A retrospective study of 3D laparoscopy and 2D laparoscopy in transabdominal preperitoneal (TAPP) for unilateral inguinal hernia in elderly patients.

This study aims to explore the safety and efficacy of 3D laparoscopy in elderly patients undergoing transabdominal preperitoneal (TAPP) surgery for inguinal hernia. Patients were divided into two groups based on the laparoscopic equipment used during surgery. Clinical data preoperatively, intraoperatively, and postoperatively were collected and subjected to statistical analysis. From January 2020 to August 2023, a total of 127 patients with primary unilateral inguinal hernia were evaluated in this study, 61 in the 3D TAPP group and 66 in the 2D TAPP group. There were no statistically significant differences in baseline data, including average age, gender distribution, BMI index, hernia type, hernia defect size and location, comorbidities, and usage of anticoagulant drugs between the two groups (P > 0.05). In terms of operative indicators, the 3D group showed shorter mean operation time (51.61 ± 7.16 min vs. 78.59 ± 13.51 min, P < 0.001), mean mesh placement time (6.07 ± 1.40 min vs. 9.77 ± 1.21 min, P < 0.001), and mean peritoneal suture time (7.34 ± 1.85 min vs. 9.73 ± 1.32 min, P < 0.001) compared to the 2D group. However, there were no statistically significant differences in mean blood loss, postoperative pain scores, postoperative hospital stay, and total hospital costs between the two groups (P > 0.05). The incidence of postoperative complications did not differ significantly between the two groups (P > 0.05). No adverse reactions such as dizziness or nausea were reported by surgeons during the procedures in either group. Three-dimensional laparoscopy in TAPP surgery provides high-definition, three-dimensional surgical images, reducing the difficulty of operations and effectively shortening the operation time.

Integrated plasma proteomics and N-glycoproteomics reveals alterations in the N-glycosylation in Chinese hepatocellular carcinoma patients.

Hepatocellular carcinoma (HCC) is a life-threatening disease that presents diagnostic challenges due to the absence of reliable biomarkers. Recently, plasma proteomics and glycoproteomics have emerged as powerful tools for identifying potential diagnostic biomarkers for various diseases. In this study, we conducted a comprehensive proteomic and glycoproteomic analysis of plasma samples from 11 HCC patients and 11 healthy control (HC) individuals. We identified 20 differentially expressed (DE) proteins and 32 DE intact glycosylated peptides (IGPs) that can effectively differentiate between HCC patients and HC samples. Our findings demonstrate that IGP profiles had better predictive power than protein profiles for screening HCC. Pathways associated with DE proteins and IGPs were identified. It was reported that the protein expression level of galectin 3 binding protein (LGALS3BP) and its N-linked glycosylation at the N398 and N551 sites might serve as valuable candidates for HCC diagnosis. These results highlight the importance of N-glycoproteomics in advancing our understanding of HCC and suggest possible candidates for the future diagnosis of this disease.

CD206+ M2-like macrophages protect against intervertebral disc degeneration partially by targeting R-spondin-2.

OBJECTIVE: This study aimed to explore the specific function of M2 macrophages in intervertebral disc degeneration (IDD). METHODS: Intervertebral disc (IVD) samples from normal (n = 4) and IDD (n = 6) patients were collected, and the expression of M2-polarized macrophage marker, CD206, was investigated using immunohistochemical staining. Nucleus pulposus cells (NPCs) in a TNF-α environment were obtained, and a mouse caudal IVD puncture model was established. Mice with Rheb deletions, specifically in the myeloid lineage, were generated and subjected to surgery-induced IDD. IDD-induced damage and cell apoptosis were measured using histological scoring, X-ray imaging, immunohistochemical staining, and TdT-mediated dUTP nick end labeling (TUNEL) assay. Finally, mice and NPCs were treated with R-spondin-2 (Rspo2) or anti-Rspo2 to investigate the role of Rspo2 in IDD. RESULTS: Accumulation of CD206 in human and mouse IDD tissues was detected. Rheb deletion in the myeloid lineage (RheBcKO) increased the number of CD206+ M2-like macrophages (mean difference 18.6% [15.7-21.6%], P < 0.001), decreased cell apoptosis (mean difference -15.6% [-8.9 to 22.2%], P = 0.001) and attenuated the IDD process in the mouse IDD model. NPCs treated with Rspo2 displayed increased extracellular matrix catabolism and apoptosis; co-culture with a conditioned medium derived from RheBcKO mice inhibited these changes. Anti-Rspo2 treatment in the mouse caudal IVD puncture model exerted protective effects against IDD. CONCLUSIONS: Promoting CD206+ M2-like macrophages could reduce Rspo2 secretion, thereby alleviating experimental IDD. Rheb deletion may help M2-polarized macrophages accumulate and attenuate experimental IDD partially by inhibiting Rspo2 production. Hence, M2-polarized macrophages and Rspo2 may serve as therapeutic targets for IDD.

