The Role of Beta 2 Integrins in Macrophage Migration During Resolution of Inflammation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3600-3600 ◽  
Author(s):  
Yevgeniya Kushchayeva ◽  
Darya Mishchuk ◽  
Tatiana Ugarova
Keyword(s):  
Lymph Nodes ◽  
Conflicts Of Interest ◽  
Fold Increase ◽  
Initial Population ◽  
Blood Monocytes ◽  
Resident Cell ◽  
Low Levels ◽  
Inflammatory Macrophages ◽  
Β2 Integrins

Abstract Abstract 3600 Poster Board III-537 The mobilization of blood monocytes and their differentiation into macrophages during the immune-inflammatory response helps to prepare the tissue for resolution. During the resolution phase of inflammation macrophages do not die locally: some cells emigrate by draining lymphatics whereas some remain at the site of inflammation. The major myelo-monocytic integrin αMβ2 (Mac-1, CD11b/CD18), together with two related integrins αDβ2 (CD11d/CD18) and αXβ2 (CD11c/CD18), mediate critical adhesive reactions of monocyte/macrophages. However, the roles of these adhesion receptors in control of macrophage retention at sites of inflammation and their emigration to lymph nodes are unclear. Using a mouse model of sterile peritonitis induced by thioglycollate injection, we examined the dynamics of macrophage β2 integrins during the resolution phase of inflammation. Macrophages were defined by FACS analyses as a population of cells expressing αMβ2high, αDβ2+ and CD115+. The initial population of resident β2, positive for βDβ2 and negative for αXβ2. The thioglycollate-challenged mice showed a ∼4-fold increase in macrophages on day 3 followed by a progressive decrease to normal resident cell numbers by day 13. Expression of αMβ2 on macrophages on day 3 decreased by 2.5-fold as a result of dilution of the initial population of αMβ2high resident macrophages by infiltrating blood monocytes expressing αMβ2low. However, after day 3, the density of αMβ2 on macrophages gradually increased and by day 13 returned to the high levels characteristic of resident macrophages. By contrast, expression of αDβ2 and αXβ2 on inflammatory macrophages increased by 2-fold by day 6-9 compared to that on resident macrophages and then returned to the resident levels by day 3. Thus, although the number of macrophages decreased from day 3 to day 9 by several fold, the population of macrophages which remained in the peritoneum was enriched in cells expressing the high levels of αMβ2 and αDα2. Tracking migration of fluorescently labeled peritoneal cells demonstrated that a population of macrophages which leaves the inflamed peritoneum and enters lymph nodes consists of cells expressing low levels of αMβ2 and αDβ2. These data suggested that upregulation of β2 integrins, especially αMβ2, may be responsible for the retention of macrophages in the peritoneum. Indeed, the rate of macrophage emigration from the peritoneum in the αMβ2-deficient mice was significantly higher than that in wild-type mice. The results indicate that macrophage emigration from the inflamed site is controlled by the level of integrin αMβ2 and αDβ2 with low expressors being migratory and high expressors remaining in the peritoneum. The data also highlight the importance of integrins αDβ2 and αXβ2 as specific markers of inflammatory macrophages. Disclosures: No relevant conflicts of interest to declare.

scholarly journals A specific role of integrin Mac-1 in accelerated macrophage efflux to the lymphatics

Blood ◽  
2005 ◽  
Vol 106 (9) ◽  
pp. 3234-3241 ◽  
Author(s):  
Chunzhang Cao ◽  
Daniel A. Lawrence ◽  
Dudley K. Strickland ◽  
Li Zhang
Keyword(s):  
Lymph Nodes ◽  
Blood Circulation ◽  
Cell Activation ◽  
Acute Inflammation ◽  
Blood Monocytes ◽  
Genetic Inactivation ◽  
Inflammatory Site ◽  
Peritoneal Mesothelium ◽  
Inflammatory Macrophages

Abstract In response to injury, monocytes migrate to the site of inflammation, where they differentiate into macrophages and participate in various biologic processes. However, their fate during the resolution of acute inflammation is not fully understood. Here, we show that inflammatory macrophages do not die locally by apoptosis; rather, they migrate across the peritoneal mesothelium to the lymphatics, through which they further migrate to the lymph nodes and to the blood circulation. Macrophage efflux is enhanced considerably on cell activation, and such accelerated macrophage migration is dependent specifically on integrin Mac-1, and can be blocked by addition of its antagonist. Thus, genetic inactivation of Mac-1 in mice inhibits the accelerated macrophage efflux from the inflammatory site to the lymphatics, but it does not compromise the accumulation of blood monocytes into the inflammatory site. Together, our study demonstrates that Mac-1 is involved specifically in the efflux of activated macrophages to the lymphatics, suggesting that Mac-1 may play an important role in the removal of local inflammatory macrophages and in their subsequent migration to the lymph nodes, a process that is critical to the development of the adaptive immunity.

