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IntroductionVariants of uncertain significance (VUS) surged with affordable genetic testing, posing challenges for determining pathogenicity. We examine the pathogenicity of a novel VUS P93S in Annexin A11 (ANXA11) - an amyotrophic lateral sclerosis/frontotemporal dementia-associated gene - in a corticobasal syndrome kindred. Established ANXA11 mutations cause ANXA11 aggregation, altered lysosomal-RNA granule co-trafficking, and transactive response DNA binding protein of 43 kDa (TDP-43) mis-localization.MethodsWe described the clinical presentation and explored the phenotypic diversity of ANXA11 variants. P93S's effect on ANXA11 function and TDP-43 biology was characterized in induced pluripotent stem cell-derived neurons alongside multiomic neuronal and microglial profiling.ResultsANXA11 mutations were linked to corticobasal syndrome cases. P93S led to decreased lysosome colocalization, neuritic RNA, and nuclear TDP-43 with cryptic exon expression. Multiomic microglial signatures implicated immune dysregulation and interferon signaling pathways.DiscussionThis study establishes ANXA11 P93S pathogenicity, broadens the phenotypic spectrum of ANXA11 mutations, underscores neuronal and microglial dysfunction in ANXA11 pathophysiology, and demonstrates the potential of cellular models to determine variant pathogenicity.HighlightsANXA11 P93S is a pathogenic variant. Corticobasal syndrome is part of the ANXA11 phenotypic spectrum. Hybridization chain reaction fluorescence in situ hybridization (HCR FISH) is a new tool for the detection of cryptic exons due to TDP-43-related loss of splicing regulation. Microglial ANXA11 and related immune pathways are important drivers of disease. Cellular models are powerful tools for adjudicating variants of uncertain significance.

Fluazuron is a novel veterinary pour-on antitick formulation which can be applied simultaneously with bovine reproduction management strategies. Considering the economic importance of the livestock industry in many countries, it is important to know whether antiparasitics such as fluazuron may cause embryonic loss. The aim of this study was to evaluate the toxicological effect of fluazuron on bovine oocytes during in vitro maturation. The best fluazuron concentrations were determined in a preliminary experiment on Chinese hamster ovary (CHO)-K1 cells and further used to compare fluazuron toxicity in both study models. Results of the annexin V and alkaline single cell gel electrophoresis assays demonstrated that fluazuron caused cytotoxicity and genotoxicity in bovine cumulus cells at all the concentrations tested (50, 75 and 100 μg fluazuron/mL). The evaluation of cortical granules and mitochondria distribution showed that cytoplasmic maturation was not affected by fluazuron treatment. However, a decrease in metaphase II + polar body, degenerate oocytes as well as disorganized chromatin in polar body were observed at all concentrations tested. Whereas the fertilization process was not altered by 50 μg/mL fluazuron, the embryo development rate decreased significantly. No significant differences were observed in any of the oxidative stress parameters assessed. This study contributes to a better understanding of fluazuron in bovines, suggesting that the antiparasitic may affect bovine reproduction and might cause embryo loss. • Fluazuron induces genotoxicity in CHO-K1 and genotoxicity and apoptosis in cumulus cells. • Fluazuron reduces metaphase II and polar body (PB) formation, degenerates oocytes, and disorganizes PB chromatin. • Embryo development rate decreased significantly after cumulus-oocyte cells exposure to 50 μg fluazuron/mL. • Fluazuron does not alter the oxidative stress parameters assessed in bovine cumulus cells and oocytes. [ABSTRACT FROM AUTHOR]

Introduction: Pulmonary arterial hypertension (PAH) is a serious complication of systemic lupus erythematosus (SLE) with increased mortality. A prothrombotic state may contribute to pathogenesis of SLE-PAH. Extracellular vesicles (EVs) are known to be associated with thrombosis. Here, we investigated circulating EVs and their associations with SLE-PAH. Methods: Eighteen SLE-PAH patients, 36 SLE-non-PAH patients, and 36 healthy controls (HCs) were enrolled. Flow cytometry was used to analyze circulating EVs from leukocytes (LEVs), red blood cells (REVs), platelets (PEVs), endothelial cells (EEVs), and Annexin V+ EVs with membrane phosphatidylserine (PS) exposure. Results: Plasma levels of all EV subgroups were elevated in SLE patients with or without PAH compared to HCs. Furthermore, plasma Annexin V+ EVs, LEVs, PEVs, REVs, EEVs, and Annexin V+ REVs were significantly elevated in SLE-PAH patients compared to SLE-non-PAH patients. Additionally, PAH patients with moderate/high SLE showed a significant increase in LEVs, PEVs, REVs, Annexin V+ EVs, and Annexin V+ REVs compared to SLE-non-PAH patients. However, PAH patients with inactive/mild SLE only exhibited elevations in Annexin V+ EVs, REVs, and Annexin V+ REVs. In the SLE-PAH patients, EEVs were positively correlated with pulmonary arterial systolic pressure, while PEVs and EEVs were positively correlated with right ventricular diameter. Moreover, the receiver operating characteristic curve indicated that Annexin V+ EVs, LEVs, PEVs, REVs, EEVs and Annexin V+ REVs could predict the presence of PAH in SLE patients. Importantly, multivariate logistic regression analysis showed that circulating levels of LEVs or REVs, anti-nRNP antibody, and serositis were independent risk factors for PAH in SLE patients. Discussion: Findings reveal that specific subgroups of circulating EVs contribute to the hypercoagulation state and the severity of SLE-PAH. Higher plasma levels of LEVs or REVs may serve as biomarkers for SLE-PAH. [ABSTRACT FROM AUTHOR]

AbstractBackgroundObjectiveMethodsResultsConclusionsTriptolide (TP) has been confirmed to possess many beneficial functions including anti-inflammation and immunosuppression.The present study aimed to explore the potential involvement of IL-2/IL-2R pathway in the immunosuppressive activities of TP.Cultured CTLL-2 cells were utilized to evaluate the potential benefits of TP. Then cell viability was determined by CCK-8 assay, IFN-γ level by ELISA assay, Annexin V-FITC/PI double-staining and CD25 expression by flow cytometry, and protein expression by western blotting. Additionally, rhIL-2–driven lymphocytes following ConA activation were investigated. The interactions of TP with IL-2 and IL-2Rα were investigated by binding assays and molecular dynamics simulations.TP treatment attenuated IFN-γ level and cell viability in both rhIL-2–induced CTLL-2 cells and rhIL-2–driven splenic lymphocytes. TP treatment increased cellular apoptosis/necrosis and cleaved PARP-1 level, while suppressed c-Myc level in rhIL-2–induced CTLL-2 cells. Additionally, TP treatment reduced CD25 expression on CTLL-2 cell surface. Notably, the phosphorylation protein levels in IL-2R signaling pathways were inhibited by TP exposure prior to rhIL-2 stimulation. SPR and BLI assays verified TP that directly bound to rhIL-2 and rmIL-2Rα, respectively. Molecular simulations suggested that TP bound at the interface of IL-2 and IL-2Rα near the hydrophobic patch composed of F62, L92 on IL-2 and L23, I46, V139 on IL-2Rα, resulting in decreased binding free energy between IL-2 and IL-2Rα.These findings collectively emphasized that TP interfered IL-2/IL-2Rα interactions, down-regulated IL-2Rα expression, and inhibited IL-2R signaling pathways activation, thereby leading to the immune cells being desensitized to rhIL-2 and exhibiting immunosuppressive properties. [ABSTRACT FROM AUTHOR]

Feline infectious peritonitis (FIP), a serious infectious disease in cats, has become a challenging problem for pet owners and the industry due to the lack of effective vaccinations and medications for prevention and treatment. Currently, most natural compounds have been proven to have good antiviral activity. Hence, it is essential to develop efficacious novel natural compounds that inhibit FIPV infection. Our study aimed to screen compounds with in vitro anti-FIPV effects from nine natural compounds that have been proven to have antiviral activity and preliminarily investigate their mechanisms of action. In this study, the CCK-8 method was used to determine the maximum noncytotoxic concentration (MNTC), 50% cytotoxic concentration (CC50), and 50% effective concentration (EC50) of natural compounds on CRFK cells and the maximum inhibition ratio (MIR) of the compounds inhibit FIPV. The effect of natural compounds on FIPV-induced apoptosis was detected via Annexin V-FITC/PI assay. Network pharmacology (NP), molecular docking (MD), and 4D label-free quantitative (4D-LFQ) proteomic techniques were used in the joint analysis the mechanism of action of the screened natural compounds against FIPV infection. Finally, Western blotting was used to validate the analysis results. Among the nine natural compounds, baicalin had good antiviral effects, with an MIR > 50% and an SI > 3. Baicalin inhibited FIPV-induced apoptosis. NP and MD analyses showed that AKT1 was the best target of baicalin for inhibiting FIPV infection. 4D-LFQ proteomics analysis showed that baicalin might inhibit FIPV infection by modulating the PI3K-AKT pathway and the apoptosis pathway. The WB results showed that baicalin promoted the expression of EGFR, PI3K, and Bcl-2 and inhibited the expression of cleaved caspase 9 and Bax. This study found that baicalin regulated the PI3K-AKT pathway and the apoptosis pathway in vitro and inhibited FIPV-induced apoptosis, thus exerting anti-FIPV effects. [ABSTRACT FROM AUTHOR]

Intervertebral disc degeneration (IVDD) is a chronic degenerative disease involving the aging and loss of proliferative capacity of nucleus pulposus cells (NPCs), processes heavily dependent on mitochondrial dynamics and autophagic flux. This study finds that the absence of BCL2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3) is associated with senescence‐related NPC degeneration, disrupting mitochondrial quality control. Bone marrow mesenchymal stem cells (BMSCs) have multidirectional differentiation potential and produce extracellular vesicles containing cellular activators. Therefore, in this study, BMSCs are induced under hypoxic stimulation to deliver BNIP3‐rich extracellular vesicles to NPCs, thereby alleviating aging‐associated mitochondrial autophagic flux, promoting damaged mitochondrial clearance, and restoring mitochondrial quality control. Mechanistically, BNIP3 is shown to interact with the membrane‐bound protein annexin A2 (ANXA2), enabling the liberation of the transcription factor EB (TFEB) from the ANXA2‐TFEB complex, promoting TFEB nuclear translocation, and regulating autophagy and lysosomal gene activation. Furthermore, a rat model of IVDD is established and verified the in vivo efficacy of the exosomes in repairing disc injuries, delaying NPC aging, and promoting extracellular matrix (ECM) synthesis. In summary, hypoxia‐induced BMSC exosomes deliver BNIP3‐rich vesicles to alleviate disc degeneration by activating the mitochondrial BNIP3/ANXA2/TFEB axis, providing a new target for IVDD treatment. [ABSTRACT FROM AUTHOR]

