Regular exercise suppresses steatosis-associated liver cancer development by degrading E2F1 and c-Myc via circadian gene upregulation.

Regular exercise is believed to suppress cancer progression. However, the precise molecular mechanisms by which exercise prevents cancer development remain unclear. In this study, using a steatosis-associated liver cancer mouse model, we found that regular exercise at a speed of 18 m/min for 20 min daily suppressed liver cancer development. To explore the underlying mechanisms, we examined the gene expression profiles in the livers of the exercise and non-exercise groups. The expressions of circadian genes, such as Per1 and Cry2, were upregulated in the exercise group. As circadian rhythm disruption is known to cause various diseases, including cancer, improving circadian rhythm through exercise could contribute to cancer prevention. We further found that the expression of a series of E2F1 and c-Myc target genes that directly affect the proliferation of cancer cells was downregulated in the exercise group. However, the expression of E2F1 and c-Myc was transcriptionally unchanged but degraded at the post-translational level by exercise. Cry2, which is regulated by the Skp1-Cul1-FBXL3 (SCFFBXL3) ubiquitin ligase complex by binding to FBXL3, can form a complex with E2F1 and c-Myc, which we think is the mechanism to degrade them. Our study revealed a previously unknown mechanism by which exercise prevents cancer development.

NitraTh epitope-based neoantigen vaccines for effective tumor immunotherapy.

Neoantigen vaccines represent an emerging and promising strategy in the field of tumor immunotherapy. Despite their potential, designing an effective neoantigen vaccine remains a challenge due to the current limitations in predicting CD4+ T cell epitopes with high accuracy. Here, we introduce a novel approach to neoantigen vaccine design that does not rely on computational prediction of CD4+ T cell epitopes. Utilizing nitrated helper T cell epitope containing p-nitrophenylalanine, termed "NitraTh epitope," we have successfully engineered a series of tumor neoantigen vaccines capable of eliciting robust neoantigen-specific immune responses. With the help of NitraTh epitope, even mutations with low predicted affinity for MHC class I molecules were successfully induced to elicit neoantigen-specific responses. In H22 cell allograft and patient-derived xenograft (PDX) liver cancer mouse models, the NitraTh epitope-based neoantigen vaccines significantly suppressed tumor progression. More strikingly, through single-cell sequencing we found that the NitraTh epitope-based neoantigen vaccines regulate macrophage reprogramming and modulate macrophages to decrease the levels of the immunosuppressive molecule prostaglandin E2 (PGE2), which in turn reshapes the tumor immunosuppressive microenvironment. In summary, NitraTh epitope-based neoantigen vaccines possess the dual effects of potently activating neoantigen-specific immunity and alleviating immunosuppression, potentially providing a new paradigm for the design of tumor neoantigen vaccines.

Programing the onset of ovarian stimulation: From early follicular phase start to oral contraceptive (OC) pill, to luteal phase E2, Duo-Stim and random start OS protocols.

Ovarian stimulation has been the single most efficient measure ever taken in ART for improving outcome by harvesting multiple oocytes and ultimately, embryos. Today, ovarian stimulation protocols consist in administrating exogenous gonadotropins in order to override the natural mechanisms which controls the ovulatory quota to one in humans. For practicality issues, there has been numerous attempts to control, or 'program', when ovarian stimulation are initiated in order to improve functionality and in turn efficacy for ART programs. The different options for controlling the onset of ovarian stimulation currently available are discussed here, as well as the novel possibility of using progestins for blocking premature ovulation.

Revealing novel insights into the enhancement of quality in black tea processing through microbial intervention.

Black tea is highly favored by consumers worldwide, with enzymatic reactions being recognized as a pivotal factor influencing tea quality. The role of microorganisms in shaping the composition of black tea has emerged as a focus of research due to their involvement in enzyme catalysis and metabolic processes. In this study, full-length amplicon sequencing combined with qPCR more accurately reflected microbial profile, and Pantoea, Pseudomonas, Paucibacter, and Cladosporium were identified as the main microbial genera. Moreover, by comprehensively analyzing color, aroma, and taste components over time in black tea samples, correlations were established between the dominant genus and various quality factors. Notably, peroxidase activity levels, total soluble sugar content, and tea pigments concentration exhibited significant associations with the dominant genus. Consequently, this microbiological perspective facilitated the exploration of driving factors for improving black tea quality while establishing a theoretical foundation for quality control in industrial production.

CDT1, transcriptionally regulated by E2F2, promotes lung adenocarcinoma progression.

