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1.
Zhonghua Yi Xue Za Zhi ; 101(40): 3329-3337, 2021 Nov 02.
Artigo em Chinês | MEDLINE | ID: mdl-34758534

RESUMO

Objective: To explore the effect and possible mechanism of circ_0023990 on the radiosensitivity of thyroid cancer cells. Methods: qRT-PCR was used to detect the expression of circ_0023990 in the cancer tissues of 55 patients with thyroid cancer and thyroid cancer cell lines (TPC-1, KTC-1, FTC-133 and CAL-62), and the relationship between the expression of circ_0023990 in cancer tissues and the clinical characteristics of the patients were analyzed. Thyroid cancer cells TPC-1 and KTC-1 were divided into sh-circ_0023990 group, sh-NC group, sh-circ_0023990+anti-miR-873-5p group, sh-circ_0023990+anti-miR-NC group, miR-873-5p group, miR-NC group, miR-873-5p+pcDNA-ANXA2 group and miR-873-5p+pcDNA group, and then clone formation experiment was used to detect cell radiosensitivity. After each group of cells was irradiated with 4Gy radiation, the expression of γH2AX protein in the cells was detected by Western Blot. The dual luciferase reporter gene experiment verified the targeting relationship between circ_0023990 and miR-873-5p or miR-873-5p and ANXA2. Results: The expression of circ_0023990 in thyroid cancer tissues was higher than that in normal tissues (2.15±0.09 vs. 0.97±0.05, P<0.05), and its expression was closely related to tumor size, lymph node metastasis and TNM staging of patients with thyroid cancer (P<0.05). The expression of circ_0023990 in thyroid cancer cell lines (TPC-1, KTC-1, FTC-133 and CAL-62) were higher than that of normal thyroid cells HTori-3 (3.16±0.38, 2.63±0.28, 1.82±0.24, 1.71±0.22 vs. 1.00±0.10, all P<0.05). The survival scores of TPC-1 and KTC-1 cells in the sh-circ_0023990 group were significantly lower than those in the sh-NC group (P<0.05), and the sensitization ratios were 2.482, 1.643; The survival scores of TPC-1 and KTC-1 cells in the sh-circ_0023990+anti-miR-873-5p group were higher than those in the sh-circ_0023990+anti-miR-NC group (P<0.05), and the sensitization ratios were 0.305, 0.441, respectively. The survival scores of TPC-1 and KTC-1 cells in the miR-873-5p group were lower than those in the miR-NC group (P<0.05), and the sensitization ratios were 2.044, 1.653 respectively. The survival scores of TPC-1 and KTC-1 cells in the miR-873-5p+pcDNA-ANXA2 group was higher than that in the miR-873-5p+pcDNA group (P<0.05), and the sensitization ratios were 0.496, 0.686, respectively. The expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+sh-circ_0023990 group were higher than that in the 4 Gy+sh-NC group (2.68±0.27 vs. 1.87±0.25, 2.46±0.19 vs. 1.77±0.14; all P<0.05), but the expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+sh-circ_0023990+anti-miR-873-5p group were lower than that in the 4 Gy+sh-circ_0023990+anti-miR-NC group (1.13±0.09 vs. 1.69±0.09, 1.11±0.08 vs. 1.60±0.08; both P<0.05). The expression of γH2AX protein in TPC-1 and KTC-1 cells in the 4 Gy+miR-873-5p group were higher than that in the 4 Gy+miR-NC group (2.35±0.16 vs. 1.84±0.14, 2.26±0.12 vs. 1.77±0.13; both P<0.05), but the expression of γH2AX protein in TPC-1 and KTC-1 cells of the 4 Gy+miR-873-5p+pcDNA-ANXA2 group were lower than that in the 4 Gy+miR-873-5p+pcDNA group (1.96±0.12 vs. 2.41±0.12, 1.92±0.07 vs. 2.28±0.12; both P<0.05). circ_0023990 targeted the negative regulation of miR-873-5p, and ANXA2 was the target gene of miR-873-5p. Conclusion: circ_0023990 was highly expressed in thyroid cancer tissues and cell lines, and it may promote the radiotherapy resistance of thyroid cancer cells in vivo through regulating miR-873-5p/ANXA2 axis.


Assuntos
Anexina A2 , MicroRNAs , Neoplasias da Glândula Tireoide , Linhagem Celular Tumoral , Proliferação de Células , Humanos , MicroRNAs/genética , Tolerância a Radiação/genética , Neoplasias da Glândula Tireoide/genética
2.
Int J Mol Sci ; 22(20)2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-34681689

RESUMO

Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) has infected >235 million people and killed over 4.8 million individuals worldwide. Although vaccines have been developed for prophylactic management, there are no clinically proven antivirals to treat the viral infection. Continuous efforts are being made all over the world to develop effective drugs but these are being delayed by periodic outbreak of mutated SARS-CoV-2 and a lack of knowledge of molecular mechanisms underlying viral pathogenesis and post-infection complications. In this regard, the involvement of Annexin A2 (AnxA2), a lipid-raft related phospholipid-binding protein, in SARS-CoV-2 attachment, internalization, and replication has been discussed. In addition to the evidence from published literature, we have performed in silico docking of viral spike glycoprotein and RNA-dependent RNA polymerase with human AnxA2 to find the molecular interactions. Overall, this review provides the molecular insights into a potential role of AnxA2 in the SARS-CoV-2 pathogenesis and post-infection complications, especially thrombosis, cytokine storm, and insulin resistance.