Choriocarcinoma is a malignant cancer that belongs to gestational trophoblastic neoplasia (GTN). Herein, serum metabolomic analysis was performed on 29 GTN patients and 30 healthy individuals to characterize the metabolic variations during GTN progression. Ultimately 24 differential metabolites (DMs) were identified, of which, Equol was down-regulated in GTN patients, whose VIP score is the 3rd highest among the 24 DMs. As an intestinal metabolite of daidzein, the anticancer potential of Equol has been demonstrated in multiple cancers, but not choriocarcinoma. Hence, human choriocarcinoma cell lines JEG-3 and Bewo were used and JEG-3-derived subcutaneous xenograft models were developed to assess the effect of Equol on choriocarcinoma. The results suggested that Equol treatment effectively suppressed choriocarcinoma cell proliferation, induced cell apoptosis, and reduced tumorigenesis. Label-free quantitative proteomics showed that 136 proteins were significantly affected by Equol and 20 proteins were enriched in Gene Ontology terms linked to protein degradation. Tripartite motif containing 21 (TRIM21), a E3 ubiquitin ligase, was up-regulated by Equol. Equol-induced effects on choriocarcinoma cells could be reversed by TRIM21 inhibition. Annexin A2 (ANXA2) interacted with TRIM21 and its ubiquitination was modulated by TRIM21. We found that TRIM21 was responsible for proteasome-mediated degradation of ANXA2 induced by Equol, and the inhibitory effects of Equol on the malignant behaviors of choriocarcinoma cells were realized by TRIM21-mediated down-regulation of ANXA2. Moreover, β-catenin activation was inhibited by Equol, which also depended on TRIM21-mediated down-regulation of ANXA2. Taken together, Equol may be a novel candidate for the treatment for choriocarcinoma. [ABSTRACT FROM AUTHOR]

The maternal -fetal interface is essential for successful embryo implantation and subsequent placental development. Many studies have showed that the maternal-fetal interface is rich in the annexin A (AnxA) family, and that AnxA family not only regulates the biological behavior of trophoblast cells at the maternal-fetal interface, but also participates in the metaphase reaction. Therefore, the aberrant expression of the AnxA family is associated with a range of adverse pregnancy outcomes, such as recurrent implantation failure and recurrent pregnancy loss. Clarifying the role of the AnxA family at the maternal-fetal interface will help to understand better the mechanism of action of AnxA leading to adverse pregnancy outcomes, and to provide new targets for the treatment of related diseases. The possible roles of the AnxA family at the maternal -fetal interface and in the related adverse pregnancy are summarized in this review, to show a potential direction of the treatment of adverse pregnancy outcomes. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD) is a rising global burden, affecting one in four adults. Despite the increasing prevalence of NAFLD, the exact cellular and molecular mechanisms remain unclear, and effective therapeutic strategies are still limited. In vitro models of NAFLD are critical to understanding the pathogenesis and searching for effective therapies; thus, we evaluated the effects of free fatty acids (FFAs) on NAFLD hallmarks and their association with the modulation of Annexin A2 (ANXA2) and Keratin 17 (KRT17) in HepG2 cells. Our results show that oleic and palmitic acids can differentially induce intracellular lipid accumulation, cell death, and promote oxidative stress by increasing lipid peroxidation, protein carbonylation, and antioxidant defense depletion. Moreover, a markedly increased expression of inflammatory cytokines demonstrated the activation of inflammation pathways associated with lipotoxicity and oxidative stress. ANXA2 overexpression and KRT17 nuclear translocation were also observed, supporting the role of both molecules in the progression of liver disease. Taken together, these data provide insights into the interplay between ANXA2 and KRT17 in NAFLD, paving the way for understanding molecular mechanisms involved with the disease and developing new therapeutic strategies. [ABSTRACT FROM AUTHOR]

Despite unquestionable advances in therapy, melanoma is still characterized by a high mortality rate. For years, high expectations have been raised by compounds of natural origin as a component of pharmacotherapy, particularly by triterpenes found in the bark of birch trees. In this study, 3,4-seco-dammara-4(29),20(21),24(25)-trien-3-oic acid (SDT) was isolated from buds of silver birch and its mechanisms of cell death induction, including apoptosis and autophagy, were determined. Cytotoxicity of SDT was evaluated by the cell viability test and clonogenic assay, whereas induction of apoptosis and autophagy was determined by annexin V staining and Western blot. The results revealed dose- and time-dependent reductions in viability of melanoma cells. Treatment of cells for 48 h led to an increase in the percentage of annexin V-positive cells, activation of caspase-8, caspase-9, and caspase-3, and cleavage of PARP, confirming apoptosis. Simultaneously, it was found that SDT increased the level of autophagy marker LC3-II and initiator of autophagy beclin-1. Pretreatment of cells with caspase-3 inhibitor or autophagy inhibitor significantly reduced the cytotoxicity of SDT and revealed that both apoptosis and autophagy contribute to a decrease in cell viability. These findings suggest that 3,4-seco-dammaranes may become a promising group of natural compounds for searching for anti-melanoma agents. [ABSTRACT FROM AUTHOR]

Diabetic retinopathy (DR) is a leading cause of blindness, yet its molecular mechanisms are unclear. Extracellular vesicles (EVs) contribute to dysfunction in DR, but the characteristics and functions of vitreous EVs are unclear. This study investigated the inflammatory properties of type 2 diabetic (db) vitreous EVs. EVs isolated from the vitreous of db and non-db donors were used for nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), immunogold staining, Western blotting, and proteomic analysis by mass spectrometry. Intracellular uptake of vitreous EVs by differentiated macrophages was evaluated using ExoGlow membrane labeling, and the impact of EVs on macrophage (THP-1) activation was assessed by cytokine levels using RT-qPCR. NTA and TEM analysis of db and non-db vitreous EVs showed non-aggregated EVs with a heterogeneous size range below 200 nm. Western blot detected EV markers (Alix, Annexin V, HSP70, and Flotillin 1) and an upregulation of Cldn5 in db EVs. While the db EVs were incorporated into macrophages, treatment of THP-1 cells with db EVs significantly increased mRNA levels of TNFα and IL-1β compared to non-db EVs. Proteomic and gene enrichment analysis indicated pro-inflammatory characteristics of db EVs. Our results suggest a potential involvement of EC-derived Cldn5+ EVs in triggering inflammation, offering a novel mechanism involved and presenting a possible therapeutic avenue for DR. [ABSTRACT FROM AUTHOR]

Objective: To verify whether phosphoglycerate mutase 1 (PGAM1) allosteric inhibitor could enhance the sensitivity of colon cancer cells to oxaliplatin (OXA). Methods: OXA sensitivity and PGAM1 expression levels of four consensus molecular subgroups (CMS) of colon cancer cells were analyzed according to genomics of drug sensitivity in cancer (GDSC）and cancer cell line encyclopedia (CCLE) databases. CCK8 assay and Annexin V/PI staining were used to investigate the effect of PGAM1 allosteric inhibitor HKB99 on the proliferation and apoptosis of human colon cancer cell lines. CCK8 assay were performed to investigate the effect of HKB99 combined with OXA on the proliferation of colon cancer DLD-1 cells and cooperativity index was analyzed according to the result. Results: Firstly, GDSC and CCLE databases results showed that CMS1 and CMS4 colon cancer cells were less sensitive to OXA than CMS2 and CMS3 cells in terms of the sensitive cells proportion. Compared with, the expression of PGAM1 was higher in CMS1 and CMS4 cells than in CMS1 and CMS3 cells (P＜0. 05). HKB99 significantly inhibits the proliferation of SW480, SCUN2B and DLD-1 cells, with an IC50 of approximately 1μM. Annexin V/PI staining results showed that apoptosis of DLD-1 cells was significantly increased compared with Control group after HKB99 treatment (P＜0. 01). Results of CCK8 assay result showed that 1. 5μM HKB99 significantly enhanced the inhibitory effect of OXA on colon cancer DLD-1 cells compared with control group (IC50: 0. 37 vs 3. 26μM). The cooperativity index analysis result showed that the synergistic coefficient of OXA with HKB99 was less than 1 at the concentration between 1. 56μM and 25μM. Conclusions: In this study, we found that CMS1 and CMS4 colon cancer cells are less sensitive to OXA and higher level of PGAM1compared with CMS2 and CMS3 cells. An allosteric inhibitor of PGAM1, HKB99, enhances the sensitivity of colon cancer cells to OXA, which suggests that HKB99 may enhance the sensitivity of OXA chemotherapy and provide a potential treatment strategy for patients with individualized chemotherapy in colon cancer. [ABSTRACT FROM AUTHOR]

Proteostasis mechanisms, such as proteotoxic-stress response and autophagy, are increasingly recognized for their roles in influencing various cancer hallmarks such as tumorigenesis, drug resistance, and recurrence. However, the precise mechanisms underlying their coordination remain not fully elucidated. The aim of this study is to investigate the molecular interplay between Hsp70 and autophagy in lung adenocarcinoma cells and elucidate its impact on the outcomes of anticancer therapies in vitro. For this purpose, we utilized the human lung adenocarcinoma A549 cell line and genetically modified it by knockdown of Hsp70 or HSF1, and the H1299 cell line with knockdown or overexpression of Hsp70. In addition, several treatments were employed, including treatment with Hsp70 inhibitors (VER-155008 and JG-98), HSF1 activator ML-346, or autophagy modulators (SAR405 and Rapamycin). Using immunoblotting, we found that Hsp70 negatively regulates autophagy by directly influencing AMPK activation, uncovering a novel regulatory mechanism of autophagy by Hsp70. Genetic or chemical Hsp70 overexpression was associated with the suppression of AMPK and autophagy. Conversely, the inhibition of Hsp70, genetically or chemically, resulted in the upregulation of AMPK-mediated autophagy. We further investigated whether Hsp70 suppression-mediated autophagy exhibits pro-survival- or pro-death-inducing effects via MTT test, colony formation, CellTiter-Glo 3D-Spheroid viability assay, and Annexin/PI apoptosis assay. Our results show that combined inhibition of Hsp70 and autophagy, along with cisplatin treatment, synergistically reduces tumor cell metabolic activity, growth, and viability in 2D and 3D tumor cell models. These cytotoxic effects were exerted by substantially potentiating apoptosis, while activating autophagy via rapamycin slightly rescued tumor cells from apoptosis. Therefore, our findings demonstrate that the combined inhibition of Hsp70 and autophagy represents a novel and promising therapeutic approach that may disrupt the capacity of refractory tumor cells to withstand conventional therapies in NSCLC. [ABSTRACT FROM AUTHOR]

