Annexin A1 Research Articles

Proteomics Study of Mesenchymal Stem Cell-Like Cells Obtained from Tumor Microenvironment of Patients with Malignant and Benign Salivary Gland Tumors.

Objective Salivary gland tumors (SGTs) show some aggressive and peculiar clinicopathological behaviors that might be related to the components of the tumor microenvironment, especially mesenchymal stem cells (MSCs)-associated proteins. However, the role of MSCs-related proteins in SGTs tumorigenesis is poorly understood. This study aimed to isolate and characterize MSCs from malignant and benign tumor tissues and to identify differentially expressed proteins between these two types of MSCs. Materials and Methods In this experimental study, MSC-like cells derived from benign (pleomorphic adenoma, n=5) and malignant (mucoepidermoid carcinoma, n=5) tumor tissues were verified by fluorochrome antibodies and flow cytometric analysis. Differentially expressed proteins were identified using two-dimensional polyacrylamide gel electrophoresis (2DE) and Mass spectrometry. Results Results showed that isolated cells strongly expressed characteristic MSCs markers such as CD44, CD73, CD90, CD105, and CD166, but they did not express or weakly expressed CD14, CD34, CD45 markers. Furthermore, the expression of CD24 and CD133 was absent or near absent in both isolated cells. Results also discovered overexpression of Annexin A4 (Anxa4), elongation factor 1-delta (EF1-D), FK506 binding protein 9 (FKBP9), cytosolic platelet-activating factor acetylhydrolase type IB subunit beta (PAFAH1B), type II transglutaminase (TG2), and s-formylglutathione hydrolase (FGH) in MSCs isolated from the malignant tissues. Additionally, heat shock protein 70 (Hsp70), as well as keratin, type II cytoskeletal 7 (CK-7), were found to be overexpressed in MSCs derived from the benign ones. ConclusionMalignant and benign SGTs probably exhibit a distinct pattern of tissue proteins that are most likely related to the metabolic pathway. However, further studies in a large number of patients are required to determine the applicability of identified proteins as new targets for cancer therapy.

Contribution of plasma, placental, inflammatory and pro-resolving mediators in labor induction

IntroductionThe onset of labor is regulated by endocrine, nervous and immunological factors. This study was designed to determine the inflammatory and pro-resolving mediator levels in plasma and placenta of women undergoing labor induction in late-term pregnancy. MethodHealthy pregnant women admitted for delivery or labor induction were included. TNF, IL-1β, IL-6, IL-8, and IL-10 were quantified by ELISA in plasmatic and placental samples and Annexin A1 (ANXA1) in the placenta by Western Blotting, and immunofluorescence to CD15+ antibody. The data were analyzed using the Wilcoxon test and Spearman correlation. The p-value was significant when <0.05. ResultsThere was a higher concentration of IL-8 was found in the amniotic plaque (p = 0.042) and IL-10 (p < 0.001) in the trophoblast of patients with spontaneous labor. Greater ANXA1 density in the trophoblast was also observed in those with induction failure. There was a positive correlation of ANXA1 density in trophoblast induction duration with (r = 0.580) and with the IL-6 level in amniotic plaque (r = 0.517), and a positive correlation between labor duration and density of ANXA1 was identified in the trophoblast (r = 0.419). An increase was identified in CD15+ cell immunocapturing among the groups with spontaneous labor compared to the group with induction failure (p < 0.001). DiscussionThe inflammatory process in labor involves both maternal and fetal participation. Induction failure is associated with higher levels of ANXA1 in the trophoblast.

Transcriptomic Analysis of Annexin A10 and Chemosensitivity in Gastric Adenocarcinoma Cells