Development of cancer biomarker heat shock protein 90α certified reference material using two different isotope dilution mass spectrometry techniques.

Heat shock protein 90α (HSP90α) has been regarded as an important indicator for judging tumor metastasis and prognosis due to its significant upregulation in various tumors. Therefore, the accurate quantification of HSP90α is of great significance for clinical diagnosis and therapy of cancers. However, the lack of HSP90α certified reference material (CRM) leads to the accuracy and consistency of quantification methods not being effectively evaluated. Besides, quantitative results without traceability make comparisons between different studies difficult. In this study, an HSP90α solution CRM was developed from the recombinant protein raw material. The recombinant protein is a dimer, and the purity of the CRM candidate reached 96.71%. Both amino acid analysis-isotope dilution mass spectrometry (AAA-IDMS) and unique peptide analysis-isotope dilution mass spectrometry (UPA-IDMS) were performed to measure the content of HSP90α in the solution CRM candidate, and the certified value was assessed to be 66.2 ± 8.8 µg/g. Good homogeneity of the CRM was identified, and the stability examination suggested that the CRM was stable for at least 4 months at - 80 °C and for 7 days at 4 °C. With traceability to SI unit (kg), this CRM has potential to help establish a metrological traceability chain for quantification of HSP90α, which will make the quantification results standardized and comparable regardless of the quantitative methods.

Comparative analysis of laparoscopic choledocholithiasis and ERCP treatment after cholecystectomy.

OBJECTIVE: To compare the overall efficacy of laparoscopic common bile duct exploration(LCBDE) with endoscopic retrograde cholangiopancreatography (ERCP ) after cholecystectomy. METHODS: From January 2017 to July 2021, Seventy patients with Choledocholithiasis after cholecystectomy who were admitted to our hospital were selected and divided into ERCP and LCBDE groups. comparison of baseline characteristics, clinical efficacy and postoperative complications between the ERCP and LCBDE. RESULTS: ①The overall efficacy rate of LCBDE was 97.1%, while the overall efficacy rate in the ERCP group was 76.6%. The LCBDE group demonstrated a significantly higher overall effective rate compared to the ERCP group, with a statistically significant difference (p < 0.05). ②The preoperative and postoperative complications of the LCBDE group were visibly lower than the other group (P < 0.05). The postoperative time to oral intake, postoperative ventilation time, length of hospital stay, and hospital costs were higher in the ERCP group compared to the LCBDE group, with a statistically significant difference (P < 0.05). CONCLUSION: In the treatment of common bile duct stones after cholecystectomy, LCBDE is a superior choice compared to ERCP in terms of stone diameter, quantity, clearance rate, and hospital costs.

Nanoliter atmospheric pressure photoionization-mass spectrometry for direct bioanalysis of polycyclic aromatic hydrocarbons.

Polycyclic aromatic hydrocarbons (PAHs) are a class of low-polarity environmental contaminants that have severe carcinogenic effects and have drawn worldwide attention. However, there remain challenges for current mass spectrometric ionization techniques in the analysis of low-polarity compounds in small-volume biosamples, such as single cells. In this work, we developed a nanoliter atmospheric pressure photoionization (nano-APPI) source and optimized its parameters for the detection of PAHs in small-volume samples. We evaluated the ionization performance of the source in direct and auxiliary gas-assisted photoionization modes and analyzed different PAH compounds as well as spiked biosamples. By combining the advantages of nano-electrospray ionization (nano-ESI) and atmospheric pressure photoionization (APPI), our newly developed nano-APPI source achieved high sensitivity for the analysis of PAHs down to the fmol level. Compared to conventional atmospheric pressure chemical ionization (APCI), the detection limit of PAHs was increased by 1-2 orders of magnitude. By optimizing various parameters, we achieved highly efficient ionization of PAHs, effective analysis of PAHs in mixed components, and sensitive detection of low-abundance PAHs in single-cell samples. Our optimized nano-APPI source was successfully applied for the sensitive analysis of PAHs in complex biological samples. Based on our study, we believe that nano-APPI holds great promise for toxicological studies on complex biological samples. The present work has implications for improving the detection sensitivity of low-polarity environmental contaminants and advancing the field of MS-based analysis of small-volume biosamples.