Rabies virus causes an estimated 59,000 annual fatalities worldwide and promising therapeutic treatments are necessary to develop. In this study, affinity tag-purification mass spectrometry was employed to delineate RABV glycoprotein and host protein interactions, and PDIA3/ERP57 was identified as a potential inhibitor of RABV infection. PDIA3 restricted RABV infection with follow mechanisms: PDIA3 mediated the degradation of RABV G protein by targeting lysine 332 via the selective macroautophagy/autophagy pathway; The PDIA3 interactor, AP3B1 (adaptor related protein complex 3 subunit beta 1) was indispensable in PDIA3-triggered selective degradation of the G protein; Furthermore, PDIA3 competitively bound with NCAM1/NCAM (neural cell adhesion molecule 1) to block RABV G, hindering viral entry into host cells. PDIA3 190-199 aa residues bound to the RABV G protein were necessary and sufficient to defend against RABV. These results demonstrated the therapeutic potential of biologics that target PDIA3 or utilize PDIA3 190-199 aa peptide to treat clinical rabies.Abbreviation: aa: amino acids; ANXA2: annexin A2; AP-MS: affinity tag purification-mass spectrometry; AP3B1: adaptor related protein complex 3 subunit beta 1; ATP6V1A: ATPase H+ transporting V1 subunit A; ATP6V1H: ATPase H+ transporting V1 subunit H; BafA1: bafilomycin A1; CHX: cycloheximide; co-IP: co-immunoprecipitation; DDX17: DEAD-box helicase 17; DmERp60: drosophila melanogaster endoplasmic reticulum p60; EBOV: Zaire ebolavirus virus; EV: empty vector; GANAB: glucosidase II alpha subunit; G protein: glycoprotein; GRM2/mGluR2: glutamate metabotropic receptor 2; HsPDIA3: homo sapiens protein disulfide isomerase family A member 3; IAV: influenza virus; ILF2: interleukin enhancer binding factor 2; KO: knockout; MAGT1: magnesium transporter 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MmPDIA3: mus musculus protein disulfide isomerase associated 3; NCAM1/NCAM: neural cell adhesion molecule 1; NGFR/p75NTR: nerve growth factor receptor; NGLY1: N-glycanase 1; OTUD4: OTU deubiquitinase 4; PDI: protein disulfide isomerase; PPIs: protein-protein interactions; RABV: rabies virus; RUVBL2: RuvB like AAA ATPase 2; SCAMP3: secretory carrier membrane protein 3; ScPdi1: Saccharomyces cerevisiae s288c protein disulfide isomerase 1; SLC25A6: solute carrier family 25 member 6; SQSTM1/p62: sequestosome 1; VSV: vesicular stomatitis virus. [ABSTRACT FROM AUTHOR]

Background: Acute myeloid leukemia (AML) is a heterogeneous ailment in both biological and clinical concepts. Numerous efforts have been devoted to discover natural compounds for combating cancer, which showed great potential in cancer management. Methylsulfonylmethane (MSM), an organosulfur dietary supplement, is utilized for improving various clinical conditions, particularly osteoarthritis. MSM can exert antitumor activity in a wide range of cancers. Objectives: The molecular mechanisms of action underlying antileukemic activity of MSM remain unclear. In this regard, we aimed to investigate the anticancer properties of MSM on human AML cell lines (U937 and HL60) with focus on underlying cell death mechanism. Methods: Anticancer activity of the MSM was examined employing MTT assay, Annexin V‐PE/7AAD staining, caspase3/7 activity test, and real‐time qPCR. Both cell lines were treated with different concentrations (50–400 mM) of MSM for 24 h. Pretreatment of the cells with a caspase inhibitor (i.e., Z‐VAD‐fmk) was performed for the assessment of apoptosis induction. Results: The results of MTT assay revealed that in both cell lines, the MSM markedly reduced cell viability in comparison to the control cells. Additionally, findings of Annexin V‐7AAD staining revealed that MSM induced apoptosis and activated caspase 3/7 in both cell lines markedly. Real‐time quantitative PCR results also supported the induction of apoptosis in AML cells. MSM altered the expression levels of various apoptotic genes (BAX, BAD, and BIM). Conclusion: Overall, our results indicated that MSM could induce apoptosis in AML cell lines in a dose‐dependent manner, which therefore could be utilized as an antileukemic agent. [ABSTRACT FROM AUTHOR]

Macrophages play pivotal roles in the regulation of inflammatory responses and tissue repair, making them a prime target for inflammation alleviation. However, the accurate and efficient macrophages targeting is still a challenging task. Motivated by the efficient and specific removal of apoptotic cells by macrophages efferocytosis, a novel biomimetic liposomal system called Effero‐RLP (Efferocytosis‐mediated Red blood cell hybrid Liposomes) is developed which incorporates the membrane of apoptotic red blood cells (RBCs) with liposomes for the purpose of highly efficient macrophages targeting. Rosiglitazone (ROSI), a PPARγ agonist known to attenuate macrophage inflammatory responses, is encapsulated into Effero‐RLP as model drug to regulate macrophage functions in DSS‐induced colitis mouse model. Intriguingly, the Effero‐RLP exhibits selective and efficient uptake by macrophages, which is significantly inhibited by the efferocytosis blocker Annexin V. In animal models, the Effero‐RLP demonstrates rapid recognition by macrophages, leading to enhanced accumulation at inflammatory sites. Furthermore, ROSI‐loaded Effero‐RLP effectively alleviates inflammation and protects colon tissue from injury in the colitis mouse model, which is abolished by deletion of macrophages from mice model. In conclusion, the study highlights the potential of macrophage targeting using efferocytosis biomimetic liposomes. The development of Effero‐RLP presents novel and promising strategies for alleviating inflammation. [ABSTRACT FROM AUTHOR]

AIM: To explore the influence of ethyl (2, 4, 6-trimethylbenzoyl) phenylphosphinate (TPOL) on cell apoptosis and its potential mechanism. METHODS: HEK293T cells sensitive to TPOL were treated with different concentrations of TPOL with or without exposure to light radiation, before treatment with various inhibitors, N-acetyl-L-cysteine (NAC), pifithrin-α and Z-DVED-FMK. Cell viability was measured by CCK-8 assay. Annexin V/propidium iodide staining was used to count the number of apoptotic cells. DCFH-DA staining was used to detect reactive oxygen species (ROS) levels, and JC-1 staining was used to assess mitochondrial membrane potential by flow cytometry. The expression of apoptosis-related proteins and cell cycle-regulated molecules was measured by Western blot. RESULTS: TPOL enhanced the apoptosis of HEK293T cells in a dose-dependent manner (P<0. 05), with a decrease in Bcl-2 and increases in Bax and cytochrome C (Cyto C), followed by up-regulation of activated caspase-9 and caspase-3, and the cleavage of PARP (P<0. 05). The TPOL-enhanced cleavage of caspase-3 and PARP was rescued by Z-DVED-FMK (P<0. 01). TPOL also led to a rapid increase in ROS, a reduction in mitochondrial membrane potential, and the release of Cyto C (P< 0. 01), all of which could be reversed by the ROS scavenger NAC. Moreover, the TPOL-caused alterations in p21, p27, Rb, and CDK2 were also recovered by the p53 inhibitor pifithrin-α(P<0. 05). The TPOL-induced changes in Bax, Bcl-2, cleaved caspase-9, activated caspase-3, and cleaved PARP were subsequently rescued by pretreatment with pifithrin-α (P<0. 05). CONCLUSION: TPOL can induce cellular apoptosis with ROS-mediated mitochondrial membrane damage through the activation of a ROS-dependent p53/p21/p27/Rb/Bax/Cyto C/caspase-mediated signal axis. [ABSTRACT FROM AUTHOR]

Despite progress in the management of patients with retinal vascular and degenerative diseases, there is still an unmet clinical need for safe and effective therapeutic options with novel mechanisms of action. Recent mechanistic insights into the pathogenesis of retinal diseases with a prominent vascular component, such as retinal vein occlusion (RVO), diabetic retinopathy (DR) and wet age-related macular degeneration (AMD), may open up new treatment paradigms that reach beyond the inhibition of vascular endothelial growth factor (VEGF). Phosphatidylserine (PS) is a novel lipid target that is linked to the pathophysiology of several human diseases, including retinal diseases. PS acts upstream of VEGF and complement signaling pathways. Annexin A5 is a protein that targets PS and inhibits PS signaling. This review explores the current understanding of the potential roles of PS as a target and Annexin A5 as a therapeutic. The clinical development status of Annexin A5 as a therapeutic and the potential utility of PS-Annexin A5 as a theranostic pairing in retinal vascular conditions in particular is described. [ABSTRACT FROM AUTHOR]