Gastric cancer (GC) is the third-leading cause of cancer-related deaths worldwide; thus, novel diagnostic and therapeutic biomarkers are needed. Annexin A10 (ANXA10) is a calcium- and phospholipid-binding protein. As far as we are aware, there are no reports describing the detailed functions of ANXA10 in GC. Therefore, we investigated the downstream mRNA variation and the effects of ANXA10 on chemoresistance in GC cell lines. ANXA10 knockout GC cell lines were generated, and we performed functional analyses, chemosensitivity drug testing, and microarray analyses. Additionally, immunohistochemistry for ANXA10 was performed on 40 patients with GC who had received 5-fluorouracil (5-FU)-based chemotherapy to compare their prognosis and clinicopathological factors. ANXA10 knockout GC cells showed significantly increased proliferation, invasion, and sensitivity to 5-FU. The overall survival of ANXA10-positive cases was considerably lower than that of ANXA10-negative cases in GC patients who received 5-FU-based chemotherapy. Microarray analysis revealed candidate pathways regulated by ANXA10 and claudin 1 (CLDN1), keratin 80 (KRT80), RANBP2-type and C3HC4-type zinc finger containing 1 (RBCK1), and solute carrier family 7 member 5 (SLC7A5) genes. ANXA10 knockout increased the susceptibility of GC cell lines to 5-FU; ANXA10 may be a predictive indicator for response to 5-FU treatment in GC cases. ANXA10 may be involved in the pathogenesis of GC, in collaboration with CLDN1, KRT80, RBCK1, and SLC7A5.

Tonsillar swabs and sputum predict SLPI- and AnxA2 expression in tonsils: A prospective study on smoking dependent SLPI- and AnxA2-expression, and tonsillar HPV infection.

Previous retrospective studies have elucidated a correlation between secretory leucocyte protease inhibitor (SLPI) and Annexin A2 (AnxA2), patient smoking status and tonsillar human papilloma virus (HPV) status. The current study assessed these parameters prospectively and to the best of our knowledge, analyzed SLPI-/AnxA2-expression for the first time in tonsillar swabs and sputum. Samples were obtained from 52 patients with tonsillar squamous cell carcinoma and 163 patients with tonsillar hyperplasia (H; n=56) and chronic or recurrent tonsillitis (CRT; n=107). HPV-DNA, SLPI and AnxA2 gene expression was analyzed in sputum, tonsillar swabs and tissue by performing reverse transcription-quantitative PCR. Results were compared with smoking status, revealing that smoking resulted in significantly increased SLPI gene expression in all biomaterials of all cases. SLPI-gene expression was significantly decreased in all HPV-DNA-positive samples (tissue/swab/sputum), while AnxA2 was significantly increased in all HPV-DNA-positive samples. Results from swabs and sputum were able to predict SLPI- and AnxA2 gene expression of the corresponding tonsil. The current prospective study confirmed previous retrospective results underlining this hypothesis: Smoking enhances SLPI-expression, preventing HPV-binding to AnxA2. HPV-binding to AnxA2 appears essential for successful cell-entry. SLPI/AnxA2-gene expression in swabs and sputum reflect their expression in tonsillar tissue. Accordingly, a positive AnxA2/SLPI-ratio in sputum/swabs could possibly be used to reduce HPV-associated carcinogenesis, by performing tonsillectomy or HPV-vaccination in patients with positive AnxA2/SLPI-ratios.

Annexin A1 Promotes Reparative Angiogenesis and Ameliorates Neuronal Injury in Ischemic Retinopathy

PurposeRetinal ischemia is the main reason for vision threatening. Inflammation and aberrant angiogenesis play an important role in the pathogenesis of ischemia. Annexin A1 (ANXA1) is an endogenous protein modulating anti-inflammatory processes, and its therapeutic potential has been reported in a range of inflammatory diseases. However, the effect of ANXA1 on ischemic retinal injury has not been examined.MethodsExpression of ANXA1 was assessed by real-time PCR and western blotting, and location of ANXA1 was evaluated by immunofluorescence staining in retina of OIR. The activation of ANXA1 was assayed in HRECs after hypoxia stimuli. The effect of ANXA1 on vascularization of OIR mouse through quantification vaso-obliteration (VO) and neovascularization (NV), as well as expression of relevant angiogenic factors and inflammatory cytokines was compared between wild type and ANXA1 deficiency mice. We also investigated the effect of ANXA1 on retinal neuronal degeneration as measured by electroretinography (ERG) and OCT.ResultsIn retinas of OIR, the expression of ANXA1 significantly increased and located in inner retinal layers. ANXA1 was induced in HRECs after hypoxic stimuli. Furthermore, ANXA1 deficiency increased pro-angiogenic and pro-inflammatory cytokines. Ablation of ANXA1 suppressed aortic outgrowth and retinal reparative revascularization and promoted pathological NV to exacerbate retinal dysfunction after ischemia injury.ConclusionsANXA1 inhibits angiogenic and inhibits pro-inflammatory cytokines and promotes reparative angiogenesis, thus exhibits neuronal protective function in ischemic retinopathy.