CDT1, a gene that shows excessive expression in various malignancies, functions as a pivotal regulator of replication licensing. In this study, we observed a positive correlation in expression between CDT1 and E2F2 among patients with lung adenocarcinoma (LUAD). Our findings substantiated that E2F2 directly interacted with the promoter region of CDT1, as confirmed by ChIP-qPCR assays, and depletion of E2F2 resulted in a downregulation of CDT1 expression in LUAD cell lines by gene interference technology. Furthermore, we identified an upregulation of CDT1 mRNA level in Chinese LUAD samples. Notably, in the loss-of-function assays, depletion of CDT1 in LUAD cell lines inhibited cell proliferation, migration, and invasion. Concurrently, it promoted cell apoptosis and induced G0/G1 phase arrest using MTT, flow cytometry, and Transwell assays, reinforcing its role as an oncogene.Furthermore, enhanced tumor ablation was determined in a CDT1-downregulated LUAD tumor-bearing nude mouse model. Collectively, our results strongly suggest that E2F2 positively regulates CDT1 expression and actively participates in the progression of lung adenocarcinoma, thereby providing valuable insights into identifying novel therapeutic targets for LUAD treatment.

A recombinant pseudorabies virus surface - displaying the classical swine fever E2 protein induces specific antibodies rapidly.

Pseudorabies virus (PRV) and classical swine fever virus (CSFV) are both economically important pathogens threatening the pig industry in many countries. The triple-gene-deleted variant of PRV, herein referred to as rPRVTJ-delgE/gI/TK, has exhibited pronounced efficacy and safety profiles. This underscores its viability as a prospective vaccine vector. However, the generation of specific anti-E2 antibodies necessitates elevated immunization doses and extended durations when the extracellular domain of the E2 protein of CSFV is secreted via the recombinant rPRVTJ-delgE/gI/TK vector. To enhance the presentation of exogenous antigens by antigen-presenting cells (APCs), we engineered the E2 protein expressed on the surface of PRV particles in this study. The recombinant virus expressing the E2 protein with a heterogonous transmembrane domain was generated in the backbone of rPRVTJ-delgE/gI/TK and designated as rPRVTJ-UL44-E2. The E2 gene was fused to the 3' terminus of the UL44 gene utilizing P2A, a self-cleaving peptide sequence. The electron microscopy showed that the E2 protein was anchored on the surface of the viral particles of rPRVTJ-delgE/gI/TK-E2. The insertion of the E2 gene did not alter the native biological characteristics of the viral vector. Rabbits immunized with 107 median tissue culture infective doses (TCID50) of rPRVTJ-UL44-E2 exhibited a rapid seroconversion to anti-E2 specific antibodies within 7 days post-immunization (dpi). All the rabbits immunized with the rPRVTJ-UL44-E2 had generated antibodies specific to E2 prior to the administration of the booster immunization. However, the immunized rabbits were not protected from the CSFV C-strain challenge. Nevertheless, this strategy has notably achieved rapid induction of E2-specific non-neutralizing antibodies. These findings provide insights that the design of rPRVTJ-UL44-E2 requires optimization, thereby indicating a promising avenue for augmenting vaccine-induced immune responses.

Homoharringtonine promotes non-small-cell lung cancer cell death via modulating HIF-1α/ERß/E2F1 feedforward loop.

OBJECTIVES: Hypoxia conditions promote the adaptation and progression of non-small-cell lung cancer (NSCLC) via hypoxia-inducible factors (HIF). HIF-1α may regulate estrogen receptor ß (ERß) and promote the progression of NSCLC. The phytochemical homoharringtonine (HHT) exerts strong inhibitory potency on NSCLC, with molecular mechanism under hypoxia being elusive. METHODS: The effects of HHT on NSCLC growth were determined by cell viability assay, colony formation, flow cytometry, and H460 xenograft models. Western blotting, molecular docking program, site-directed mutagenesis assay, immunohistochemical assay, and immunofluorescence assay were performed to explore the underlying mechanisms of HHT-induced growth inhibition in NSCLC. KEY FINDINGS: HIF-1α/ERß signaling-related E2F1 is highly expressed and contributes to unfavorable survival and tumor growth. The findings in hypoxic cells, HIF-1α overexpressing cells, as well as ERß- or E2F1-overexpressed and knockdown cells suggest that the HIF-1α/ERß/E2F1 feedforward loop promotes NSCLC cell growth. HHT suppresses HIF-1α/ERß/E2F1 signaling via the ubiquitin-proteasome pathway, which is dependent on the inhibition of the protein expression of HIF-1α and ERß. Molecular docking and site-directed mutagenesis revealed that HHT binds to the GLU305 site of ERß. HHT inhibits cell proliferation and colony formation and promotes apoptosis in both NSCLC cells and xenograft models. CONCLUSION: The formation of the HIF-1α/ERß/E2F1 feedforward loop promotes NSCLC growth and reveals a novel molecular mechanism by which HHT induces cell death in NSCLC.

Knockdown of PAIP1 Inhibits Breast Cancer Cell Proliferation by Regulating Cyclin E2 mRNA Stability.