Evolution of Mass Spectrometry Instruments and Techniques for Blood Proteomics.

Mass spectrometry (MS)-based blood proteomics is a crucial research focus in identifying disease biomarkers. Blood serum or plasma is the most commonly used sample for such analysis; however, it presents challenges due to the complexity and dynamic range of protein abundance. Despite these difficulties, the development of high-resolution MS instruments has made comprehensive investigation of blood proteomics possible. The evolution of time-of-flight (TOF) or Orbitrap MS instruments has played a significant role in the field of blood proteomics. These instruments are now among the most prominent techniques for blood proteomics due to their sensitivity, selectivity, fast response, and stability. For optimal results, it is necessary to eliminate high-abundance proteins from the blood sample, to maximize the depth coverage of the blood proteomics analysis. This can be achieved through various methods, including commercial kits, chemically synthesized materials, and MS technologies. This paper reviews recent advancements in MS technology and its remarkable applications in biomarker discovery, particularly in the areas of cancer and COVID-19 studies.

CK2α causes stemness and chemotherapy resistance in liver cancer through the Hedgehog signaling pathway.

BACKGROUND: Liver cancer is one of the major causes of cancer-related deaths globally. Cancer cell stemness and chemotherapy resistance contribute to the high mortality. Although evidence indicates that the alpha subunit of protein kinase 2 (CK2α) is involved in several human cancers, its function in liver cancer remains unknown. In the present study, we aimed to elucidate the role of CK2α in liver cancer. METHODS: We examined the role of CK2α regulation in stemness and chemotherapy resistance capacity of liver cancer cells. MTT assays, tumor sphere formation assays, RT-PCR, flow cytometry, Western blotting assay, clonogenicity assay, matrigel invasion assay and bioinformatics were conducted in this study. RESULTS: CK2α expression in the liver cancer tissues was notably upregulated compared with that in the corresponding non-tumorous tissues. The overexpression of CK2α promoted tumor sphere formation, increased the percentage of CD133(+) and side population cells, caused the resistance of liver cancer cells to 5-FU treatment, increased the expression levels of NANOG, OCT4, SOX2, Gli1 and Ptch1, and enhanced the ability of CD133(+) cell clone formation and invasion. Consistently, the downregulation of CK2α had the opposite effects. CK2α silencing inhibited the Hedgehog pathway by reducing the expression of Gli1 and Ptch1. Mechanistically, CK2α regulation on liver cancer cell stemness and chemotherapy resistance was found to be involved in the Hedgehog signaling pathway. CONCLUSIONS: Our study may bring some new insights into the occurrence of liver cancer. Furthermore, these findings suggest that targeting CK2α may be a novel therapeutic strategy for patients with liver cancer.

Lateral Flow Immunoassay Based on Time-Resolved Fluorescence Microspheres for Rapid and Quantitative Screening CA199 in Human Serum.

Carbohydrate antigen 199 (CA199) is a serum biomarker which has certain value and significance in the diagnosis, prognosis, treatment, and postoperative monitoring of cancer. In this study, a lateral flow immunoassay based on europium (III) polystyrene time-resolved fluorescence microspheres (TRFM-based LFIA), integrated with a portable fluorescence reader, has been successfully establish for rapid and quantitative analysis of CA199 in human serum. Briefly, time-resolved fluorescence microspheres (TRFMs) were conjugated with antibody I (Ab1) against CA199 as detection probes, and antibody II (Ab2) was coated as capture element, and a "TRFMs-Ab1-CA199-Ab2" sandwich format would form when CA199 was detected by the TRFM-based LFIA. Under the optimal parameters, the detection limit of the TRFM-based LFIA for visible quantitation with the help of an ultraviolet light was 4.125 U/mL, which was four times lower than that of LFIA based on gold nanoparticles. Additionally, the fluorescence ratio is well linearly correlated with the CA199 concentration (0.00-66.0 U/mL) and logarithmic concentration (66.0-264.0 U/mL) for quantitative detection. Serum samples from 10 healthy people and 10 liver cancer patients were tested to confirm the performances of the point-of-care application of the TRFM-based LFIA, 20.0 U/mL of CA199 in human serum was defined as the threshold for distinguishing healthy people from liver cancer patients with an accuracy of about 60%. The establishment of TRFM-based LFIA will provide a sensitive, convenient, and efficient technical support for rapid screening of CA199 in cancer diagnosis and prognosis.