Potential regulatory mechanism of TNF-α/TNFR1/ANXA1 in glioma cells and its role in glioma cell proliferation.

The purpose of this study was to explore the regulatory mechanism of Annexin A1 (ANXA1) in glioma cells in the inflammatory microenvironment induced by tumour necrosis factor α (TNF-α) and its effects on glioma cell proliferation. CCK-8 analysis demonstrated that TNF-α stimulation promotes rapid growth in glioma cells. Changes in tumour necrosis factor receptor 1 (TNFR1) and ANXA1 expression in glioma cells stimulated with TNF-α were revealed through western blot analysis and immunofluorescence staining. Coimmunoprecipitation analysis revealed that ANXA1 interacts with TNFR1. Moreover, we found that ANXA1 promotes glioma cell growth by activating the p65 and Akt signalling pathways. Finally, immunohistochemistry analysis showed an obvious correlation between ANXA1 expression and Ki-67 in glioma tissues. In summary, our results indicate that the TNF-α/TNFR1/ANXA1 axis regulates the proliferation of glioma cells and that ANXA1 plays a regulatory role in the inflammatory microenvironment.

Knockdown of Annexin A2 Enhances Radiosensitivity by Increasing G2/M-Phase Arrest, Apoptosis and Activating the p38 MAPK-HSP27 Pathway in Nasopharyngeal Carcinoma.

Annexin A2 (ANXA2) has been found to be involved in cancer proliferation, metastasis and prognosis; however, its exact role in nasopharyngeal carcinoma (NPC) radioresistance remains unknown. We found that ANXA2 expression was correlated with prognosis in NPC patients, and longer overall survival in NPC patients with low ANXA2 expression than those with high ANXA2 expression. ANXA2 knockdown increased the radiosensitivity in radioresistant NPC cells, and ANXA2 overexpression decreased the radiosensitivity in NPC cells. Knocking-down ANXA2 expression increased the irradiation-induced apoptosis of radioresistant NPC cells, and ANXA2 overexpression decreased the irradiation-induced apoptosis of NPC cells. ANXA2 knockdown induced G2/M phase arrest in NPC cells post-irradiation, and ANXA2 overexpression abrogated G2/M phase arrest in NPC cells post-irradiation. ANXA2 overexpression resulted in inhibition of the p38 MAPK-HSP27 pathway, while ANXA2 knockdown resulted in activation of the p38 MAPK-HSP27 pathway. In addition, ANXA2 knockdown increased the radiosensitivity of the xenografted tumors in nude mice. Our data demonstrate that knockdown of Annexin A2 enhanced radiosensitivity in NPC by increasing G2/M-phase arrest, apoptosis and activating the p38 MAPK-HSP27 pathway. ANXA2 may be a promising target used to overcome radioresistance in NPC.

Targeting the Annexin A1-FPR2/ALX pathway for host-directed therapy in dengue disease.

Host immune responses contribute to dengue's pathogenesis and severity, yet the possibility that failure in endogenous inflammation resolution pathways could characterise the disease has not been contemplated. The pro-resolving protein Annexin A1 (AnxA1) is known to counterbalance overexuberant inflammation and mast cell (MC) activation. We hypothesised that inadequate AnxA1 engagement underlies the cytokine storm and vascular pathologies associated with dengue disease. Levels of AnxA1 were examined in the plasma of dengue patients and infected mice. Immunocompetent, interferon (alpha and beta) receptor one knockout (KO), AnxA1 KO, and formyl peptide receptor 2 (FPR2) KO mice were infected with dengue virus (DENV) and treated with the AnxA1 mimetic peptide Ac2-26 for analysis. In addition, the effect of Ac2-26 on DENV-induced MC degranulation was assessed in vitro and in vivo. We observed that circulating levels of AnxA1 were reduced in dengue patients and DENV-infected mice. Whilst the absence of AnxA1 or its receptor FPR2 aggravated illness in infected mice, treatment with AnxA1 agonistic peptide attenuated disease manifestationsatteanuated the symptoms of the disease. Both clinical outcomes were attributed to modulation of DENV-mediated viral load-independent MC degranulation. We have thereby identified that altered levels of the pro-resolving mediator AnxA1 are of pathological relevance in DENV infection, suggesting FPR2/ALX agonists as a therapeutic target for dengue disease.

