Apigenin suppresses epithelial-mesenchymal transition in high glucose-induced retinal pigment epithelial cell by inhibiting CBP/p300-mediated histone acetylation.

Epithelial mesenchymal transition (EMT) is a critical process implicated in the pathogenesis of retinal fibrosis and the exacerbation of diabetic retinopathy (DR) within retinal pigment epithelium (RPE) cells. Apigenin (AP), a potential dietary supplement for managing diabetes and its associated complications, has demonstrated inhibitory effects on EMT in various diseases. However, the specific impact and underlying mechanisms of AP on EMT in RPE cells remain poorly understood. In this study, we have successfully validated the inhibitory effects of AP on high glucose-induced EMT in ARPE-19 cells and diabetic db/db mice. Notably, our findings have identified CBP/p300 as a potential therapeutic target for EMT in RPE cells and have further substantiated that AP effectively downregulates the expression of EMT-related genes by attenuating the activity of CBP/p300, consequently reducing histone acetylation alterations within the promoter region of these genes. Taken together, our results provide novel evidence supporting the inhibitory effect of AP on EMT in RPE cells, and highlight the potential of specifically targeting CBP/p300 as a strategy for inhibiting retinal fibrosis in the context of DR.

Antigout effects and mechanisms of total flavonoids from prunus tomentosa.

BACKGROUND: In recent years, hyperuricemia and acute gouty arthritis have become increasingly common, posing a serious threat to public health. Current treatments primarily involve Western medicines with associated toxic side effects. OBJECTIVE: This study aims to investigate the therapeutic effects of total flavones from Prunus tomentosa (PTTF) on a rat model of gout and explore the mechanism of PTTF's anti-gout action through the TLR4/NF-κB signaling pathway. METHODS: We measured serum uric acid (UA), creatinine (Cr), blood urea nitrogen (BUN), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1ß), and interleukin-6 (IL-6) levels using an enzyme-linked immunosorbent assay (ELISA). Histopathological changes were observed using HE staining, and the expression levels of relevant proteins were detected through Western blotting. RESULTS: After PTTF treatment, all indicators improved significantly. PTTF reduced blood levels of UA, Cr, BUN, IL-1ß, IL-6, and TNF-α, and decreased ankle swelling. CONCLUSIONS: PTTF may have a therapeutic effect on animal models of hyperuricemia and acute gouty arthritis by reducing serum UA levels, improving ankle swelling, and inhibiting inflammation. The primary mechanism involves the regulation of the TLR4/NF-κB signaling pathway to alleviate inflammation. Further research is needed to explore deeper mechanisms.

Cell-free DNA assay for malignancy classification of high-risk lung nodules.

OBJECTIVE: Although low-dose computed tomography has been proven effective to reduce lung cancer-specific mortality, a considerable proportion of surgically resected high-risk lung nodules were still confirmed pathologically benign. There is an unmet need of a novel method for malignancy classification in lung nodules. METHODS: We recruited 307 patients with high-risk lung nodules who underwent curative surgery, and 247 and 60 cases were pathologically confirmed malignant and benign lung lesions, respectively. Plasma samples from each patient were collected before surgery and performed low-depth (5×) whole-genome sequencing. We extracted cell-free DNA characteristics and determined radiomic features. We built models to classify the malignancy using our data and further validated models with 2 independent lung nodule cohorts. RESULTS: Our models using one type of profile were able to distinguish lung cancer and benign lung nodules at an area under the curve metrics of 0.69 to 0.91 in the study cohort. Integrating all the 5 base models using cell-free DNA profiles, the cell-free DNA-based ensemble model achieved an area under the curve of 0.95 (95% CI, 0.92-0.97) in the study cohort and 0.98 (95% CI, 0.96-1.00) in the validation cohort. At a specificity of 95.0%, the sensitivity reached 80.0% in the study cohort. With the same threshold, the specificity and sensitivity had similar performances in both validation cohorts. Furthermore, the performance of area under the curve reached 0.97 in both the study and validation cohorts when considering the radiomic profile. CONCLUSIONS: The cell-free DNA profiles-based method is an efficient noninvasive tool to distinguish malignancies and high-risk but pathologically benign lung nodules.