Primary closure after laparoscopic common bile duct exploration is safe and feasible for patients with non-severe acute cholangitis.

BACKGROUND: The safety and feasibility of primary closure after laparoscopic common bile duct exploration (LCBDE) have been confirmed in elective settings. However, the suitability of primary closure after LCBDE in the treatment of patients with non-severe acute cholangitis in emergency settings remains unclear. The aim of the present study was to explore the safety and feasibility of LCBDE with primary closure in patients with non-severe acute cholangitis. METHODS: Consecutive patients with choledocholithiasis combined with gallbladder stones treated by LCBDE with primary closure at our institution from January 2015 to April 2021 were retrospectively reviewed. These patients were divided into two groups: emergency group (patients with non-severe acute cholangitis) and elective group (patients without acute cholangitis). The demographic and perioperative data of the two groups were compared. RESULTS: One hundred twenty-two patients received LCBDE combined with primary closure during this period, including 70 in the emergency group and 52 in the elective group. Baseline characteristics were balanced in both groups, except for higher levels of white blood cells (WBC), C-reactive protein (CRP), total bilirubin, alkaline phosphatase (ALP), and albumin in the emergency group. No postoperative mortality occurred in either group. Compared to the elective group, the emergency group had a longer operation time (P = 0.011), and more estimated blood loss (P < 0.001). No significant differences were found between the two groups in terms of conversion (2.9% vs. 0.0%, P = 0.507), use of baskets (84.2% vs. 78.8%, P = 0.481), use of electrohydraulic lithotripsy (EHL) (2.9% vs. 1.9%, P = 1.000), or postoperative hospital stay (P = 0.214). The incidence of postoperative complications was comparable between the two groups. During the follow-up period, none of the patients experienced biliary stricture, and 1 case of stone recurrence occurred in the elective group. CONCLUSIONS: LCBDE with primary closure for choledocholithiasis patients with non-severe acute cholangitis has the equivalent efficacy and morbidity to elective surgery. Primary closure after LCBDE is a safe and feasible option for choledocholithiasis patients with non-severe acute cholangitis.

Analysis of low-abundance molecules in complex matrices by quadrupole-linear ion trap mass spectrometry using a simultaneous fragmentation and accumulation strategy.

RATIONALE: Fast and sensitive analysis of low-abundance molecules in complex matrices has always been a challenge in chemical and biological applications. Mass spectrometry (MS) has been widely used in the fields of chemical and biological analysis due to its unparalleled specificity and sensitivity. However, the MS signals consistently deteriorate in the presence of matrices. Demands for more sensitive and efficient methods to analyze those low-abundance molecules in chemical and biological systems are in urgent need. METHODS: Based on a home-made quadrupole-linear ion trap (Q-LIT) mass spectrometer, a simultaneous fragmentation and accumulation strategy was developed to improve the sensitivity of the analysis for the low-abundance molecules in complex matrices. Ions were filtered by the quadrupole into the LIT. The precursor ions were fragmented and the product ions were isolated and accumulated in the LIT simultaneously. The fragmentation, isolation and accumulation processes were conducted at the same time. The accumulation time could be controlled to accumulate sufficient product ions. RESULTS: With this strategy, the signal intensity of targeted molecules could be increased by 2-8 times and by increasing the accumulation time, this could be further enhanced. Those interferences induced by isomers and matrices can be reduced by using our method. We further applied our method to the quantification and analysis of biological samples. Tryptic digested peptides of myoglobin (Mb) were successfully detected by our method. CONCLUSIONS: We have established a new method with great advantages in the detection of molecules in complex matrices. The application of this method promises better results in the bioanalytical area, especially for the analysis of substances in complex matrices in the future.