Simple Summary: The greater wax moth, Galleria mellonella, is a significant pest in apiculture. This investigation explores the individual and combined effects of the ectoparasite Bracon hebetor (B.h.) and the entomopathogenic nematode Steinernema carpocapsae (S.c.) on G. mellonella larvae. We found that control larvae exhibited higher glutathione peroxidase (GPx) and glutathione S-transferase (GST) activities compared to those treated with B.h., S.c., or the B.h. + S.c. combination. Conversely, malondialdehyde (MDA) levels displayed an opposite trend. Superoxide dismutase (SOD) activity was reduced in the B.h. and S.c. groups but significantly higher in the combined treatment. Cytochrome P450 activity increased in response to parasitism by B. hebetor. The Annexin V-FITC assay revealed decreased cell viability in parasitized groups (B.h. 79.4%, S.c. 77.3%, B.h. + S.c. 70.1%) compared to controls. DNA damage analysis demonstrated significant differences between groups, and SEM observations confirmed severe cuticle abnormalities or malformations in G. mellonella larvae. The greater wax moth, Galleria mellonella, is a significant pest in apiculture and a well-established model organism for immunological and ecotoxicological studies. This investigation explores the individual and combined effects of the ectoparasite Bracon hebetor (B.h.) and the entomopathogenic nematode Steinernema carpocapsae (S.c.) on G. mellonella larvae. We evaluated the activity of oxidative stress enzymes, including superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione S-transferase (GST), malondialdehyde (MDA) levels, cytochrome P450 activity, cell viability using Annexin V-FITC, DNA damage via comet assay, and larval morphology through scanning electron microscopy (SEM). Control larvae exhibited higher GPx and GST activities compared to those treated with B.h., S.c., or the B.h. + S.c. combination. Conversely, MDA levels displayed the opposite trend. SOD activity was reduced in the B.h. and S.c. groups but significantly higher in the combined treatment. Cytochrome P450 activity increased in response to parasitism by B. hebetor. The Annexin V-FITC assay revealed decreased cell viability in parasitized groups (B.h. 79.4%, S.c. 77.3%, B.h. + S.c. 70.1%) compared to controls. DNA damage analysis demonstrated significant differences between groups, and SEM observations confirmed severe cuticle abnormalities or malformations in G. mellonella larvae. These findings highlight the complex interactions between B. hebetor, S. carpocapsae, and their host, G. mellonella. Additionally, they illuminate the intricate physiological responses triggered within the host larvae. [ABSTRACT FROM AUTHOR]

Background: GP‐2250, a novel analog of taurultam (TRLT), has emerged as a potent anti‐neoplastic drug; however, the mechanisms underlying its effects are not well understood. Here, we investigated the mechanism of action and the biological effects of GP‐2250 using in vitro and in vivo models. Methods: We carried out a series of in vitro (MTT assay, Annexin V/PI assay, colony formation assay, reverse‐phase protein array [RPPA], and HRLC/IC analysis) to determine the biological activity of GP‐2250 and investigate the mechanism of action. In vivo experiments were carried out to determine the therapeutic efficacy of GP‐2250 alone and in combination with standard‐of‐care drugs (e.g., paclitaxel, cisplatin, topotecan, and poly ADP‐ribose polymerase [PARP] inhibitors). Results: We investigated the cytotoxic effect of GP‐2250 in 10 ovarian cancer cell lines and found GP‐2250 combined with a PARP inhibitor had the greatest synergy. RPPA revealed that GP‐2250 inhibited hypoxia‐inducible factor‐1α, AKT, and mammalian target of rapamycin (mTOR) activation and expression. High‐resolution mass spectrometry revealed that hexokinase2 activity and protein expression were significantly reduced by GP‐2250 exposure. Furthermore, GP‐2250 reduced glycolysis and ATP synthesis in cancer cells. An in vivo pharmacodynamic experiment using the OVCAR8 mouse model demonstrated that 500 mg/kg GP‐2250 was effective in downregulating AKT and mTOR activation and expression. In the in vivo therapy experiment using an orthotopic mouse model, a combination of GP‐2250 with either PARP inhibitors or bevacizumab showed a significant reduction of tumor weights and nodules compared to those treated with a vehicle, control IgG groups, or monotherapy groups. Conclusions: Taken together, our data indicate that GP‐2250 exerts profound effects on tumor metabolism and, in combination with PARP inhibitors or bevacizumab, showed promising anti‐tumor efficacy. These findings could have implications for the clinical development of GP‐2250. [ABSTRACT FROM AUTHOR]

Colorectal cancer (CRC) is the third most common cancer in the world. Despite considerable improvements in the treatment of this cancer, further research to discover novel and more effective agents is ongoing. In this study, possible cytotoxic and apoptotic properties of six benzothiazolopyrimidine derivatives were studied. To assess the IC50 values of these agents, MTT assay was performed on HCT 116, CT26, and NIH/3T3 cells. Moreover, cell death mechanism induced by studied compounds was evaluated by PI/annexin V staining. Then, based on molecular docking results and in vitro experiments, the compounds with the highest anticancer properties were further analyzed in vivo in a mouse model of CRC. MTT results indicated that BTP(1) and BTP(4) had the highest selective cytotoxicity on colorectal cancer cells. Furthermore, flow cytometry results demonstrated a considerable increase in the percentage of the early apoptotic cells in BTP(1)‐ and BTP(4)‐treated groups. In vivo studies confirmed the antitumor properties of the two compounds by a significant regression in tumor size of BTP(1)‐ and BTP(4)‐treated mice compared to control groups. Histopathological examination of tumor tissues showed an increased number of apoptotic cells in these two groups compared to the control animals. Additionally, hematoxylin and eosin staining of the spleen and liver of treated mice did not exhibit considerable tissue damage. Thus, BTP(1) and BTP(4) can be considered promising agents in the treatment of colorectal cancer, although further experiments are required to assess their mechanism of action before their application in clinical studies. [ABSTRACT FROM AUTHOR]

Background: MutT homolog 1 (MTH1) sanitizes oxidized dNTP pools to promote the survival of cancer cells and its expression is frequently upregulated in cancers. Polyubiquitination stabilizes MTH1 to facilitate the proliferation of melanoma cells, suggesting the ubiquitin system controls the stability and function of MTH1. However, whether ubiquitination regulates MTH1 in gastric cancers has not been well defined. This study aims to investigate the interaction between MTH1 and a deubiquitinase, USP9X, in regulating the proliferation, survival, migration, and invasion of gastric cancer cells. Methods: The interaction between USP9X and MTH1 was evaluated by co-immunoprecipitation (co-IP) in HGC-27 gastric cancer cells. siRNAs were used to interfere with USP9X expression in gastric cancer cell lines HGC-27 and MKN-45. MTT assays were carried out to examine the proliferation, propidium iodide (PI) and 7-AAD staining assays were performed to assess the cell cycle, Annexin V/PI staining assays were conducted to examine the apoptosis, and transwell assays were used to determine the migration and invasion of control, USP9X-deficient, and USP9X-deficient plus MTH1-overexpressing HGC-27 and MKN-45 gastric cancer cells. Results: Co-IP data show that USP9X interacts with and deubiquitinates MTH1. Overexpression of USP9X elevates MTH1 protein level by downregulating its ubiquitination, while knockdown of USP9X has the opposite effect on MTH1. USP9X deficiency in HGC-27 and MKN-45 cells causes decreased proliferation, cell cycle arrest, extra apoptosis, and defective migration and invasion, which could be rescued by excessive MTH1. Conclusion: USP9X interacts with and stabilizes MTH1 to promote the proliferation, survival, migration and invasion of gastric cancer cells. [ABSTRACT FROM AUTHOR]

Introduction: Exosomes produced by the protozoan parasite Leishmania (LeishEXO) are well-established drivers of virulence, though mechanisms underlying their exacerbation of experimental leishmaniasis remain elusive. Expression of Annexin A1 (ANXA1), a protein implicated in exosome-mediated pathologies and viral internalization, has been shown to correlate with cutaneous leishmaniasis severity. Given ANXA1's regulation of myeloid cells - the canonical hosts for Leishmania - we studied the potential role of ANXA1 and its receptors FPR1/2 in exerting LeishEXO's effects. Methods: Murine and in vitro ANXA1-/- models were used to study the generation of protective TH1 responses during experimental L. major infection with and without LeishEXO. Recruitment of inflammatory cells was assessed using a peritoneal cell recruitment assay and immunophenotyping, and production of inflammatory mediators was measured using a cytokine and chemokine array. Treatment of experimental models with FPR2 antagonist WRW4 and FPR1/2 agonist WKYMVm was used to delineate the role of the FPR/ANXA1 axis in LeishEXO-mediated hyperpathogenesis. Results: We established that ANXA1 deficiency prohibits LeishEXO-mediated pathogenesis and myeloid cell infection, with minimal alterations to adaptive and innate immune phenotypes. FPR2 blockade with WRW4 similarly inhibited leishmanial hyperpathogenesis, while direct activation of FPRs with WKYMVm enhanced infection and recapitulated the LeishEXO-mediated phenotype. This research describes LeishEXO's utilization of the ANXA1/FPR axis to facilitate parasitic internalization and pathogenesis, which may be leveraged in the development of therapeutics for leishmaniasis. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a well-documented bacterial contaminant in platelet concentrates (PCs), a blood component used to treat patients with platelet deficiencies. This bacterium can evade routine PC culture screening and cause septic transfusion reactions. Here, we investigated the gene expression modulation within the PC niche versus trypticase soy media (TSB) of S. aureus CBS2016-05, a strain isolated from a septic reaction, in comparison to PS/BAC/317/16/W, a strain identified during PC screening. RNA-seq analysis revealed upregulation of the capsule biosynthesis operon (capA-H), surface adhesion factors (sasADF), clumping factor A (clfA), protein A (spa), and anaerobic metabolism genes (pflAB, nrdDG) in CBS2016-05 when grown in PCs versus TSB, implying its enhanced pathogenicity in this milieu, in contrast to the PS/BAC/317/16/W strain. Furthermore, we investigated the impact of S. aureus CBS2016-05 on platelet functionality in spiked PCs versus non-spiked PC units. Flow cytometry analyses revealed a significant decrease in glycoprotein (GP) IIb (CD41) and GPIbα (CD42b) expression, alongside increased P-selectin (CD62P) and phosphatidylserine (annexin V) expression in spiked PCs compared to non-spiked PCs (p = 0.01). Moreover, spiked PCs exhibited a drastic reduction in MitoTrack Red FM and Calcein AM positive platelets (87.3% vs. 29.4%, p = 0.0001 and 95.4% vs. 24.7%, p = 0.0001) in a bacterial cell density manner. These results indicated that S. aureus CBS2016-05 triggers platelet activation and apoptosis, and compromises mitochondrial functionality and platelet viability, in contaminated PCs. Furthermore, this study enhanced our understanding of the effects of platelet-bacteria interactions in the unique PC niche, highlighting S. aureus increased pathogenicity and deleterious effect on platelet functionality in a strain specific manner. Our novel insights serve as a platform to improve PC transfusion safety. [ABSTRACT FROM AUTHOR]