A novel imprinted locus on bovine chromosome 18 homologous with human chromosome 16q24.1.

Genomic imprinting is an epigenetic regulation mechanism in mammals resulting in the parentally dependent monoallelic expression of genes. Imprinting disorders in humans are associated with several congenital syndromes and cancers and remain the focus of many medical studies. Cattle is a better model organism for investigating human embryo development than mice. Imprinted genes usually cluster on chromosomes and are regulated by different methylation regions (DMRs) located in imprinting control regions that control gene expression in cis. There is an imprinted locus on human chromosome 16q24.1 associated with congenital lethal developmental lung disease in newborns. However, genomic imprinting on bovine chromosome 18, which is homologous with human chromosome 16 has not been systematically studied. The aim of this study was to analyze the allelic expressions of eight genes (CDH13, ATP2C2, TLDC1, COTL1, CRISPLD2, ZDHHC7, KIAA0513, and GSE1) on bovine chromosome 18 and to search the DMRs associated gene allelic expression. Three transcript variants of the ZDHHC7 gene (X1, X2, and X5) showed maternal imprinting in bovine placentas. In addition, the monoallelic expression of X2 and X5 was tissue-specific. Five transcripts of the KIAA0513 gene showed tissue- and isoform-specific monoallelic expression. The CDH13, ATP2C2, and TLDC1 genes exhibited tissue-specific imprinting, however, COTL1, CRISLPLD2, and GSE1 escaped imprinting. Four DMRs, established after fertilization, were found in this region. Two DMRs were located between the ZDHHC7 and KIAA0513 genes, and two were in exon 1 of the CDH13 and ATP2C2 genes, respectively. The results from this study support future studies on the molecular mechanism to regulate the imprinting of candidate genes on bovine chromosome 18.

RECQL4 Inhibits Radiation-Induced Tumor Immune Awakening via Suppressing the cGAS-STING Pathway in Hepatocellular Carcinoma.

Many patients with hepatocellular carcinoma (HCC) respond poorly to radiotherapy despite remarkable advances in treatment. A deeper insight into the mechanism of sensitivity of HCC to this therapy is urgently required. It is demonstrated that RECQL4 is upregulated in the malignant cells of patients with HCC. Elevated RECQL4 levels reduce the sensitivity of HCC to radiotherapy by repairing radiation-induced double-stranded DNA (dsDNA) fragments. Mechanistically, the inhibitory effect of RECQL4 on radiotherapy is due to the reduced recruitment of dendritic cells and CD8+ T cells in the tumor microenvironment (TME). RECQL4 disrupts the radiation-induced transformation of the TME into a tumoricidal niche by inhibiting the cGAS-STING pathway in dendritic cells. Knocking out STING in dendritic cells can block the impact of RECQL4 on HCC radiosensitivity. Notably, high RECQL4 expressions in HCC is significantly associated with poor prognosis in multiple independent cohorts. In conclusion, this study highlights how HCC-derived RECQL4 disrupts cGAS-STING pathway activation in dendritic cells through DNA repair, thus reducing the radiosensitivity of HCC. These findings provide new perspectives on the clinical treatment of HCC.

A novel role for the ROS-ATM-Chk2 axis mediated metabolic and cell cycle reprogramming in the M1 macrophage polarization.

