Limited prognostic role of routine serum markers (AP, CEA, LDH and NSE) in oligorecurrent prostate cancer patients undergoing PSMA-radioguided surgery.

INTRODUCTION: We evaluated the prognostic role of pre-salvage prostate-specific membrane antigen-radioguided surgery (PSMA-RGS) serum levels of alkaline phosphatase (AP), carcinoembryonic antigen (CEA), lactate dehydrogenase (LDH), and neuron-specific enolase (NSE). MATERIALS AND METHODS: Patients who consecutively underwent PSMA-RGS for prostate cancer (PCa) oligorecurrence between January 2019 and January 2022 were selected. Biomarkers were assessed one day before surgery. Cox regression and logistic regression models tested the relationship between biochemical recurrence-free survival (BFS), 6- and 12-month biochemical recurrence (BCR), and several independent variables, including biomarkers. RESULTS: 153 consecutive patients were analyzed. In the univariable Cox regression analysis, none of the biomarkers achieved predictor status (AP: hazard ratio [HR] = 1.03, 95% CI 0.99, 1.01; p = 0.19; CEA: HR = 1.73, 95% CI 0.94, 1.21; p = 0.34; LDH: HR = 1.01, 95% CI 1.00, 1.01; p = 0.05; NSE: HR = 1.02, 95% CI 0.98, 1.06; p = 0.39). The only independent predictor of BFS was the number of positive lesions on PSMA PET (HR = 1.17, 95% CI 1.02, 1.30; p = 0.03). The number of positive lesions was confirmed as independent predictor for BCR within 6 and 12 months (BCR < 6 months: odds ratio [OR] = 1.1, 95% CI 1.0, 1.3; p = 0.04; BCR < 12 months: OR = 1.1, 95% CI 1.0, 1.3; p = 0.04). CONCLUSION: The assessment of AP, CEA, LDH, and NSE before salvage PSMA-RGS showed no prognostic impact. Further studies are needed to identify possible predictors that will optimize patient selection for salvage PSMA-RGS.

Application Value of CYFRA21-1 Combined with NSE, CEA, and SCC-Ag in Lung Cancer.

BACKGROUND: This study aims to investigate the application value of serum cytokeratin 19 fragment (CYFRA21-1) combined with nerve-specific enolase (NSE), carcinoembryonic antigen (CEA), and squamous cell carcinoma antigen (SCC-Ag) in the diagnosis of lung cancer (LC). METHODS: A total of 831 cases of LC, 360 cases of benign lung disease (BLD) and 102 healthy controls, were enrolled. The data were processed using SPSS, GraphPad Prism, and MedCalc software. RESULTS: The tumor marker (TM) levels in the LC and BLD groups were significantly higher than those in the control group; the CYFRA21-1, NSE, and CEA levels in the patients with LC were higher than in those with BLD. In particular, the increase was predominantly observed for the levels of CEA and CYFRA21-1 in adenocarcinoma (LUAD), CYFRA21-1 and SCC-Ag in squamous cell carcinoma (LUSC), and NSE in small cell carcinoma (SCLC). The CYFRA21-1, NSE, and CEA levels were significantly higher in stage IV than in other stages in LC. Univariate binary logistic analysis showed that increased levels of all four TMs were risk factors for BLD and LC. The area under the curve (AUC) of CYFRA21-1 was most effective in distinguishing patients with BLD or LC from the controls and in distinguishing patients with BLD and LC. The AUCs of combined CYFRA21-1, NSE, and CEA were increased to 0.755, 0.922, and 0.783, respectively, with no significant difference with the AUC of the four combined tests. In the histological classification, the best predictors were CEA, for LUAD, CYFRA21-1 for LUSC, and NSE for SCLC. Moreover, the expression levels of CYFRA21-1, NSE, and CEA significantly decreased after each treatment course. CONCLUSIONS: The combined assay of CYFRA21-1, NSE, and CEA addresses the aspects of accuracy, sensitivity, specificity, and economic cost and should be considered as a potential diagnostic test in LC.

ENO1 promotes trophoblast invasion regulated by E2F8 in recurrent miscarriage.