Assuntos
Anexina A2/metabolismo , COVID-19/patologia , Anexina A2/química , COVID-19/virologia , Síndrome da Liberação de Citocina/metabolismo , Síndrome da Liberação de Citocina/patologia , Humanos , Simulação de Acoplamento Molecular , RNA Polimerase Dependente de RNA/química , RNA Polimerase Dependente de RNA/metabolismo , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Trombose/metabolismo , Trombose/patologia , Internalização do Vírus
3.
Arthritis Res Ther ; 23(1): 266, 2021 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-34702315

RESUMO

BACKGROUND: Connective tissue growth factor (CTGF)-induced angiogenesis is a crucial factor in rheumatoid arthritis (RA), but CTGF-interacting protein and related molecular mechanism of their interaction have not been fully elucidated. METHODS: CTGF-interacting proteins were identified through the LC-MS/MS analysis of the Co-IP products from fibroblast-like synoviocyte (FLS) lysates, and the interaction between CTGF and annexin A2 (ANXA2) was further confirmed through Co-IP and BiFC assay. The binding domain, mutant, mechanism, and angiogenesis function were assessed by homology modeling, molecular docking, MTT, cell scratch, tube formation, and chick chorioallantoic membrane (CAM) assays. Additionally, severe combined immunodeficiency (SCID) mouse co-implantation model was constructed to confirm the effect of ANXA2/CTGF-TSP1 in the process of RA in vivo. RESULTS: ANXA2 was identified and verified as an interaction partner of CTGF for the first time by Co-IP and LC-MS/MS analysis. Co-localization of CTGF and ANXA2 was observed in RA-FLS, and direct interaction of the TSP-1 domain of CTGF and ANXA2 was determined in HEK293T cells. The spatial conformation and stable combination of the ANXA2/CTGF-TSP1 complex were assessed by homology modeling in the biomimetic environment. The function of the ANXA2/CTGF-TSP1 complex was proved on promoting FLS proliferation, migration, and angiogenesis in vitro and deteriorating FLS invasion and joint damage in SCID mice. CONCLUSIONS: TSP-1 is the essential domain in CTGF/ANXA2 interaction and contributes to FLS migration and pannus formation, inducing the process of RA.


Assuntos
Anexina A2 , Artrite Reumatoide , Animais , Anexina A2/genética , Cromatografia Líquida , Fator de Crescimento do Tecido Conjuntivo/genética , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Células HEK293 , Humanos , Camundongos , Simulação de Acoplamento Molecular , Pannus , Espectrometria de Massas em Tandem
4.
Arthritis Res Ther ; 23(1): 265, 2021 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-34696809

RESUMO

BACKGROUND: The development and optimization of therapies for rheumatoid arthritis (RA) is currently hindered by a lack of methods for early non-invasive monitoring of treatment response. Annexin A2, an inflammation-associated protein whose presence and phosphorylation levels are upregulated in RA, represents a potential molecular target for tracking RA treatment response. METHODS: LS301, a near-infrared dye-peptide conjugate that selectively targets tyrosine 23-phosphorylated annexin A2 (pANXA2), was evaluated for its utility in monitoring disease progression, remission, and early response to drug treatment in mouse models of RA by fluorescence imaging. The intraarticular distribution and localization of LS301 relative to pANXA2 was determined by histological and immunohistochemical methods. RESULTS: In mouse models of spontaneous and serum transfer-induced inflammatory arthritis, intravenously administered LS301 showed selective accumulation in regions of joint pathology including paws, ankles, and knees with positive correlation between fluorescent signal and disease severity by clinical scoring. Whole-body near-infrared imaging with LS301 allowed tracking of spontaneous disease remission and the therapeutic response after dexamethasone treatment. Histological analysis showed preferential accumulation of LS301 within the chondrocytes and articular cartilage in arthritic mice, and colocalization was observed between LS301 and pANXA2 in the joint tissue. CONCLUSIONS: We demonstrate that fluorescence imaging with LS301 can be used to monitor the progression, remission, and early response to drug treatment in mouse models of RA. Given the ease of detecting LS301 with portable optical imaging devices, the agent may become a useful early treatment response reporter for arthritis diagnosis and drug evaluation.