Reactive oxygen species (ROS) play a pivotal role in macrophage-mediated acute inflammation. However, the precise molecular mechanism by which ROS regulate macrophage polarization remains unclear. Here, we show that ROS function as signaling molecules that regulate M1 macrophage polarization through ataxia-telangiectasia mutated (ATM) and cell cycle checkpoint kinase 2 (Chk2), vital effector kinases in the DNA damage response (DDR) signaling pathway. We further demonstrate that Chk2 phosphorylates PKM2 at the T95 and T195 sites, promoting glycolysis and facilitating macrophage M1 polarization. In addition, Chk2 activation increases the Chk2-dependent expression of p21, inducing cell cycle arrest for subsequent macrophage M1 polarization. Finally, Chk2-deficient mice infected with lipopolysaccharides (LPS) display a significant decrease in lung inflammation and M1 macrophage counts. Taken together, these results suggest that inhibiting the ROS-Chk2 axis can prevent the excessive inflammatory activation of macrophages, and this pathway can be targeted to develop a novel therapy for inflammation-associated diseases and expand our understanding of the pathophysiological functions of DDR in innate immunity.

Preoperative supine time for adrenal venous sampling: a prospective randomized controlled trial.

BACKGROUND: Primary aldosteronism (P.A.) is the most common form of secondary hypertension, accounting for 5% of hypertensive patients and 17-23% of patients with resistant hypertension. Compared to primary hypertension, P.A. is more prone to cause severe organ damage and even early death. Adrenal venous sampling (AVS) is a practical confirmatory test for subtyping aldosterone-producing adenoma and bilateral adrenal hyperplasia, helping physicians to make an accurate decision between surgery or medication. According to guidelines, supine in bed before AVS is recommended for a desirable result of AVS. However, investigations about the most optimal preoperative supine time before AVS are lacking. METHODS/DESIGN: This is a multi-center prospective randomized controlled study. One hundred twenty patients diagnosed as P.A. and willing for AVS examination will be included. Participants will be randomly allocated to a 15-min supine time group or 2-h supine time group. The primary outcome is the degree of biochemical remission (serum potassium and orthostatic ARR). The secondary outcomes are degrees of clinical remission (blood pressure, type and dose of antihypertensive drugs), the technical success rate, and the adverse event of AVS (selective index ≥ 2 is considered successful surgery without corticotropin stimulation). DISCUSSION: P.A. is an intractable public health problem, and many techniques including AVS have been developed to identify this disease correctly. This study will help to understand whether the length of preoperative supine time would affect the diagnostic efficacy of AVS and thus help to formulate a more reasonable AVS procedure. TRIAL REGISTRATION: ClinicalTrials.gov NCT05658705. Registered on 10 September 2022.

Identification of Fasudil as a collaborator to promote the anti-tumor effect of lenvatinib in hepatocellular carcinoma by inhibiting GLI2-mediated hedgehog signaling pathway.

Lenvatinib is a frontline tyrosine kinase inhibitor for patients with advanced hepatocellular carcinoma (HCC). However, just 25% of patients benefit from the treatment, and acquired resistance always develops. To date, there are neither effective medications to combat lenvatinib resistance nor accurate markers that might predict how well a patient would respond to the lenvatinib treatment. Thus, novel strategies to recognize and deal with lenvatinib resistance are desperately needed. In the current study, a robust Lenvatinib Resistance index (LRi) model to predict lenvatinib response status in HCC was first established. Subsequently, five candidate drugs (Mercaptopurine, AACOCF3, NU1025, Fasudil, and Exisulind) that were capable of reversing lenvatinib resistance signature were initially selected by performing the connectivity map (CMap) analysis, and fasudil finally stood out by conducting a series of cellular functional assays in vitro and xenograft mouse model. Transcriptomics revealed that the co-administration of lenvatinib and fasudil overcame lenvatinib resistance by remodeling the hedgehog signaling pathway. Mechanistically, the feedback activation of EGFR by lenvatinib led to the activation of the GLI2-ABCC1 pathway, which supported the HCC cell's survival and proliferation. Notably, co-administration of lenvatinib and fasudil significantly inhibited IHH, the upstream switch of the hedgehog pathway, to counteract GLI2 activation and finally enhance the effectiveness of lenvatinib. These findings elucidated a novel EGFR-mediated mechanism of lenvatinib resistance and provided a practical approach to overcoming drug resistance in HCC through meaningful drug repurposing strategies.

Salvianolic acid B-loaded polydopamine-modified hollow mesoporous organic silica nanoparticles for treatment of breast cancer metastasis via suppressing cancer-associated fibroblasts.