Recurrent miscarriage (RM) is related to the dysfunction of extravillous trophoblast cells (EVTs), but the comprehensive mechanisms remain largely unexplored. We analyzed single-cell RNA sequencing (scRNA-seq), bulk RNA sequencing and microarray datasets obtained from Gene Expression Omnibus (GEO) database to explore the hub genes in the mechanisms of RM. We identified 1724 differentially expressed genes (DEGs) in EVTs from the RM, and they were all expressed along the trajectory of EVTs. These DEGs were associated with hypoxia and glucose metabolism. Single-cell Regulatory Network Inference and Clustering (SCENIC) analysis revealed that E2F transcription factor (E2F) 8 (E2F8) was a key transcription factor for these DEGs. And the expression of ENO1 can be positively regulated by E2F8 via RNA sequencing analysis. Subsequently, we performed immunofluorescence assay (IF), plasmid transfection, western blotting, chromatin immunoprecipitation (ChIP), real-time quantitative polymerase chain reaction (qRT-PCR), and transwell assays for validation experiments. We found that the expression of alpha-Enolase 1 (ENO1) was lower in the placentas of RM. Importantly, E2F8 can transcriptionally regulate the expression of ENO1 to promote the invasion of trophoblast cells by inhibiting secreted frizzled-related protein 1/4 (SFRP1/4) to activate Wnt signaling pathway. Our results suggest that ENO1 can promote trophoblast invasion via an E2F8-dependent manner, highlighting a potential novel target for the physiological mechanisms of RM.

Meta-analysis of evaluating neuron specific enolase as a serum biomarker for sepsis-associated encephalopathy.

INTRODUCTION: Brain dysfunction in sepsis is known as Sepsis-associated encephalopathy (SAE), which often results in severe cognitive and neurological sequelae and increases the risk of death. Neuron specific enolase (NSE) may serve as an important neurocritical biomarker for detection and longitudinal monitoring in SAE patients. Our Meta-analysis aimed to explore the diagnostic and prognostic value of serum NSE in SAE patients. Currently, no systematic Review and Meta-analysis have been assessed that NSE as a biomarker of SAE. METHODS: The study protocol was registered in the PROSPERO database (CRD42023398736) and adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. We conducted a systematic review and Meta-analysis to evaluate the serum NSE's diagnostic accuracy for SAE and prognostic strength for probability of death of septic patients. We systematic searched electronic bibliographic databases from PubMed, MEDLINE, Web of Science, Embase, Cochrane databases, CNKI, CQVIP, and WFSD. QUADAS-2 assessment tool was used to evaluate quality and risk of bias of the selected studies. Subgroup analyses, funnel plots, sensitivity analyses were also carried out. Review Manager version 5.4 and Stata16.0. was used for statistical analysis. RESULTS: This Meta-analysis included 22 studies with 1361 serum samples from SAE patients and 1580 serum samples from no-encephalopathy septic (NE) patients. The Meta-analysis showed that individuals with SAE had higher serum NSE level than NE controls (SMD 1.93 (95 % CI 1.51-2.35), P < 0.00001). In addition, there are 948 serum samples from survival septic patients and 446 serum samples from non-survival septic patients, septic patients with survival outcomes had lower serum NSE levels than those with death outcomes (SMD -1.87 (95 % CI -2.43 to -1.32), P < 0.00001). CONCLUSION: Our Meta-analysis reveals a significant association between elevated NSE concentrations and the increased likelihood of concomitant SAE and mortality during septic patients. This comprehensive analysis will equip ICU physicians with up-to-date insights to accurately identify patients at risk of SAE and implement appropriate intervention strategies to mitigate morbidity and improve neurological outcomes. However, it is important to note that the presence of substantial heterogeneity among studies poses challenges in determining the most effective discrimination cutoff values and optimal sampling collection time.

Novel immunosensor based on electrochemiluminescence inner filter effect and static quenching between fibrillary Ag-MOGs and SiO2@PANI@AuNPs for enabling the sensitive detection of neuron-specific enolase.

Metal-organic gels (MOGs) are unique supramolecular gels that are convenient to synthesize. In this work, a cathodic electrochemiluminescence (ECL) system based on Ag-MOGs as a luminophore and K2S2O8 as a co-reactor was developed. The ECL spectrum of the Ag-MOGs overlapped significantly with the strong UV-Vis spectrum of the SiO2@PANI@AuNPs, which effectively quenched the ECL luminescence of the Ag-MOGs. Relying on the inner filter effect between Ag-MOGs and SiO2@PANI@AuNPs, a novel ECL-IFE immunosensor was developed for the detection of neuron-specific enolase (NSE). Under optimal conditions, the ECL signal of the immunosensor displayed excellent linearity over the NSE concentration range of 10 fg/mL-100 ng/mL. The limit of detection (LOD) was 2.6 fg/mL (S/N = 3) with a correlation coefficient R2 of 0.9975. The ECL immunosensor also exhibited excellent stability and reproducibility for the detection of NSE. The results reported provide a feasible concept for the development analytical methods for the detection of other clinically relevant biomarkers.