Assuntos
Anexina A2 , Artrite Experimental , Artrite Reumatoide , Animais , Artrite Experimental/diagnóstico por imagem , Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/diagnóstico por imagem , Artrite Reumatoide/tratamento farmacológico , Condrócitos , Camundongos , Imagem Óptica , Tirosina
5.
BMC Cancer ; 21(1): 1039, 2021 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-34530774

RESUMO

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) remains a treatment-refractory malignancy with poor prognosis. It is urgent to identify novel and valid biomarkers to predict the progress and prognosis of PDAC. The S100A family have been identified as being involved in cell proliferation, migration and differentiation progression of various cancer types. However, the expression patterns and prognostic values of S100As in PDAC remain to be analyzed. METHODS: We investigated the transcriptional expressions, methylation level and prognostic value of S100As in PDAC patients from the Oncomine, GEPIA2, Linkedomics and cBioPortal databases. Real-time PCR was used to detect the expressions of S100A2/4/6/10/14/16 in four pancreatic cancer cell lines and pancreatic cancer tissues from PDAC patients undergoing surgery. To verify the results further, immunohistochemistry was used to measure the expression of S100A2/4/6/10/14/16 in 43 PDAC patients' tissue samples. The drug relations of S100As were analyzed by using the Drugbank database. RESULTS: The results suggested that, the expression levels of S100A2/4/6/10/14/16 were elevated to PDAC tissues than in normal pancreatic tissues, and the promoter methylation levels of S100A S100A2/4/6/10/14/16 in PDAC (n = 10) were lower compared with normal tissue (n = 184) (P < 0.05). In addition, their expressions were negatively correlated with PDAC patient survival. CONCLUSIONS: Taken together, these results suggest that S100A2/4/6/10/14/16 might be served as prognostic biomarkers for survivals of PDAC patients.


Assuntos
Adenocarcinoma/metabolismo , Biomarcadores Tumorais/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Neoplasias Pancreáticas/metabolismo , Proteínas S100/metabolismo , Adenocarcinoma/mortalidade , Anexina A2/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Carcinoma Ductal Pancreático/mortalidade , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Fatores Quimiotáticos/metabolismo , Bases de Dados Genéticas , Progressão da Doença , Humanos , Pâncreas/metabolismo , Neoplasias Pancreáticas/mortalidade , Prognóstico , RNA Mensageiro/metabolismo , Proteína A6 Ligante de Cálcio S100/metabolismo , Proteína A4 de Ligação a Cálcio da Família S100/metabolismo , Proteínas S100/genética , Transcrição Genética
6.
Int J Mol Sci ; 22(17)2021 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-34502195

RESUMO

Signal transducers and activators of transcription 3 (STAT3) acts as a transcriptional signal transducer, converting cytokine stimulation into specific gene expression. In tumor cells, aberrant activation of the tyrosine kinase pathway leads to excessive and continuous activation of STAT3, which provides further signals for tumor cell growth and surrounding angiogenesis. In this process, the tumor-associated protein Annexin A2 interacts with STAT3 and promotes Tyr705 phosphorylation and STAT3 transcriptional activation. In this study, we found that (20S) ginsenoside Rh2 (G-Rh2), a natural compound inhibitor of Annexin A2, inhibited STAT3 activity in HepG2 cells. (20S) G-Rh2 interfered with the interaction between Annexin A2 and STAT3, and inhibited Tyr705 phosphorylation and subsequent transcriptional activity. The inhibitory activity of STAT3 leaded to the negative regulation of the four VEGFs, which significantly reduced the enhanced growth and migration ability of HUVECs in co-culture system. In addition, (20S)G-Rh2 failed to inhibit STAT3 activity in cells overexpressing (20S)G-Rh2 binding-deficient Annexin A2-K301A mutant, further proving Annexin A2-mediated inhibition of STAT3 by (20S)G-Rh2. These results indicate that (20S)G-Rh2 is a potent inhibitor of STAT3, predicting the potential activity of (20S)G-Rh2 in targeted therapy applications.


Assuntos
Anexina A2/antagonistas & inibidores , Carcinoma Hepatocelular/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Ginsenosídeos/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Fator de Transcrição STAT3/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Anexina A2/genética , Anexina A2/metabolismo , Apoptose , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Ciclo Celular , Movimento Celular , Proliferação de Células , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Células Tumorais Cultivadas , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo
7.
Sci Rep ; 11(1): 10663, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34471143

RESUMO

Early life stress (ELS) causes long-lasting changes in gene expression through epigenetic mechanisms. However, little is known about the effects of ELS in adulthood, specifically across different age groups. In this study, the epigenetic modifications of p11 expression in adult mice subjected to ELS were investigated in different stages of adulthood. Pups experienced maternal separation (MS) for 3 h daily from postnatal day 1 to 21. At young and middle adulthood, behavioral test, hippocampal p11 expression levels, and levels of histone acetylation and methylation and DNA methylation at the hippocampal p11 promoter were measured. Middle-aged, but not young adult, MS mice exhibited increased immobility time in the forced swimming test. Concurrent with reduced hippocampal p11 levels, mice in both age groups showed a decrease in histone acetylation (AcH3) and permissive histone methylation (H3K4me3) at the p11 promoter, as well as an increase in repressive histone methylation (H3K27me3). Moreover, our results showed that the expression, AcH3 and H3Kme3 levels of p11 gene in response to MS were reduced with age. DNA methylation analysis of the p11 promoter revealed increased CpG methylation in middle-aged MS mice only. The results highlight the age-dependent deleterious effects of ELS on the epigenetic modifications of p11 transcription.