Background. Still, cancer remains to be one of the main causes of death globally. Sesquiterpene lactones (SLs) appeared to have remarkable pharmacologic properties, particularly as anticancer. This study evaluated the cytotoxicity and apoptogenic potential of Janerin and its underlying mechanism in HL60, THP‐1, and Jurkat leukemia cell lines vs. normal peripheral blood mononuclear cells (PBMCs). Methods. The viability of leukemia and PBMCs following treatment with Janerin (2.5–20 μM) and doxorubicin (2 μM, as the positive control) for 48 h was determined via the resazurin assay. The apoptotic cells were determined by annexin V and propidium iodide test. Also, the genes expressions involved in apoptosis were detected by real‐time PCR. Results. Janerin reduced cell viability in leukemia cells in a dose‐dependent manner with no significant toxicity toward normal PBM cells. Janerin significantly increased apoptosis in leukemia cells after 48 h of treatment. In these cells, the expressions of p21, p53, CASP3, CASP9, and Bax/Bcl2 ratio were significantly elevated, whereas CASP8 remained unchanged (p < 0.01). It was suggested that the intrinsic pathway was the mechanism by which Janerin induced apoptosis in HL60, THP‐1, and Jurkat leukemia cells in a time‐ and dose‐dependent manner. Conclusion. The findings imply that Janerin might be a suitable substitute for doxorubicin in leukemia patients. [ABSTRACT FROM AUTHOR]

In recent years, in the development of emerging immunotherapy, B7‐H3 is also termed as CD276 and has become a novel chimeric antigen receptor (CAR)‐T target against glioma and other tumours, and aroused extensive attention. However, B7‐H3 has three isoforms (2, 3 and 4Ig) with the controversial expression and elusive function in tumour especially glioma. The current study mainly focuses on the regulatory factors and related mechanisms of generation of different B7‐H3 isoforms. First, we have determined that 2Ig is dominant in glioma with high malignancy, and 4Ig is widely expressed, whereas 3Ig shows negative expression in all glioma. Next, we have further found that RNA binding protein annexin A2 (ANXA2) is essential for B7‐H3 isoform maintenance, but fail to determine the choice of 4Ig or 2Ig. RNA methyltransferase NOP2/Sun RNA methyltransferase 2 (NSUN2) and 5‐methylcytosine reader Y‐box binding protein 1 (YBX1) facilitate the production of 2Ig. Our findings have uncovered a series of factors (ANXA2/NSUN2/YBX1) that can determine the alternative generation of different isoforms of B7‐H3 in glioma. Our result aims to help peers gain a clearer understanding of the expression and regulatory mechanisms of B7H3 in tumour patients, and to provide better strategies for designing B7H3 as a target in immunotherapy. [ABSTRACT FROM AUTHOR]

Simple Summary: Glucotoxicity is a condition that leads to β-cell death, driving a decrease in insulin secretion and leading to hyperglycemia and, eventually, diabetes. Here, we investigate whether connexin Cx30.2 could protect against glucotoxicity-induced apoptosis in β cells. We found that RIN-m5F β cells exposed to high glucose exhibited increased Cx30.2 protein expression. Interestingly, decreased expression of Cx30.2 by specific siRNAs resulted in augmented β-cell death, suggesting that Cx30.2 has an important role in maintaining cell survival during glucotoxicity. Overall, Cx30.2 could be an attractive pharmacological target to avoid high glucose-induced β-cell apoptosis. Glucotoxicity may exert its deleterious effects on pancreatic β-cell function via a myriad of mechanisms, leading to impaired insulin secretion and, eventually, type 2 diabetes. β-cell communication requires gap junction channels to be present among these cells. Gap junctions are constituted by transmembrane proteins of the connexins (Cxs) family. Two Cx genes have been identified in β cells, Cx36 and Cx30.2. We have found evidence that the glucose concentration on its own is sufficient to regulate Cx30.2 gene expression in mouse islets. In this work, we examine the involvement of the Cx30.2 protein in the survival of β cells (RIN-m5F). Methods: RIN-m5F cells were cultured in 5 mM D-glucose (normal) or 30 mM D-glucose (high glucose) for 24 h. Cx30.2 siRNAs was used to downregulate Cx30.2 expression. Apoptosis was measured by means of TUNEL, an annexin V staining method, and the cleaved form of the caspase-3 protein was determined using Western blot. Results: High glucose did not induce apoptosis in RIN-m5F β cells after 24 h; interestingly, high glucose increased the Cx30.2 total protein levels. Moreover, this work found that the downregulation of Cx30.2 expression in high glucose promoted apoptosis in RIN-m5F cells. Conclusion: The data suggest that the upregulation of Cx30.2 protects β cells from hyperglycemia-induced apoptosis. Furthermore, Cx30.2 may be a promising avenue of therapeutic investigation for the treatment of glucose metabolic disorders. [ABSTRACT FROM AUTHOR]

Objective and Design: Annexin A1 (ANXA1) plays a role in maintaining intestinal hemostasis, especially following mucosal inflammation. The published data about ANXA1 was derived from experimental animal models where there is an overlapping between epithelial and immune cells. There is no in vitro gut epithelial model that can assess the direct effect of ANXA1 on the gut epithelium. Methods: We developed high-throughput stem-cell-based murine epithelial cells and bacterial lipopolysaccharides (LPS) were used to induce inflammation. The impact of ANXA1 and its functional part (Ac2-26) was evaluated in the inflamed model. Intestinal integrity was assessed by the transepithelial electrical resistance (TEER), and FITC-Dextran permeability. Epithelial junction proteins were assessed using confocal microscopy and RT-qPCR. Inflammatory cytokines were evaluated by RT-qPCR and ELISA. Results: LPS challenge mediated a damage in the epithelial cells as shown by a drop in the TEER and an increase in FITC-dextran permeability; reduced the expression of epithelial junctional proteins (Occludin, ZO-1, and Cadherin) and increased the expression of the gut leaky protein, Claudin − 2. ANXA1 and Ac2-26 treatment reduced the previous damaging effects. In addition, ANXA1 and Ac2-26 inhibited the inflammatory responses mediated by the LPS and increased the transcription of the anti-inflammatory cytokine, IL-10. Conclusion: ANXA1 and Ac2-26 directly protect the epithelial integrity by affecting the expression of epithelial junction and inflammatory markers. The inflamed gut model is a reliable tool to study intestinal inflammatory diseases, and to evaluate the efficacy of potential anti-inflammatory drugs and the screening of new drugs that could be candidates for inflammatory bowel disease. [ABSTRACT FROM AUTHOR]

Major facilitator superfamily domain containing 12 (MFSD12) regulates lysosomal cysteine import and promotes the proliferation and survival of melanoma cells. However, the expression and function of MFSD12 in other cancers, particularly in lung cancer, remain unclear. The expression of MFSD12 across various types of cancers and corresponding control tissues was examined using TIMER. MFSD12 expression in lung adenocarcinoma (LUAD) and its correlation with distinct clinicopathological features of LUAD patients were analyzed with UALCAN. The correlation between MFSD12 expression and survival of LUAD patients was assessed using the R package, survival, and the relationship between MFSD12 expression and immune infiltration status in LUAD was investigated using CIBERSORT. In addition, MFSD12 expression was knocked down in PC9 LUAD cells and their proliferation, capacity for expansion, cell cycle, apoptosis, and migration/invasion were evaluated through CCK-8 assays, colony formation assays, 7-AAD staining, Annexin V/PI staining, and Transwell assays, respectively. The stemness of these PC9 cells was determined through Western blotting, flow cytometry, and tumor sphere formation assays. MFSD12 mRNA levels were significantly elevated in multiple types of cancers, including LUAD. MFSD12 expression was also positively correlated with cancer stage, nodal metastasis, and infiltration of various immune cells in LUAD, and high MFSD12 levels predicted poor survival among LUAD patients. Knockdown of MFSD12 in PC9 cells resulted in decreased proliferation, attenuated colony formation capacity, cell cycle arrest, elevated apoptosis, impaired migration/invasion, and reduced stemness in PC9 cells. MFSD12 is an oncogene in LUAD. [ABSTRACT FROM AUTHOR]

Annexins, a group of Ca2+-dependent phospholipid-binding proteins, exert diverse roles in neuronal development, normal central nervous system (CNS) functioning, neurological disorders, and CNS tumors. This paper reviews the roles of individual annexins (A1-A13) in these contexts. Annexins possess unique structural and functional features, such as Ca2+-dependent binding to phospholipids, participating in membrane organization, and modulating cell signaling. They are implicated in various CNS processes, including endocytosis, exocytosis, and stabilization of plasma membranes. Annexins exhibit dynamic roles in neuronal development, influencing differentiation, proliferation, and synaptic formation in CNS tissues. Notably, annexins such as ANXA1 and ANXA2 play roles in apoptosis and blood-brain barrier (BBB) integrity. Neurological disorders, including Alzheimer's disease, multiple sclerosis, and depression, involve annexin dysregulation, influencing neuroinflammation, blood-brain barrier integrity, and stress responses. Moreover, annexins contribute to the pathogenesis of CNS tumors, either promoting or suppressing tumor growth, angiogenesis, and invasion. Annexin expression patterns vary across different CNS tumor types, providing potential prognostic markers and therapeutic targets. This review underscores the multifaceted roles of annexins in the CNS, highlighting their importance in normal functioning, disease progression, and potential therapeutic interventions. [ABSTRACT FROM AUTHOR]