Polyadenylate-binding protein-interacting protein 1 (PAIP1) is a protein that modulates translation initiation in eukaryotic cells. Studies have shown that PAIP1 was overexpressed in various type of cancers, and drives cancer progression by promoting cancer cell proliferation, invasion, and migration. In our previous study, we identified that PAIP1 was overexpressed in breast cancer, and the expression was correlated with poor prognosis. However, the biological function of PAIP1 in breast cancer has not been clearly understood. In this study, we constructed PAIP1 specifically silenced breast cancer cells. Then, cell proliferation, cell cycle distribution, and apoptosis were detected in PAIP1 knockdown cells. RNA-seq analysis was performed to predict the downstream target of PAIP1, and the molecular mechanism was explored. As a results, we found that knockdown of PAIP1 repressed cell proliferation, induced cell cycle arrest, and triggers apoptosis. Xenograft mouse model showed that knockdown of PAIP1 inhibits cell growth in vivo. RNA-seq predicted that CCNE2 mRNA was one of the downstream targets of PAIP1. In addition, we identified that knockdown of PAIP1-inhibited cell proliferation through modulating cyclin E2 expression. Mechanically, knockdown of PAIP1 reduces the expression of cyclin E2 by regulating the mRNA stability of cyclin E2. Moreover, in breast cancer tissues, we found that the expression of PAIP1 was positively correlated with cyclin E2. Taken together, our findings establish the role and mechanism of PAIP1 in breast cancer progression, indicating that PAIP1 would be a new therapeutic target for breast cancer treatment.

Risk Factors for E2SKAPE Infections and Mortality Among Liver Transplant Recipients.

PURPOSE: Infections caused by Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp (ESKAPE) plus Escherichia coli (E2SKAPE), in particular multidrug-resistant (MDR) E2SKAPE infections, occur frequently and pose a life-threatening to liver transplant (LT) recipients. To prevent E2SKAPE infections and improve the prognosis of LT recipients, the identification of risk factors for E2SKAPE infections and mortality is necessary. METHODS: E2SKAPE pathogens were isolated and identified from clinical samples following standard microbiological procedures. All episodes of E2SKAPE infections and mortality documented among LT recipients were analyzed. FINDINGS: A total of 83 episodes of E2SKAPE infections, including 75 (90.4%) episodes of MDR-E2SKAPE infections, occurred in 23.1% (53/229) of LT recipients. E. faecium was the dominant causative bacterium (37/83; 44.6%). The most common site of infections was the urinary tract (14/53; 26.4%). Sixteen (7%) patients died within 2 months after LT, and 7 deaths were E2SKAPE infections-related. Multivariate logistic regression analysis revealed that female sex [odds ratio (OR) = 3.665, 95% confidence interval (CI): 1.614-8.321, P = 0.002], duration of surgery ≥ 400 min [OR = 2.328, 95%CI: 1.151-4.707, P = 0.019], intraoperative red blood cell (RBC) transfusion ≥ 12U [OR = 2.542, 95%CI: 1.218-5.306, P = 0.013] and indwelling urethral catheter use ≥ 3 days [OR = 3.96, 95%CI: 1.309-11.981, P = 0.015] were independent risk factors for E2SKAPE infections after LT, and that only exposure to more than 2 intravenous antibiotics post-LT [OR = 0.318, 95%CI: 0.15-0.674, P = 0.003] was negatively associated with acquisition of E2SKAPE infections. The predictors of crude mortality included female sex [OR = 4.822, 95%CI: 1.299-17.904, P = 0.019], creatinine on day 3 post-LT > 1.5 mg/dL [OR = 11.014, 95%CI: 2.985-40.637, P < 0.001], mechanical ventilation post-LT [OR = 10.724, 95%CI: 2.695-42.676, P = 0.001] and recipients with E2SKAPE infections [OR = 4.112, 95%CI: 1.169-14.47, P = 0.028]. IMPLICATIONS: A high incidence of E2SKAPE infections was noted in the early post-LT period. The most common infection site was the urinary tract, and the dominant pathogenic bacterium was E. faecium. Female sex, prolonged surgery time, massive RBC transfusion, or delayed urethral catheter removal were associated with E2SKAPE infections. Only exposure to more than 2 intravenous antibiotics post-LT was negatively related to the acquisition of E2SKAPE infections. The predictors of mortality included female sex, creatinine on day 3 post-LT>1.5 mg/dL, mechanical ventilation post-LT, and recipients with E2SKAPE infections.

High concentration of estrogen resulted by COH may affect the secretion of pro-angiogenic factors in uNK cells by downregulating the expression of IL-11 in decidual stromal cells.