Assuntos
Experiências Adversas da Infância/psicologia , Anexina A2/genética , Metilação de DNA/genética , Proteínas S100/genética , Estresse Psicológico/genética , Acetilação , Animais , Ilhas de CpG/genética , Epigênese Genética/genética , Hipocampo/fisiopatologia , Histonas/genética , Humanos , Privação Materna , Camundongos , Regiões Promotoras Genéticas/genética
8.
Biomolecules ; 11(7)2021 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-34356598

RESUMO

S100A10, a member of the S100 family of Ca2+-binding proteins, is a widely distributed protein involved in many cellular and extracellular processes. The best recognized role of S100A10 is the regulation, via interaction with annexin A2, of plasminogen conversion to plasmin. Plasmin, together with other proteases, induces degradation of the extracellular matrix (ECM), which is an important step in tumor progression. Additionally, S100A10 interacts with 5-hydroxytryptamine 1B (5-HT1B) receptor, which influences neurotransmitter binding and, through that, depressive symptoms. Taking this into account, it is evident that S100A10 expression in the cell should be under strict control. In this work, we summarize available literature data concerning the physiological stimuli and transcription factors that influence S100A10 expression. We also present our original results showing for the first time regulation of S100A10 expression by grainyhead-like 2 transcription factor (GRHL2). By applying in silico analysis, we have found two highly conserved GRHL2 binding sites in the 1st intron of the gene encoding S100A10 protein. Using chromatin immunoprecipitation (ChIP) and luciferase assays, we have shown that GRHL2 directly binds to these sites and that this DNA region can affect transcription of S100A10.


Assuntos
Anexina A2 , Simulação por Computador , Proteínas de Ligação a DNA , Regulação Neoplásica da Expressão Gênica , Modelos Biológicos , Proteínas de Neoplasias , Neoplasias , Proteínas S100 , Fatores de Transcrição , Anexina A2/biossíntese , Anexina A2/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Proteínas S100/biossíntese , Proteínas S100/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
9.
Anticancer Res ; 41(8): 4089-4092, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34281879

RESUMO

BACKGROUND/AIM: Serum-derived macrophage activating factor (serum-MAF) is expected to have adjuvant effects through rapid phagocytic activation, which depends on F-actin accumulation in multi-layered membrane ruffles induced within 5 min after serum-MAF addition. This study aimed to elucidate the importance of annexin A2, which is a multifunctional Ca2+-binding protein related to cytoskeletal membrane dynamics, in serum-MAF signalling. MATERIALS AND METHODS: Annexin A2 and F-actin localizations were analyzed via immunostaining and confocal microscopy. Using EGTA as chelator, the role of Ca2+ in serum-MAF signalling was examined. RESULTS: Annexin A2 was found to translocate from the cytosol to the cell cortex within 30 s of serum-MAF stimulation. Ca2+ chelation inhibited the translocation of annexin A2, frill-like structure formation, and phagocytic activation by serum-MAF. CONCLUSION: Annexin A2 and Ca2+ were responsible for the rapid phagocytic activation by serum-MAF. This study provides an understanding of phagocytic activation in macrophages, which could be beneficial for cancer immunotherapy.


Assuntos
Anexina A2/metabolismo , Ativação de Macrófagos , Fatores Ativadores de Macrófagos/sangue , Macrófagos/fisiologia , Actinas/metabolismo , Humanos , Fagocitose , Células THP-1
10.
Int J Mol Sci ; 22(13)2021 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202091

RESUMO

As a cell surface tissue plasminogen activator (tPA)-plasminogen receptor, the annexin A2 (A2) complex facilitates plasmin generation on the endothelial cell surface, and is an established regulator of hemostasis. Whereas A2 is overexpressed in hemorrhagic disease such as acute promyelocytic leukemia, its underexpression or impairment may result in thrombosis, as in antiphospholipid syndrome, venous thromboembolism, or atherosclerosis. Within immune response cells, A2 orchestrates membrane repair, vesicle fusion, and cytoskeletal organization, thus playing a critical role in inflammatory response and tissue injury. Dysregulation of A2 is evident in multiple human disorders, and may contribute to the pathogenesis of various inflammatory disorders. The fibrinolytic system, moreover, is central to wound healing through its ability to remodel the provisional matrix and promote angiogenesis. A2 dysfunction may also promote tissue fibrogenesis and end-organ fibrosis.


