or CT26 (CT26/hPDL1) tumor transplants were established to assess the in vivo antitumor activity of HK010. The pharmacokinetics (PK) and toxicity of HK010 were evaluated in cynomolgus monkeys. Results HK010 was generated as an Fc-muted immunoglobulin (Ig)G4 PD-L1x4-1BB bispecific antibody (BsAb) with a distinguished Fab/antigen complex structure, and maintained a high affinity for human PD-L1 (KD: 2.27 nM) and low affinity for human 4-1BB (KD: 493 nM) to achieve potent PD-1/PD-L1 blockade and appropriate 4-1BB agonism. HK010 exhibited synergistic antitumor activity by blocking the PD-1/PD-L1 signaling pathway and stimulating the 4-1BB signaling pathway simultaneously, and being strictly dependent on the PD-L1 receptor in vitro and in vivo. In particular, when the dose was decreased to 0.3 mg/kg, HK010 still showed a strong antitumor effect in a humanized mouse model bearing MC38/hPDL1 tumors. Strikingly, HK010 treatment enhanced antitumor immunity and induced durable antigen-specific immune memory to prevent rechallenged tumor growth by recruiting CD8+ T cells and other lymphocytes into tumor tissue and activating tumor-infiltrating lymphocytes. Moreover, HK010 not only did not induce nonspecific production of proinflammatory cytokines but was also observed to be well tolerated in cynomolgus monkeys in 5 week repeated-dose (5, 15, or 50 mg/kg) and single-dose (75 or 150 mg/kg) toxicity studies. Conclusion We generated an Fc-muted anti-PD-L1x4-1BB BsAb, HK010, with a distinguished structural interaction with PD-L1 and 4-1BB that exhibits a synergistic antitumor effect by blocking the PD-1/PD-L1 signaling pathway and stimulating the 4-1BB signaling pathway simultaneously. It is strictly dependent on the PD-L1 receptor with no systemic toxicity, which may offer a new option for cancer immunotherapy.

Bird monitoring is an important approach to studying the diversity and abundance of birds, especially during migration, as it can provide core data for bird conservation purposes. The previous methods for bird number estimation are largely based on manual counting, which suffers from low throughput and a high error rate. In this study, we aimed to provide an alternative bird-counting method from video datasets by using five available ImageJ methods: Particle Analyzer, Find Maxima, Watershed segmentation, TrackMate, and trainable WEKA segmentation. The numbers of birds and their XY coordinates were extracted from videos to conduct a side-by-side comparison with the manual counting results, and the three important criteria of the sensitivity, precision, and F1 score were calculated for the performance evaluation. From the tests, which we conducted for four different cases with different bird numbers or flying patterns, TrackMate had the best overall performance for counting birds and pinpointing their locations, followed by Particle Analyzer, Find Maxima, WEKA, and lastly, Watershed, which showed low precision in most of the cases. In summary, five ImageJ-based counting methods were compared in this study, and we validated that TrackMate obtains the best performance for bird counting and detection.

Introduction: Hepatocellular carcinoma (HCC) is the most common primary liver cancer, characterized by high mortality rate. In clinical practice, several makers of liver cancer, such as VEGFR1, FGFR1 and PDGFRα, were identified and their potentials as a therapeutic target were explored. However, the unsatisfied treatment results emphasized the needs of new therapeutic targets.Methods: 112 HCC patients samples were obtained to evaluate the expression of LRRC41, SOX9, CD44, and EPCAM in HCC, combined with prognosis analysis. A DEN-induced HCC rat model was constructed to verify the expression of LRRC41 and SOX9 in HCC and lung metastasis tissues. Immune score evaluation was analysized by bioinformatics methods. Network pharmacology was performed to explored the potential FDA-approved drugs targeting LRRC41.Results: Through analysis of the Timer database and tissue micro-array, we confirmed that LRRC41 was over-expressed in HCC and exhibited a significant positive correlation with recurrence and metastasis. Immunohistochemistry staining of human HCC tissue samples revealed significant upregulation of LRRC41, SOX9, CD44, and EPCAM, with LRRC41 showing a positive correlation with SOX9, CD44, and EPCAM expression. UALCAN database analysis indicated that LRRC41 and SOX9 contribute to poor prognosis whereas CD44 and EPCAM did not demonstrate the same significance. Furthermore, analysis of a DEN-induced HCC rat model confirmed the significantly elevated expression of LRRC41 and SOX9 in HCC and lung metastasis tissues. Drug sensitivity analysis and molecular docking targeting LRRC41 identified several FDA-approved drugs, which may have potential antitumor effects on HCC by targeting LRRC41.Conclusion: Our findings highlight the role of LRRC41 overexpression in promoting HCC progression and its association with a poor prognosis. Drug sensitivity analysis and molecular docking shows several FDA-approved drugs may be potential therapeutic targets for HCC. Targeting LRRC41 may hold promise as a potential therapeutic strategy for HCC.

