OBJECTIVETo study the expression of annexin II (AnnII) and the fibrinolytic activity in NB4 cells and their alterations in the presence of arsenic trioxide (ATO) and daunorubicin (DNR).

METHODSLeukemia cell line NB4 was treated with ATO or DNR for 24 ∼ 72 h. Cell surface expression of AnnII and its mRNA were analysed by flow cytometry and real time PCR, respectively, the fibrinolytic activity by chromogenic assay.

RESULTSCompared with other acute leukemia cell lines, the expression of AnnII on untreated NB4 cells was relatively higher. The AnnII positive cell rates on NB4, HL-60, K562, and A3 cells were (94.5 ± 1.6)%, (40.1 ± 2.1)%, (36.3 ± 1.5)% and (11.8 ± 2.5)%, respectively. The fibrinolytic activity of NB4 cells was the greatest with a A value of 0.68 ± 0.02. The fibrinolytic activity of NB4 cells was obviously decreased by ATO, DNR or monoclonal antibody against AnnII, being decreased by 60.4%, 35.8% and 26.0% of the pretreatment level, respectively. The expressions of AnnII and its mRNA in NB4 cells were decreased dramatically after ATO and DNR treated for 48 h. Annexin II positive cells rate were (55.46 ± 4.72)% and (27.00 ± 6.18)%, respectively.

CONCLUSIONNB4 cells have strong ability to enhance the catalytic efficiency of the t-PA-dependent plasminogen activation and AnnII on the cell membrane contributes to this activity. Its high fibrinolytic activity can be corrected by ATO and DNR through down-regulating AnnII.

Annexin A2 ; Apoptosis ; Daunorubicin ; HL-60 Cells ; Humans ; Leukemia ; metabolism

OBJECTIVETo explore effect of all-trans retinoic acid(ATRA) on annexin Ⅱ expression in NB4 cells and to analyze the luciferase activity of annexinⅡ promoter in condition of ATRA-induced treatment.

METHODSNB4 cells were cultured in vitro, the transcriptional or translational expression levels of Annexin Ⅱ in NB4 cells treated with 1 µmol/L ATRA at different time points were detected by RT-PCR or Western blot respectively. Annexin Ⅱ-promoter was constructed, the recombinant plasmids pGL4.15 -Annexin Ⅱ -promoter were transfected into NB4 cells with electroporation, and after being treated with 1 µmol/L ATRA for 24 hours the luciferase acttivity of Annexin Ⅱ promoter was determined by luciferase activity assay.

RESULTSThe transcriptional expression of Annexin Ⅱ was down-regulated after 48 h. The translation expression of Annexin Ⅱ was slowly weakened after 24 h, and it was seriously reduced after 48 h. Further, Luciferase activity of AnnexinⅡ promoter in NB4 cells treated with 1 µmol/L ATRA was down-regulated, and showed a decreased tendency at indicated time points.

CONCLUSIONAll-trans retinoic acid can induce the down-regulation of AnnexinⅡ expression on the membrane of NB4 cells, and the activity of Annexin Ⅱpromoter is down-regulated too. This study provide a basis for further study of molecular mechanism.

Annexin A2 ; Cell Differentiation ; Cell Line, Tumor ; Down-Regulation ; Humans ; Tretinoin

OBJECTIVE: The aim of this study was to determine annexin II expression in cervical cancer. METHODS: In Ewha Womans University Mokdong Hospital, normal and cervical cancer tissues were obtained from healthy women (n=11), from patients with cervical intraepithelial neoplasia (CIN, n=11) and from patients with cervical cancer (n=33). The expressions of annexin II mRNA and protein were examined by quantitative competitive-PCR and by western blot analysis. Annexin II mRNA and protein expressions were examined with repects to the clinical characteristics including tumor sizes and cancer stages. RESULTS: The expression of annexin II mRNA in cervical cancer was higher than that in the normal cervix, and CIN (p<0.05). Annexin II mRNA expression was not correlated with cervical cancer stage, or size of the tumor (p>0.05). The expression of annexin II protein in cervical cancer was higher than that in CIN but its expression was decreased according to the cervical cancer stage. CONCLUSION: Our results suggest that overexpression of annexin II mRNA and protein may be a biologic marker of cervical carcinogenesis.
Annexin A2* ; Biomarkers ; Blotting, Western ; Carcinogenesis ; Cervical Intraepithelial Neoplasia ; Cervix Uteri ; Female ; Humans ; RNA, Messenger ; Uterine Cervical Neoplasms*

OBJECTIVETo investigate the effects of bortezomib on the migration of endothelial cells and the expression of angiogenesis-related molecules, and explore the mechanism of its antiproliferation of tumor cells.

METHODSCell count kit CCK-8 was used to detect the relative proliferation activity of cells after treated by bortezomib at different concentrations for 12 h and 24 h, respectively. Transwell model was uesd to detect the migration rate of cells. Expression levels of VEGF and Annexin A2 genes were determined by real-time quantitative PCR. Annexin A2 protein was validated by Western blot.

