Downregulation of KLF5 by EBER1 via the ERK signaling pathway in EBV-positive nasopharyngeal carcinoma cells: implications for latent EBV infection.

Nasopharyngeal carcinoma (NPC) carcinogenesis and malignant transformation are intimately associated with Epstein-Barr virus (EBV) infection. A zinc-fingered transcription factor known as Krüppel-like factor 5 (KLF5) has been shown to be aberrantly expressed in a number of cancer types. However, little is known about the regulatory pathways and roles of KLF5 in EBV-positive NPC. Our study found that KLF5 expression was significantly lower in EBV-positive NPC than in EBV-negative NPC. Further investigation revealed that EBER1, which is encoded by EBV, down-regulates KLF5 via the extracellular signal-regulated kinase (ERK) signalling pathway. This down-regulation of KLF5 by EBER1 contributes to maintaining latent EBV infection in NPC. Furthermore, we uncovered the biological roles of KLF5 in NPC cells. Specifically, KLF5 may influence the cell cycle, prevent apoptosis, and encourage cell migration and proliferation - all of which have a generally pro-cancer impact. In conclusion, these findings offer novel strategies for EBV-positive NPC patients' antitumour treatment.

PRODH safeguards human naive pluripotency by limiting mitochondrial oxidative phosphorylation and reactive oxygen species production.

Naive human embryonic stem cells (hESCs) that resemble the pre-implantation epiblasts are fueled by a combination of aerobic glycolysis and oxidative phosphorylation, but their mitochondrial regulators are poorly understood. Here we report that, proline dehydrogenase (PRODH), a mitochondria-localized proline metabolism enzyme, is dramatically upregulated in naive hESCs compared to their primed counterparts. The upregulation of PRODH is induced by a reduction in c-Myc expression that is dependent on PD0325901, a MEK inhibitor routinely present in naive hESC culture media. PRODH knockdown in naive hESCs significantly promoted mitochondrial oxidative phosphorylation (mtOXPHOS) and reactive oxygen species (ROS) production that triggered autophagy, DNA damage, and apoptosis. Remarkably, MitoQ, a mitochondria-targeted antioxidant, effectively restored the pluripotency and proliferation of PRODH-knockdown naive hESCs, indicating that PRODH maintains naive pluripotency by preventing excessive ROS production. Concomitantly, PRODH knockdown significantly slowed down the proteolytic degradation of multiple key mitochondrial electron transport chain complex proteins. Thus, we revealed a crucial role of PRODH in limiting mtOXPHOS and ROS production, and thereby safeguarding naive pluripotency of hESCs.

Betaine Alleviates High-Fat Diet Induced Excessive Lipid Deposition in Gibel Carp Hepatopancreas and L8824 Cells by Enhancing VLDL Secretion through HNF4α/MTTP Pathway.

Betaine, a methyl donor, plays a crucial role in lipid metabolism. Previous studies have shown that appropriate betaine supplementation in a high-fat diet reduces triglycerides (TG) of serum and hepatopancreas in fish. However, the underlying mechanism remains unclear. This study examined whether betaine can enhance the secretion of very low-density lipoprotein (VLDL) and sought to identify the specific mechanisms through which this enhancement occurs. A lipid accumulation model was established in gibel carp and L8824 cells using a high-fat diet and oleic acid, respectively. Different doses of betaine (1, 4, and 16 g/kg in the diet; 400 µmol in cell culture) were administered, and measurements were taken for lipid deposition, gene expression of HNF4α, MTTP, and ApoB, as well as the regulation of Mttp and Apob promoters by HNF4α. The results showed that betaine supplementation mitigated lipid droplet accumulation, TG levels, and VLDL production induced by the high-fat diet in gibel carp hepatopancreas and L8824 cells. Moreover, betaine not only increased VLDL content in the cell culture supernatant but also reversed the inhibitory effects of the high-fat diet on protein expression of MTTP, ApoB, and HNF4α in both gibel carp hepatopancreas and L8824 cells. Additionally, HNF4α exhibits transactivating activity on the promoter of Mttp in gibel carp. These findings suggest that betaine supplementation exerts its effects through the HNF4α/MTTP/ApoB pathway, promoting the assembly and secretion of VLDL and effectively reducing lipid accumulation in the hepatopancreas of farmed gibel carp fed a high-fat diet.