Impaired Hydroxylation of 5-Methylcytosine In TET2 mutated Patients with Myeloid Malignancies

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1-1
Author(s):  
Anna Jankowska ◽  
Myunggon Ko ◽  
Yun Huang (equal contribution) ◽  
Utz J. Pape ◽  
Hadrian Szpurka ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Genomic Dna ◽  
Conflicts Of Interest ◽  
Methylation Pattern ◽  
Myeloid Malignancies ◽  
Cpg Sites ◽  
Low Levels ◽  
Methylation Patterns ◽  
The Impact

Abstract Abstract 1 TET2 mutations are frequently found across broad spectrum of myeloid malignancies but how these mutations contribute to diseases is still unknown. Preliminary results from our laboratory have suggested that TET2 converts 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) and consequently, the levels of 5-hmC may be lower in genomes of mutant bone marrow cells. To facilitate study of TET2 function we developed a blot assay to detect 5-hmC in genomic DNA with a specific antiserum to 5-hmC. In a second improved assay with increased sensitivity and precision, we treated genomic DNA with bisulfite in order to convert 5-hmC to cytosine 5-methylenesulfonate (CMS) and measured 5-hmC levels indirectly using a specific anti-CMS serum. Based on the results of this technique we demonstrate here for the first time that indeed TET2 mutations in predicted catalytic residues and other positions compromised TET2 function. We studied 102 patients with various myeloid malignancies (4/28 MDS, 14%, 26/48 MDS/MPN, 54% and 1/4 MPN, 2% and primary 2/11 AML 18% and 3/11 sAML, 27% TET2 mutants, respectively) and compared to wt cases or controls (N=17). Mutations were found throughout the entire coding region and were mostly inactivating (33/45 TET2 mutations). The levels of 5-hmC in genomic DNA from TET2 mutants were significantly decreased in comparison to wt cases and controls (p=4.5e-08 and p=1.8e-09, respectively). Particularly low levels of 5-hmC were found in patients with homozygous (UPD)/hemizygous (deletion) TET2 mutations and those with biallelic mutations. Surprisingly, 18% of all TET2 WT patients also showed low levels of genomic 5-hmC (despite normal TET2 mRNA expression), suggesting that these patients may carry not yet identified variants/lesions in TET2 or other partner proteins involved in TET2-mediated catalysis. To further investigate the impact of TET2 mutations associated with myeloid malignancies we also introduced 9 different missense mutations corresponding to those found in patients into murine Tet2 cells; severe loss of enzymatic activity was observed in 7/9 cases as measured by greatly diminished 5-hmC levels. To study the role of Tet2 in normal hematopoiesis we depleted Tet2 in C57BL/6 mice by retrovirus-mediated transduction of shRNA against Tet2. Tet2 depletion is associated with skewing of hematopoietic differentiation towards the monocyte/macrophage lineage. To further investigate the function of TET2 we transduced the myeloid THP-1 cell line with lentiviral vector containing TET2 cDNA (TET2+) or an empty vector. This manipulation allowed us to select clones showing 19-fold increase in TET2 mRNA expression without significantly alterations of proliferation kinetics. Using this model we studied the impact of TET2 overexpression on resultant methylation pattern of CpG sites. We have applied Illumina Infinium HumanMethylation27 arrays (27,5K CpG sites/14.4K genes). Overexpression of TET2 resulted in a distinct promoter methylation patterns with 169 altered CpG sites with difference of averaged β>0.5 (considered significant as compared to control). Among these differentially methylated loci, 27 promoters were significantly hypomethylated while 42 were hypermethylated as compared to control cells. Change in methylation pattern observed through overexpression of TET2 in vitro prompted us to analyze methylation patterns in patients with and without TET2 mutations or those with decreased 5-hmC levels. Using methylation arrays a total of 62 cases were analyzed. When patients were grouped based on the levels of 5hmC, an associated methylation signature can be clearly discerned with 2512 differentially methylated loci and distinct skewing towards hypomethylation (2510 sites; e.g., TMEM102, ABCC11) vs. hypermethylation (2 sites, AIM2 and SP140), consistent with the observation made in the TET2+ cells line. In sum, our results provide strong evidence for TET2 as the first mutated gene in myeloid malignancies that is involved in conversion of 5-mC to 5-hmC in DNA, indicating the novel role of TET2 in a substantial component of epigenetic deregulation in myeloid malignancies. Disclosures: No relevant conflicts of interest to declare.