Inhibition of STAT6 Activation by AS1517499 Inhibits Expression and Activity of PPARγ in Macrophages to Resolve Acute Inflammation in Mice.

Signal transducer and activator of transcription 6 (STAT6) promotes an anti-inflammatory process by inducing the development of M2 macrophages. We investigated whether modulating STAT6 activity in macrophages using AS1517499, the specific STAT6 inhibitor, affects the restoration of homeostasis after an inflammatory insult by regulating PPARγ expression and activity. Administration of AS1517499 suppressed the enhanced STAT6 phosphorylation and nuclear translocation observed in peritoneal macrophages after zymosan injection. In addition, AS1517499 delayed resolution of acute inflammation as evidenced by enhanced secretion of pro-inflammatory cytokines, reduced secretion of anti-inflammatory cytokines in PLF and supernatants from peritoneal macrophages, and exaggerated neutrophil numbers and total protein levels in PLF. We demonstrate temporal increases in annexin A1 (AnxA1) protein and mRNA levels in peritoneal lavage fluid (PLF), peritoneal macrophages, and spleen in a murine model of zymosan-induced acute peritonitis. In vitro priming of mouse bone marrow-derived macrophages (BMDM) and peritoneal macrophages with AnxA1 induced STAT6 activation with enhanced PPARγ expression and activity. Using AS1517499, we demonstrate that inhibition of STAT6 activation delayed recovery of PPARγ expression and activity, as well as impaired efferocytosis. Taken together, these results suggest that activation of the STAT6 signaling pathway mediates PPARγ expression and activation in macrophages to resolve acute inflammation.

Phosphoproteome Analysis Identifies a Synaptotagmin-1-Associated Complex Involved in Ischemic Neuron Injury

Cerebral stroke is one of the leading causes of death in adults worldwide. However, the molecular mechanisms of stroke-induced neuron injury are not fully understood. Here, we obtained phosphoproteomic and proteomic profiles of the acute ischemic hippocampus by LC–MS/MS analysis. Quantitative phosphoproteomic analyses revealed that the dysregulated phosphoproteins were involved in synaptic components and neurotransmission. We further demonstrated that phosphorylation of Synaptotagmin-1 (Syt1) at the Thr112 site in cultured hippocampal neurons aggravated oxygen-glucose deprivation–induced neuronal injury. Immature neurons with low expression of Syt1 exhibit slight neuronal injury in a cerebral ischemia model. Administration of the Tat-Syt1T112A peptide protects neurons against cerebral ischemia-induced injury in vitro and in vivo. Surprisingly, potassium voltage-gated channel subfamily KQT member 2 (Kcnq2) interacted with Syt1 and Annexin A6 (Anxa6) and alleviated Syt1-mediated neuronal injury upon oxygen-glucose deprivation treatment. These results reveal a mechanism underlying neuronal injury and may provide new targets for neuroprotection after acute cerebral ischemia onset.

A quantitative proteomic analysis reveals the potential roles of PRDX3 in neurite outgrowth in N2a-APPswe cells

Alzheimer's disease (AD) is characterized by amyloid plaques and neurofibrillary tangles accompanied by progressive neurite loss. Mitochondria play pivotal roles in AD development. PRDX3 is a mitochondrial peroxide reductase critical for H2O2 scavenging and signal transduction. In this study, we found that PRDX3 knockdown (KD) in the N2a-APPswe cell line promoted retinoic acid (RA)-induced neurite outgrowth but did not reduce the viability of cells damaged by tert-butyl hydroperoxide (TBHP). We found that knocking down PRDX3 expression induced dysregulation of more than one hundred proteins, as determined by tandem mass tag (TMT)-labeled proteomics. A Gene Ontology (GO) analysis revealed that the dysregulated proteins were enriched in protein localization to the plasma membrane, the lipid catabolic process, and intermediate filament cytoskeleton organization. A STRING analysis showed close protein–protein interactions among dysregulated proteins. The expression of Annexin A1 (ANXA1), serine (Ser)-/threonine (Thr)-protein phosphatase 2A catalytic subunit alpha isoform (PP2A) and glutathione S-transferase Mu 2 (GSTM2) was significantly upregulated in PRDX3-KD N2a-APPswe cell lines, as verified by western blotting. Our study revealed, for the first time, that PRDX3 may play important roles in neurite outgrowth and AD development.