Factors influencing blood tumor marker concentrations in the absence of neoplasia.

BACKGROUND: Tumor markers (TMs) are a heterogeneous group of molecules used in the diagnosis, prognosis and follow-up of cancer patients. During neoplastic differentiation, cells can either directly synthesize or induce the synthesis of TMs, and the release of these molecules into the bloodstream allows their quantification in biological fluids. Although very small concentrations of TMs are usually present in the serum or plasma of healthy subjects, increased concentrations may also be found in the presence of benign diseases or due to technical interference, producing false positive results. MATERIAL AND METHODS AND RESULTS: Our review analyses the causes of false positives described between January 1970 to February 2023 for the TMs most frequently used in clinical practice: α-fetoprotein (AFP), ß2-microglobulin (ß2-M), cancer antigen 15-3 (CA 15-3), cancer antigen CA 19-9 (CA 19-9), cancer antigen CA 72-4 (CA 72-4), cancer antigen 125 (CA 125), carcinoembryonic antigen (CEA), chromogranin A (CgA), choriogonadotropin (hCG), cytokeratin 19 fragment (CYFRA 21-1), neuron-specific enolase (NSE), human epididymis protein 4 (HE4), serum HER2 (sHER2), squamous cell carcinoma antigen (SCCA), protein induced by vitamin K absence-II (PIVKA-II), Pro-gastrin-releasing peptide (Pro-GRP), prostate-specific antigen (PSA), Protein S-100 (S-100) and thyroglobulin (Tg). A total of 247 references were included. CONCLUSIONS: A better understanding of pathophysiological processes and other conditions that affect the concentration of TMs might improve the interpretation of results and their clinical application.

Glycolytic enzyme Enolase-1 regulates insulin gene expression in pancreatic ß-cell.

Enolase-1 (Eno1) plays a critical role in regulating glucose metabolism; however, its specific impact on pancreatic islet ß-cells remains elusive. This study aimed to provide a preliminary exploration of Eno1 function in pancreatic islet ß-cells. The findings revealed that the expression of ENO1 mRNA in type 2 diabetes donors was significantly increased and positively correlated with HbA1C and negatively correlated with insulin gene expression. A high level of Eno1 in human insulin-secreting rat INS-1832/13 cells with co-localization with intracellular insulin proteins was accordingly observed. Silencing of Eno1 using siRNA or inhibiting Eno1 protein activity with an Eno1 antagonist significantly reduced insulin secretion and insulin content in ß-cells, while the proinsulin/insulin content ratio remained unchanged. This reduction in ß-cells function was accompanied by a notable decrease in intracellular ATP and mitochondrial cytochrome C levels. Overall, our findings confirm that Eno1 regulates the insulin secretion process, particularly glucose metabolism and ATP production in the ß-cells. The mechanism primarily involves its influence on insulin production, suggesting that Eno1 represents a potential target for ß-cell protection and diabetes treatment.

Identification of host protein ENO1 (alpha-enolase) interacting with Cryptosporidium parvum sporozoite surface protein, Cpgp40.

BACKGROUND: Cryptosporidium parvum is an apicomplexan zoonotic parasite causing the diarrheal illness cryptosporidiosis in humans and animals. To invade the host intestinal epithelial cells, parasitic proteins expressed on the surface of sporozoites interact with host cells to facilitate the formation of parasitophorous vacuole for the parasite to reside and develop. The gp40 of C. parvum, named Cpgp40 and located on the surface of sporozoites, was proven to participate in the process of host cell invasion. METHODS: We utilized the purified Cpgp40 as a bait to obtain host cell proteins interacting with Cpgp40 through the glutathione S-transferase (GST) pull-down method. In vitro analysis, through bimolecular fluorescence complementation assay (BiFC) and coimmunoprecipitation (Co-IP), confirmed the solid interaction between Cpgp40 and ENO1. In addition, by using protein mutation and parasite infection rate analysis, it was demonstrated that ENO1 plays an important role in the C. parvum invasion of HCT-8 cells. RESULTS: To illustrate the functional activity of Cpgp40 interacting with host cells, we identified the alpha-enolase protein (ENO1) from HCT-8 cells, which showed direct interaction with Cpgp40. The mRNA level of ENO1 gene was significantly decreased at 3 and 24 h after C. parvum infection. Antibodies and siRNA specific to ENO1 showed the ability to neutralize C. parvum infection in vitro, which indicated the participation of ENO1 during the parasite invasion of HCT-8 cells. In addition, we further demonstrated that ENO1 protein was involved in the regulation of cytoplasmic matrix of HCT-8 cells during C. parvum invasion. Functional study of the protein mutation illustrated that ENO1 was also required for the endogenous development of C. parvum. CONCLUSIONS: In this study, we utilized the purified Cpgp40 as a bait to obtain host cell proteins ENO1 interacting with Cpgp40. Functional studies illustrated that the host cell protein ENO1 was involved in the regulation of tight junction and adherent junction proteins during C. parvum invasion and was required for endogenous development of C. parvum.