RESULTSAfter treated with bortezomib at concentrations of 2.5, 5.0 and 10 nmol/L for 12h, respectively, the HMEC-1 cell proliferation activity was 1.004 ± 0.002, 0.793 ± 0.021 and 0.874 ± 0.062, respectively, being no statistical difference from that of control group (1.000) P < 0.05); while the migration rates of them were 0.697 ± 0.060, 0.597 ± 0.090 and 0.874 ± 0.062, respectively, being significantly lower than that of control group (1.000) (P < 0.05) and so did for the expression of VEGF and Annexin A2 genes. After treated with 5 nmol/L bortezomib for 12 h, the Annexin A2 and VEGF gene relative expression level of HMEC-1 cells was 0.540 ± 0.001 and 0.793 ± 0.153, respectively, being of statistical difference from that of control group (1.000) P < 0.05). The conspicuous downregulation of Annexin A2 protein was also confirmed by Western Blot.

CONCLUSIONSBortezomib can inhibit migration of endothelial cell HMEC-1 by downregulating the expression of VEGF and Annexin A2, displaying a new mechanism of bortezomib for inhibition of tumor proliferation.

Annexin A2 ; metabolism ; Bortezomib ; Cell Proliferation ; drug effects ; Endothelial Cells ; metabolism ; Humans ; RNA, Messenger ; genetics ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVETo investigate the effects of Annexin II (AnxA2) gene silencing by siRNA on proliferation and invasive potential of lymphoma cell line Jurkat cells.

METHODSA synthesized siRNA duplex targeting to AnxA2 was transfected into Jurkat cells. Transfection efficiency was analyzed by real-time PCR and flow cytometry. MTT assay for cell proliferation and transwell plates for invasive potential were performed.

RESULTSCompared with the negative controls, the cell proliferation inhibitory rate of the AnxA2 siRNA transfected Jurkat cells was significantly increased at 24 h, 48 h and 72 h [(17.4 +/- 2.3)%, (22.4 +/- 3.8)%, (37.6 +/- 1.5)% vs (-1.3 +/- 5.1)%, (-5.5 +/- 4.4)%, (-10.8 +/- 5.5)%, respectively, P<0.05]. The cell invasive potential of the transfected Jurkat cells was inhibited remarkably at 48 h (11.3 +/- 4.2 vs 54.3 +/- 8.7, P<0.01).

CONCLUSIONAnxA2 gene silenced by siRNA can inhibit the proliferation and the invasive potential of Jurkat cells remarkably.

Annexin A2 ; genetics ; metabolism ; Cell Proliferation ; Chemotaxis ; genetics ; Gene Silencing ; Humans ; Jurkat Cells ; RNA, Small Interfering ; genetics ; Transfection

This study was purposed to investigate the apoptosis-inducing effect of Annexin A2 gene (AnxA2) on multiple myeloma (MM) cells and its mechanisms. The human MM cell lines U266 and RPMI8226 were transfected by using siRNA targeting at AnxA2; the expressions of AnxA2 mRNA and protein in the siRNA-transfected cells were detected by real-time PCR and Western blot, respectively; the cell apoptosis was assayed by flow cytometry. The results showed that silencing AnxA2 gene by siRNA resulted in decreased expressions of AnxA2 gene and protein, increased apoptosis of U266 and RPMI8226 cell lines (P < 0.05), at the same time resulted in down-regulation of apoptosis-related gene expressions including p65NF-κB, IL-2, IL-6 (P < 0.05), and up-regulation of P53 gene expression (P < 0.05). It is concluded that the AnxA2 silence plays a promoting role in apoptosis of MM cell lines U266 and RPMI8226.
Annexin A2 ; genetics ; Apoptosis ; genetics ; Cell Line, Tumor ; Humans ; Multiple Myeloma ; genetics ; pathology ; RNA, Small Interfering ; genetics

BACKGROUND: Fluoranthene (FR) is a common environmental pollutant that exists in a complex mixture with other polycyclic aromatic hydrocarbons (PAHs). We identified biomarkers for monitoring FR exposure and investigated the rescue effect of FR-induced cellular toxicity via aryl hydrocarbon receptor (AHR) antagonist activity in bone marrow derived mesenchymal stem cells (BM-MSCs).METHODS: Morphological changes, viability, and rescue effects of an AHR antagonist (CH223191) were examined in BM-MSCs after exposure to FR. Cytotoxic effects were assayed using the tetrazolium-based colorimetric assay. Apoptosis was measured by annexin V and propidium iodide dye-based flowcytometry assay, mitochondrial membrane potential assay, and nuclear DNA fragmentation assay. Molecular signaling pathways of apoptosis and autophagy were investigated using immunoblotting. Proteomics were performed in order to reveal the spectra of cellular damage and identify biomarkers for FR exposure.RESULTS: Exposing BM-MSCs to FR (IC₅₀=50 µM) induced cell death and morphological changes, while the AHR antagonist showed rescue effects. Autophagy was activated and mitochondrial membrane potential was decreased. Proteomic analysis identified 48 deregulated proteins (26 upregulated and 22 downregulated). Among them, annexin A6, pyruvate kinase, UDP-glucose dehydrogenase, and phospholipase A2 could be potential biomarkers for FR exposure.CONCLUSION: The exposure of BM-MSCs to FR induced remarkable alterations in cellular biology and the proteome, allowing for identification of novel biomarkers for FR exposure. Furthermore, AHR antagonists might be able to prevent cellular damage due to FR exposure.
Annexin A5 ; Annexin A6 ; Apoptosis ; Autophagy ; Biomarkers ; Bone Marrow ; Cell Death ; DNA Fragmentation ; Immunoblotting ; Membrane Potential, Mitochondrial ; Mesenchymal Stromal Cells ; Oxidoreductases ; Phospholipases A2 ; Polycyclic Hydrocarbons, Aromatic ; Propidium ; Proteome ; Proteomics ; Pyruvate Kinase ; Receptors, Aryl Hydrocarbon