High Annexin A10 expression is correlated with poor prognosis in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is the third-leading cause of cancer-related death. Owing to its poor prognosis, new molecular biomarkers for PDAC are needed. Annexin A10 (ANXA10) is a calcium-/phospholipid-binding protein belonging to the annexin family of proteins. ANXA10 is not only associated with gastric phenotypes, but also acts an independent prognostic factor in several cancers. However, the role of ANXA10 in PDAC remains unknown. Therefore, we examined the relationship between ANXA10 and the prognosis of PDAC. We analyzed the expression of ANXA10 using data from public databases, and performed immunohistochemistry analysis for 81 PDAC cases. We then investigated the relationship between ANXA10 expression and clinicopathological features. ANXA10 was detected in 47 of 81 PDAC cases (58%). High expression of ANXA10 was significantly related to poor overall survival (OS; p=0.011). Univariate analysis of OS revealed three prognostic parameters: tumor grade (p=0.046), perineural invasion (p=0.017), and ANXA10 expression (p=0.012). Multivariate analysis indicated that ANXA10 expression (p<0.01) alone was a prognostic factor in PDAC cases. Our findings suggest that ANXA10 expression is an independent prognostic factor in PDAC cases and shows promise as a new biomarker in PDAC.

Annexin A10 Expression as a Novel Prognostic Marker in Lung Adenocarcinoma.

Annexin A10 (ANXA10) is a member of the annexin family and a calcium-dependent phospholipid-binding protein. The aim of this study was to clarify the clinical significance of ANXA10 expression in lung adenocarcinoma. ANXA10 expression was immunohistochemically examined in surgical specimens of lung adenocarcinoma obtained from 74 consecutive patients who underwent complete resection from January 2014 to December 2014. Expression of ANXA10 was down-regulated in A549 cells via siRNA transfection and the effect of ANXA10 on cell migration was assessed by the wound healing assay. Expression of ANXA10 was examined by immunocytochemistry and polymerase chain reaction. High ANXA10 expression was significantly correlated with poor overall survival (p=0.00705). Multivariate analysis with the Cox proportional hazard model demonstrated that ANXA10 expression was an independent prognostic factor. Cell migration was suppressed in ANXA10-down-regulated A549 cell lines. ANXA10 has a role in cancer cell migration and high ANXA10 expression is a novel prognostic marker in lung adenocarcinoma.

Clinicopathological study and immunohistochemical analysis of expression of annexin A5 and apelin in human placentae of gestational diabetes mellitus

Background and aimsGestational diabetes mellitus (GDM) is one of the commonest medical complications of pregnancy. Annexin A5 (ANXA5) is a protein, found in apical surfaces of syncytiotrophoblasts, which prevents fetal and placental vascular thrombosis in GDM. Apelin is a bioactive peptide which has been linked to GDM. The aim of the present study was to correlate macroscopic as well as microscopic changes and immunohistochemical expression of ANXA5 and apelin in placentae of GDM with maternal and neonatal clinical features and also to compare the results with those in matched controls. MethodsThis prospective observational study was undertaken for a period of one year from April 2020 to March 2021. It comprised of 42 patients of GDM. Gross features, microscopic features and intensity and grade of expression of ANXA5 and Apelin were analyzed in placentae of GDM. ResultsMorphological changes detected in GDM placentae included increased immature villi (16 cases, 38%), increased syncytial knots (36, 86%), perivillous fibrin deposition (20, 48%), fibrosis of villous stroma (20, 48%), presence of nucleated red blood cells (12, 28.5%) and hypervascularity (34, 81%). The extent of histopathological changes noted in GDM placentae was significantly higher than that in matched controls. GDM placentae showed significantly reduced expression of ANXA5 and Apelin in terms of grade and intensity when compared with matched controls. Reduced expression (mild intensity) of ANXA5 was noted in 22 GDM cases (52.3%) whereas apelin expression was of weak intensity in 26 (61.9%) cases. Among GDM patients, statistically significant association was noted between ANXA5 intensity and neonatal resuscitation, apelin grade and preterm birth as well as low birth weight and apelin intensity and requirement of treatment in sick neonatal care unit. ConclusionThe placental expression of the proteins, ANXA5 and Apelin, is altered in GDM though their exact pathogenetic mechanisms are yet to be understood. They can be targets for development of prophylactic and therapeutic agents in future.

