[This corrects the article DOI: 10.1016/j.apsb.2022.05.019.].

OBJECTIVES: To analyze the differentially expressed exosomal miRNAs in patients with chronic heart failure (CHF) complicated by hyperuricemia (HUA) and explore their potential as novel diagnostic molecular markers and their target genes. METHODS: This study was conducted among 30 CHF patients with HUA (observation group) and 30 healthy volunteers (control group) enrolled between September, 2020 and September, 2023. Peripheral blood samples were collected from 6 CHF patients with HUA for analyzing exosomal miRNAs by high-throughput sequencing, and the results were validated in the remaining 24 patients using qRT-PCR. GO and KEGG enrichment analyses were performed to predict the the target genes of the identified differential miRNAs. We also validated the differentially expressed miRNAs by animal experiment. RESULTS: A total of 42 differentially expressed exosomal miRNAs were detected in observation group by high-throughput sequencing; among them, miR-27a-5p was significantly upregulated (P=0.000179), and miR-139-3p was significantly downregulated (P=0.000058). In the 24 patients with both CHF and PUA, qRT-PCR validated significant upregulation of miR-27a-5p (P=0.004) and downregulation of miR-139-3p (P=0.005) in serum exosomes. When combined, miR-27a-5p and miR-139-3p had a maximum area under the curve (AUC) of 0.899 (95% CI: 0812-0.987) for predicting CHF complicated by HUA. GO and KEGG enrichment analyses suggested that the differential expressions of miR-27a-5p and miR-139-3p was associated with the activation of the AMPK-mTOR signaling pathway to activate the autophagic response. We obtained the same conclusion from animal experiment. CONCLUSIONS Upregulated exosomal miR-27a-5p combined with downregulated exosomal miR-139-3p expression can serve as a novel molecular marker for diagnosis of CHF complicated by HUA, and their differential expression may promote autophagy in cardiomyocytes by activating the AMPK-mTOR signaling pathway.
Humans ; Hyperuricemia/diagnosis* ; Heart Failure/genetics* ; MicroRNAs/metabolism* ; Exosomes/metabolism* ; Chronic Disease ; Male ; Female ; Middle Aged ; Animals

OBJECTIVES: To explore the effects of cannabidiol on endoplasmic reticulum stress and neuronal apoptosis in rats with multiple concussions (MCC). METHODS: SD rats were randomized into sham group, MCC group, 1% tween20 (TW) treatment group, and low-dose (10 mg/kg) and high-dose (40 mg/kg) cannabidiol treatment groups. In all but the sham group, MCC models were established using a metal pendulum percussion device, after which the rats received daily intraperitoneal injections of the corresponding agents for 2 weeks. The expressions of PERK, eIF2α, ATF4, CHOP, TRIB3, p-Akt and pro-caspase-3 in the brain tissue of the rats were detected with qRT-PCR, Western blotting and immunofluorescence staining. The core targets of cannabidiol in treatment of traumatic brain injury (TBI) were identified by network pharmacology analysis, and molecular docking was carried out to simulate the interaction of cannabidiol with the factors related to endoplasmic reticulum stress and apoptosis. RESULTS: Compared with the sham-operated rats, the rat models of MCC showed significantly increased mRNA expressions of PERK, eIF2α and CHOP and protein expressions of PERK, eIF2α, ATF4, CHOP, TRIB3, p-AKT and pro-caspase-3 in the cerebral cortex. CBD treatment, especially at the high dose, obviously increased the expression of p-Akt and lowered the expression levels of the other factors tested in the rat models. Network pharmacology analysis indicated interactions of the core targets of CBD with the factors related to endoplasmic reticulum stress and TBI, and molecular docking study showed a high binding energy of CBD with multiple factors pertaining to endoplasmic reticulum stress and apoptosis. CONCLUSIONS MCC induce endoplasmic reticulum stress and apoptosis in rat brain tissues, for which CBD, especially at a high dose, provides neuroprotective effects by inhibiting endoplasmic reticulum stress and cell apoptosis.
Animals ; Endoplasmic Reticulum Stress/drug effects* ; Apoptosis/drug effects* ; Rats, Sprague-Dawley ; Activating Transcription Factor 4/metabolism* ; Transcription Factor CHOP/metabolism* ; Rats ; Eukaryotic Initiation Factor-2/metabolism* ; Signal Transduction/drug effects* ; eIF-2 Kinase/metabolism* ; Cannabidiol/pharmacology* ; Neurons/metabolism* ; Brain Concussion/metabolism* ; Male ; Molecular Docking Simulation