Comprehensive Analysis Based on the TCGA Database Identified SCIN as a Key DNA Methylation-Driver Gene in Epstein-Barr Virus-Associated Gastric Cancer.

An important feature of EBV-associated gastric cancer (EBVaGC) is extensive methylation of viral and host genomes. This study aims to analyze DNA methylation-driven genes (DMDG) in EBVaGC through bioinformatics methods, providing an important bioinformatics basis for the differential diagnosis and treatment of potential methylation biomarkers in EBVaGC. We downloaded the mRNA expression profiles and methylation datasets of EBVaGC and EBV-negative gastric cancer (EBVnGC) through the TCGA database to screen methylated-differentially expressed genes (MDEGs). DNA methylation-driver genes were identified based on MethylMix algorithm and key genes were further identified by LASSO regression and Random Forest algorithm. Then, we performed gene enrichment analysis for key genes and validated them by GEO database. Gene expression differences in EBVaGC and EBVnGC cell lines was determined by quantitative real-time PCR (qRT-PCR) and western blotting and in GT38 cell and SNU719 cell which all treated by 5-Aza-CdR. Finally, the effect of key gene on the migration and proliferation capacity of EBVaGC cells was determined by Transwells assay and Cell counting Kit-8 (CCK-8) assay. We obtained a total of 687 hypermethylation-low expression genes (Hyper-LGs) and further obtained 53 DNA methylation-driver genes based on the MethylMix algorithm. A total of six key genes (SCIN, ETNK2, PCDH20, PPP1R3C, MATN2, and HOXA5) were identified by LASSO regression and Random Forest algorithm. Among them, SCIN expression was significantly lower in EBVaGC cell lines than in EBVnGC cell lines, and its expression was significantly recovered in EBVaGC cell lines treated with 5-Aza-CdR. Overexpression of SCIN can promote the proliferation and migration capacity of EBVaGC cells. Our study will provide some bioinformatics basis for the study of EBVaGC-related methylation. SCIN may be used as potential methylation biomarkers for the diagnosis and treatment of EBVaGC.

Prophylactic hyperthermic intraperitoneal chemotherapy in T4 colorectal cancer: Can it improve the oncologic prognosis? - A propensity score matching study.

BACKGROUND: Some studies show that cytoreductive surgery with hyperthermic intraperitoneal chemotherapy (HIPEC) may improve overall survival and is a possible curative treatment for selected colorectal cancer (CRC) patients with restricted peritoneal metastasis (PM). The value of HIPEC in preventing PM of CRC is still controversial. MATERIALS AND METHODS: In this retrospective propensity score matching (PSM) cohort study, all patients with cT4N0-2M0 undergoing treatment at a single institution in China (2014-2018) were reviewed. The 3-year disease-free survival (DFS) was set as the primary outcome, and the 3-year PM rate was also analyzed. RESULTS: 220 patients were included in this study for analysis. After 1:3 PSM: HIPEC (n = 45) and No HIPEC (n = 135). Through analysis, it was found that prophylactic HIPEC correlated to better DFS [hazard ratio (HR) 0.43, 95 % confidence interval (CI) 0.19-0.95; p = 0.037], and N2 stage correlated to worse DFS [HR 1.97, 95 % CI 1.09-3.56; p = 0.025]. For laparoscopic surgery subgroup analyses, 3-year PM rate of patients with laparoscopic surgery was 13.8 % in No HIPEC group, and 2.6 % in HIPEC group (p = 0.070). Besides, no post-operative death occurred, the anastomotic leakage rate was 2.2 % in HIPEC group and 0.7 % in the control group (p = 0.439). CONCLUSIONS: Prophylactic HIPEC may improve the prognosis in patients with cT4N0-1M0 CRC, but not in cT4N2M0 CRC, and it does not significantly increase surgery-related complications. Laparoscopic surgery followed by HIPEC for T4 stage CRC may not increase risk of PM.

Sequential infection of human norovirus and Salmonella enterica resulted in higher mortality and ACOD1/IRG1 upregulation in zebrafish larvae.