Primary closure versus T-tube drainage after laparoscopic common bile duct exploration in patients with non-severe acute cholangitis.

Although the feasibility of T-tube drainage after emergency laparoscopic common bile duct exploration (LCBDE) has been reported, the safety and effectiveness of primary closure (PC) after LCBDE in patients with non-severe acute cholangitis (AC) remain uncertain. This study aimed to investigate the safety and feasibility of PC after LCBDE in patients with non-severe AC. Consecutive choledocholithiasis patients with non-severe AC who were treated with a laparoscopic approach at our institution between January 2014 and March 2021 were enrolled. These patients were divided into two groups (PC group and T-tube group) based on the way of closure of the common bile duct. The baseline characteristics and perioperative data between the two groups were compared. A total of 230 patients who underwent LCBDE met the inclusion criteria, and there were 94 patients in the PC group and 126 patients in the T-tube group. Baseline data were balanced between the two groups, except that there was less acute cholecystitis in the PC group than in the T-tube group (P = 0.027). Compared to the T-tube group, the PC group had a shorter operation time (P < 0.001), less estimated blood loss (P < 0.001), less use of electrohydraulic lithotripsy (EHL) (P = 0.001), shorter time of drainage removal (P < 0.001) and postoperative hospital stay (P < 0.001) and residual stones (P = 0.029). There was no significant difference between the two groups in terms of conversion (4.3 vs. 4.4%, P = 1.000), intraoperative transfusion (0.0 vs. 0.7%, P = 1.000), use of basket (71.2 vs. 69.9%, P = 0.816), postoperative bleeding (1.1 vs. 0.7%, P = 1.000), biliary leakage (4.3 vs. 3.7%, P = 1.000), incision infection (1.1 vs. 2.2%, P = 0.649), pneumonia (2.1 vs. 1.4%, P = 1.000), or cholangitis (1.1 vs. 2.9%, P = 0.651). No postoperative mortality occurred in either group. During the follow-up period, no biliary stricture occurred in the two groups, and two patients in the T-tube group were found to have stone recurrence. PC after LCBDE in choledocholithiasis patients with non-severe AC shows superior clinical outcomes to T-tube drainage in terms of the operation time, estimated blood loss, time of drainage removal, postoperative hospital stay, and residual stones. PC is a safe and feasible treatment for choledocholithiasis patients with non-severe AC after LCBDE.

Direct analysis of hydroxylated polycyclic aromatic hydrocarbons in biological samples with complex matrices using polarity-reversed nanoelectrospray ionization.

RATIONALE: Polycyclic aromatic hydrocarbons (PAHs) are a class of environmental contaminants with carcinogenic effect drawing worldwide attention. PAHs can be converted into hydroxylated PAHs (OH-PAHs) through metabolic processes. Thus, they are commonly considered as an important class of biomarkers of PAH exposure. However, direct analysis of related metabolites of these environmental pollutants in biological samples using mass spectrometry is still challenging because of matrix effect and ion suppression during nanoelectrospray ionization (nano-ESI). METHODS: In our previous work, a polarity-reversed nanoelectrospray ionization (PR-nESI) technique was developed for the analysis of biomolecules in complex matrices. In this work, we further optimized PR-nESI for direct and sensitive analysis of OH-PAHs in different samples under severe salt interference in negative polarity. RESULTS: Compared with conventional nano-ESI, the optimized PR-nESI method realized sensitive detection of 1-naphthol in samples with a concentration of salt up to millimolar level. The signal-to-noise ratio (S/N) of OH-PAHs was increased by 1-2 orders of magnitude compared with conventional nano-ESI. Six different OH-PAHs were successfully detected with high S/N ratio using PR-nESI. PR-nESI was further successfully applied in the analysis of OH-PAHs in spiked fetal blood serum, human urine, and single-cell samples. For environmentally exposed subjects, the detections of OH-PAHs in single-cell samples and urines from human smokers were successfully conducted. CONCLUSION: The optimized PR-nESI method was successfully applied for the sensitive analysis of OH-PAHs in complex biological samples with severe salt effects. Based on the present study, PR-nESI can have a promising prospect for the sensitive analysis of other metabolites of environmental pollutants in negative polarity.