OBJECTIVE: Drug Delivery System was constructed using dopamine-coated organic-inorganic hybrid hollow mesoporous organic silica nanoparticles (HMON-PDA) as drug carriers and salvianolic acid B (SAB) as a model drug. Then, we further investigated whether it can inhibit lung metastasis of breast cancer by inhibiting cancer-associated fibroblasts (CAFs). METHODS: The organic-inorganic hybrid hollow mesoporous organic silica nanoparticles (HMON) were prepared. The particle size, zeta potential, and polydispersion coefficient were characterized. High-performance liquid chromatography was used to determine the effect of different feed ratios of HMON and SAB on drug loading rate. Then, SAB-loaded HMON were modified by polydopamine, which is called SAB@HMON-PDA. Cell viability was detected by MTT assay. The migration of 4T1 cells was investigated by wound healing experiment, and the invasion of 4T1 cells was detected by the transwell method. Finally, the mouse breast cancer lung metastasis models were used to explore whether SAB@HMON-PDA can inhibit lung metastasis of breast cancer by inhibiting CAFs. RESULTS: The obtained nanoparticles have hollow spherical structure. The average particle sizes of HMON, SAB@HMON, and SAB@HMON-PDA were 143.5 ± 0.03, 138.3 ± 0.02, and 172.3 ± 0.18 nm, respectively. The zeta potentials were -44.33±0.15, -41.4 ± 1.30, and -24.13±0.47 mV, respectively. When the ratio of HMON to SAB was 2:1, the drug loading rate reached (18.37±0.04)%. In addition, the prepared SAB@HMON-PDA responded to release SAB under acidic and GSH conditions. The prepared SAB@HMON-PDA could inhibit the migration and invasion of 4T1 cells. The results showed that SAB@HMON-PDA and SAB could inhibit lung metastasis of breast cancer in mice, and SAB@HMON-PDA had a more significant inhibitory effect than SAB. CONCLUSION: We successfully prepared SAB@HMON-PDA with the dual response of pH and GSH. SAB@HMON-PDA can inhibit the migration and invasion of 4T1 cells, and the effect is more significant than free SAB. This inhibitory effect may be related to the inhibition of CAFs. In vivo experiments demonstrated that SAB@HMON-PDA can inhibit lung metastasis of breast cancer by inhibiting CAFs, and its effect was more significant than that of free SAB.

Si-Ni-San inhibits hepatic Fasn expression and lipid accumulation in MAFLD mice through AMPK/p300/SREBP-1c axis.

BACKGROUND: Soothing the liver and regulating qi is one of the core ideas of traditional Chinese medicine (TCM) in the treatment of fatty liver. Si-Ni-San (SNS) is a well-known herbal formula in TCM for liver soothing and qi regulation in fatty liver treatment. However, its efficacy lacks modern scientific evidence. PURPOSE: This study was aimed to investigate the impact of SNS on metabolic associated fatty liver disease (MAFLD) in mice and explore the underlying molecular mechanisms, particularly its effects on lipid metabolism in hepatocytes. METHODS: The therapeutic effect of SNS was evaluated using in vivo and in vitro models of high-fat/high-cholesterol (HFHC) diet-induced mice and palmitic acid (PA)-induced hepatocytes, respectively. Molecular biological techniques such as RNA-sequencing (RNA-seq), co-immunoprecipitation (co-IP), and western blotting were employed to elucidate the molecular mechanism of SNS in regulating lipid metabolism in hepatocytes. RESULTS: Our findings revealed that SNS effectively reduced lipid accumulation in the livers of HFHC diet-induced mice and PA-induced hepatocytes. RNA-seq analysis demonstrated that SNS significantly down-regulated the expression of fatty acid synthase (Fasn) in the livers of HFHC-fed mice. Mechanistically, SNS inhibited Fasn expression and lipid accumulation by activating adenosine monophosphate (AMP)-activated protein kinase (AMPK). Activation of AMPK suppressed the activity of the transcriptional coactivator p300 and modulated the protein stability of sterol regulatory element-binding protein-1c (SREBP-1c). Importantly, p300 was required for the inhibition of Fasn expression and lipid accumulation by SNS. Furthermore, SNS activated AMPK by decreasing adenosine triphosphate (ATP) production in hepatocytes. CONCLUSION: This study provided novel evidence on the regulatory mechanisms underlying the effects of SNS on Fasn expression. Our findings demonstrate, for the first time, that SNS exerts suppressive effects on Fasn expression through modulation of the AMPK/p300/SREBP-1c axis. Consequently, this regulatory pathway mitigates excessive lipid accumulation and ameliorates MAFLD in mice.