Assuntos
Anexina A2/genética , Suscetibilidade a Doenças , Fibrinólise/genética , Fibrose/etiologia , Inflamação/etiologia , Animais , Anexina A2/metabolismo , Doenças Autoimunes/etiologia , Doenças Autoimunes/metabolismo , Biomarcadores , Fibrose/metabolismo , Hemostasia/genética , Humanos , Imunidade , Inflamação/metabolismo , Especificidade de Órgãos , Regeneração
12.
Int J Mol Sci ; 22(9)2021 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-34064456

RESUMO

Primary Sjögren's syndrome (pSS) is a complex heterogeneous disease characterized by a wide spectrum of glandular and extra-glandular manifestations. In this pilot study, a SWATH-MS approach was used to monitor extracellular vesicles-enriched saliva (EVs) sub-proteome in pSS patients, to compare it with whole saliva (WS) proteome, and assess differential expressed proteins between pSS and healthy control EVs samples. Comparison between EVs and WS led to the characterization of compartment-specific proteins with a moderate degree of overlap. A total of 290 proteins were identified and quantified in EVs from healthy and pSS patients. Among those, 121 proteins were found to be differentially expressed in pSS, 82% were found to be upregulated, and 18% downregulated in pSS samples. The most representative functional pathways associated to the protein networks were related to immune-innate response, including several members of S100 protein family, annexin A2, resistin, serpin peptidase inhibitors, azurocidin, and CD14 monocyte differentiation antigen. Our results highlight the usefulness of EVs for the discovery of novel salivary-omic biomarkers and open novel perspectives in pSS for the identification of proteins of clinical relevance that could be used not only for the disease diagnosis but also to improve patients' stratification and treatment-monitoring. Data are available via ProteomeXchange with identifier PXD025649.


Assuntos
Vesículas Extracelulares/metabolismo , Redes Reguladoras de Genes , Proteoma/genética , Saliva/metabolismo , Síndrome de Sjogren/genética , Adulto , Idoso , Anexina A2/genética , Anexina A2/metabolismo , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/metabolismo , Biomarcadores/metabolismo , Proteínas Sanguíneas/genética , Proteínas Sanguíneas/metabolismo , Estudos de Casos e Controles , Vesículas Extracelulares/química , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/metabolismo , Masculino , Espectrometria de Massas/métodos , Pessoa de Meia-Idade , Projetos Piloto , Mapeamento de Interação de Proteínas , Proteoma/classificação , Proteoma/metabolismo , Proteômica/instrumentação , Proteômica/métodos , Resistina/genética , Resistina/metabolismo , Proteínas S100/genética , Proteínas S100/metabolismo , Saliva/química , Serpinas/genética , Serpinas/metabolismo , Síndrome de Sjogren/diagnóstico , Síndrome de Sjogren/metabolismo , Síndrome de Sjogren/patologia
13.
Mol Cell Biochem ; 476(10): 3563-3575, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34018148

RESUMO

Lung cancer is still a main cause of cancer-related death worldwide. Non-small-cell lung cancer (NSCLC) accounts for the majority of lung cancers, and gefitinib is an effective targeted drug for NSCLC. It is important to explore the underlying molecular mechanisms of gefitinib resistance to provide new treatment strategies and to improve the prognosis of gefitinib-resistant NSCLC patients. This study aimed to examine the role of filamin A (FLNA) in acquired resistance to gefitinib in NSCLC, and identify ANXA2 (annexin A2), one of calcium-dependent phospholipid-binding proteins, as its corresponding regulatory factor. First, we established resistant cells via long-term exposure to gefitinib to analyse the association between FLNA and gefitinib resistance. Through quantitative real-time polymerase chain reaction (qRT-PCR), Cell Counting Kit-8 (CCK-8), western blotting (WB), and flow cytometry assays, we evaluated the role of FLNA. The effect of FLNA knockdown or overexpression was analysed not only in cell lines but also in mouse models. We verified the FLNA-interacting protein through coimmunoprecipitation (CoIP) experiments and found that the downstream signalling pathway was regulated by FLNA and its interacting protein. Finally, the upstream transcription factor was identified by chromatin immunoprecipitation (ChIP). Increased FLNA expression induced gefitinib resistance. Knockdown of FLNA restored gefitinib sensitivity and induced apoptosis in vivo and in vitro. FLNA and ANXA2 cooperatively led to the activation of the Wnt pathway, which was closely linked to gefitinib resistance. Subsequently, SP1 promoted transcriptional activation of FLNA to regulate gefitinib resistance. We determined that FLNA serves as a regulator of gefitinib resistance in NSCLC and found that FLNA and ANXA2 together induced gefitinib resistance by activating the Wnt pathway.