Human norovirus (HNoVs) and Salmonella are both very important foodborne pathogens with mixed infection of HNoV and Salmonella reported clinically. With the use of model organism zebrafish (Danio rerio), it was observed that the sequential infection of HNoVs and Salmonella caused lower survival rates (12.5 ± 4.2%) than the single-pathogen infection by Salmonella (31.6 ± 7.3%, P < 0.05) or HNoVs (no mortality observed). Gene expression study with the use of RT-PCR and global transcriptomic analysis revealed that the mortality of zebrafish larvae was very likely due to the harmful inflammatory responses. Specifically, it was noted that the genes encoding aconitate decarboxylase 1 (ACOD1), also known as immunoresponsive gene 1 (IRG1), were significantly upregulated in the sequentially infected zebrafish larvae. The expression of acod1 could lead to mitochondrial reactive oxygen species (ROS) production. The ROS levels were indeed higher in sequentially infected zebrafish larvae than the single-pathogen infected ones (P < 0.05). An immersion treatment of glutathione or citraconate did not affect the microbial loads of HNoVs and Salmonella but significantly reduced the ROS levels and protected the zebrafish larvae by inducing higher survival rates in the sequentially infected zebrafish larvae (P < 0.05). Taken together, this study accumulated new knowledge over the function of ACOD1/IRG1 pathway in infectious diseases, and proposed possible treatment strategies accordingly.

Comparison of Intraoperative Endplate Injury between Mini-Open Lateral Lumbar Interbody Fusion (LLIF) and Transforaminal Lumbar Interbody Fusion (TLIF) and Analysis of Risk Factors: A Retrospective Study.

OBJECTIVE: The study aimed to compare the incidence of intraoperative endplate injury in patients who underwent Transforaminal interbody fusion (TLIF) and mini-open lumbar interbody fusion (LLIF) surgery. The independent risk factors related to endplate injury in LLIF procedure were analyzed. METHODS: A total of 199 patients who underwent LLIF (n = 106) or TLIF (n = 93) surgery from June 2019 to September 2021 were reviewed. The endplate injury was assessed by postoperative sagittal CT scan. A binary logistic analysis model were used to identify independent risk factors related to LLIF endplate injury based on univariate analysis. RESULTS: There was an obvious difference in the occurrence of intraoperative endplate injury between LLIF (42/106, 39.6%) and TLIF group (26/93, 28%), although it did not reach the significant level. L1 CT value (OR = 0.985, 95% CI = 0.972-0.998), cage position (OR = 3.881, 95% CI = 1.398-10.771) and height variance (OR = 1.263, 95% CI = 1.013-1.575) were independent risk factors for endplate injury in LLIF procedure. According to the cage settlement patterns, there 5 types of A to E. The severity of the facet joint degeneration was positively related to the occurrence of endplate injury. CONCLUSIONS: The incidence of intraoperative endplate injury is higher in LLIF than in TLIF procedures. Low bone quantity, cage posterior position and larger height variance are risk factors to induce endplate injury in LLIF surgery. The facet joint degeneration may be related to severe endplate injuries and even fractures.

RBFOX1 and Working Memory: From Genome to Transcriptome Revealed Posttranscriptional Mechanism Separate From Attention-Deficit/Hyperactivity Disorder.

Background: Many psychiatric disorders share a working memory (WM) impairment phenotype, yet the genetic causes remain unclear. Here, we generated genetic profiles of WM deficits using attention-deficit/hyperactivity disorder samples and validated the results in zebrafish models. Methods: We used 2 relatively large attention-deficit/hyperactivity disorder cohorts, 799 and 776 cases, respectively. WM impairment was assessed using the Rey Complex Figure Test. First, association analyses were conducted at single-variant, gene-based, and gene-set levels. Deeper insights into the biological mechanism were gained from further functional exploration by bioinformatic analyses and zebrafish models. Results: Genomic analyses identified and replicated a locus with rs75885813 as the index single nucleotide polymorphism showing significant association with WM defects but not with attention-deficit/hyperactivity disorder. Functional feature exploration found that these single nucleotide polymorphisms may regulate the expression level of RBFOX1 through chromatin interaction. Further pathway enrichment analysis of potential associated single nucleotide polymorphisms revealed the involvement of posttranscription regulation that affects messenger RNA stability and/or alternative splicing. Zebrafish with functionally knocked down or genome-edited rbfox1 exhibited WM impairment but no hyperactivity. Transcriptome profiling of rbfox1-defective zebrafish indicated that alternative exon usages of snap25a might partially lead to reduced WM learning of larval zebrafish. Conclusions: The locus with rs75885813 in RBFOX1 was identified as associated with WM. Rbfox1 regulates synaptic and long-term potentiation-related gene snap25a to adjust WM at the posttranscriptional level.