Lipocortin represents a family of similar Ca++ depentent phospholipid-binding proteins capable of blocking the activity of phospholipase A2 (PLA2) in vitro. Generally, these proteins are believed to inhibit the release of arachidonic acid from photopholipids and the formation of lipid mediators such as prostaglandin, leukotriene, and platelet activating factor. Lipocortin 1, initially identified as a glucocorticoid- responsive protein in macrophages and neutrophils has been implicated in transmembrane signal transduction during growth factor-mediated cell proliferation and transformation. To define the synthesis and its regulation, we investigated the expression of lipocortin 1 in both the mRNA and protein level in U937 cell line in the presence of several differentiation factors. The results were as follows. 1. The expression of lipocortin 1 and its mRNA was increased during TPA-induced differentiation of U937 cells to maximum of 2-fold and 5-fold respectively. Both the protein and mRNA levels decreased after 48 hours. 2. With the treatment with IFN-gamma, the expression of CD16 was increased. However, the protein and mRNA levels of lipocortin 1 were, not changed significantly. 3. Neither the dexamethasone or hydrocortisone have any effects on the expression of lipocortin 1 in both TPA-differentiated and undifferentiated U937 cells. The results from this study would give a light on defining the functional role of lipocortin 1 in macro-moncycle cell lineage and possibly some informative clues for the pathogenic mechanisms of the inflammatory diseases.
Annexin A1* ; Annexins* ; Arachidonic Acid ; Cell Lineage ; Cell Proliferation ; Dexamethasone ; Humans ; Hydrocortisone ; Macrophages ; Neutrophils ; Phospholipases A2 ; Platelet Activating Factor ; RNA, Messenger ; Signal Transduction ; U937 Cells*

OBJECTIVE: Granulocyte-macrophage colony-stimulating factor (GM-CSF) facilitates mammalian embryonic development and implantation. However, its biological function after implantation is not elucidated. The aim of this study is to assess the changes of gene expression by GM-CSF in human trophoblast obtained in early pregnancy. METHODS: Human trophoblast obtained in early pregnancy was cultured with or without GM-CSF. The difference of gene expression was evaluated with microarray and selected genes were reevaluated with real-time reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Microarray analysis revealed that the expressions of 468 genes were increased while those of 40 genes were decreased by GM-CSF. These genes were evaluated according to the known biologic pathways. The regulation of actin cytoskeleton and focal adhesion pathways were mostly influenced by GM-CSF. Annexin A2, thymosin-like 3, vimentin, myogenin, ACK1, and tensin1 genes were selected for real-time RT-PCR. The increased expressions of of vimentin and ACK1, and decreased expressions of tensin1 were confirmed by real-time RT-PCR. CONCLUSION: GM-CSF activates focal adhesion pathway in human trophoblast by increasing the expression of vimentin and ACK1, and decreasing the expression of tensin1.
Actin Cytoskeleton ; Annexin A2 ; Embryonic Development ; Female ; Focal Adhesions ; Gene Expression ; Granulocyte-Macrophage Colony-Stimulating Factor ; Humans ; Microarray Analysis ; Myogenin ; Pregnancy ; Trophoblasts ; Vimentin

Ischemia-reperfusion injury is complicated with multiple injury pathways. If a particular agent is used to restore blood flow and prevent cell death, many damaged neurons may come back to life. However, for stroke victims, there are no effective curative therapeutic approaches available other than thrombolytic treatment. The efficacy of neuroprotective agents are limited by low diffusion, narrow time window, strict dose titration, and lack of confidence at the preclinical level. NXY-059 reflects the fundamental limitation of neuroprotectant. There are recent attempts to overcome these limitations, with use of annexin A2, fingolimod, hydrogen, nitrite, and more. By covering two components, this report reviews what we have recently learned. In addition, it sheds light on some of the newer issues in clinical application.
Annexin A2 ; Benzenesulfonates ; Cell Death ; Diffusion ; Hydrogen ; Light ; Multiple Trauma ; Neurons ; Neuroprotective Agents ; Propylene Glycols ; Reperfusion Injury ; Sphingosine ; Stroke ; Fingolimod Hydrochloride