OBJECTIVE: High serum estrogen concentrations after controlled ovarian hyperstimulation (COH) and fresh embryo transfers are associated with the increased risk of pregnancy complications resulting from aberrant placentation. Uterine natural killer (uNK) cells are important for establishment of pregnancy and normal placentation. It has been found that the proliferation and function of uNK cells are compromised by COH. However, the underlying role of high concentration of estrogen following COH in the abnormalities of uNK cells is poorly understood. METHODS: Expression of cytokines and immunophenotype study of uNK was performed by flow cytometry analysis. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to quantify RNA expression; Western blot was performed to quantify protein levels. RESULTS: The secretion level of pro-angiogenic factors in uNK cells is significantly reduced by co-culture with decidual stromal cells (DSCs) induced by high estrogen. It was discovered that COH and supraphysiologic levels of estrogen downregulated IL-11 in decidual tissue of mice. Additionally, we found that the downregulation of IL-11 is a major factor contributing to the downregulation of VEGF and PLGF in uNK cells. Moreover, we found that uNK cells may acquire IL-11Rα sequentially during differentiation and that only a portion of uNK cells are IL-11Rα positive. Lastly, we discovered that IL-11 may regulate VEGF and PLGF secretion in uNK cells via the ERK signaling pathway. CONCLUSION: These results suggested the downregulation of IL-11 expression in DSCs caused by high estrogen levels affects the secretion of pro-angiogenic factors in uNK cells, which provided an explanation for the pregnancy complications caused by COH.

Inhibition of 15-prostaglandin dehydrogenase attenuates acetaminophen-induced liver injury via suppression of apoptosis in liver endothelial cells.

Hepatic microvascular disruption caused by injury to liver sinusoidal endothelial cells (LSECs) is an aggravating factor for drug-induced liver injury (DILI). It is suggested that prostaglandin E2 (PGE2) may be able to attenuate LSEC injury. However, it is also known that 15-keto PGE2, a metabolite of PGE2 produced by 15-prostaglandin dehydrogenase (15-PGDH) that is not a ligand of PGE2 receptors, suppresses inflammatory acute liver injury as a ligand of peroxisome proliferator-activated receptor γ. In this study, we aimed to understand whether 15-PGDH activity is essential for preventing DILI by suppressing hepatic microvascular disruption in a mouse model of acetaminophen (APAP)-induced liver injury. To inhibit 15-PGDH activity prior to APAP-induced LSEC injury, we administered the 15-PGDH inhibitor, SW033291, 1 h before and 3 h after APAP treatment. We observed that LSEC injury preceded hepatocellular injury in APAP administered mice. Hepatic endogenous PGE2 levels did not increase up till the initiation of LSEC injury but rather increased after hepatocellular injury. Moreover, hepatic 15-PGDH activity was downregulated in APAP-induced liver injury. The inhibition of 15-PGDH attenuated LSEC injury and subsequently hepatic injury by inhibiting apoptosis in APAP administered mice. Our in vitro studies also suggested that PGE2 inhibited APAP-induced apoptosis via the EP4/PI3K pathway in endothelial cells. Therefore, a decrease in 15-PGDH activity would be beneficial for preventing APAP-induced liver injury by attenuating LSEC injury.

Transcription factor E2F4 facilitates SUMOylation to promote HCC progression through interaction with LIN9.

SUMOylation plays a crucial role in numerous cellular biological and pathophysiological processes associated with human disease; however, the mechanisms regulating the genes involved in SUMOylation remain unclear. In the present study, E2F transcription factor 4 (E2F4) was identified as an E2F member related to hepatocellular carcinoma (HCC) progression by public database analysis. It was found that E2F4 promoted the proliferation and invasiveness of HCC cells via SUMOylation using Soft agar and Transwell migration assays. Mechanistically, it was demonstrated that E2F4 upregulated the transcript and protein expression levels of baculoviral IAP repeat containing 5, cell division cycle associated 8 and DNA topoisomerase II α using western blotting. Furthermore, the interaction between E2F4 with lin­9 DREAM multi­vulva class B core complex component (LIN9) was explored by co­immunoprecipitation, immunofluorescence co­localization and bimolecular fluorescence complementation assays. Moreover, it was demonstrated that E2F4 promoted the progression of HCC cells via LIN9. Rescue experiments revealed that LIN9 facilitated the SUMOylation and proliferation of HCC cells, which was prevented by knocking down E2F4 expression. In conclusion, the findings of the present study indicated that E2F4 plays a major role in the proliferation of HCC cells and may be a potential therapeutic target in the future.

Chemical tools for probing the Ub/Ubl conjugation cascades.