Identification of plasminogen-binding sites in Streptococcus suis enolase that contribute to bacterial translocation across the blood-brain barrier.

Streptococcus suis is an emerging zoonotic pathogen that can cause invasive disease commonly associated with meningitis in pigs and humans. To cause meningitis, S. suis must cross the blood-brain barrier (BBB) comprising blood vessels that vascularize the central nervous system (CNS). The BBB is highly selective due to interactions with other cell types in the brain and the composition of the extracellular matrix (ECM). Purified streptococcal surface enolase, an essential enzyme participating in glycolysis, can bind human plasminogen (Plg) and plasmin (Pln). Plg has been proposed to increase bacterial traversal across the BBB via conversion to Pln, a protease which cleaves host proteins in the ECM and monocyte chemoattractant protein 1 (MCP1) to disrupt tight junctions. The essentiality of enolase has made it challenging to unequivocally demonstrate its role in binding Plg/Pln on the bacterial surface and confirm its predicted role in facilitating translocation of the BBB. Here, we report on the CRISPR/Cas9 engineering of S. suis enolase mutants eno261, eno252/253/255, eno252/261, and eno434/435 possessing amino acid substitutions at in silico predicted binding sites for Plg. As expected, amino acid substitutions in the predicted Plg binding sites reduced Plg and Pln binding to S. suis but did not affect bacterial growth in vitro compared to the wild-type strain. The binding of Plg to wild-type S. suis enhanced translocation across the human cerebral microvascular endothelial cell line hCMEC/D3 but not for the eno mutant strains tested. To our knowledge, this is the first study where predicted Plg-binding sites of enolase have been mutated to show altered Plg and Pln binding to the surface of S. suis and attenuation of translocation across an endothelial cell monolayer in vitro.

Novel electrochemical platform based on C3N4-graphene composite for the detection of neuron-specific enolase as a biomarker for lung cancer.

Lung cancer remains the leading cause of cancer mortality worldwide. Small cell lung cancer (SCLC) accounts for 10-15% of cases and has an overall 5-years survival rate of only 15%. Neuron-specific enolase (NSE) has been identified as a useful biomarker for early SCLC diagnosis and therapeutic monitoring. This work reports an electrochemical immunosensing platform based on a graphene-graphitic carbon nitride (g-C3N4) nanocomposite for ultrasensitive NSE detection. The g-C3N4 nanosheets and graphene nanosheets were synthesized via liquid exfoliation and integrated through self-assembly to form the nanocomposite. This nanocomposite was used to modify screen-printed carbon electrodes followed by covalent immobilization of anti-NSE antibodies. The unique properties of the graphene-g-C3N4 composite facilitated efficient antibody loading while also enhancing electron transfer efficiency and electrochemical response. Systematic optimization of experimental parameters was performed. The immunosensor exhibited a wide linear detection range of 10 pg/mL to 100 ng/mL and low limit of detection of 3 pg/mL for NSE along with excellent selectivity against interferences. Real serum matrix analysis validated the applicability of the developed platform for sensitive and accurate NSE quantifica-tion at clinically relevant levels. This novel graphene-g-C3N4 nanocomposite based electro-chemical immunoassay demonstrates great promise for early diagnosis of SCLC.

Ultrasensitive detection of NSE employing a novel electrochemical immunosensor based on a conjugated copolymer.