Assuntos
Anexina A2/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Filaminas/metabolismo , Gefitinibe/farmacologia , Neoplasias Pulmonares/metabolismo , Proteínas de Neoplasias/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Anexina A2/genética , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Filaminas/genética , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Proteínas de Neoplasias/genética
14.
Biomed Pharmacother ; 140: 111744, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34049223

RESUMO

Endometrial cancer (EC) is the most common neoplasm of the female reproductive tract in the developed world. Patients usually are diagnosed in early stage having a good prognosis. However, up to 20-25% of patients are diagnosed in advanced stages and have a higher risk of recurrence, making the prognosis worse. Previously studies identified ANXA2 as a predictor of recurrent disease in EC even in low risk patients. Furthermore, Circulating Tumor Cells (CTC) released from the primary tumor into the bloodstream, are plasticity entities responsible of the process of metastasis, becoming into an attractive clinical target. In this work we validated ANXA2 expression in CTC from high-risk EC patients. After that, we modelled in vitro and in vivo the tumor cell attachment of ANXA2-expressing CTC to the endothelium and the homing for the generation of micrometastasis. ANXA2 overexpression does not provide an advantage in the adhesion process of CTC, but it could be playing an important role in more advanced steps, conferring a greater homing capacity. We also performed a high-throughput screening (HTS) for compounds specifically targeting ANXA2, and selected Daunorubicin as candidate hit. Finally, we validated Daunorubicin in a 3D transendothelial migration system and also in a in vivo model of advanced EC, demonstrating the ability of Daunorubicin to inhibit the proliferation of ANXA2-overexpressing tumor cells.


Assuntos
Anexina A2/metabolismo , Neoplasias do Endométrio/metabolismo , Neoplasias do Endométrio/patologia , Animais , Anexina A2/genética , Adesão Celular , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Daunorrubicina/farmacologia , Daunorrubicina/uso terapêutico , Neoplasias do Endométrio/tratamento farmacológico , Neoplasias do Endométrio/genética , Endotélio/fisiologia , Feminino , Ensaios de Triagem em Larga Escala , Humanos , Biópsia Líquida , Camundongos , Modelos Biológicos , Células Neoplásicas Circulantes
15.
Life Sci ; 279: 119645, 2021 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-34043991

RESUMO

AIMS: Norcantharidin (NCTD) exhibits antitumor, anti-inflammatory, and anti-fibrosis properties, which makes NCTD an attractive candidate for the treatment of pathological scars. This study was designed to investigate the potential effects of NCTD on fibroblast proliferation and explore the underlying mechanisms. MATERIALS AND METHODS: First, cell viability and cell apoptosis were evaluated to determine the effects of NCTD on human skin fibroblasts, at 10, 50, and 100 µM. To explore the mechanism, bioinformatics analyses, chromatin immunoprecipitation, RNA immunoprecipitation, and RNA pulldown assays, and luciferase reporter assays were performed to verify the relationships among NCTD, signal transducer and activator of transcription 3 (STAT3), annexin A2 pseudogene 2 (ANXA2P2), and ubiquitin-associated protein 2-like (UBAP2L) mRNA in fibroblasts. Loss-of-function experiments were performed to investigate the roles played by STAT3, ANXA2P2, and UBAP2L in the proliferation and apoptosis of fibroblasts. KEY FINDINGS: We found that NCTD administration induced fibroblast apoptosis and inhibited fibroblast proliferation in a dose-dependent manner. Mechanistically, NCTD inhibited ANXA2P2 transcription through the inhibition of STAT3 phosphorylation. Subsequently, ANXA2P2 was found to enhance the physical interaction between UBAP2L mRNA and lin-28 homolog B (LIN28B), which increased the stability and levels of UBAP2L mRNA. Loss-of-function assays demonstrated that ANXA2P2 and UBAP2L knockdown induced fibroblast apoptosis and suppressed fibroblast proliferation. SIGNIFICANCE: In conclusion, we confirmed that NCTD inhibits fibroblast proliferation by inhibiting the STAT3/ANXA2P2/UBAP2L axis, which suggested that NCTD could represent a new candidate for the treatment of pathological scars.


Assuntos
Anexina A2/metabolismo , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Proteínas de Transporte/metabolismo , Proliferação de Células , Fibroblastos/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Estabilidade de RNA/efeitos dos fármacos , Proteínas de Ligação a RNA/metabolismo , Anexina A2/genética , Antineoplásicos/farmacologia , Apoptose , Proteínas de Transporte/genética , Células Cultivadas , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Pseudogenes , Proteínas de Ligação a RNA/genética
16.
Mol Med Rep ; 24(1)2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33955501