Deciphering the immune modulation through deep transcriptomic profiling and therapeutic implications of DNA damage repair pattern in hepatocellular carcinoma.

AIMS: DNA damage repair (DDR) plays a pivotal role in hepatocellular carcinoma (HCC), driving oncogenesis, progression, and therapeutic response. However, the mechanisms of DDR mediated immune cells and immuno-modulatory pathways in HCC are yet ill-defined. METHODS: Our study introduces an innovative deep machine learning framework for precise DDR assessment, utilizing single-cell RNA sequencing (scRNA-seq) and bulk RNA-seq data. Single-cell RNA sequencing data were obtained and in total 85,628 cells of primary or post-immunotherapy cases were analyzed. Large-scale HCC datasets, including 1027 patients in house together with public datasets, were used for 101 machine-learning models and a novel DDR feature was derived at single-cell resolution (DDRscore). Druggable targets were predicted using the reverse phase protein array (RPPA) proteomic profiling of 169 HCC patients and RNA-seq data from 22 liver cancer cell lines. RESULTS: Our investigation reveals a dynamic interplay of DDR with natural killer cells and B cells in the primary HCC microenvironment, shaping a tumor-promoting immune milieu through metabolic programming. Analysis of HCC post-immunotherapy demonstrates elevated DDR levels that induces epithelial-mesenchymal transition and fibroblast-like transformation, reshaping the fibrotic tumor microenvironment. Conversely, attenuated DDR promotes antigen cross-presentation by dendritic cells and CD8+ T cells, modulating the inflammatory tumor microenvironment. Regulatory network analysis identifies the CXCL10-CXCR3 axis as a key determinant of immunotherapeutic response in low DDR HCC, potentially regulated by transcription factors GATA3, REL, and TBX21. Using machine learning techniques by combining bulk RNA-seq data in house together with public datasets, we introduce DDRscore, a robust consensus DDR scoring system to predict overall survival and resistance to PD-1 therapy in HCC patients. Finally, we identify BRAF as a potential therapeutic target for high DDRscore patients. CONCLUSION: Our comprehensive findings advance our understanding of DDR and the tumor microenvironment in HCC, providing insights into immune regulatory mechanisms mediated via DDR pathways.

[Mutational Signatures Analysis of Micropapillary Components and Exploration of ZNF469 Gene in Early-stage Lung Adenocarcinoma with Ground-glass Opacities].