Desialylated Platelets Are Cleared From the Circulation by the Hepatic Asialoglycoprotein Receptor,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3257-3257
Author(s):  
Renata Grozovsky ◽  
Silvia Giannini ◽  
Karin M. Hoffmeister
Keyword(s):  
Bone Marrow ◽  
Conflicts Of Interest ◽  
Fold Increase ◽  
Asialoglycoprotein Receptor ◽  
Regulatory Mechanisms ◽  
Wild Type ◽  
Cell Counts ◽  
Blood Cell Counts ◽  
Terminal Galactose

Abstract Abstract 3257 The regulatory mechanisms of platelet homeostasis remain elusive. We investigated here the role of hepatic asialoglycoprotein receptor (a.k.a. Ashwell-Morell receptor) in platelet clearance. Mice lacking the hepatic asialoglycoprotein receptor Asgpr2 subunit had increased platelet survivals (T1/2 = 49.5±2h) when compared to wild type (WT, T1/2 = 31±4h) mice. Consequently, Asgpr2−/− mice had platelet counts increased by ∼20%, compared to WT, with increased terminal galactose exposure, as demonstrated using the galactose specific lectin RCA1. Bone marrow and spleen megakaryocyte numbers were reduced by ∼15% and ∼20% in Asgpr2−/− mice, compared to WT mice. Sialidase (NA, Clostidium perfringens, 50mU/mice) maximally desialylated circulating platelets when injected intravenously, as evidenced by increased RCA1 binding. Sialidase injection resulted in a ∼60% depletion of circulating platelets after 24h in Asgpr2−/− mice, compared to >90% in WT mice, indicating that desialylated platelets were partially removed by Asgpr1/2. In contrast to platelets, red blood cell counts were unaffected by sialidase treatment. Sialidase injection for 72h resulted in a 2.3-fold and 1.2-fold increase in megakaryocyte numbers in the spleen and bone marrow of WT mice, respectively, but not in Asgpr2−/− mice. In contrast to sialidase treatment, injections of rabbit anti-mouse platelet serum (RAMPS) depleted >95% of circulating platelets and increased by 70% bone marrow, but not spleen MK numbers in both WT and Asgpr2−/− mice. The data shows that removal of desialylated, i.e, senescent, platelets by the hepatic Ashwell-Morell receptor differs to that of antibody-mediated platelet clearance. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Production of Monocyte Chemoattractant Protein 1 in Tuberculosis Patients

1998 ◽  
Vol 66 (5) ◽  
pp. 2319-2322 ◽  
Author(s):  
YuanGuang Lin ◽  
JianHua Gong ◽  
Ming Zhang ◽  
Wanfen Xue ◽  
Peter F. Barnes
Keyword(s):  
Immune Response ◽  
Mycobacterium Tuberculosis ◽  
Inflammatory Response ◽  
Lymph Nodes ◽  
Blood Monocytes ◽  
Monocyte Chemoattractant Protein 1 ◽  
Tuberculosis Patients ◽  
Monocyte Chemoattractant ◽  
Monocyte Chemoattractant Protein

ABSTRACT To investigate the role of monocyte chemoattractant protein 1 (MCP-1) in the immune response to Mycobacterium tuberculosis, we studied MCP-1 production in tuberculosis patients. CD14+ blood monocytes from tuberculosis patients spontaneously expressed higher levels of MCP-1 mRNA and protein than CD14+ monocytes from healthy tuberculin reactors. MCP-1 production in lymph nodes from tuberculosis patients was also markedly increased. These findings suggest that MCP-1 can contribute to the antimycobacterial inflammatory response by attracting monocytes and T lymphocytes.