Conjugation of ubiquitin (Ub) and structurally related ubiquitin-like proteins (Ubl's), essential for many cellular processes, employs muti-step reactions orchestrated by specific E1, E2 and E3 enzymes. The E1 enzyme activates the Ub/Ubl C-terminus in an ATP-dependent process that results in the formation of a thioester linkage with the E1 active site cysteine. The thioester activated Ub/Ubl is transferred to the active site of an E2 enzyme which then interacts with an E3 enzyme to promote conjugation to the target substrate. The E1-E2-E3 enzymatic cascades utilize labile intermediates, extensive conformational changes, and vast combinatorial diversity of short-lived protein-protein complexes to conjugate Ub/Ubl to various substrates in a regulated manner. In this review, we discuss various chemical tools and methods used to study the consecutive steps of Ub/Ubl activation and conjugation, which are often too elusive for direct studies. We focus on methods developed to probe enzymatic activities and capture and characterize stable mimics of the transient intermediates and transition states thereby providing insights into fundamental mechanisms in the Ub/Ubl conjugation pathways.

Actively changing the narrative: An exploration of culturally grounded parenting and social skills.

Recognizing culturally salient aspects of socialization practices and understanding how these practices support culturally valued aspects of development is an integral component in conducting anti-racist research and validating the lived experiences of minoritized families. With this aim, we explored how Active Direction, an observational rating of an African American approach to parenting measured during mother-child interactions at age 2.5 (n = 172), supported social skills and emotion regulation for children living in a Southwestern metropolitan area of the United States concurrently, in kindergarten (n = 109), and in 1st grade (n = 108). Descriptive findings indicated few significant associations between Active Direction and socials skills or emotion regulation. Exploratory analyses, which included traditional parenting behavior measures of Sensitivity and Intrusiveness, also indicated limited significant relations between any measure of parenting and child skills. However, moderation analyses indicated that high levels of Active Direction attenuated the effects of sensitivity on aspects of child social skills. The lack of significant findings across the current study highlight how extant measures-of child social skills and parentings behaviors-are not performing as expected within these African American families.

Reconocer aspectos culturalmente salientes de las prácticas de socialización y comprender cómo estas prácticas apoyan aspectos del desarrollo culturalmente valorados, es un componente integral para llevar a cabo la investigación antiracista y darle validez a las experiencias vividas de familias vistas como minorías. Con este propósito, exploramos cómo Activa Direccción, una evaluación de observación de un acercamiento afroamericano a la crianza medido durante las interacciones madre­niño a la edad de 2.5 (n = 172), apoyaba las habilidades sociales y la regulación de la emoción para niños que vivían en un área metropolitana de los Estados Unidos, de manera concurrente, en el kinder (n = 109) y en el primer grado (n = 108). Los resultados descriptivos señalaron pocas asociaciones significativas entre Activa Direccción y las habilidades sociales o la regulación de la emoción. Los análisis exploratorios, los cuales incluyen medidas de Sensibilidad y de Entremetimiento en cuanto al comportamiento de crianza tradicional, también señalaron limitadas relaciones significativas entre cualquier medida de crianza y las habilidades sociales. Sin embargo, los análisis de moderación señalaron que altos niveles de Activa Dirección atenúan los efectos de la Sensibilidad sobre los aspectos de las habilidades sociales del niño. La falta de resultados significativos, a lo largo del presente estudio, subraya hasta qué punto las medidas ­de habilidades sociales del niño y de comportamientos de crianza­ no están actuando de la manera esperada dentro de estas familias afroamericanas.

[NRF2 mediated redox stress in arsenic induced human keratinocytes malignant transformation].

OBJECTIVE: To explore the role of nuclear transcription factor E2-related factor 2(NRF2)-mediated reductive stress in arsenite induced malignant transformation in human keratinocytes. METHODS: HaCaT cells and fluorescent labeled mitochondrial glutathione HaCaT cells(Mito-Grx1-roGFP2 HaCaT) were cultured to 35 passages in medium containing 0.0 and 1.0 µmol/L NaAsO_2 to establish a model of malignant transformation of cells. Cellular and mitochondrial reduced glutathione/oxidized glutathione(GSH/GSSG) and reduced coenzyme II/oxidized coenzyme II(NADPH/NADP~+) ratios were measured in HaCaT cells. Cell doubling time, cell migration ability, soft agar clone formation ability and GSH/GSSG at different times in the 0 passage, the early stage(1st, 7th and 14th passages) and later stage(21st, 28th and 35th passages) were measured in Mito-Grx1-roGFP2 HaCaT cells. NaAsO_2 induced malignant transformation cells were transfected with NRF2 siRNA, and detected the expression level of NRF2 and the redox-related indexes and malignant transformation indexes. RESULTS: Compared with the control group, the GSH/GSSG ratio in 1.0 µmol/L NaAsO_2 treated HaCaT cells significantly decreased in the 1st and 7th generations, but significantly increased after the 21st generation, and the NADPH/NADP~+ ratio significantly increased in the 1st, 14th, 21st, 28th and 35th generations; The levels of GSH/GSSG in mitochondria significantly increased from 1st to 35th generation, and the levels of NADPH/NADP~+ in mitochondria significantly increased at 1st, 7th, 21st, 28th and 35th generation. After continuous treatment of Mito-Grx1-roGFP2 HaCaT cells with 0.0 or 1.0 µmol/L NaAsO_2 to 35 passages, the doubling time of cells treated with 1.0 µmol/L NaAsO_2 was significantly shortened, the cell migration rate was increased greatly, and more clones with larger volumes than the control cells formed. The GSH/GSSG ratio in mitochondria of Mito-Grx1-roGFP2 HaCaT cells showed a significant decrease in the 1st generation and increased from the 7th generation onwards(all P<0.05). After transfection of NaAsO_2 treated cells with NRF2 siRNA, the levels of hydrogen peroxide and superoxide increased compared with the siRNA controls. The levels of cell and mitochondrial NADPH/NADP~+ and GSH/GSSG decreased and the level of mitochondrial GSH/GSSG in Mito-Grx1-roGFP2 HaCaT cells decreased. Cell doubling time increased, cell migration rate and soft agar clone formation ability decreased(all P<0.05). The malignant phenotype was reversed. CONCLUSION: In the early stage(1st, 7th and 14th passages) of NaAsO_2 treated HaCaT cells, oxidative stress occurred with continuous high NRF2 expression. Later(21st, 28th and 35th passages), NRF2 induced reductive stress, leading to malignant transformation.