BACKGROUND: In China, lung cancer remains the cancer with the highest incidence and mortality rate. Among early-stage lung adenocarcinomas (LUAD), the micropapillary (MPP) component is prevalent and typically exhibits high aggressiveness, significantly correlating with early metastasis, lymphatic infiltration, and reduced five-year survival rates. Therefore, the study is to explore the similarities and differences between MPP and non-micropapillary (non-MPP) components in malignant pulmonary nodules characterized by GGOs in early-stage LUAD, identify unique mutational features of the MPP component and analyze the relationship between the ZNF469 gene, a member of the zinc-finger protein family, and the prognosis of early-stage LUAD, as well as its correlation with immune infiltration. METHODS: A total of 31 malignant pulmonary nodules of LUAD were collected and dissected into paired MPP and non-MPP components using microdissection. Whole-exome sequencing (WES) was performed on the components of early-stage malignant pulmonary nodules. Mutational signatures analysis was conducted using R packages such as maftools, Nonnegative Matrix Factorization (NMF), and Sigminer to unveil the genomic mutational characteristics unique to MPP components in invasive LUAD compared to other tumor tissues. Furthermore, we explored the expression of the ZNF469 gene in LUAD using The Cancer Genome Atlas (TCGA) database to investigate its potential association with the prognosis. We also investigated gene interaction networks and signaling pathways related to ZNF469 in LUAD using the GeneMANIA database and conducted Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Lastly, we analyzed the correlation between ZNF469 gene expression and levels of immune cell infiltration in LUAD using the TIMER and TISIDB databases. RESULTS: MPP components exhibited a higher number of genomic variations, particularly the 13th COSMIC (Catalogue of Somatic Mutations in Cancer) mutational signature characterized by the activity of the cytidine deaminase APOBEC family, which was unique to MPP components compared to non-MPP components in tumor tissues. This suggests the potential involvement of APOBEC in the progression of MPP components in early-stage LUAD. Additionally, MPP samples with high similarity to APOBEC signature displayed a higher tumor mutational burden (TMB), indicating that these patients may be more likely to benefit from immunotherapy. The expression of ZNF469 was significantly upregulated in LUAD compared to normal tissue, and was associated with poor prognosis in LUAD patients (P<0.05). Gene interaction network analysis and GO/KEGG enrichment analysis revealed that COL6A1, COL1A1, COL1A2, TGFB2, MMP2, COL8A2 and C2CD4C interacted with ZNF469 and were mainly involved in encoding collagen proteins and participating in the constitution of extracellular matrix. ZNF469 expression was positively correlated with immune cell infiltration in LUAD (P<0.05). CONCLUSIONS: The study has unveiled distinctive mutational signatures in the MPP components of early-stage invasive LUAD in the Asian population. Furthermore, we have identified that the elevated expression of mutated ZNF469 impacts the prognosis and immune infiltration in LUAD, suggesting its potential as a diagnostic and prognostic biomarker in LUAD.

Exploring the potential of calcium-based biomaterials for bone regeneration in dentistry: a systematic review.

INTRODUCTION: Regenerative medicine emerged as a promising strategy for addressing bone defects, with several bone grafts currently being used, including autografts, allografts, xenografts and alloplasts. Calcium-based biomaterials (CaXs), a well-known class of synthetic materials, have demonstrated good biological properties and are being investigated for their potential to facilitate bone regeneration. This systematic review evaluates the current clinical applications of CaXs in dentistry for bone regeneration. EVIDENCE ACQUISITION: A comprehensive search was conducted to collect information about CaXs and their applications in the dental field over the last ten years. The search was limited to relevant articles published in peer-reviewed journals. EVIDENCE SYNTHESIS: A total of 72 articles were included in this scoping review, with eight studies related to periodontology, 63 in implantology and three in maxillofacial surgery respectively. The findings suggest that CaXs hold promise as an alternative intervention for minor bone regeneration in dentistry. CONCLUSIONS: Calcium-based biomaterials have shown potential as a viable option for bone regeneration in dentistry. Further research is warranted to fully understand their efficacy and safety in larger bone defects. CaXs represent an exciting avenue for researchers and clinicians to explore in their ongoing efforts to advance regenerative medicine.

Essential oil extracted from Quzhou Aurantii Fructus prevents acute liver failure through inhibiting lipopolysaccharide-mediated inflammatory response.