Adenosine prevents phorbol ester injury in rabbit lungs: role of leukotrienes and TNF

1991 ◽  
Vol 71 (5) ◽  
pp. 1949-1955 ◽  
Author(s):  
J. D. Bradley ◽  
P. B. Zanaboni ◽  
R. O. Webster ◽  
L. J. Baudendistel ◽  
T. E. Dahms
Keyword(s):  
Lung Injury ◽  
Tumor Necrosis ◽  
Fold Increase ◽  
Blood Components ◽  
Myristate Acetate ◽  
Synthesis Inhibitor ◽  
Low Levels ◽  
Circulating Levels ◽  
Necrosis Factor

The objective of this study was to determine whether adenosine (ADO) prevents phorbol myristate acetate- (PMA) induced lung injury by modulating peptidoleukotrienes (LT) and/or tumor necrosis factor (TNF) production. PMA significantly increased pulmonary vascular resistance (PVR, 275 +/- 4 to 447 +/- 30 cmH2O.1–1.min) and microvascular filtration coefficient.(Kf, 0.024 +/- 0.002 to 0.040 +/- 0.006 g.min-1.cmH2O-1) in isolated blood-perfused rabbit lungs. ADO (5 mumol/min) blocked the increases in PVR (257 +/- 9 to 283 +/- 26) and Kf (0.028 +/- 0.005 to 0.018 +/- 0.002). After PMA (30 min), perfusate levels of LTC4 + LTD4 increased by 15.3 +/- 2.1 pg/ml; LTE4 increased by 15.1 +/- 4.1 pg/ml. ADO reduced the increase in LTC4 + LTD4 to 2.7 +/- 6.1 pg/ml, but total LT increased by 31.9 +/- 16.6 pg/ml, implying that ADO enhanced the conversion of LTC4 and LTD4 to LTE4. MK-886 (L663,536), an LT synthesis inhibitor, blocked the increase in total LT (6.1 +/- 13.9 pg/ml) but did not reduce the PMA-induced increase in Kf (0.022 +/- 0.003 to 0.035 +/- 0.005) or PVR (238 +/- 11 to 495 +/- 21). After PMA administration, perfusate TNF levels were not different from the 10-fold increase observed in control experiments and were not reduced by ADO or MK-886. TNF production was independent of perfusate blood components and presumably due to low levels of endotoxin in the perfusate (70–90 ng/ml). These results indicate that ADO does not protect against PMA-induced acute lung injury by altering circulating levels of LT or TNF.

scholarly journals The Use of Lymphocyte Function–Associated Antigen (Lfa)-1–Deficient Mice to Determine the Role of Lfa-1, Mac-1, and α4 Integrin in the Inflammatory Response of Neutrophils

2001 ◽  
Vol 194 (2) ◽  
pp. 219-226 ◽  
Author(s):  
Robert B. Henderson ◽  
Lina H.K. Lim ◽  
Philippe A. Tessier ◽  
Felicity N.E. Gavins ◽  
Meg Mathies ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Neutrophil Recruitment ◽  
Lymphocyte Function ◽  
Adhesion Receptor ◽  
Low Levels ◽  
First Time ◽  
Deficient Mice ◽  
Β2 Integrins

After injury or infection, neutrophils rapidly migrate from the circulation into tissues by means of an orderly progression of adhesion receptor engagements. Neutrophils have been previously considered to use selectins exclusively to roll on vessels before an adhesion step mediated by the β2 integrins, lymphocyte function–associated antigen (LFA)-1, and Mac-1. Here we use LFA-1−/− mice, function blocking monoclonal antibodies, and intravital microscopy to investigate the roles of LFA-1, Mac-1, and α4 integrins in neutrophil recruitment in vivo. For the first time, we show that LFA-1 makes a contribution to neutrophil rolling by stabilizing the transient attachment or tethering phase of rolling. In contrast, Mac-1 does not appear to be important for either rolling or firm adhesion, but instead contributes to emigration from the vessel. Blocking Mac-1 in the presence of LFA-1 significantly reduces emigration, suggesting cooperation between these two integrins. Low levels of α4β1 integrin can be detected on neutrophils from LFA-1+/+ and −/− mice. These cells make use of α4β1 during the rolling phase, particularly in the absence of LFA-1. Thus LFA-1 and α4β1, together with the selectins, are involved in the rolling phase of neutrophil recruitment, and, in turn, affect the later stages of the transmigration event.