Background: The intestinal wound healing process is a complex event of three overlapping phases: exudative, proliferative, and remodeling. Although some mechanisms have been extensively described, the intestinal healing process is still not fully understood. There are some similarities but also some differences compared to other tissues. The aim of this systematic review was to summarize all studies with knockout (KO) experimental models in bowel anastomoses, underline any recent knowledge, and clarify further the cellular and molecular mechanisms of the intestinal healing process. A systematic review protocol was performed. Materials and methods: Medline, EMBASE, and Scopus were comprehensively searched. Results: a total of eight studies were included. The silenced genes included interleukin-10, the four-and-one-half LIM domain-containing protein 2 (FHL2), cyclooxygenase-2 (COX-2), annexin A1 (ANXA-1), thrombin-activatable fibrinolysis inhibitor (TAFI), and heparin-binding epidermal growth factor (HB-EGF) gene. Surgically, an end-to-end bowel anastomosis was performed in the majority of the studies. Increased inflammatory cell infiltration in the anastomotic site was found in IL-10-, annexin-A1-, and TAFI-deficient mice compared to controls. COX-1 deficiency showed decreased angiogenesis at the anastomotic site. Administration of prostaglandin E2 in COX-2-deficient mice partially improved anastomotic leak rates, while treatment of ANXA1 KO mice with Ac2-26 nanoparticles reduced colitis activity and increased weight recovery following surgery. Conclusions: our findings provide new insights into improving intestinal wound healing by amplifying the aforementioned genes using appropriate gene therapies. Further research is required to clarify further the cellular and micromolecular mechanisms of intestinal healing. [ABSTRACT FROM AUTHOR]

Purpose: Nasopharyngeal carcinoma (NPC) is an aggressive head and neck disease with a high incidence of distant metastases. Enlargeosomes are cytoplasmic organelles marked by, desmoyokin/AHNAK. This study aimed to evaluate the expression of AHNAK in NPC and its effect on enlargeosomes and to investigate the correlation between AHNAK expression levels and clinical NPC patient characteristics. Methods: Primary nasopharyngeal carcinoma (NPC) and NPC specimens were evaluated by analyzing public data, and immunohistochemistry. Systematic in vitro and in vivo experiments were performed using different NPC-derived cell lines and mouse models. Results: In this study, we detected AHNAK and Annexin A2(ANXA2), a protein coating the surface of enlargeosomes, in NPC samples. We found that AHNAK was down-regulated. Down-regulation of AHNAK was associated with poor overall survival in NPC patients. Moreover, transcription factor FOSL1-mediated transcriptional repression was responsible for the low expression of AHNAK by recruiting EZH2. Whereas Annexin A2 was upregulated in human NPC tissues. Upregulation of Annexin A2 was associated with lymph node metastasis and distant metastasis in NPC patients. Functional studies confirmed that silencing of AHNAK enhanced the growth, invasion, and metastatic properties of NPC cells both in vitro and in vivo. In terms of mechanism, loss of AHNAK led to an increase of annexin A2 protein level in NPC cells. Silencing ANXA2 restored NPC cells' migrative and invasive ability upon loss of AHNAK. Conclusion: Here, we report AHNAK as a tumor suppressor in NPC, which may act through annexin A2 oncogenic signaling in enlargeosome, with potential implications for novel approaches to NPC treatment. [ABSTRACT FROM AUTHOR]

Annexin A1 (AnxA1) is a glucocorticoid-inducible protein and an important endogenous modulator of inflammation. However, its effect in the endometrial microenvironment is poorly explained. This study aimed to evaluate the role of endogenous AnxA1 in an endometritis mouse model induced by lipopolysaccharide (LPS). Female C57BL/6 wild-type (WT) and AnxA1−/− mice were divided into two groups: SHAM and LPS. To induce endometritis, mice received a vaginal infusion of 50 μL of LPS (1 mg/mL) dissolved in phosphate-buffered saline. After 24 h, the mice were euthanized, and blood and uteri samples were collected. The endometrium inflammatory scores were significantly increased in the LPS-treated group. AnxA1−/− mice from the LPS group demonstrated a significant increase in the number of degranulated mast cell levels compared to AnxA1−/− SHAM mice. The Western blotting analysis revealed that a lack of AnxA1 promoted the upregulation of NLRP3 and pro-IL-1β in the acute endometritis animal model compared to WT LPS animals. LPS-induced endometritis increased the number of blood peripheral leukocytes in both WT and AnxA1−/− mice compared with SHAM group mice (p < 0.001). AnxA1−/− mice also showed increased plasma levels of IL-1β (p < 0.01), IL-6, IL-10, IL-17, and TNF-α (p < 0.05) following LPS-induced endometritis. In conclusion, a lack of endogenous AnxA1 exacerbated the inflammatory response in an endometritis model via NLRP3 dysregulation, increased uterine mast cell activation, and plasma pro-inflammatory cytokine release. [ABSTRACT FROM AUTHOR]

Background: Throughout the storage of blood, the red cells undergo alterations known as "storage lesions," which involve shape changes and the formation of microparticles (MPs). Studies of the formation of red cell microparticles (RMPs) emphasize the prospective application of RMPs as a quality control measure in the preparation and storage of blood components in the future. In the present study, twenty packed RBC units in citrate phosphate dextrose adenine-1 (CPDA1) were collected from volunteers and stored for 35 days. Over 35 days of storage, samples were collected at six distinct time points weekly and evaluated for the presence of RMPs. MPs were separated by the ultracentrifugation method. Electron microscopy was used to characterize the morphology and size of the isolated microparticles, and flow cytometry was performed to determine the percentage of RMPs that expressed glycophorin A (CD235a) and Annexin V antigens. RMPs' procoagulant activity (PCA) was assessed using a plasma recalcification test. RMP concentration in accordance with ABO blood grouping was assessed by using various types of donated blood groups. Results: RMPs progressively increased over storage. The procoagulant activity (PCA) exhibited a significant increase during storage, as evidenced by a shorter plasma recalcification time (P value = 0.001). A significant negative correlation (P value = 0.001) between plasma recalcification time and Annexin V-positive microparticles, as well as a dual-positive Annexin V/CD235a population, was identified, indicating a strong correlation between the direct quantitative assay by flowcytometry and the functional assay through the PCA. Conclusion: RMPs increase on storage with increased PCA. Finding ways to reduce these microparticles in packed RBC units is crucial for reducing the risk of transfusion-related coagulopathy. [ABSTRACT FROM AUTHOR]

Acute liver injury, there is a risky neurological condition known as hepatic encephalopathy (HE). Herbacetin is a glycosylated flavonoid with many pharmacological characteristics. The purpose of this study was to assess the ability of herbacetin to protect against the cognitive deficits associated with thioacetamide (TAA) rat model and delineate the underlying behavioral and pharmacological mechanisms. Rats were pretreated with herbacetin (20 and 40 mg/kg) for 30days. On 30th day, the rats were injected with TAA (i.p. 350 mg/kg) in a single dose. In addition to a histpathological studies, ultra-structural architecture of the brain, liver functions, oxidative stress biomarkers, and behavioral tests were evaluated. Compared to the TAA-intoxicated group, herbacetin improved the locomotor and cognitive deficits, serum hepatotoxicity indices and ammonia levels. Herbacetin reduced brain levels of malodialdeyde, glutamine synthetase (GS), tumor necrosis factor- alpha (TNF-α), interleukin 1 B (IL-1β), annexin v, and increased brain GSH, Sirtuin 1 (SIRT1), and AMP-activated kinase (AMPK) expression levels. Also, herbacetin improve the histopathological changes and ultra- structure of brain tissue via attenuating the number of inflammatory and apoptotic cells. Herbacetin treatment significantly reduced the toxicity caused by TAA. These findings suggest that herbacetin might be taken into account as a possible neuroprotective and cognitive enhancing agent due to its ability to reduce oxidative stress, inflammation and apoptosis associated with TAA. [ABSTRACT FROM AUTHOR]

Opuntia ficus-indica (L.) Mill. belongs to the Cactaceae family and the genus Opuntia; it is a succulent plant that adapts to extreme climatic conditions. The aerial part of the plant consists of the cladodes, morphological changes of branches that appear green, are covered with thorns, and are essential to reduce excessive perspiration of water. The composition of cladodes is very varied, and the main constituents are water, fibers, polysaccharides, proteins, fatty acids, vitamins, sterols, minerals, and polyphenols. Polyphenols are responsible for many beneficial activities for human health, such as antioxidant, anti-inflammatory, anticancer, and nutritional properties. The purpose of this manuscript was to compare the properties of cladodes belonging to the same plant but with different stages of maturity. Relative extracts were tested both in vitro and on a cell line and antioxidant and anti-apoptotic properties were found. The antioxidant activity was tested by the Oxygen Radical Absorbance Capacity (ORAC) test, the 1,1-diphenyl-2-picrylhydrazil (DPPH) test, and the measurement of cellular accumulation of reactive oxygen species (ROS). Anti-apoptotic activity was evaluated by the annexin/PI assay and measurement of caspases 9 and 3 expression. The results obtained showed that the extracts considered possess antioxidant and anti-apoptotic properties. However, the different stages of maturity of cladodes are essential for the performance of both functions. In addition, important variations were made in the dissolution of the extracts that brought greater safety in their use. In conclusion, this manuscript provides further information on cladodes of Opuntia ficus-indica, which can be used as adjuvants in many human pathologies. [ABSTRACT FROM AUTHOR]

Background: Colorectal cancer (CRC) is among the most common cancers worldwide. In spite of promising improvements, it has a relatively poor prognosis. Therefore, there is a crucial need for discovering novel and efficient therapies. This study aimed to investigate the potential of phycocyanin as a therapeutic agent for CRC, considering its antioxidant and anti-inflammatory properties. Methods: The study used HT-29 and HUVEC cell lines. Cells were exposed to different concentrations of phycocyanin for 24 h. The cytotoxic effects of the phycocyanin on HUVEC and HT-29 cells were evaluated using MTT. Following the treatment, RNA extraction was conducted using TRI reagent. Afterward, real-time PCR (RT-PCR) was performed on Bcl-2, Bax, Akt, PTEN, and ß-actin to study their expression levels. The phycocyanin-treated cells were also stained with Annexin V-FITC to detect any apoptosis and necrosis. Results: Following the MTT assay, the number of living HT-29 cells significantly decreased, with no toxic effect on HUVEC cells. RT-PCR results indicated a significant increase in PTEN and Bax expressions in HT-29 cells due to treatment with phycocyanin, while there was no change in the expression of Bcl-2 and AKT. Flow cytometry on HT-29 cells showed induction of apoptosis and necrosis in 8.79% and 2.9% of cells, respectively, while no necrosis or apoptosis was observed in normal cells. Conclusion: Phycocyanin has a potential for treating colon cancer by affecting AKT/PTEN and Bax/Bcl- 2 and inducing apoptotic genes (Bax and PTEN) while having minimal side effects. Therefore, it could be a promising therapeutic agent for colon cancer. [ABSTRACT FROM AUTHOR]