Quzhou Aurantii Fructus (QAF) has a long history as a folk medicine and food for the treatment of liver diseases. While our earlier study provided evidence of hepatoprotective properties contained within the flavonoids and limonins constituents in QAF, the potential preventative effects afforded by essential oil components present within QAF remains enigmatic. In this study, we prepared Quzhou Aurantii Fructus essential oil (QAFEO) and confirmed its anti-inflammatory effects on liver inflammation through experimentation on lipopolysaccharide and D-galactosamine (LPS/D-GalN) induced acute liver failure (ALF) mouse models. Using RNA-sequence (RNA-seq) analysis, we found that QAFEO prevented ALF by systematically blunting the pathways involved in response to LPS and toll-like receptor signaling pathways. QAFEO effectively suppressed the phosphorylation of tank-binding kinase 1 (TBK1), TGF-beta activated kinase 1 (TAK1), interferon regulatory factor 3 (IRF3), and the activation of mitogen activated kinase-like protein (MAPK) and nuclear factor-kappa B (NF-κB) pathways in vivo and in vitro. Importantly, QAFEO substantially reduced myeloid differentiation primary response gene 88 (MyD88)- toll-like receptor 4 (TLR4) interaction levels. Moreover, 8 compounds from QAFEO could directly bind to REAL, TAK1, MyD88, TBK1, and IRF3. Taken together, the results of our study support the notion that QAFEO exerts a hepatoprotective effect through inhibiting LPS-mediated inflammatory response.

Comprehensive analysis of P2Y family genes expression, immune characteristics, and prognosis in pan-cancer.

BACKGROUND: P2Y receptors are a family of G protein-coupled receptor genes that have an important function in cancer development and metastasis. However, systematic studies have not been conducted on human tumors. This study attempted to explore the role of P2Y family genes (P2Ys) in pan-cancer. METHODS: Gene expression and clinical data were downloaded from The Cancer Genome Alas dataset. Gene differential expression, mutation, prognosis, tumor microenvironment (TME) (containing immune cells infiltration, Estimate/immune/stromal scores, immune checkpoints, immune and molecular subtypes, DNA repair genes and methyltransferase), clinical correlation, protein-protein interaction network and functional enrichment analysis were performed. In addition, experiments such as western blots were performed for validation. RESULTS: Eight P2Ys were differentially expressed in most tumor and normal tissues, and their abnormal expression in a variety of cancers could significantly reduce the survival rate of patients. Expression levels of P2Ys, especially P2Y6, P2Y12, P2Y13, P2Y14, were correlated significantly with immune cells, immune checkpoint genes, immune and molecular subtypes and Estimate/immune/stromal scores in a variety of cancers such as uveal melanoma, liver hepatocellular carcinoma, stomach adenocarcinoma, colorectal cancer (CRC), prostate adenocarcinoma, breast invasive carcinoma and uterine corpus endometrial carcinoma (all p < 0.05). P2Ys play an important role in TME and are involved in immune regulation. In addition, enrichment analysis and western blots showed that the levels of P2Y2 and P2Y6 expression regulate the Akt/GSK-3ß/ß-catenin pathway in CRC, thereby affecting epithelial-to-mesenchymal transition. CONCLUSION: P2Ys may be used as potential pan-cancer biomarkers in prognosis and immunology. They may also be new targets for tumor immunotherapy, which has wide clinical implications.

Energy stress-induced circZFR enhances oxidative phosphorylation in lung adenocarcinoma via regulating alternative splicing.

BACKGROUND: Circular RNAs (circRNAs) contribute to multiple biological functions and are also involved in pathological conditions such as cancer. However, the role of circRNAs in metabolic reprogramming, especially upon energy stress in lung adenocarcinoma (LUAD), remains largely unknown. METHODS: Energy stress-induced circRNA was screened by circRNA profiling and glucose deprivation assays. RNA-seq, real-time cell analyzer system (RTCA) and measurement of oxygen consumption rate (OCR) were performed to explore the biological functions of circZFR in LUAD. The underlying mechanisms were investigated using circRNA pull-down, RNA immunoprecipitation, immunoprecipitation and bioinformatics analysis of alternative splicing. Clinical implications of circZFR were assessed in 92 pairs of LUAD tissues and adjacent non-tumor tissues, validated in established patient-derived tumor xenograft (PDTX) model. RESULTS: CircZFR is induced by glucose deprivation and is significantly upregulated in LUAD compared to adjacent non-tumor tissues, enhancing oxidative phosphorylation (OXPHOS) for adaptation to energy stress. CircZFR is strongly associated with higher T stage and poor prognosis in patients with LUAD. Mechanistically, circZFR protects heterogeneous nuclear ribonucleoprotein L-like (HNRNPLL) from degradation by ubiquitination to regulate alternative splicing, such as myosin IB (MYO1B), and subsequently activates the AKT-mTOR pathway to facilitate OXPHOS. CONCLUSION: Our study provides new insights into the role of circRNAs in anticancer metabolic therapies and expands our understanding of alternative splicing.