Abstract Metastasis involves epithelial‐to‐mesenchymal transition (EMT), a process that is regulated by complex gene networks, where their deliberate disruption may yield a promising outcome. However, little is known about mechanisms that coordinate these metastasis‐associated networks. To address this gap, hub genes with broad engagement across various human cancers by analyzing the transcriptomes of different cancer cell types undergoing EMT are identified. The oncogenic signaling adaptor protein tyrosine 3‐monooxygenase/tryptophan 5‐monooxygenase activation protein gamma (YWHAG) is ranked top for its clinical relevance and impact. The cellular kinome and transcriptome data are surveyed to construct the regulome of YWHAG, revealing stress responses and metabolic processes during cancer EMT. It is demonstrated that a YWHAG‐dependent cytoprotective mechanism in the regulome is embedded in EMT‐associated networks to protect cancer cells from oxidative catastrophe through enhanced autophagy during EMT. YWHAG deficiency results in a rapid accumulation of reactive oxygen species (ROS), delayed EMT, and cell death. Tumor allografts show that metastasis potential and overall survival time are correlated with the YWHAG expression level of cancer cell lines. Metastasized tumors have higher expression of YWHAG and autophagy‐related genes than primary tumors. Silencing YWHAG diminishes primary tumor volumes, prevents metastasis, and prolongs the median survival period of the mice.

Abstract Uncontrolled and excessive progression of liver fibrosis is thought to be the prevalent pathophysiological cause of liver cirrhosis and hepatocellular cancer, and there are currently no effective antifibrotic therapeutic options available. Intercellular communication and cellular heterogeneity in the liver are involved in the progression of liver fibrosis, but the exact nature of the cellular phenotypic changes and patterns of interregulatory remain unclear. Here, we performed single‐cell RNA sequencing on nonparenchymal cells (NPCs) isolated from normal and fibrotic mouse livers. We identified eight main types of cells, including endothelial cells, hepatocytes, dendritic cells, B cells, natural killer/T (NK/T) cells, hepatic stellate cells (HSCs), cholangiocytes and macrophages, and revealed that macrophages and HSCs exhibit the most variance in transcriptional profile. Further analyses of HSCs and macrophage subpopulations and ligand–receptor interaction revealed a high heterogeneity characterization and tightly interregulated network of these two groups of cells in liver fibrosis. Finally, we uncovered a profibrotic Thbs1+ macrophage subcluster, which expands in mouse and human fibrotic livers, activating HSCs via PI3K/AKT/mTOR signaling pathway. Our findings decode unanticipated insights into the heterogeneity of HSCs and macrophages and their intercellular crosstalk at a single‐cell level, and may provide potential therapeutic strategies in liver fibrosis.

Accurate structural and functional imaging is vital for the diagnosis and prognosis of urinary system diseases. Fluorescence bioimaging in the second near-infrared spectral region (NIR-II, 1000–1700 nm) has shown advantages of higher spatial resolution, deeper penetration, and finer signal-to-background ratio (SBR) compared to the conventional fluorescence imaging methods but limited to its clinical inapplicability. Herein, we first report in vivo NIR-II fluorescence imaging of the urinary system enabled by a clinically approved and renal excretable dye methylene blue (MB), which cannot only achieve clear invasive/non-invasive urography but also noninvasively detect renal function efficiently. These results demonstrate that MB assisted NIR-II fluorescence imaging holds a great promise for structural and functional imaging of the urinary system both clinically and preclinically.

Arabidopsis H3K9 methyltransferases directly interact with ASYMMETRIC LEAVES1 (AS1) and AS2 to repress KNOTTED-LIKE FROM ARABIDOPSIS THALIANA 1 (KNAT1) and KNAT2 in leaf development.