In the current study a simple and highly specific label-free impedimetric neuron specific enolase (NSE) immunosensor based on a copolymer matrix-coated disposable electrode was designed and tested. The copolymer matrix was prepared using a very conductive EDOT monomer and semi-conductive thiophene-bearing epoxy groups (ThEp), and the combination of the two monomers enhanced the conductivity and protein loading capacity of the electrode surface. The P(ThEp-co-EDOT) copolymer matrix was prepared via a drop-casting process and anti-NSE recognition biomolecules were immobilized directly on the epoxy groups of the copolymer. After the coupling of NSE molecules on the P(ThEp-co-EDOT) copolymer matrix-coated electrode surface, the charge transfer resistance (Rct) of the biosensor changed dramatically. These changes in Rct were proportional to the NSE molecule amounts captured by anti-NSE molecules. Under optimized experimental conditions, the increment in the Rct value was proportional to the NSE concentration over a range of 0.01 to 25 pg mL-1 with a detection limit (LOD) of 2.98 × 10-3 pg mL-1. This copolymer-coated electrode provided a lower LOD than the other biosensors. In addition, the suggested electrochemical immuno-platform showed good selectivity, superior reproducibility, long-term stability, and high recovery of NSE in real serum (95.64-102.20%) and saliva (95.28-105.35%) samples. These results showed that the present system had great potential for electrochemical biosensing applications.

Lower versus higher blood pressure targets in comatose patients resuscitated from out-of-hospital cardiac arrest-Protocol for a secondary Bayesian analysis of the box trial.

BACKGROUND: The management of blood pressure targets during intensive care after out-of-hospital cardiac arrest (OHCA) remains a topic of debate. The blood Pressure and Oxygenation Targets After OHCA (BOX) trial explored the efficacy of two different blood pressure targets in 789 patients during intensive care after OHCA. In the primary frequentist analysis, no statistically significant differences were found for neurological outcome after 90 days. METHODS: This protocol outlines secondary Bayesian analyses of 365-day all-cause mortality and two secondary outcomes: neurological outcome after 365 days, and plasma neuron-specific enolase, a biomarker of brain injury, after 48 h. We will employ adjusted Bayesian logistic and linear regressions, presenting results as relative and absolute differences with 95% confidence intervals. We will use weakly informative priors for the primary analyses, and skeptical and evidence-based priors (where available) in sensitivity analyses. Exact probabilities for any benefit/harm will be presented for all outcomes, along with probabilities of clinically important benefit/harm (risk differences larger than 2%-points absolute) and no clinically important differences for the binary outcomes. We will assess whether heterogeneity of treatment effects on mortality is present according to lactate at admission, time to return of spontaneous circulation, primary shockable rhythm, age, hypertension, and presence of ST-elevation myocardial infarction. DISCUSSION: This secondary analysis of the BOX trial aim to complement the primary frequentist analysis by quantifying the probabilities of beneficial or harmful effects of different blood pressure targets. This approach seeks to provide clearer insights for researchers and clinicians into the effectiveness of these blood pressure management strategies in acute medical conditions, particularly focusing on mortality, neurological outcomes, and neuron-specific enolase.

Interrogating l-fuconate dehydratase with tartronate and 3-hydroxypyruvate reveals subtle differences within the mandelate racemase-subgroup of the enolase superfamily.

Enzymes of the enolase superfamily share a conserved structure and a common partial reaction (i.e., metal-assisted, Brønsted base-catalyzed enol(ate) formation). The architectures of the enolization apparatus at the active sites of the mandelate racemase (MR)-subgroup members MR and l-fuconate dehydratase (FucD) are almost indistinguishable at the structural level. Tartronate and 3-hydroxypyruvate (3-HP) recognize the enolization apparatus and can be used to interrogate the active sites for differences that may not be apparent from structural data. We report a circular dichroism-based assay of FucD activity that monitors the change in ellipticity at 216 nm (Δ[Θ]S-P = 8985 ± 87 deg cm2 mol-1) accompanying the conversion of l-fuconate to 2-keto-3-deoxy-l-fuconate. Tartronate was a linear mixed-type inhibitor of FucD (Ki = 8.4 ± 0.7 mM, αKi = 63 ± 11 mM), binding 18-fold weaker than l-fuconate, compared with 2-fold weaker binding of tartronate by MR relative to mandelate. 3-HP irreversibly inactivated FucD (kinact/KI = 0.018 ± 0.002 M-1s-1) with an efficiency that was ∼4.6 × 103-fold less than that observed with MR. The inactivation arose predominantly from modifications at multiple sites and Tris-HCl, but not l-fuconate, afforded protection against inactivation. Similar to the reaction of 3-HP with MR, 3-HP modified the Brønsted base catalyst (Lys 220) at the active site of FucD, which was facilitated by the Brønsted acid catalyst His 351. Thus, the interactions of tartronate and 3-HP with MR and FucD revealed differences in binding affinity and reactivity that differentiated between the enzymes' enolization apparatuses.