Saikosaponin A triggers cell ferroptosis in hepatocellular carcinoma by inducing endoplasmic reticulum stress-stimulated ATF3 expression.

Ferroptosis is a type of nonapoptotic necrotic cell death characterized by iron-dependent lipid peroxidation. Saikosaponin A (SsA), a natural bioactive triterpenoid saponin extracted from Radix Bupleuri, has shown potent antitumor activity against various tumors. However, the underlying mechanism of the antitumor activity of SsA remains unclear. Here, we discovered that SsA induced HCC cell ferroptosis in vitro and in vivo. Using RNA-sequence analysis, we found that SsA mainly affected the glutathione metabolic pathway and inhibited the expression of cystine transporter solute carrier family 7 member 11 (SLC7A11). Indeed, SsA increased intracellular malondialdehyde (MDA) and iron accumulation, while it decreased the levels of reduced glutathione (GSH) in HCC. Deferoxamine (DFO), ferrostatin-1 (Fer-1) and GSH could rescue SsA-induced cell death, whereas Z-VAD-FMK was found ineffective in inhibiting SsA-induced cell death in HCC. Importantly, our result indicated that SsA induced the expression of activation transcription factor 3 (ATF3). SsA-induced cell ferroptosis and suppression of SLC7A11 are dependent on ATF3 in HCC. Moreover, we revealed that SsA induced ATF3 upregulation via activation of endoplasmic reticulum (ER) stress. Taken together, our findings support that ATF3-dependent cell ferroptosis mediated the antitumor effects of SsA, opening the possibility to explore SsA as a ferroptosis inducer in HCC.

Enhanced Detection of Landmark Minimal Residual Disease in Lung Cancer Using Cell-free DNA Fragmentomics.

Currently, approximately 30%-55% of the patients with non-small cell lung cancer (NSCLC) develop recurrence due to minimal residual disease (MRD) after receiving surgical resection of the tumor. This study aims to develop an ultrasensitive and affordable fragmentomic assay for MRD detection in patients with NSCLC. A total of 87 patients with NSCLC, who received curative surgical resections (23 patients relapsed during follow-up), enrolled in this study. A total of 163 plasma samples, collected at 7 days and 6 months postsurgical, were used for both whole-genome sequencing (WGS) and targeted sequencing. WGS-based cell-free DNA (cfDNA) fragment profile was used to fit regularized Cox regression models, and leave-one-out cross-validation was further used to evaluate models' performance. The models showed excellent performances in detecting patients with a high risk of recurrence. At 7 days postsurgical, the high-risk patients detected by our model showed an increased risk of 4.6 times, while the risk increased to 8.3 times at 6 months postsurgical. These fragmentomics determined higher risk compared with the targeted sequencing-based circulating mutations both at 7 days and 6 months postsurgical. The overall sensitivity for detecting patients with recurrence reached 78.3% while using both fragmentomics and mutation results from 7 days and 6 months postsurgical, which increased from the 43.5% sensitivity by using only the circulating mutations. The fragmentomics showed great sensitivity in predicting patient recurrence compared with the traditional circulating mutation, especially after the surgery for early-stage NSCLC, therefore exhibiting great potential to guide adjuvant therapeutics. Significance: The circulating tumor DNA mutation-based approach shows limited performance in MRD detection, especially for landmark MRD detection at an early-stage cancer after surgery. Here, we describe a cfDNA fragmentomics-based method in MRD detection of resectable NSCLC using WGS, and the cfDNA fragmentomics showed a great sensitivity in predicting prognosis.