Reduced Expression of Annexin A6 Induces Metabolic Reprogramming That Favors Rapid Fatty Acid Oxidation in Triple-Negative Breast Cancer Cells.

Simple SummaryThe expression status of Annexin A6 (AnxA6) has been shown to influence tumor growth, metastasis, and resistance to therapeutic intervention, but the contribution of this tumor suppressor in the metabolic adaptation of basal-like (AnxA6-low) versus mesenchymal-like (AnxA6-high) TNBC subsets remains unclear. The downregulation of AnxA6 in TNBC cells attenuated mitochondrial respiration, glycolytic flux, and cellular ATP production capacity, resulting in a quiescent metabolic phenotype. The overexpression or chronic lapatinib induced the expression of AnxA6 in AnxA6-low TNBC cells and reversed the quiescent phenotype to a more lipogenic/glycolytic phenotype. Interestingly, AnxA6-depletion was associated with rapid fatty acid uptake and oxidation (lypolytic metabolic phenotype) but decreased lipid droplet accumulation. Our data suggest that the expression status of AnxA6 in TNBC cells is associated with distinct metabolic adaptations of basal-like and mesenchymal-like TNBC subsets in response to cellular stress and/or therapeutic intervention.The ability of cancer cells to alter their metabolism is one of the major mechanisms underlying rapid tumor progression and/or therapeutic resistance in solid tumors, including the hard-to-treat triple-negative breast cancer (TNBC) subtype. Here, we assessed the contribution of the tumor suppressor, Annexin A6 (AnxA6), in the metabolic adaptation of basal-like (AnxA6-low) versus mesenchymal-like (AnxA6-high), as well as in lapatinib-resistant TNBC cells. Using model basal-like and mesenchymal-like TNBC cell lines, we show that TNBC cells also exhibit metabolic heterogeneity. The downregulation of AnxA6 in TNBC cells generally attenuated mitochondrial respiration, glycolytic flux, and cellular ATP production capacity resulting in a quiescent metabolic phenotype. We also show that AnxA6 depletion in mesenchymal-like TNBC cells was associated with a rapid uptake and mitochondrial fatty acid oxidation and diminished lipid droplet accumulation and altered the lipogenic metabolic phenotype of these cells to a lypolytic metabolic phenotype. The overexpression or chronic lapatinib-induced upregulation of AnxA6 in AnxA6-low TNBC cells reversed the quiescent/lypolytic phenotype to a more lipogenic/glycolytic phenotype with gluconeogenic precursors as additional metabolites. Collectively, these data suggest that the expression status of AnxA6 in TNBC cells underlies distinct metabolic adaptations of basal-like and mesenchymal-like TNBC subsets in response to cellular stress and/or therapeutic intervention and suggest AnxA6 as a biomarker for metabolic subtyping of TNBC subsets.

ZBM‐H‐induced activation of GRP78 ATPase promotes apoptosis via annexin A7 in A549 lung cancer cells

Hypochlorous acid (HOCl) is an essential signal for the regulation of cancer cell fate, including autophagy and apoptosis. HOCl regulated autophagy by affecting the oxidation modification of glucose-regulated protein 78 (GRP78) and the activity of GRP78 ATPase. The mechanism of GRP78 ATPase in cell apoptosis has however not yet been clarified. Here we reported that ZBM-H, as a probe of HOCl, was able to directly bind to GRP78 in the presence or absence of ATP. Following ZBM-H treatment, the interaction between GRP78 and annexin A7 (ANXA7) was promoted, and this was accompanied by increased phosphorylation of integrin β4 (ITGB4). In addition, ZBM-H enhanced the phosphorylation of ANXA7. ABO, an inhibitor of ANXA7, inhibited ZBM-H-induced ITGB4 phosphorylation and apoptosis, while ANXA7 activator SEC had opposite effect. Collectively, these data provide new evidence for the mechanism by which ZBM-H-induced activation of GRP78 ATPase regulates apoptosis of A549 lung cancer cells.