Enolase of Streptococcus suis serotype 2 promotes biomolecular condensation of ribosomal protein SA for HBMECs apoptosis.

BACKGROUND: Ribosomal protein SA (RPSA) of human brain microvascular endothelial cells (HBMECs) can transfer from the cytosol to the cell surface and act as a receptor for some pathogens, including Streptococcus suis serotype 2 (SS2), a zoonotic pathogen causing meningitis in pigs and humans. We previously reported that SS2 virulence factor enolase (ENO) binds to RPSA on the cell surface of HBMECs and induces apoptosis. However, the mechanism that activates RPSA translocation to the cell surface and induces ENO-mediated HBMEC apoptosis is unclear. RESULTS: Here, we show that RPSA localization and condensation on the host cell surface depend on its internally disordered region (IDR). ENO binds to the IDR of RPSA and promotes its interaction with RPSA and vimentin (VIM), which is significantly suppressed after 1,6-Hexanediol (1,6-Hex, a widely used tool to disrupt phase separation) treatment, indicating that ENO incorporation and thus the concentration of RPSA/VIM complexes via co-condensation. Furthermore, increasing intracellular calcium ions (Ca2+) in response to SS2 infection further facilitates the liquid-like condensation of RPSA and aggravates ENO-induced HBMEC cell apoptosis. CONCLUSIONS: Together, our study provides a previously underappreciated molecular mechanism illuminating that ENO-induced RPSA condensation activates the migration of RPSA to the bacterial cell surface and stimulates SS2-infected HBMEC death and, potentially, disease progression. This study offers a fresh avenue for investigation into the mechanism by which other harmful bacteria infect hosts via cell surfaces' RPSA.

Neuron-specific enolase at admission as a predictor for stroke volume, severity and outcome in ischemic stroke patients: a prognostic biomarker review.

An ideal blood biomarker for stroke should provide reliable results, enable fast diagnosis, and be readily accessible for practical use. Neuron-specific enolase (NSE), an enzyme released after neuronal damage, has been studied as a marker for brain injury, including cerebral infarction. However, different methodologies and limited sample sizes have restricted the applicability of any potential findings. This work aims to determine whether NSE levels at Emergency Department (ED) admission correlate with stroke severity, infarcted brain volume, functional outcome, and/or death rates. A systematic literature review was performed using PubMed, Embase, and Scopus databases. Each reviewer independently assessed all published studies identified as potentially relevant. All relevant original observational studies (cohort, case-control, and cross-sectional studies) were included. Eleven studies (1398 patients) met the inclusion criteria. Among these, six studies reported a significant correlation between NSE levels and stroke severity, while only one found no association. Four studies indicated a positive relationship between infarcted brain volume assessed by imaging and NSE levels, in contrast to the findings of only one study. Four studies identified an association related to functional outcome and death rates, while three others did not reach statistical significance in their findings. These data highlight that NSE levels at ED admissions proved to be a promising tool for predicting the outcome of ischemic stroke patients in most studies. However, they presented high discrepancies and low robustness. Therefore, further research is necessary to establish and define the role of NSE in clinical practice.

USP21 deubiquitinates and stabilizes HSP90 and ENO1 to promote aerobic glycolysis and proliferation in cholangiocarcinoma.

Deubiquitylating enzymes (DUBs) play an essential role in targeted protein degradation and represent an emerging therapeutic paradigm in cancer. However, their therapeutic potential in cholangiocarcinoma (CCA) has not been explored. Herein, based on The Cancer Genome Atlas (TCGA) and The Gene Expression Omnibus (GEO) databases, we found that ubiquitin-specific protease 21 (USP21) was upregulated in CCA, high USP21 level was associated with poor prognosis. In vivo and in vitro, we identified USP21 as a master regulator of CCA growth and maintenance, which directly interacted with deubiquitinates and stabilized the heat shock protein 90 (HSP90) through K48-linked deubiquitination, and in turn, this stabilization increased HIF1A expression, thus upregulating key glycolytic enzyme genes ENO2, ENO3, ALDOC, ACSS2, and then promoted aerobic glycolysis, which provided energy for CCA cell proliferation. In addition, USP21 could directly stabilize alpha-Enolase 1 (ENO1) to promote aerobic glycolysis. Furthermore, increased USP21 level enhanced chemotherapy resistance to the gemcitabine-based regimen. Taken together, we identify a USP21-regulated aerobic glycolysis mechanism that involves the USP21/HSP90/HIF1A axis and USP21/ENO1 axis in CCA tumorigenesis, which could serve as a potential target for the treatment of CCA.