Ursodeoxycholic acid (UDCA) is a naturally occurring, low-toxicity, and hydrophilic bile acid (BA) in the human body that is converted by intestinal flora using primary BA. Solute carrier family 7 member 11 (SLC7A11) functions to uptake extracellular cystine in exchange for glutamate, and is highly expressed in a variety of human cancers. Retroperitoneal liposarcoma (RLPS) refers to liposarcoma originating from the retroperitoneal area. Lipidomics analysis revealed that UDCA was one of the most significantly downregulated metabolites in sera of RLPS patients compared with healthy subjects. The augmentation of UDCA concentration (≥25 μg/mL) demonstrated a suppressive effect on the proliferation of liposarcoma cells. [¹⁵N₂]-cystine and [¹³C₅]-glutamine isotope tracing revealed that UDCA impairs cystine uptake and glutathione (GSH) synthesis. Mechanistically, UDCA binds to the cystine transporter SLC7A11 to inhibit cystine uptake and impair GSH de novo synthesis, leading to reactive oxygen species (ROS) accumulation and mitochondrial oxidative damage. Furthermore, UDCA can promote the anti-cancer effects of ferroptosis inducers (Erastin, RSL3), the murine double minute 2 (MDM2) inhibitors (Nutlin 3a, RG7112), cyclin dependent kinase 4 (CDK4) inhibitor (Abemaciclib), and glutaminase inhibitor (CB839). Together, UDCA functions as a cystine exchange factor that binds to SLC7A11 for antitumor activity, and SLC7A11 is not only a new transporter for BA but also a clinically applicable target for UDCA. More importantly, in combination with other antitumor chemotherapy or physiotherapy treatments, UDCA may provide effective and promising treatment strategies for RLPS or other types of tumors in a ROS-dependent manner.

Traditional in vitro models have inevitable limitations and there are significant differences in assessing drug efficacy and side effects as compared to human trials.Organ-on-a-chip technology simulates human organs in a physiological environment and functional chip with a high fidelity physiological or pathophysiological level,offering great innovative prospects for drug development.This paper mainly introduces the research progress and application of organ chip from the perspective of various systems in vivo.At the same time,the limitations of the current devel-opment process of organ chip and the future development direction are proposed.

More and more in-depth tumor mechanism research and clinical precision treatment have put forward higher requirements for the technology for the establishment of tumor models.More and more tumor organoids have been applied.This model is able to reproduce the genomic,transcriptome,proteome and other omics features of pa-rental tumors without heterogeneity found in cell line models.Based on these characteristics,tumor organoids have gradually become a representative preclinical model,facilitating the translation of new research results and precision treatment of patients.This article summarizes the latest progress of tumor organoids on R&D of technology and appli-cation,focusing on the current methods of establishing tumor organoids,opportunities and challenges in clinical and research application.

Objective To establish breast cancer organoids for a long time and to characterize their molecular ex-pression and test their sensitivity to chemotherapy drugs.Methods Breast cancer specimens were digested with col-lagenase to release cancer cells for organoid culture.The organoids were characterized by morphology and ER,PR,HER-2,CK expression by technology of histoimmunofluorescence.Among them,three were tested for paclitaxel,doxorubicin and cisplatin sensitivity.Results A total of 44 cases of breast cancer organoids were established,all of which showed dense spherical-like growth and ER,PR,HER-2,CK,matching their clinical counterpart.Breast cancer organoids were sensitive to chemotherapy drugs paclitaxel,doxorubicin and cisplatin.Conclusions Organoid model,as a new in vitro may reproduce the pathophysiology features with heterogeneity.

Objective To establish human and mouse pancreatic cancer cell strains stably expressing Cas9 protein,green fluorescent protein,red fluorescent proteins,luciferase-tdTomato,and to validate the activity of luciferase and gene editing of Cas9 function for pancreatic cancer research using luciferase and CRISPR/Cas9 system.Methods In human pancreatic cancer cells(AsPC-1,CFPAC-1,HPAC,BxPC-3,HS 766T,MIA PaCa-2,PANC-1,and SW 1990),and mouse pancreatic cancer cell(Pan02),the cells were infected with Cas9-expressing plas-mid pLv-EF1α-Cas9m1.1-Puro,and single-cell clones were selected for culture and expansion.After extracting the total protein,Western blot verified the expression level of Cas9;Infected with fluorescent protein expression plasmids pLv-EF1α-EGFP,pLv-EF1α-mCherry,pLv-EF1α-tdTomato,pLv-EF1α-Luc2-tdT,and selected single cell clones stably expressing fluorescent proteins were cultured and amplified under fluorescence microscope.Cas9 stable expression cell line was selected to be infected with pLv-EF1α-Luc2-tdT,and the mono-clonal culture of stable expression of fluorescent proteins was selected for expansion under fluorescence micro-scope.One of the cell lines were selected to be infected with Lv-EF1a-mCherry,and the mCherry-positive cells were sorted out by flow cytometry,and then the guide RNA of mCherry gene was then infected by lentivirus to tar-get the mCherry gene,and after cell expansion,mCherry knockdown was detected by fluorescence microscope observation and flow cytometry;5 BALB/c Nude mice were subcutaneously inoculated with MIA PaCa-2-Luc2-tdT cells(1.0×107/cells each),and imaged in vivo after 36 days.Results 48 human pancreatic cancer cell strains with stable Cas9 expression were screened(including 23 cells expressing Cas9m1.1,25 cells expressing Cas9m1.1-Luc2-tdT),33 pancreatic cancer cell strains with stable expression of fluorescent proteins were screened(8 cells expressing EGFP,7 expressing mCherry,and 9 each expressing Luc2-tdT and tdTomato).Cells expressing mCherry and Cas9 were infected with mCherry gRNA and mCherry was knocked down.In vivo imaging showed that both bioluminescence and fluorescence luminescence were present in MIA PaCa-2 cells ex-pressing Luc2-tdT.Conclusions 33 pancreatic cancer cell strains with stable expression of fluorescent proteins are successfully established,in which the Luc2-tdT-expressing cell strains have luciferase activity;48 pancreatic cancer cell strains with stable expression of Cas9 are successfully established,and the Cas9 protein has gene edi-ting activity,gene editing activity varies depending on the original cell strains.