Siniperca chuatsi rhabdovirus (SCRV) is a major strain of viral fish virus resulting in multiple transmissions and devastating damage in aquaculture. Currently, there are no available approved therapeutics. In this study, we screened and identified a novel curcumin analog (EF-24) for evaluating its in vitro anti-SCRV properties and potential molecular mechanisms. Present results demonstrated that EF-24 could strongly delay the occurrence of cytopathic effects (CPEs) in epithelioma papulosum cyprinid cells (EPCs) and inhibit SCRV replication and viral nucleoprotein expression in the early stages of infection by the time-of-addition assay. Furthermore, flow cytometry analysis after Annexin V-FITC/PI double staining and immunofluorescence microscopy observation after JC-1 incubation showed that EF-24 downregulated cell mitochondrial apoptosis induced by SCRV. The enzymatic activities of caspase-3 and caspase-9 were also reduced after EF-24 treatment, indicating that EF-24 may protect cells from SCRV infection by decreasing mitochondrial intrinsic apoptosis in infected cells. Collectively, we demonstrated for the first time that the curcumin analog EF-24 possesses antiviral ability against SCRV, suggesting its potential for effective control of fish rhabdovirus spreading. [ABSTRACT FROM AUTHOR]

The unclear pathogenesis of chronic itch originating from several systemic disorders poses challenges to clinical intervention. Recent studies recapitulate the spinal neurocircuits associated with neuroinflammation and synaptic plasticity responsible for pruriceptive sensations. The resolution of nociception and inflammation by Annexin 1 (ANXA1) has been identified. Given that pain and itch share many neural mechanisms, we employed two mice models of chronic itch to study the underlying targets and therapeutic potential of ANXA1, comprising allergic contact dermatitis-induced itch and cholestatic itch. Herein, we report that spinal expression of ANXA1 is down-regulated in mice with dermatitis-induced itch and cholestatic itch. Repetitive injections of ANXA1-derived peptide Ac2-26 (intrathecal, 10 μg) reduce itch-like scratching behaviors following dermatitis and cholestasis. Single exposure to Ac2-26 (intrathecal, 10 μg) alleviates the established itch phenotypes. Moreover, systemic delivery of Ac2-26 (intravenous, 100 μg) is effective against chronic dermatitis-induced itch and cholestatic itch. Strikingly, Ac2-26 therapy inhibits transferrin receptor 1 over-expression, iron accumulation, cytokine IL-17 release and the production of its receptor IL-17R, as well as astrocyte activation in the dorsal horn of spinal cord in mouse with dermatitis and cholestasis. Pharmacological intervention with iron chelator deferoxamine impairs chronic itch behaviors and spinal iron accumulation after dermatitis and cholestasis. Also, spinal IL-17/IL-17R neutralization attenuates chronic itch. Taken together, this current research indicates that ANXA1 protects against the beginning and maintenance of long-term dermatitis-induced itch and cholestatic itch, which may occur via the spinal suppression of IL-17-mediated neuroinflammation, astrocyte activation and iron overload. [ABSTRACT FROM AUTHOR]

• Spatial architecture and composition of the ECM differ across genetically distinct forms of muscular dystrophy, especially with respect to Annexin A6 protein deposition. • Matrix from dystrophin-mediated muscular dystrophy inhibits myoblast movement. • Matrix from dysferlin-deficient muscular dystrophy promotes myoblast motility and fusion potential. • Annexin A6 was sufficient to enhance myoblast motility. • Recombinant annexin A6 promotes myoblast differentiation. Extracellular matrix (ECM) pathologic remodeling underlies many disorders, including muscular dystrophy. Tissue decellularization removes cellular components while leaving behind ECM components. We generated "on-slide" decellularized tissue slices from genetically distinct dystrophic mouse models. The ECM of dystrophin- and sarcoglycan-deficient muscles had marked thrombospondin 4 deposition, while dysferlin-deficient muscle had excess decorin. Annexins A2 and A6 were present on all dystrophic decellularized ECMs, but annexin matrix deposition was excessive in dysferlin-deficient muscular dystrophy. Muscle-directed viral expression of annexin A6 resulted in annexin A6 in the ECM. C2C12 myoblasts seeded onto decellularized matrices displayed differential myoblast mobility and fusion. Dystrophin-deficient decellularized matrices inhibited myoblast mobility, while dysferlin-deficient decellularized matrices enhanced myoblast movement and differentiation. Myoblasts treated with recombinant annexin A6 increased mobility and fusion like that seen on dysferlin-deficient decellularized matrix and demonstrated upregulation of ECM and muscle cell differentiation genes. These findings demonstrate specific fibrotic signatures elicit effects on myoblast activity. [Display omitted] [ABSTRACT FROM AUTHOR]

Microglial efferocytosis plays a crucial role in facilitating and sustaining homeostasis in the central nervous system, and it is involved in neuropsychiatric disorders. How microglial efferocytosis is affected under the condition of major depressive disorder (MDD) remains elusive. In this study, we hypothesized that microglial efferocytosis in the hippocampus is impaired in the chronic unpredicted mild stress (CUMS) model of MDD, which is involved in the development of MDD. Depressive-like behavior in adult male mice was induced by CUMS and confirmed by behavioral tests. Microglial efferocytosis was evaluated using immunofluorescence staining of hippocampal slices and primary microglia co-cultured with apoptotic cells. The protein and mRNA levels of phagocytosis-related molecules and inflammation-related cytokines were detected using western blotting and RT-qPCR, respectively. Annexin V was injected to mimic impairment of microglial efferocytosis. TREM2-siRNA was further used on primary microglia to examine efferocytosis-related signaling pathways. Microglia were activated and the expression of proinflammatory cytokines was increased in CUMS mice, while microglial efferocytosis and efferocytosis-related molecules were decreased. Inhibition of the TREM2/Rac1 pathway impaired microglial efferocytosis. Annexin V injection inhibited microglial efferocytosis, increased inflammation in the hippocampus and depressive-like behavior. The potential antidepressant effect of the upregulation of the TREM2/Rac1 pathway was not evaluated. Impairment of microglial efferocytosis is involved in the development of depressive-like behavior, with downregulation of the TREM2/Rac1 pathway and increased inflammation. These results may increase our understanding of the pathophysiological mechanisms associated with MDD and provide novel targets for therapeutic interventions. • Microglial efferocytosis is impaired in mice model of stress-induced depression. • Efferocytosis is reduced in proinflammatory activated microglia, with the down-regulation of TREM2/Rac1 pathway. • Blockade of microglial efferocytosis with Annexin V exacerbates the depressive-like behavior of CUMS-treated mice. [ABSTRACT FROM AUTHOR]

Nonalcoholic fatty liver disease (NAFLD) has emerged as a major chronic liver illness characterized by increase of lipid content in the liver. This study investigated the role of lauric acid to treat NAFLD in male adult Sprague Dawley rats. In this study, to induce NAFLD in the rats, a high-fat diet (HFD) was administered for eight consecutive weeks. Lauric acid groups received lauric acid (250 and 500 mg/kg; orally), concurrently with HFD for eight consecutive weeks. Lauric acid could ameliorate the serum levels of TG, TC, ALT, AST, blood glucose, and insulin. Moreover, lauric acid significantly elevated the levels of SOD, GSH, catalase, and IL-10. Additionally, it lowered the hepatic levels of MDA, ROS, MPO, 4-HNE, interleukin (IL)-1β, and tumor necrosis factor (TNF-α). Furthermore, lauric acid significantly up-regulated the hepatic expression of IRS1, AMPK, PI3K, and SIRT1 genes. In parallel, lauric acid could improve the histopathological picture of the liver and reduce the liver apoptosis via decreasing the expression of annexin V (Anx V). Finally, our data proposed that lauric acid could be an effective candidate for the NAFLD treatment. [ABSTRACT FROM AUTHOR]

Hepatitis C virus (HCV) infection is tightly connected to the lipid metabolism with lipid droplets (LDs) serving as assembly sites for progeny virions. A previous LD proteome analysis identified annexin A3 (ANXA3) as an important HCV host factor that is enriched at LDs in infected cells and required for HCV morphogenesis. To further characterize ANXA3 function in HCV, we performed proximity labeling using ANXA3- BioID2 as bait in HCV-infected cells. Two of the top proteins identified proximal to ANXA3 during HCV infection were the La-related protein 1 (LARP1) and the ADP ribosylation factorlike protein 8B (ARL8B), both of which have been previously described to act in HCV particle production. In follow-up experiments, ARL8B functioned as a pro-viral HCV host factor without localizing to LDs and thus likely independent of ANXA3. In contrast, LARP1 interacts with HCV core protein in an RNA-dependent manner and is translocated to LDs by core protein. Knockdown of LARP1 decreased HCV spreading without altering HCV RNA replication or viral titers. Unexpectedly, entry of HCV particles and E1/E2-pseudotyped lentiviral particles was reduced by LARP1 depletion, whereas particle production was not altered. Using a recombinant vesicular stomatitis virus (VSV)ΔG entry assay, we showed that LARP1 depletion also decreased entry of VSV with VSV, MERS, and CHIKV glycoproteins. Therefore, our data expand the role of LARP1 as an HCV host factor that is most prominently involved in the early steps of infection, likely contributing to endocytosis of viral particles through the pleiotropic effect LARP1 has on the cellular translatome. [ABSTRACT FROM AUTHOR]

A new series of 2H‐chromene‐based hydrazones (10 a–w, 12 a–d, and 14 a–d) were efficiently synthesized by the reaction of substituted 2H‐chromene aldehydes (8 a–w) with various hydrazines (9, 11, and 13) in absolute ethanol under microwave irradiation as well as by lemon juice mediated conventional heating. The structures of synthesized target molecules were identified by spectroscopic analysis methods (1H NMR, 13C NMR, FT‐IR, and HRMS). The in vitro anticancer activity of the new 2H‐chromene‐based hydrazones were evaluated on the three breast cancer cell lines (MCF‐7, MDA MB‐231, and MDA MB‐468) by MTT assay. Among these synthesized compounds, 10 r and 12 b exhibited the most potent antiproliferative activities with IC50 values at 9.12±0.45 μM and 10.71±0.67 μM respectively, against MCF‐7 cell. Flowcytometric analysis revealed that these two potent compounds, 10 r and 12 b arrested the cell cycle at the G2/M phase and induced apoptosis of proteins in a dose‐dependent manner. It was further confirmed by Hoechst staining and Annexin V‐FITC assay. Additionally, compounds 10 r and 12 b were examined by in silico molecular docking, which revealed that compound 10 r displayed excellent binding affinity energy of around −10 kcal/mol with human HER2 receptor. Also, the ADME studies validated the hybrid compounds showing promising physicochemical, pharmacokinetic, and drug‐like attributes. Broadly, these results underscored the potential of 2H‐chromene‐based hydrazone derivatives as promising anticancer agents. [ABSTRACT FROM AUTHOR]