Sulforaphane improves post-resuscitation myocardial dysfunction by inhibiting cardiomyocytes ferroptosis via the Nrf2/IRF1/GPX4 pathway.

BACKGROUND: Ferroptosis is an important type of cell death contributing to myocardial dysfunction induced by whole body ischemia reperfusion following cardiac arrest (CA) and resuscitation. Sulforaphane (SFN), known as the activator of the nuclear factor E2-related factor 2 (Nrf2), has been proven to effectively alleviate regional myocardial ischemia reperfusion injury. The present study was designed to investigate whether SFN could improve post-resuscitation myocardial dysfunction by inhibiting cardiomyocytes ferroptosis and its potential regulatory mechanism. METHODS AND RESULTS: An in vivo pig model of CA and resuscitation was established. Hypoxia/reoxygenation (H/R)-stimulated AC16 cardiomyocytes was constructed as an in vitro model to simulate the process of CA and resuscitation. In vitro experiment, SFN reduced ferroptosis-related ferrous iron, lipid reactive oxygen species, and malondialdehyde, increased glutathione, and further promoted cell survival after H/R stimulation in AC16 cardiomyocytes. Mechanistically, the activation of Nrf2 with the SFN decreased interferon regulatory factor 1 (IRF1) expression, then reduced its binding to the promoter of glutathione peroxidase 4 (GPX4), and finally recovered the latter's transcription after H/R stimulation in AC16 cardiomyocytes. In vivo experiment, SFN reversed abnormal expression of IRF1 and GPX4, inhibited cardiac ferroptosis, and improved myocardial dysfunction after CA and resuscitation in pigs. CONCLUSIONS: SFN could effectively improve myocardial dysfunction after CA and resuscitation, in which the mechanism was potentially related to the inhibition of cardiomyocytes ferroptosis through the regulation of Nrf2/IRF1/GPX4 pathway.

Spinster homolog 2/S1P signaling ameliorates macrophage inflammatory response to bacterial infections by balancing PGE2 production.

BACKGROUND: Mitochondria play a crucial role in shaping the macrophage inflammatory response during bacterial infections. Spinster homolog 2 (Spns2), responsible for sphingosine-1-phosphate (S1P) secretion, acts as a key regulator of mitochondrial dynamics in macrophages. However, the link between Spns2/S1P signaling and mitochondrial functions remains unclear. METHODS: Peritoneal macrophages were isolated from both wild-type and Spns2 knockout rats, followed by non-targeted metabolomics and RNA sequencing analysis to identify the potential mediators through which Spns2/S1P signaling influences the mitochondrial functions in macrophages. Various agonists and antagonists were used to modulate the activation of Spns2/S1P signaling and its downstream pathways, with the underlying mechanisms elucidated through western blotting. Mitochondrial functions were assessed using flow cytometry and oxygen consumption assays, as well as morphological analysis. The impact on inflammatory response was validated through both in vitro and in vivo sepsis models, with the specific role of macrophage-expressed Spns2 in sepsis evaluated using Spns2flox/floxLyz2-Cre mice. Additionally, the regulation of mitochondrial functions by Spns2/S1P signaling was confirmed using THP-1 cells, a human monocyte-derived macrophage model. RESULTS: In this study, we unveil prostaglandin E2 (PGE2) as a pivotal mediator involved in Spns2/S1P-mitochondrial communication. Spns2/S1P signaling suppresses PGE2 production to support malate-aspartate shuttle activity. Conversely, excessive PGE2 resulting from Spns2 deficiency impairs mitochondrial respiration, leading to intracellular lactate accumulation and increased reactive oxygen species (ROS) generation through E-type prostanoid receptor 4 activation. The overactive lactate-ROS axis contributes to the early-phase hyperinflammation during infections. Prolonged exposure to elevated PGE2 due to Spns2 deficiency culminates in subsequent immunosuppression, underscoring the dual roles of PGE2 in inflammation throughout infections. The regulation of PGE2 production by Spns2/S1P signaling appears to depend on the coordinated activation of multiple S1P receptors rather than any single one. CONCLUSIONS: These findings emphasize PGE2 as a key effector of Spns2/S1P signaling on mitochondrial dynamics in macrophages, elucidating the mechanisms through which Spns2/S1P signaling balances both early hyperinflammation and subsequent immunosuppression during bacterial infections.