Objective To explore the value of lipoprotein-associated phospholipase A2(Lp-PLA2)in the diagnostic grading and prog-nostic assessment of pneumonia-associated acute respiratory distress syndrome(p-ARDS).Methods The study was a prospective ob-servational study.Fifty-seven patients with p-ARDS admitted to the ICU ward of Tianjin Hospital from January 2022 to August 2023 were included as the research subjects.Twenty-six pneumonia patients admitted to the general respiratory ward during the same period and 10 healthy individuals undergoing medical examinations were selected as the control group.Their serum samples were collected,and the samples from p-ARDS and pneumonia patients were obtained within 24 hours of admission.The levels of serum Lp-PLA2,in-terleukin 6(IL-6),and IL-8 were detected using the Luminex? multiplex test kit.The baseline data and laboratory test results,inclu-ding routine blood parameters,biochemical markers,C-reactive protein(CRP),procalcitonin(PCT),and D-dimer at admission,were collected from the patients with p-ARDS or pneumonia.The levels of serum Lp-PLA2 were compared by grouping based on clinical diagnosis,severity of ARDS,and clinical outcomes on day 28 after admission.The diagnostic and prognostic value of serum Lp-PLA2 in p-ARDS was evaluated by plotting the receiver operating characteristic(ROC)curve,Spearman correlation analysis,and Logistic regression analysis.Results The levels of serum Lp-PLA2 in the p-ARDS group([233.67±83.49]ng/mL)were significantly higher than that in the pneumonia group([150.86±39.48]ng/mL,P＜0.05),while those in the pneumonia group were significantly higher than that in the healthy control group([150.86±39.48]ng/mL vs[92.07±12.89]ng/mL,P＜0.05).The analysis results of the ROC curve showed that serum Lp-PLA2 had a better ability to distinguish p-ARDS from pneumonia than indicators such as IL-6,IL-8,CRP,and PCT,with an area under the ROC curve(AUCROC)of 0.781(95％CI:0.685-0.878).The diagnostic value of serum Lp-PLA2 combined with D-dimer was higher,with an AUCROC of 0.897(95％CI:0.832-0.963).Subgroup analysis found that as lung inju-ry worsened,the levels of serum Lp-PLA2 increased,and that serum Lp-PLA2 levels were negatively correlated with the PaO2/FiO2 ra-tio in p-ARDS patients(r=-0.549)and positively correlated with the sequential organ failure assessment(SOFA)scores at admission(r=0.412).The levels of serum Lp-PLA2 in the death group of p-ARDS were significantly higher than that in the survival group([314.5±43.1]ng/mL vs[174.9±48.9]ng/mL,P＜0.001).Logistic regression analysis showed that after adjusting for the SOFA score,serum Lp-PLA2 was independently associated with the mortality risk on day 28 after admission(OR=1.099,95％CI:1.026-1.178,P=0.007).Similar results were obtained after adjusting for IL-8 or the PaO2/FiO2 ratio.Conclusion Serum Lp-PLA2 may be used as a biomarker to aid in the diagnostic grading and prognostic assessment of p-ARDS.

Objective:To explore the impact of scutellarin on the levels of phosphatidylinositol 3-kinase(P13K)and phosphorylated P13K(p-PI3K)in activated microglia.Methods:A rat cerebral ischemia model was made by middle cerebral artery occlusion(MCAO)method.SD rats were randomly divided into three groups:sham operation group(Sham),cerebral ischemia group(MCAO),and cerebral ischemia with scutellarin group(MCAO+S).A model of ischemia-hypoxia injury of BV2 microglia was established by glucose-oxygen deprivation(OGD)and randomly assigned as control group(Control),OGD group and OGD+scutellarin group(OGD+S).Changes in PI3K and p-PI3K expres-sion in microglia were assessed using double immunofluorescence staining and Western Blot.Results:Immunofluores-cence staining and Western Blot analyses revealed a significant increase in p-PI3K levels in activated microglia both in vivo and in vitro(P＜0.05).Furthermore,treatment with scutellarin led to a further elevation in p-PI3K(P＜0.05).However,there were no significant alterations observed in PI3 K expression among the groups(P＞0.05).Conclusion:Scutellarin may play a positive role in the treatment of cerebral ischemia by up-regulating the phosphorylation level of P13K in activated microglia.