Annexin A1 Is a Potential Prognostic Marker for, and Enhances the Metastasis of, Cholangiocarcinoma.

Objective:Annexin A1 (ANXA1) is a calcium-dependent phospholipid-binding protein which contributes to proliferation, cancer progression and metastasis. Overexpression of ANXA1 is closely associated with metastasis in numerous types of cancer. Cholangiocarcinoma (CCA) is a bile-duct cancer which has high rates of metastasis. Previously, we demonstrated up-regulation of ANXA1 in a highly metastatic CCA cell line (KKU-213AL5). Here, we investigated the functions of ANXA1 in the progression of CCA cell lines and evaluated its clinical impacts in human CCA tissues. Methods: Effects of ANXA1 on metastatic potential of CCA cell lines were evaluated using cell-proliferation, clonogenic, migration and invasion assays. The expression of ANXA1 in 44 intrahepatic human CCA tissues was investigated using immunohistochemistry (IHC). The association of ANXA1 with clinicopathological features of CCA patients was analyzed. Results:Silencing of ANXA1 expression using siRNA significantly decreased cell proliferation, colony formation, cell migration and invasion in the KKU-213AL5 cell line. IHC results showed low expression of ANXA1 in normal bile ducts in the non-tumor area. In contrast, high expression of ANXA1 in human CCA tissues was associated with advanced tumor stage, tumor size and presence of lymph-node metastasis. Conclusion:These findings strongly imply that ANXA1 contributes to the progression of CCA. ANXA1 can serve as a potential prognostic marker for CCA. Ablation of ANXA1 action may be an alternative strategy to prevent metastasis of CCA.

Methylprednisolone up-regulates annexin A1 (ANXA1) to inhibit the inflammation, apoptosis and oxidative stress of cigarette smoke extract (CSE)-induced bronchial epithelial cells, a chronic obstructive pulmonary disease in vitro model, through the formyl peptide receptor 2 (FPR2) receptors and the adenosine 5’-monophosphate (AMP)-activated protein kinase (AMPK) pathway

ABSTRACT Chronic obstructive pulmonary disease (COPD) is a progressive degenerative disease, of which smoking is the main causer. We carried out this study with the aim of exploring the underlying mechanism of methylprednisolone (MP) treating the COPD. To stimulate COPD in vitro, cigarette smoke extract (CSE)was employed to induce human bronchial epithelial cells BEAS-2B. With the help of MTT and Tunel assays, the viability and apoptosis of BEAS-2B cells after indicated treatment were assessed. The levels of inflammatory response and oxidative stress were determined by the changes of markers basing on their commercial kits. Additionally, annexin A1 (ANXA1) expressions at both protein and mRNA levels were assessed with Western blot and Reverse transcription‑quantitative PCR (RT-qPCR). Moreover, the expressions of apoptosis- and formyl peptide receptor 2 (FPR2) receptors and the adenosine 5’-monophosphate (AMP)-activated protein kinase (AMPK) pathway-related proteins were determined with Western blot., related proteins and proteins. As a result, MP up-regulated the ANXA1 expression in CSE-induced BEAS-2B cells. MP enhanced the viability but suppressed the apoptosis, inflammatory response and oxidative stress of CSE-induced BEAS-2B cells via regulating FPR2/AMPK pathway, while ANXA1 knockdown exhibited oppositive effects on them. In conclusion, MP up-regulated ANXA1 to inhibit the inflammation, apoptosis and oxidative stress of BEAS-2B cells induced by CSE, alleviating COPD through suppressing the FPR2/AMPK pathway.