AB045. E2F6 promotes temozolomide resistance in glioblastoma by enrichment in CARD6 and POSTN promoter region.

BACKGROUND: Glioblastoma (GBM) is the most aggressive primary malignant brain tumor. Temozolomide (TMZ) is the most used first-line chemotherapeutic agent for GBM after surgery, but acquired resistance to TMZ frequently leads to treatment failure and is a major challenge in the clinical treatment of GBM. Increasing evidence suggests that E2F transcription factor 6 (E2F6) is associated with a variety of tumor malignant biological behaviors and drug resistance, but its biological function and underlying molecular mechanisms in GBM are unknown. METHODS: The study investigated the levels of E2F6 in both TMZ-sensitive and TMZ-resistant GBM cells and tissues using Western blotting and immunofluorescence assays. In vitro experiments were conducted to explore the impact of E2F6 on TMZ resistance and glioma stem cell stemness. These experiments included Western blotting, colony formation assay, flow cytometry assay, and TdT-mediated dUTP nick-end labeling (TUNEL) assay. Bioinformatic analyses were conducted to investigate the mechanism behind the high expression of E2F6 in TMZ-resistant cells and its correlation with caspase recruitment domain 6 (CARD6) and disulfide-linked cell adhesion protein (POSTN). The study employed bioinformatic analyses, messenger RNA (mRNA) sequencing, chromatin immunoprecipitation sequencing assay, immunofluorescence, quantitative real-time polymerase chain reaction (qRT-PCR), and Western blotting. To examine the function of E2F6, an intracranial xenograft tumor mouse model was used for in vivo experiments. RESULTS: It was found that CARD6 and POSTN were significantly associated with TMZ resistance and survival of GBM patients. E2F6 was up-regulated in TMZ-resistant cells and tissues. Knockdown of E2F6 down-regulated the expression of CARD6, promoted TMZ-induced apoptosis, and enhanced chemo-sensitivity, whereas its overexpression significantly increased TMZ resistance in vitro and in vivo. In addition, E2F6 can promote TMZ resistance through stem-like properties acquisition. We identified a signaling pathway related to E2F6 and POSTN, which maintains the self-renewal of GBM stem cells (GSCs). E2F6 concentrates in the promoter region of POSTN, thereby regulating the expression of GSCs-related genes cluster of differentiation 133 (CD133), Nestin, and sex-determining region Y-box 2 (SOX2), which may be involved in tumor metabolism and drug resistance processes. Down-regulation of E2F6 down-regulated the expression of POSTN and inhibited tumor growth in nude mice. CONCLUSIONS: These results suggest that the E2F6-CARD6/POSTN signaling axis regulates the malignant biological behaviors of GBM and TMZ resistance. These findings are expected to provide promising therapeutic targets for CARD6 overcoming GBM TMZ resistance.

AB067. New drug development for the use of PARP1-E2F1 transcriptional inhibitors in the treatment of glioblastoma.