Antidepressant Mechanism of Traditional Chinese Medicine Formula Xiaoyaosan in CUMS-Induced Depressed Mouse Model via RIPK1-RIPK3-MLKL Mediated Necroptosis Based on Network Pharmacology Analysis.

Background: Depression is a stress-related disorder that seriously threatens people's physical and mental health. Xiaoyaosan is a classical traditional Chinese medicine formula, which has been used to treat mental depression since ancient times. More and more notice has been given to the relationship between the occurrence of necroptosis and the pathogenesis of mental disorders. Objective: The purpose of present study is to explore the potential mechanism of Xiaoyaosan for the treatment of depression using network pharmacology and experimental research, and identify the potential targets of necroptosis underlying the antidepressant mechanism of Xiaoyaosan. Methods: The mice model of depression was induced by chronic unpredictable mild stress (CUMS) for 6 weeks. Adult C57BL/6 mice were randomly divided into five groups, including control group, chronic unpredictable mild stress group, Xiaoyaosan treatment group, necrostatin-1 (Nec-1) group and solvent group. Drug intervention performed from 4th to 6th week of modeling. The mice in Xiaoyaosan treatment group received Xiaoyaosan by intragastric administration (0.254 g/kg/d), and mice in CUMS group received 0.5 ml physiological saline. Meanwhile, the mice in Nec-1 group were injected intraperitoneally (i.p.) with Nec-1 (10 mg/kg/d), and the equivalent volume of DMSO/PBS (8.3%) was injected into solvent group mice. The behavior tests such as sucrose preference test, forced swimming test and novelty-suppressed feeding test were measured to evaluate depressive-like behaviors of model mice. Then, the active ingredients in Xiaoyaosan and the related targets of depression and necroptosis were compiled through appropriate databases, while the "botanical drugs-active ingredients-target genes" network was constructed by network pharmacology analysis. The expressions of RIPK1, RIPK3, MLKL, p-MLKL were detected as critical target genes of necroptosis and the potential therapeutic target compounds of Xiaoyaosan. Furthermore, the levels of neuroinflammation and microglial activation of hippocampus were measured by detecting the expressions of IL-1ß, Lipocalin-2 and IBA1, and the hematoxylin and eosin (H&E) stained was used to observe the morphology in hippocampus sections. Results: After 6-weeks of modeling, the behavioral data showed that mice in CUMS group and solvent group had obvious depressive-like behaviors, and the medication of Xiaoyaosan or Nec-1 could improve these behavioral changes. A total of 96 active ingredients in Xiaoyaosan which could regulate the 23 key target genes were selected from databases. Xiaoyaosan could alleviate the core target genes in necroptosis and improve the hippocampal function and neuroinflammation in depressed mice. Conclusion: The activation of necroptosis existed in the hippocampus of CUMS-induced mice, which was closely related to the pathogenesis of depression. The antidepressant mechanism of Xiaoyaosan included the regulation of multiple targets in necroptosis. It also suggested that necroptosis could be a new potential target for the treatment of depression.

CEBPG promotes acute myeloid leukemia progression by enhancing EIF4EBP1.

BACKGROUND: Acute myeloid leukemia (AML) is a myeloid neoplasm accounts for 7.6% of hematopoietic malignancies. AML is a complex disease, and understanding its pathophysiology is contributing to the improvement in the treatment and prognosis of AML. In this study, we assessed the expression profile and molecular functions of CCAAT enhancer binding protein gamma (CEBPG), a gene implicated in myeloid differentiation and AML progression. METHODS: shRNA mediated gene interference was used to down-regulate the expression of CEBPG in AML cell lines, and knockdown efficiency was detected by RT-qPCR and western blotting. The effect of knockdown on the growth of AML cell lines was evaluated by CCK-8. Western blotting was used to detect PARP cleavage, and flow cytometry were used to determine the effect of knockdown on apoptosis of AML cells. Genes and pathways affected by knockdown of CEBPG were identified by gene expression analysis using RNA-seq. One of the genes affected by knockdown of CEBPG was Eukaryotic translation initiation factor 4E binding protein 1 (EIF4EBP1), a known repressor of translation. Knockdown of EIF4EBP1 was used to assess its potential role in AML progression downstream of CEBPG. RESULTS: We explored the ChIP-Seq data of AML cell lines and non-AML hematopoietic cells, and found CEBPG was activated through its distal enhancer in AML cell lines. Using the public transcriptomic dataset, the Cancer Cell Line Encyclopedia (CCLE) and western blotting, we also found CEBPG was overexpressed in AML. Moreover, we observed that CEBPG promotes AML cell proliferation by activating EIF4EBP1, thus contributing to the progression of AML. These findings indicate that CEBPG could act as a potential therapeutic target for AML patients. CONCLUSION: In summary, we systematically explored the molecular characteristics of CEBPG in AML and identified CEBPG as a potential therapeutic target for AML patients. Our findings provide novel insights into the pathophysiology of AML and indicate a key role for CEBPG in promoting AML progression.