Nanoplastics induce neuroexcitatory symptoms in zebrafish (Danio rerio) larvae through a manner contrary to Parkinsonian's way in proteomics.

Although nanoplastics (NPs) can penetrate the blood-brain barrier and accumulate in the brain, the neurotoxicity of these particles and the mechanisms associated with their unique physio-chemical properties have yet to be sufficiently ascertained. In this study, we assessed the neuroexcitatory symptoms of zebrafish (Danio rerio) larvae treated with polystyrene (PS) NPs based on an examination of locomotory behaviour, dopamine levels, and acetylcholinesterase activity. We found that PS NPs caused oxidative stress and inhibited atoh1a expression in the cerebellum of Tg(atoh1a:dTomato) transgenic zebrafish larvae, thereby indicating damage to the central nervous system. In contrast to the Parkinson's disease (PD) like effects induced by most types of nanoparticles, such as graphene oxide, we established that PS NPs influenced the neuronal proteomic profiles of zebrafish larvae in a manner contrary to the molecular pathways characteristic of PD-like effects, which could be explained by the molecular dynamic simulation. Unlike graphene oxide nanoparticles that promote significant change in the internal structure of neuroproteins, the complex macromolecular polymers of PS NPs promoted the coalescence and increased expression of neuroproteins, thereby plausibly contributing to the neuroexcitatory symptoms observed in treated zebrafish larvae. Consequently, compared with traditional nanoparticles, we believe that the unique physio-chemical properties of NPs could be a potential factor contributing to their toxicity.

A novel postoperative nomogram and risk classification system for individualized estimation of survival among patients with parotid gland carcinoma after surgery.

BACKGROUND: Parotid gland carcinoma (PGC) is a rare but aggressive head and neck cancer, and the prognostic model associated with survival after surgical resection has not yet been established. This study aimed to construct a novel postoperative nomogram and risk classification system for the individualized prediction of overall survival (OS) among patients with resected PGC. METHODS: Patients with PGC who underwent surgery between 2004 and 2015 from the Surveillance, Epidemiology, and End Results (SEER) database were randomized into training and validation cohorts (7:3). A nomogram developed using independent prognostic factors based on the results of the multivariate Cox regression analysis. Harrell's concordance index (C-index), time-dependent area under the curve (AUC), and calibration plots were used to validate the performance of the nomogram. Moreover, decision curve analysis (DCA) was performed to compare the clinical use of the nomogram with that of traditional TNM staging. RESULTS: In this study, 5077 patients who underwent surgery for PGC were included. Age, sex, marital status, tumor grade, histology, TNM stage, surgery type, radiotherapy, and chemotherapy were independent prognostic factors. Based on these independent factors, a postoperative nomogram was developed. The C-index of the proposed nomogram was 0.807 (95% confidence interval 0.797-0.817). Meanwhile, the time-dependent AUC (> 0.8) indicated that the nomogram had a satisfactory discriminative ability. The calibration curves showed good concordance between the predicted and actual probabilities of OS, and DCA curves indicated that the nomogram had a better clinical application value than the traditional TNM staging. Moreover, a risk classification system was built that could perfectly classify patients with PGC into three risk groups. CONCLUSIONS: This study constructed a novel postoperative nomogram and corresponding risk classification system to predict the OS of patients with PGC after surgery. These tools can be used to stratify patients with high or low risk of mortality and provide high-risk patients with more directed therapies and closer follow-up.

Elevated temperature decreases cardiovascular toxicity of nanoplastics but adds to their lethality: A case study during zebrafish (Danio rerio) development.

To highlight the key role of global warming on the toxicity of contaminants, the cardiovascular toxicity of nanoparticles (NPs) was estimated in developing zebrafish (Danio rerio) at different exposure temperatures, and the toxicity mechanisms were explored via multi-omic analyses. Polystyrene NPs (50 nm) at 0.1 mg·L-1 entered zebrafish embryos at 24 h post-fertilization and caused cardiovascular toxicity in the developing zebrafish at 27 ℃. This was explained by the down-regulation of the branched-chain amino acid and insulin signaling pathways owing to induced oxidative stress. Elevated exposure temperatures promoted the accumulation of NPs in developing zebrafish, increased the levels of oxidative stress and enhanced the oxidative phosphorylation rate in mitochondria, thus resulting in an additive effect on the mortality of zebrafish larvae. Notably, elevated exposure temperatures reduced the cardiovascular toxicity of NPs, as the effective concentration of NPs for inhibiting embryonic heartbeat rate increased from 0.1 mg·L-1 at 27 ℃ to 1.0 mg·L-1 at 30 ℃. Experiments of transgenic zebrafish Tg(myl7:GFP) and multi-omic analyses revealed that elevated temperatures enhanced the myocardial contractility of larvae, thus reducing the cardiovascular toxicity of NPs. However, the health risks of enhanced myocardial contraction caused by NP exposure at elevated temperatures requires further consideration.