RESUMO

The aim of the present study was to investigate the effects of human epididymis protein 4 (HE4) on drug resistance and its underlying mechanisms. The associations among proteins were detected by immunoprecipitation and immunofluorescence assays. Then, stably transfected cell lines CAOV3­HE4­L and CAOV3­A2­L expressing HE4 short hairpin (sh)RNAs and ANXA2 shRNAs, respectively, were constructed. MTT assay, immunocytochemistry, western blotting, reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and flow cytometry were employed to examine drug sensitivity, as well as the expression and activity of P­glycoprotein (P­gp). HE4 and P­gp in epithelial ovarian cancer tissues were assessed via immunohistochemistry. MicroRNAs that targeted the P­gp gene, ABCB1, were predicted using bioinformatics methods, and their expression was evaluated by RT­qPCR. The common signaling pathways shared by HE4, ANXA2 and P­gp were selected by Gene Set Enrichment Analysis (GSEA). The interaction of HE4, ANXA2 and P­gp were confirmed. P­gp expression was positively associated with HE4 and ANXA2 expression, respectively. Moreover, it was observed that there was no significant rescue of P­gp expression in CAOV3­A2­L cells following the administration of active HE4 protein. In addition, the expression of HE4 and P­gp in ovarian cancer tissues of drug­resistant patients were higher compared with that of the drug­sensitive group (P<0.05). Furthermore, the results revealed that hsa­miR­129­5p was significantly increased accompanied by decreased HE4 or ANXA2 expression and P­gp expression in CAOV3­HE4­L and CAOV3­A2­L cells. GSEA analyses disclosed that HE4, ANXA2 and P­gp genes were commonly enriched in the signaling pathway involved in regulating the actin cytoskeleton. These results indicated that HE4 promotes P­gp­mediated drug resistance in ovarian cancer cells through the interactions with ANXA2, and the underlying mechanism may be associated with decreased expression of hsa­miR­129­5p and dysregulation of the actin cytoskeleton signaling pathway.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Anexina A2/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Epiteliais e Glandulares/genética , Neoplasias Ovarianas/genética , Proteína 2 do Domínio Central WAP de Quatro Dissulfetos/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Citoesqueleto de Actina/genética , Citoesqueleto de Actina/metabolismo , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Doxorrubicina/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/metabolismo , Neoplasias Epiteliais e Glandulares/metabolismo , Neoplasias Epiteliais e Glandulares/patologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Prognóstico , Transdução de Sinais/genética , Análise de Sobrevida , Proteína 2 do Domínio Central WAP de Quatro Dissulfetos/genética
17.
PLoS One ; 16(5): e0251240, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33989302

RESUMO

We have established a new screening system for identifying interacting proteins by combining biomolecular fluorescence complementation (BiFC) and a transposon gene trap system. This system requires creation of a bait strain that stably expresses a fusion product of part of the fluorescent monomeric Kusabira-Green (mKG) protein to a protein of interest. A PiggyBac transposon vector is then introduced into this strain, and a sequence encoding the remainder of mKG is inserted into the genome and fused randomly with endogenous genes. The binding partner can be identified by isolating cells that fluoresce when BiFC occurs. Using this system, we screened for interactors of p65 (also known as RELA), an NF-κB subunit, and isolated a number of mKG-positive clones. 5'- or 3'-RACE to produce cDNAs encoding mKG-fragment fusion genes and subsequent reconstitution assay identified PKM, HSP90AB1, ANXA2, HSPA8, and CACYBP as p65 interactors. All of these, with the exception of CACYBP, are known regulators of NF-κB. Immunoprecipitation assay confirmed endogenously expressed CACYBP and p65 formed a complex. A reporter assay revealed that CACYBP enhanced 3κB reporter activation under TNFα stimulation. This screening system therefore represents a valuable method for identifying interacting factors that have not been identified by other methods.


Assuntos
Elementos de DNA Transponíveis/genética , Mapeamento de Interação de Proteínas/métodos , Mapas de Interação de Proteínas/fisiologia , Fator de Transcrição RelA/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Anexina A2/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Linhagem Celular Tumoral , Fluorescência , Células HEK293 , Proteínas de Choque Térmico HSC70/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Células HeLa , Humanos , Microscopia de Fluorescência/métodos , Ligação Proteica , Fator de Transcrição RelA/genética
18.
Asian Pac J Cancer Prev ; 22(5): 1459-1466, 2021 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-34048174

RESUMO

OBJECTIVE: Immediate single instillation of chemotherapy following transurethral resection of bladder tumor (TURBT) is suggested for non-muscle invasive bladder cancer (NMIBC) patients. However, no study has evaluated molecular marker that was involved in intravesical recurrence (IVR) after single instillation of chemotherapy. Therefore, this study aimed to evaluate whether P-glycoprotein, multidrug resistance protein 1 (MRP1), Annexin A2 (ANXA2) or nucleophosmin (NPM) expression predicts IVR after initial TURBT and immediate single intravesical adriamycin instillation. METHODS: We retrospectively reviewed consecutive 443 patients who underwent TURBT. Of these, 54 patients who underwent initial TURBT and single instillation of adriamycin for NMIBC were included. The expressions of P-glycoprotein, MRP1, ANXA2 and NPM were evaluated immunohistochemically and were divided into 2 groups (low or high) according to the staining intensity and/or proportion of positive cells. IVR was assessed by Kaplan-Meier method. Cox`s multivaritate analyses were performed to identify independent predictors for IVR. RESULTS: Nineteen patients (35.1%) had IVR. High P-glycoprotein expression was significantly correlated with multiplicity, pT stage and high grade. High ANXA2 expression was significantly correlated with high grade. MRP1 and NPM were not correlated with any clinicopathological variables. MRP1 expression and ANXA2 expression were significantly correlated with P-glycoprotein expression. Patients with high P-glycoprotein expression had significantly worse IVR-free survival (IVRFS) than those with low P-glycoprotein expression (P =0.015). The difference in IVRFS rates between patients with high ANXA2 expression and those with low ANXA2 expression was nearly significant (P =0.057). Univariate analyses indicated multiplicity, high grade and high P-glycoprotein expression were significant predictors for IVR. Multivariate analysis indicated high grade was an independent predictor for IVR. CONCLUSIONS: High P-glycoprotein expression was associated with IVR. Further study was needed to determine significance of P-glycoprotein expression in IVR after single intravesical adriamycin instillation.