Sulfated Undaria pinnatifida polysaccharide inhibits the formation of kidney stones by inhibiting HK-2 cell damage and reducing the adhesion of nano‑calcium oxalate crystals

ObjectiveThe formation of kidney stone is closely related to cell injury and crystal adhesion. MethodThe sulfur trioxide–pyridine method was used to sulfate raw Undaria pinnatifida polysaccharide (UPP) with a molecular weight (Mw) of 8.33 kDa. Four polysaccharides with the sulfate group (–OSO3−) contents of 1.59% (UPP0), 6.03% (UPP1), 20.83% (UPP2), and 36.39% (UPP3) were obtained. The antioxidant activity of the four UPPs, the difference in oxidative damage inflicted by nano-CaOx monohydrate (nano-COM) on human proximal tubular epithelial (HK-2) cells before and after protection by UPPs, and the inhibitory effect on nano-COM adhesion were explored. ResultsStructural characterization showed that sulfation was successful. As the –OSO3− content in the UPPs was increased, the antioxidant activity and capability of the UPPs to regulate the growth of calcium oxalate (CaOx) crystals gradually increased. The damage caused by nano-COM crystals to HK-2 cells under protection by UPPs was weakened. This effect enhanced cell viability, enabled the maintenance of good cell morphology, reduced reactive oxygen species (ROS) levels, and inhibited the decrease in mitochondrial membrane potential, as well as decreased the eversion of phosphatidylserine (PS) and the expression of the adhesion proteins osteopontin (OPN), heat shock protein (HSP 90), and Annexin A1 (ANXA1). The adhesion of nano-COM to HK-2 cells was inhibited under the protection by UPPs. ConclusionUPP3 with the highest content of –OSO3− presented the best antioxidant activity and crystal regulation ability, while UPP2 with the second highest –OSO3− content showed optimal cell protection ability and crystal adhesion inhibition ability. The biological activity of UPPs was regulated by Mw and –OSO3− content. UPP2 with moderate –OSO3− content may become a potential drug for preventing CaOx stones.

Abstract WP260: The Response Of Annexin A1 In Astrocyte Activation And White Matter Injury- A New Target For Stroke In Sickle Cell Disease

Introduction: Ischemic stroke is common in sickle cell disease (SCD). White matter injuries, incorporated into silent cerebral infarcts, are major risk factors of stroke in SCD. The pathogenesis of cerebral infarction in SCD is not known. Induced thrombogenesis may cause impaired astrocyte that stimulates demyelination leading to white matter injury. Annexin A1 (Anxa1) can regulate astrocyte function by inhibiting intracellular phospholipase A2 (PLA2). We hypothesized that astrocyte activation during thrombogenic events promotes white matter injury in SCD. We tested astrocyte dysfunction and white matter injury in a clinically relevant mouse model of SCD. Methods: Cerebral Infarctions induced by de novo thrombogenesis were identified by TTC staining in transgenic sickle (SS) and control (AA) mice with human sickle and normal hemoglobin respectively. Cerebral microvessels and astrocytes were isolated from SS and AA mice. Elisa assays were used to measure thrombin, PLA2 and Anxa1. Immunofluorescence staining was performed to determine astrocyte activation and demyelination. Results: Thrombogenesis within cerebral microvessels was elevated by ~2-fold (n=5, p&lt;0.05), and cerebral infarction became prevalent in SS mice compared to AA mice (n=7, p&lt;0.001). The infarcted brain tissue from SS mice showed neuronal cell death, whereas the brain tissue from the AA mice appeared normal. The SS mice had increased glial fibrillary acid protein positive (GFAP + ) activated astrocytes with significant increase in PLA2 accumulation. Relative abundance of non-phosphorylated neurofilament H (SMI32) and myelin basic protein (MBP) (SMI32/MBP), indicating white matter injury, was substantially induced in the SS compared to the AA. Expression of Anxa1 reduced significantly in plasma as well as within astrocytes’ end feet attached to the isolated cerebral microvessels of the SS mice. Recombinant Anxa1 prophylaxis to SS mice reduced the GFAP + astrocytes, PLA2 accumulation, demyelination and infarct score (n=6, p&lt;0.01) compared to SS mice treated with vehicle. Conclusion: White matter injury in SS mice is associated with astrocyte dysfunction and lack of Anxa1, which may be developed as potential therapeutic targets to prevent and/or treat stroke in SCD.