Oncogenic viruses and host lipid metabolism: a new perspective.

As noncellular organisms, viruses do not have their own metabolism and rely on the metabolism of host cells to provide energy and metabolic substances for their life cycles. Increasing evidence suggests that host cells infected with oncogenic viruses have dramatically altered metabolic requirements and that oncogenic viruses produce substances used for viral replication and virion production by altering host cell metabolism. We focused on the processes by which oncogenic viruses manipulate host lipid metabolism and the lipid metabolism disorders that occur in oncogenic virus-associated diseases. A deeper understanding of viral infections that cause changes in host lipid metabolism could help with the development of new antiviral agents as well as potential new therapeutic targets.

KLF2 regulates stemness of human mesenchymal stem cells by targeting FGFR3.

Mesenchymal stem cells (MSCs) are an attractive source of pluripotent cells for regenerative therapy; however, maintaining stemness and self-renewal of MSCs during expansion ex vivo is challenging. For future clinical applications, it is essential to define the roles and signaling pathways that regulate the fate of MSCs. Based on our earlier finding that Krüppel-like factor 2 (KLF2) participates in maintaining stemness in MSCs, we examined further the role of this factor in intrinsic signaling pathways. Using a chromatin immunoprecipitation (ChIP)-sequence assay, we found that the FGFR3 gene is a KLF2 binding site. Knockdown of FGFR3 significantly decreased the levels of key pluripotency factors, enhanced the expression of differentiation-related genes and down-regulated colony formation of human bone marrow MSCs (hBMSCs). Using alizarin red S and oil red O staining, we found that knockdown of FGFR3 inhibited the osteogenic and adipogenic ability of MSCs under conditions of differentiation. The ChIP-qPCR assay confirmed that KLF2 interacts with the promoter regions of FGFR3. Our findings suggest that KLF2 promotes hBMSC stemness by direct regulation of FGFR. Our findings may contribute to enhanced MSC stemness by genetic modification of stemness-related genes.

In vivo Drug Screening to Identify Anti-metastatic Drugs in Twist1a-ERT2 Transgenic Zebrafish.

Here, we present an in vivo drug screening protocol using a zebrafish model of metastasis for the identification of anti-metastatic drugs. A tamoxifen-controllable Twist1a-ERT2 transgenic zebrafish line was established to serve as a platform for the identification. By crossing Twist1a-ERT2 with xmrk (a homolog of hyperactive form of the epidermal growth factor receptor) transgenic zebrafish, which develop hepatocellular carcinoma, approximately 80% of the double transgenic zebrafish show spontaneous cell dissemination of mCherry-labeled hepatocytes from the liver to the entire abdomen and tail regions in five days, through induction of epithelial to mesenchymal transition (EMT). This rapid and high-frequency induction of cell dissemination makes it possible to perform an in vivo drug screen for the identification of anti-metastatic drugs targeting metastatic dissemination of cancer cells. The protocol evaluates the suppressor effect of a test drug on metastasis in five days, by comparing the frequencies of the fish showing abdominal and distant dissemination patterns in the test drug-treated group with those in the vehicle-treated group. Our study previously identified that adrenosterone, an inhibitor for hydroxysteroid (11-beta) dehydrogenase 1 (HSD11ß1), has a suppressor effect on cell dissemination in the model. Furthermore, we validated that a pharmacologic and genetic inhibition of HSD11ß1 suppressed metastatic dissemination of highly metastatic human cell lines in a zebrafish xenotransplantation model. Taken together, this protocol opens new routes for the identification of anti-metastatic drugs. Graphical overview Timing Day 0: Zebrafish spawning Day 8: Primary tumor induction Day 11: Chemical treatment Day 11.5: Metastatic dissemination induction in the presence of a test chemical Day 16: Data analysis.