Assuntos
Anexina A2/metabolismo , Cistectomia/mortalidade , Doxorrubicina/administração & dosagem , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Recidiva Local de Neoplasia/patologia , Neoplasias da Bexiga Urinária/patologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Idoso , Antibióticos Antineoplásicos/administração & dosagem , Biomarcadores Tumorais/metabolismo , Terapia Combinada , Feminino , Seguimentos , Humanos , Masculino , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/terapia , Proteínas Nucleares/metabolismo , Prognóstico , Estudos Retrospectivos , Taxa de Sobrevida , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/terapia
19.
Environ Toxicol ; 36(8): 1576-1590, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33913242

RESUMO

The purpose of this article was to probe the mechanism underlying long noncoding RNA (lncRNA)-LINC00184 in cholangiocarcinoma development and to investigate the effects of LINC00184 on cholangiocarcinoma. We used bioinformatics to analyze the expression of LINC00184, microRNA (miR)-23b-3p and ANXA2 in cholangiocarcinoma tissues. The levels of LINC00184, miR-23b-3p, and ANXA2 were detected by qRT-PCR. Cell proliferation was tested by CCK8. Transwell assay was used to detect cell invasion and migration. The target connection between LINC00184, miR-23b-3p, or ANXA2 was probed by luciferase reporter assay. RNA pull-down method was employed to test the relationship among LINC00184/miR-23b-3p/ANXA2 in cholangiocarcinoma cells. The Pearson correlation coefficient analyzed was applied to analyze the correlation among LINC00184, miR-23b-3p, and ANXA2. LC-MS/M analysis was used to explore whether the changes of adenine metabolism was affected by LINC00184 in cholangiocarcinoma cells. We discovered that LINC00184 expression was heightened in cholangiocarcinoma patients and cells. Knockdown of LINC00184 repressed cell proliferation, invasion, migration and adenine metabolism in cholangiocarcinoma cells. miR-23b-3p was regarded as a target of LINC00184 and its depletion perversed the inhibitive influence of LINC00184 silencing on cholangiocarcinoma cells. ANXA2 was a target of miR-23b-3p and was negatively modulated by miR-23b-3p. Moreover, ANXA2 was positively modulated by LINC00184 via sponging miR-23b-3p. In short, silencing of LINC00184 suppressed cell proliferation, invasion and migration through over-expression of miR-23b-3p and reducing of ANXA2 in cholangiocarcinoma cells. These findings contribute to understanding the influences of LINC00184, miR-23b-3p, and ANXA2 on cholangiocarcinoma and provide basis for cholangiocarcinoma treatment.


Assuntos
Anexina A2 , Neoplasias dos Ductos Biliares , Colangiocarcinoma , MicroRNAs , RNA Longo não Codificante , Adenina , Ductos Biliares Intra-Hepáticos , Movimento Celular , Proliferação de Células , Humanos
20.
Int J Mol Sci ; 22(7)2021 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-33810523

RESUMO

Routine manipulation of the mouse genome has become a landmark in biomedical research. Traits that are only associated with advanced developmental stages can now be investigated within a living organism, and the in vivo analysis of corresponding phenotypes and functions advances the translation into the clinical setting. The annexins, a family of closely related calcium (Ca2+)- and lipid-binding proteins, are found at various intra- and extracellular locations, and interact with a broad range of membrane lipids and proteins. Their impacts on cellular functions has been extensively assessed in vitro, yet annexin-deficient mouse models generally develop normally and do not display obvious phenotypes. Only in recent years, studies examining genetically modified annexin mouse models which were exposed to stress conditions mimicking human disease often revealed striking phenotypes. This review is the first comprehensive overview of annexin-related research using animal models and their exciting future use for relevant issues in biology and experimental medicine.


Assuntos
Anexina A1/metabolismo , Lipídeos/química , Pesquisa Médica Translacional , Animais , Anexina A2/metabolismo , Anexina A5/metabolismo , Anti-Inflamatórios/metabolismo , Anti-Inflamatórios/farmacologia , Cálcio/química , Membrana Celular/metabolismo , Diabetes Mellitus/metabolismo , Progressão da Doença , Homeostase , Camundongos , Camundongos Knockout , Nanotecnologia , Neoplasias/metabolismo , Neovascularização Patológica , Peptídeos/química , Fenótipo , Ligação Proteica , Transporte Proteico
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