Correlation between serum galanin and neuron-specific enolase levels with EEG abnormalities in pediatric convulsive status epilepticus and the efficacy of triple drug therapy.

OBJECTIVE: This study aimed to investigate the association between serum galanin (GAL) and neuron-specific enolase (NSE) levels in children with convulsive status epilepticus (CSE) and their relationship with abnormal electroencephalogram (EEG) patterns. Additionally, the study assessed the effectiveness of a combination therapy involving midazolam, diazepam, and phenobarbital in treating CSE. PATIENTS AND METHODS: The research involved 100 children diagnosed with CSE and included a control group of 50 healthy children. Serum GAL and NSE levels were measured, and EEGs were analyzed for abnormalities in the CSE group. Comparisons were made between the healthy control group and the CSE group, particularly within the first 24 hours after persistent seizures. The severity of EEG abnormalities was correlated with GAL and NSE levels. The treatment consisted of an observation group that received the triple therapy of midazolam, diazepam, and phenobarbital, while a control group received diazepam and phenobarbital. Clinical efficacy, symptom improvement, Status Epilepticus Severity Score (STESS), and adverse reactions were evaluated. RESULTS: The results indicated elevated levels of GAL and NSE in the CSE group, with higher levels noted within 24 hours after persistent seizures. Furthermore, a positive correlation was observed between the severity of EEG abnormalities and GAL and NSE levels. The group receiving the triple therapy demonstrated superior efficacy, faster resolution of seizures and fever, reduced STESS scores, and fewer adverse reactions than the control group. In conclusion, this study highlights the positive correlation between serum GAL and NSE levels and the severity of EEG abnormalities in pediatric CSE. The triple therapy approach is effective in treating CSE, leading to improved clinical symptoms, reduced brain damage, and enhanced safety. CONCLUSIONS: The study concludes that serum GAL and NSE levels in children with convulsive status epilepticus are positively correlated with the degree of EEG abnormalities. The combination therapy involving midazolam, diazepam, and phenobarbital is effective in treating children with convulsive status epilepticus, significantly improving clinical symptoms, reducing brain damage, and ensuring safety.

Aberrant brain-heart coupling is associated with the severity of post cardiac arrest brain injury.

OBJECTIVE: To investigate autonomic nervous system activity measured by brain-heart interactions in comatose patients after cardiac arrest in relation to the severity and prognosis of hypoxic-ischemic brain injury. METHODS: Strength and complexity of bidirectional interactions between EEG frequency bands (delta, theta, and alpha) and ECG heart rate variability frequency bands (low frequency, LF and high frequency, HF) were computed using a synthetic data generation model. Primary outcome was the severity of brain injury, assessed by (i) standardized qualitative EEG classification, (ii) somatosensory evoked potentials (N20), and (iii) neuron-specific enolase levels. Secondary outcome was the 3-month neurological status, assessed by the Cerebral Performance Category score [good (1-2) vs. poor outcome (3-4-5)]. RESULTS: Between January 2007 and July 2021, 181 patients were admitted to ICU for a resuscitated cardiac arrest. Poor neurological outcome was observed in 134 patients (74%). Qualitative EEG patterns suggesting high severity were associated with decreased LF/HF. Severity of EEG changes were proportional to higher absolute values of brain-to-heart coupling strength (p < 0.02 for all brain-to-heart frequencies) and lower values of alpha-to-HF complexity (p = 0.049). Brain-to-heart coupling strength was significantly higher in patients with bilateral absent N20 and correlated with neuron-specific enolase levels at Day 3. This aberrant brain-to-heart coupling (increased strength and decreased complexity) was also associated with 3-month poor neurological outcome. INTERPRETATION: Our results suggest that autonomic dysfunctions may well represent hypoxic-ischemic brain injury post cardiac arrest pathophysiology. These results open avenues for integrative monitoring of autonomic functioning in critical care patients.

Improvement of differential diagnosis of lung cancer by use of multiple protein tumor marker combinations.