BACKGROUND: Glioblastoma (GBM) is the most malignant brain tumor and ranks among the most lethal of all human cancers, without improvements in survival over the last 30 years. Data obtained in our group suggest that PARP1, a well-known DNA-repairing protein, could also play a key role in the regulation of cell cycle through its interaction with the transcription factor E2F1. Therefore, considering that most oncogenic processes are associated with cell cycle deregulation, we hypothesized that disruption of PARP1-E2F1 interaction would provide a novel therapeutic approach to different types of cancer. METHODS: The identification of novel compounds disrupting PARP1-E2F1 interaction was carried out by combining in silico and in vitro screening, using a rational drug design. The virtual screen was performed using a molecular library of several million compounds at the selected target site, using AtomNet® (Atomwise, San Francisco, CA, USA), the first deep learning neural network for structure-based drug design and discovery. Since there is no complete structural information of the PARP1-E2F1 protein-protein interaction, a homologous structure of the BRCT domain of BRCA1 complex with the phospho-peptide (PDBID: 1T2V) was used to identify the potential binding interface of BRCT domain of PARP-1 (PDBID: 2COK) and the E2F1 protein. Top scoring compounds were clustered and filtered to obtain a final subset of 83 compounds that were incorporated to our in vitro screening, which included both transcriptional E2F1 activity and survival studies. Complete culture medium supplemented with the compounds selected in the in silico screening (10 µM) were added and incubated for 24 hours. E2F1 activity was observed by measuring luminescence. For the viability assay, the fluorescence reading was performed (excitation 544 nm and emission 590 nm). RESULTS: The in silico and in vitro screening resulted in 12 compounds that inhibited E2F1 transcriptional activity and significantly reduced cell number. The highest inhibition of both E2F1 transcriptional activity and cell growth was observed with compound 3797, which was selected for further studies. CONCLUSIONS: Both in silico and in vitro results indicate that inhibition of PARP1-E2F1 transcriptional activity may provide a new rationale for designing novel therapeutic approaches for the treatment of GBM.

The Current Role of Analytical Psychology in Maintaining Fictitious Boundaries that are Promoted through the Race Lie: A call to dismantle the virtual wall that exists through attitudes of white supremacy.

In 1928, the American Anthropological Association declared that "Anthropology provided no scientific basis for discrimination against any people on the ground of racial inferiority, religious affiliation, or linguistic heritage" (Guthrie, 1976/1998/2004, p. 30). In 1945, Jung denounced race theory as a pseudo-science. In 1950, UNESCO released its statement denouncing race. Long discredited as scientifically invalid, the race concept still holds uncanny value and significance for Americans and Europeans. In effect, the concept seems to be mysteriously linked to the limited accessibility and the limited economic support that is allotted to people of colour, internationally. This paper will explore the global implications of Jung's expressed attitude towards people of colour prior to 1945, which I identify as an attitude of white supremacy, an attitude that stands in direct contrast to the analytical ethos, as expressed by the International Association for Analytical Psychology (IAAP). This attitude may promote the continuance of racialized beliefs and behaviours within the planning and provision of care to individuals in need of medical and mental health services. It is requested that a written acknowledgment of harm be added to the works of C. G. Jung.

En 1928, l'American Anthropological Association a déclaré que « l'anthropologie ne fournissait aucun fondement scientifique pour la discrimination contre toute personne sur la base de l'infériorité raciale, de l'affiliation religieuse ou de l'héritage linguistique ¼ (Guthrie, 2004, p. 30). En 1945, Jung a dénoncé la théorie de la race comme une pseudoscience. En 1950, l'UNESCO a publié une déclaration dénonçant la notion de race. Longtemps discrédité comme scientifiquement invalide, le concept de race a toujours une valeur et une signification étranges pour les Américains et les Européens. En effet, le concept semble être mystérieusement lié à l'accessibilité limitée et au soutien économique limité qui est accordé aux personnes de couleur, à l'échelle internationale. Cette présentation explorera les implications globales de l'attitude exprimée par Jung envers les personnes de couleur avant 1945, que j'identifie comme une attitude de suprématie blanche, une attitude qui contraste directement avec l'esprit analytique, tel qu'exprimé par l'Association Internationale de Psychologie Analytique. Cette attitude risque de favoriser le maintien de croyances et de comportements racialisés dans la planification et la dispensation de soins aux personnes qui ont besoin de services médicaux et de santé mentale. Il est demandé qu'une reconnaissance écrite du préjudice causé soit ajoutée aux travaux de C. G. Jung.

En 1928, la Asociación Americana de Antropología declaró que "la antropología no proporcionaba ninguna base científica para discriminar a un pueblo sobre la base de inferioridad racial, afiliación religiosa o herencia lingüística" (Guthrie, 2004, p. 30). En 1945, Jung denunció la teoría racial como pseudociencia. En 1950, la UNESCO publicó su declaración denunciando la raza. Desacreditado hace tiempo como científicamente inválido, el concepto de raza sigue teniendo un valor y un significado sorprendente para estadounidenses y europeos. En efecto, el concepto parece estar misteriosamente vinculado a la limitada accesibilidad y al limitado apoyo económico que se asigna a las personas de color, a escala internacional. Esta presentación explorará las implicancias globales de la actitud expresada por Jung hacia la gente de color antes de 1945, la cual identifico como una actitud de supremacía blanca, una actitud que contrasta directamente con el ethos analítico, tal y como lo expresa la Asociación Internacional de Psicología Analítica. Esta actitud puede promover la continuidad de creencias y comportamientos raciales en la planificación y provisión de cuidados a individuos que necesitan servicios médicos y de salud mental. Se solicita que se añada por escrito a las obras de C. G. Jung un reconocimiento del daño.