Chronic inflammation is increasingly considered as the most important component of vascular aging, contributing to the progression of age‐related cardiovascular diseases. To delay the process of vascular aging, anti‐inflammation may be an effective measure. The anti‐inflammatory factor annexin A1 (ANXA1) is shown to participate in several age‐related diseases; however, its function during vascular aging remains unclear. Here, an ANXA1 knockout (ANXA1−/−) and an endothelial cell‐specific ANXA1 deletion mouse (ANXA1△EC) model are used to investigate the role of ANXA1 in vascular aging. ANXA1 depletion exacerbates vascular remodeling and dysfunction while upregulates age‐ and inflammation‐related protein expression. Conversely, Ac2‐26 (a mimetic peptide of ANXA1) supplementation reverses this phenomenon. Furthermore, long‐term tumor necrosis factor‐alpha (TNF‐α) induction of human umbilical vein endothelial cells (HUVECs) increases cell senescence. Finally, the senescence‐associated secretory phenotype and senescence‐related protein expression, rates of senescence‐β‐galactosidase positivity, cell cycle arrest, cell migration, and tube formation ability are observed in both ANXA1‐knockdown HUVECs and overexpressed ANXA1‐TNF‐α induced senescent HUVECs. They also explore the impact of formyl peptide receptor 2 (a receptor of ANXA1) in an ANXA1 overexpression inflammatory model. These data provide compelling evidence that age‐related inflammation in arteries contributes to senescent endothelial cells that promote vascular aging. [ABSTRACT FROM AUTHOR]

Riboflavin is a water‐soluble yellowish vitamin and is controversial regarding its effect on tumour cells. Riboflavin is a powerful photosensitizer that upon exposure to radiation, undergoes an intersystem conversion with molecular oxygen, leading to the production of ROS. In the current study, we sought to ascertain the impact of irradiated riboflavin on C6 glioblastoma cells regarding proliferation, cell death, oxidative stress and migration. First, we compared the proliferative behaviour of cells following nonradiated and radiated riboflavin. Next, we performed apoptotic assays including Annexin V and caspase 3, 7 and 9 assays. Then we checked on oxidative stress and status by flow cytometry and ELISA kits. Finally, we examined inflammatory change and levels of MMP2 and SIRT1 proteins. We caught a clear antiproliferative and cytotoxic effect of irradiated riboflavin compared to nonradiated one. Therefore, we proceeded with our experiments using radiated riboflavin. In all apoptotic assays, we observed a dose‐dependent increase. Additionally, the levels of oxidants were found to increase, while antioxidant levels decreased following riboflavin treatment. In the inflammation analysis, we observed elevated levels of both pro‐inflammatory and anti‐inflammatory cytokines. Additionally, after treatment, we observed reduced levels of MMP2 and SIRT. In conclusion, radiated riboflavin clearly demonstrates superior antiproliferative and apoptotic effects on C6 cells at lower doses compared to nonradiated riboflavin. [ABSTRACT FROM AUTHOR]

Background: Our earlier research revealed that the secreted lysyl oxidase-like 4 (LOXL4) that is highly elevated in triple-negative breast cancer (TNBC) acts as a catalyst to lock annexin A2 on the cell membrane surface, which accelerates invasive outgrowth of the cancer through the binding of integrin-β1 on the cell surface. However, whether this machinery is subject to the LOXL4-mediated intrusive regulation remains uncertain. Methods: Cell invasion was assessed using a transwell-based assay, protein-protein interactions by an immunoprecipitation-Western blotting technique and immunocytochemistry, and plasmin activity in the cell membrane by gelatin zymography. Results: We revealed that cell surface annexin A2 acts as a receptor of plasminogen via interaction with S100A10, a key cell surface annexin A2-binding factor, and S100A11. We found that the cell surface annexin A2/ S100A11 complex leads to mature active plasmin from bound plasminogen, which actively stimulates gelatin digestion, followed by increased invasion. Conclusion: We have refined our understanding of the role of LOXL4 in TNBC cell invasion: namely, LOXL4 mediates the upregulation of annexin A2 at the cell surface, the upregulated annexin 2 binds S100A11 and S100A10, and the resulting annexin A2/S100A11 complex acts as a receptor of plasminogen, readily converting it into active-form plasmin and thereby enhancing invasion. [ABSTRACT FROM AUTHOR]

Background: Cancer is among the serious health problems of the medical world, for treatment of which severe treatments are used. However, the prognosis of cancer patients is still poor. The application of NK cell-derived exosomes (NK-Exo) is a new method for cancer immunotherapy. These nanoparticles with a size range of 30-120 nm are a small model of mother cells. In this study, the anti-tumor activity of NK-Exo and LAK-Exo (activated NK cell-derived exosome) against acute myeloid leukemia (AML) is investigated in vitro. Materials and Methods: The MACS method was performed for the separation of NK cells from the buffy coats of healthy donors, and an EXOCIBE kit was used for the isolation of NK-Exo. After treating the KG-1 cell line with different doses of NK-Exo, MTT assay, and annexin V-PE were done to evaluate cell proliferation and apoptosis, respectively, and for confirmation of involved proteins, Real-Time PCR and western blotting were performed. Results: Anti-tumor activity of NK-Exo and LAK-Exo was dose- and time-dependent. Their highest activities were observed following 48 hours of incubation with 50 µg/ml exosome (p<0.0001). However, this cytotoxic activity was also seen over a short period of time with low concentrations of NK-Exo (p<0.05) and LAK-Exo (p<0.001). The cytotoxic effect of LAK-Exo on target cells was significantly higher than NK-EXO. The induction of apoptosis by different pathways was time-point dependent. Total apoptosis was 34.56% and 51.6% after 48 hours of tumor cell coculture with 50µg/ml NK-Exo and LAK-Exo, respectively. Significant expression of CASPASE3, P38, and CYTOCHROME C genes was observed in the cells treated with 50 µg/ml NK-Exo and LAKExo. Conclusion: Our study confirmed the antileukemia activity of NK-Exo against AML tumor cells in vitro. Therefore, NK-Exo can be considered as a promising and effective treatment for leukemia therapy. [ABSTRACT FROM AUTHOR]

Background: The protein annexin A6 (AnxA6) is involved in numerous membrane-related biological processes including cell migration and invasion by interacting with other proteins. The dysfunction of AnxA6, including protein expression abundance change and imbalance of post-translational modification, is tightly related to multiple cancers. Herein we focus on the biological function of AnxA6 SUMOylation in hepatocellular carcinoma (HCC) progression. Methods: The modification sites of AnxA6 SUMOylation were identified by LC-MS/MS and amino acid site mutation. AnxA6 expression was assessed by immunohistochemistry and immunofluorescence. HCC cells were induced into the epithelial-mesenchymal transition (EMT)-featured cells by 100 ng/mL 12-O-tetradecanoylphorbol-13-acetate exposure. The ability of cell migration was evaluated under AnxA6 overexpression by transwell assay. The SUMO1 modified AnxA6 proteins were enriched from total cellular proteins by immunoprecipitation with anti-SUMO1 antibody, then the SUMOylated AnxA6 was detected by Western blot using anti-AnxA6 antibody. The nude mouse xenograft and orthotopic hepatoma models were established to determine HCC growth and tumorigenicity in vivo. The HCC patient's overall survival versus AnxA6 expression level was evaluated by the Kaplan–Meier method. Results: Lys579 is a major SUMO1 modification site of AnxA6 in HCC cells, and SUMOylation protects AnxA6 from degradation via the ubiquitin-proteasome pathway. Compared to the wild-type AnxA6, its SUMO site mutant AnxA6K579R leads to disassociation of the binding of AnxA6 with RHOU, subsequently RHOU-mediated p-AKT1ser473 is upregulated to facilitate cell migration and EMT progression in HCC. Moreover, the SENP1 deSUMOylates AnxA6, and AnxA6 expression is negatively correlated with SENP1 protein expression level in HCC tissues, and a high gene expression ratio of ANXA6/SENP1 indicates a poor overall survival of patients. Conclusions: AnxA6 deSUMOylation contributes to HCC progression and EMT phenotype, and the combination of AnxA6 and SENP1 is a better tumor biomarker for diagnosis of HCC grade malignancy and prognosis. [ABSTRACT FROM AUTHOR]

Plant annexins constitute a conserved protein family that plays crucial roles in regulating plant growth and development, as well as in responses to both biotic and abiotic stresses. In this study, a total of 144 annexin genes were identified in the barley pan-genome, comprising 12 reference genomes, including cultivated barley, landraces, and wild barley. Their chromosomal locations, physical–chemical characteristics, gene structures, conserved domains, and subcellular localizations were systematically analyzed to reveal the certain differences between wild and cultivated populations. Through a cis-acting element analysis, co-expression network, and large-scale transcriptome analysis, their involvement in growth, development, and responses to various stressors was highlighted. It is worth noting that HvMOREXann5 is only expressed in pistils and anthers, indicating its crucial role in reproductive development. Based on the resequencing data from 282 barley accessions worldwide, genetic variations in thefamily were investigated, and the results showed that 5 out of the 12 identified HvMOREXanns were affected by selection pressure. Genetic diversity and haplotype frequency showed notable reductions between wild and domesticated barley, suggesting that a genetic bottleneck occurred on the annexin family during the barley domestication process. Finally, qRT-PCR analysis confirmed the up-regulation of HvMOREXann7 under drought stress, along with significant differences between wild accessions and varieties. This study provides some insights into the genome organization and genetic characteristics of the annexin gene family in barley at the pan-genome level, which will contribute to better understanding its evolution and function in barley and other crops. [ABSTRACT FROM AUTHOR]