ARV-825 Demonstrates Antitumor Activity in Gastric Cancer via MYC-Targets and G2M-Checkpoint Signaling Pathways.

OBJECTIVE: Suppression of bromodomain and extra terminal (BET) proteins has a bright prospect to treat MYC-driven tumors. Bromodomain containing 4 (BRD4) is one of the BET proteins. ARV-825, consisting of a BRD4 inhibitor conjugated with a cereblon ligand using proteolysis-targeting chimera (PROTAC) technology, was proven to decrease the tumor growth effectively and continuously. Nevertheless, the efficacy and mechanisms of ARV-825 in gastric cancer are still poorly understood. METHODS: Cell counting kit 8 assay, lentivirus infection, Western blotting analysis, Annexin V/propidium iodide (PI) staining, RNA sequencing, a xenograft model, and immunohistochemistry were used to assess the efficacy of ARV-825 in cell level and animal model. RESULTS: The messenger RNA (mRNA) expression of BRD4 in gastric cancer raised significantly than those in normal tissues, which suggested poor outcome of patients with gastric cancer. ARV-825 displayed higher anticancer efficiency in gastric cancer cells than OTX015 and JQ1. ARV-825 could inhibit cell growth, inducing cell cycle block and apoptosis in vitro. ARV-825 induced degradation of BRD4, BRD2, BRD3, c-MYC, and polo-like kinase 1 (PLK1) proteins in four gastric cancer cell lines. In addition, cleavage of caspase 3 and poly-ADP-ribose polymerase (PARP) was elevated. Knockdown or overexpression CRBN could increase or decrease, respectively, the ARV-825 IC50 of gastric cancer cells. ARV-825 reduced MYC and PLK1 expression in gastric cancer cells. ARV-825 treatment significantly reduced tumor growth without toxic side effects and downregulated the expression of BRD4 in vivo. CONCLUSIONS: High mRNA expression of BRD4 in gastric cancer indicated poor prognosis. ARV-825, a BRD4 inhibitor, could effectively suppress the growth and elevate the apoptosis of gastric cancer cells via transcription downregulation of c-MYC and PLK1. These results implied that ARV-825 could be a good therapeutic strategy to treat gastric cancer.

Direct distinction of ibuprofen and flurbiprofen enantiomers by ion mobility mass spectrometry of their ternary complexes with metal cations and cyclodextrins in the gas phase.

Enantiomeric drugs are widely used and play important roles in pharmaceuticals. Ion mobility spectrometry coupled with mass spectrometry technology provides a unique method for distinguishing the enantiomeric drugs, enantiomeric identification, and quantitation in the gas phase. In this study, enantiomeric molecules of ibuprofen and flurbiprofen were clearly recognized by forming host-guest complex ions using trapped ion mobility time-of-flight mass spectrometry. Ternary complex ions can be produced easily by electrospray ionization of the mixed solutions of ibuprofen, cyclodextrins, and CaCl2 , LiCl, or NaCl, as well as flurbiprofen, cyclodextrins, and CaCl2 . The relative contents of different chiral ibuprofens in a mixed solution were also quantitatively measured. This new method is a simple, effective, and a convenient enantioselective analysis method.

BRD4 PROTAC degrader ARV-825 inhibits T-cell acute lymphoblastic leukemia by targeting 'Undruggable' Myc-pathway genes.