BACKGROUND: Differential diagnosis of non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) in hospitalized patients is crucial for appropriate treatment choice. OBJECTIVE: To investigate the relevance of serum tumor markers (STMs) and their combinations for the differentiation of NSCLC and SCLC subtypes. METHODS: Between 2000 and 2003, 10 established STMs were assessed retrospectively in 311 patients with NSCLC, 128 with SCLC prior systemic first-line therapy and 51 controls with benign lung diseases (BLD), by automatized electrochemiluminescence immunoassay technology. Receiver operating characteristic (ROC) curves and logistic regression analyses were used to evaluate the diagnostic efficacy of both individual and multiple STMs with corresponding sensitivities at 90% specificity. Standards for Reporting of Diagnostic Accuracy (STARD guidelines) were followed. RESULTS: CYFRA 21-1 (cytokeratin-19 fragment), CEA (carcinoembryonic antigen) and NSE (neuron specific enolase) were significantly higher in all lung cancers vs BLD, reaching AUCs of 0.81 (95% CI 0.76-0.87), 0.78 (0.73-0.84), and 0.88 (0.84-0.93), respectively. By the three marker combination, the discrimination between benign and all malignant cases was improved resulting in an AUC of 0.93 (95% CI 0.90-0.96). In NSCLC vs. BLD, CYFRA 21-1, CEA and NSE were best discriminative STMs, with AUCs of 0.86 (95% CI 0.81-0.91), 0.80 (0.74-0.85), and 0.85 (0.79-0.91). The three marker combination also improved the AUC: 0.92; 95% CI 0.89-0.96). In SCLC vs. BLD, ProGRP (pro-gastrin-releasing peptide) and NSE were best discriminative STMs, with AUCs of 0.89 (95% CI 0.84-0.94) and 0.96 (0.93-0.98), respectively, and slightly improved AUC of 0.97 (95% CI 0.95-0.99) when in combination. Finally, discrimination between SCLC and NSCLC was possible by ProGRP (AUC 0.86; 95% CI 0.81-0.91), NSE (AUC 0.83; 0.78-0.88) and CYFRA 21-1 (AUC 0.69; 0.64-0.75) and by the combination of the 3 STMs (AUC 0.93; 0.91-0.96), with a sensitivity of 88% at 90% specificity. CONCLUSIONS: The results confirm the power of STM combinations for the differential diagnosis of lung cancer from benign lesions and between histological lung cancer subtypes.

Murine Traumatic Brain Injury Model Comparison: Closed Head Injury Versus Controlled Cortical Impact.

INTRODUCTION: Various murine models have been utilized to study TBI, including closed head injury (CHI) and controlled cortical impact (CCI), without direct comparison. The aim of our study was to evaluate these models to determine differences in neurological and behavioral outcomes postinjury. METHODS: Male C57B/6 mice (9-10 wk) were separated into six groups including: untouched, sham craniotomy (4 mm), CCI 0.9 mm depth of impact, CCI 1.6 mm, CCI 2.2 mm, and CHI. CCI was performed using a 3 mm impact tip at a velocity of 5 m/s, dwell time of 250 ms, and depth as noted above. CHI was completed with a centered 400 g weight drop from 1 cm height. Mice were survived to 14-d (n = 5 per group) and 30-d (n = 5 per group) respectively for histological analysis of p-tau within the hippocampus. These mice underwent Morris Water Maze memory testing and Rotarod motor testing. Serum was collected from a separate cohort of mice (n = 5 per group) including untouched, isoflurane only, CCI 1.6 mm, CHI at 1, 4, 6, and 24 h for analysis of neuron specific enolase and glial fibrillary acidic protein (GFAP) via ELISA. Laser speckle contrast imaging was analyzed prior to and after impact in the CHI and CCI 1.6 mm groups. RESULTS: There were no significant differences in Morris Water Maze or Rotarod testing times between groups at 14- or 30-d. P-tau was significantly elevated in all groups except CCI 1.6 mm contralateral and CCI 2.2 mm ipsilateral compared to untouched mice at 30-d. P-tau was also significantly elevated in the CHI group at 30 d compared to CCI 1.6 mm contralateral and CCI 2.2 mm on both sides. GFAP was significantly increased in mice undergoing CHI (9959 ± 91 pg/mL) compared to CCI (2299 ± 1288 pg/mL), isoflurane only (133 ± 75 pg/mL), and sham (86 ± 58 pg/mL) at 1-h post TBI (P < 0.0001). There were no differences in serum neuron specific enolase levels between groups. Laser doppler imaging demonstrated similar decreases in cerebral blood flow between CHI and CCI; however, CCI mice had a reduction in blood flow with craniotomy only that did not significantly decrease further with impact. CONCLUSIONS: Based on our findings, CHI leads to increased serum GFAP levels and increased p-tau within the hippocampus at 30-d postinjury. While CCI allows the comparison of one cerebral hemisphere to the other, CHI may be a better model of TBI as it requires less technical expertise and has similar neurological outcomes in these murine models.