BACKGROUND: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive disease with a high risk of induction failure and poor outcomes, with relapse due to drug resistance. Recent studies show that bromodomains and extra-terminal (BET) protein inhibitors are promising anti-cancer agents. ARV-825, comprising a BET inhibitor conjugated with cereblon ligand, was recently developed to attenuate the growth of multiple tumors in vitro and in vivo. However, the functional and molecular mechanisms of ARV-825 in T-ALL remain unclear. This study aimed to investigate the therapeutic efficacy and potential mechanism of ARV-825 in T-ALL. METHODS: Expression of the BRD4 were determined in pediatric T-ALL samples and differential gene expression after ARV-825 treatment was explored by RNA-seq and quantitative reverse transcription-polymerase chain reaction. T-ALL cell viability was measured by CCK8 assay after ARV-825 administration. Cell cycle was analyzed by propidium iodide (PI) staining and apoptosis was assessed by Annexin V/PI staining. BRD4, BRD3 and BRD2 proteins were detected by western blot in cells treated with ARV-825. The effect of ARV-825 on T-ALL cells was analyzed in vivo. The functional and molecular pathways involved in ARV-825 treatment of T-ALL were verified by western blot and chromatin immunoprecipitation (ChIP). RESULTS: BRD4 expression was higher in pediatric T-ALL samples compared with T-cells from healthy donors. High BRD4 expression indicated a poor outcome. ARV-825 suppressed cell proliferation in vitro by arresting the cell cycle and inducing apoptosis, with elevated poly-ADP ribose polymerase and cleaved caspase 3. BRD4, BRD3, and BRD2 were degraded in line with reduced cereblon expression in T-ALL cells. ARV-825 had a lower IC50 in T-ALL cells compared with JQ1, dBET1 and OTX015. ARV-825 perturbed the H3K27Ac-Myc pathway and reduced c-Myc protein levels in T-ALL cells according to RNA-seq and ChIP. In the T-ALL xenograft model, ARV-825 significantly reduced tumor growth and led to the dysregulation of Ki67 and cleaved caspase 3. Moreover, ARV-825 inhibited cell proliferation by depleting BET and c-Myc proteins in vitro and in vivo. CONCLUSIONS: BRD4 indicates a poor prognosis in T-ALL. The BRD4 degrader ARV-825 can effectively suppress the proliferation and promote apoptosis of T-ALL cells via BET protein depletion and c-Myc inhibition, thus providing a new strategy for the treatment of T-ALL.

Improving the performance of proteomic analysis via VAILase cleavage and 193-nm ultraviolet photodissociation.

Further improving the proteomic identification coverage and reliability is still challenging in the mass spectrometry (MS)-based proteomics. Herein, we combine VAILase and trypsin digestion with 193-nm ultraviolet photodissociation (UVPD) and higher-energy collision dissociation (HCD) to improve the performance of bottom-up proteomics. As VAILase exhibits high complementarity to trypsin, the proteome sequence coverage is improved obviously whether with HCD or 193-nm UVPD. The high diversity of fragment ion types produced by UVPD contributes to the improvements of identification reliability for both trypsin- and VAILase-digested peptides with an average XCorr score improvement of 10%.

Gas-phase amination of aromatic hydrocarbons by corona discharge-assisted nitrogen fixation.

This paper reports on the gas-phase amination reaction of aromatic hydrocarbons occurring under corona discharge conditions with N2 gas as the nitrogen source. The corona discharge device within an atmospheric pressure chemical ionization source was employed to achieve the plasma-assisted N2 fixation, and the coupled ion trap mass spectrometer (IT-MS) was used to detect positively charged product ions. In the model case, under APCI conditions, unusual product ions, [M + 16]+ and [M + 14]+, were observed. Based on the high resolution MS data and tandem mass spectrometric information, [M + 16]+ was confirmed to be protonated p-toluidine and [M + 14]+ was confirmed to be p-methylphenylnitrenium ion. According to the experimental results of the isotopic labelling and substituent effect, one feasible mechanism is proposed as follows. Firstly, N2 is activated by plasma caused via the corona discharge and then electrophilically attacks toluene, yielding the key intermediate, p-methylphenylnitrenium; secondly, the intermediate undergoes double-hydrogen transfer reaction to give rise to the final product ion, protonated p-toluidine. This study may provide a novel idea to explore new and green method for the synthesis of anilines.