A MiR-142-3p/EGR2 Feedback Circuitry In Human CSF-1 Driven Differentiation of Monocytes Into Macrophages

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2366-2366
Author(s):  
Brice Lagrange ◽  
Nathalie Droin ◽  
Romain Aucagne ◽  
Jérôme Paggetti ◽  
Anne Largeot ◽  
...  
Keyword(s):  
Conflicts Of Interest ◽  
Human Peripheral Blood ◽  
Human Monocytes ◽  
Blood Monocytes ◽  
Early Growth Response ◽  
Microarray Profiling ◽  
Chemical Inhibition ◽  
Regulatory Feedback Loop ◽  
Myelomonocytic Leukemia

Abstract Abstract 2366 Colony-stimulating factor-1 (CSF-1 or M-CSF) triggers the differentiation of human peripheral blood monocytes into macrophages through and integrated cytokine/transcription factors circuitry. Using microarray profiling to explore the role of microRNAs (miRNAs) in this molecular circuitry, we identified the down-regulation of miR-142-3p in human macrophages obtained from CSF-1-treated monocytes. We show that miR-142-3p is a repressor of the transcription factor EGR2 (Early Growth Response 2) through direct 3'UTR interactions. Interestingly, EGR2 binds the promoter of the pre-miR-142-3p gene to negatively regulate its expression, identifying a self-regulatory feedback loop. Enforced expression of miR-142-3p in primary human monocytes as well as decreased expression of miR-142-3p observed in monocytes from patients with a chronic myelomonocytic leukemia further assess the link between miR-142-3p and EGR2 expression in these cells. A chemical inhibition of the Src kinase family prevents the regulation loop induced by CSF-1. Thus, our study uncovers an EGR2/miR-142-3p circuitry which regulates CSF-1 driven differentiation of human monocytes into macrophages. Disclosures: No relevant conflicts of interest to declare.

Factor VIII Region 1811–1818 Is Involved in Factor IXa Binding and Factor VIIIa Stability

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3357-3357
Author(s):  
Esther Bloem ◽  
Henriet Meems ◽  
Maartje van den Biggelaar ◽  
Koen Mertens ◽  
Alexander B Meijer
Keyword(s):  
Factor Viii ◽  
Half Life ◽  
Conflicts Of Interest ◽  
Fold Increase ◽  
Factor Ix ◽  
The Other ◽  
Factor V ◽  
Fluorescent Intensity ◽  
A2 Domain

Abstract Abstract 3357 Previously, we identified a role for the lysine residue couple 1967/1968 in the stability of activated factor VIII (FVIIIa). Using tandem mass tags (TMT 126/127) in combination with mass spectrometry, we identified lysine residues involved in the interaction between the A2 domain and the rest of heterodimer (A1/A3-C1-C2) of FVIIIa (Bloem et al., J Biol Chem 2012;287:5575–83). Upon FVIII activation and A2 domain dissociation, the highest increase in surface exposure occurred for the lysine couple 1967/1968, and functional studies confirmed the role thereof in FVIIIa stability. In addition to lysines 1967/1968 also other lysines displayed an increased surface exposure, including those in positions 1804, 1808, 1813 and 1818. The A3 domain region 1803–1818 has previously been implicated in interactions with ligands such as activated factor IX (FIXa). As such, one might expect increased surface exposure due to FVIII activation. On the other hand, A2 domain dissociation may have rearranged the A3 domain surface in this region. The relation between FIXa assembly and A2 domain retention was therefore explored in the present study. To unravel the role of region 1803–1818 in FVIIIa stability and FIXa binding, either region 1803–1810 or 1811–1818 was replaced by the corresponding regions of the homologous factor V. Additionally, as Asn1810 is N-linked glycosylated and this glycan is maintained in both chimeras, a variant that lacks this glycan (N1810C) was investigated. Factor × activation kinetics were used to investigate the apparent FIXa binding affinity of the FVIII variants. FXa generation was assessed on 15% phosphatidyl serine (PS) containing membranes. FIXa titration experiments showed that the affinity for the 1811–1818 variant is reduced (apparent Kd from 1.3 nM to 2.4 nM). Removal of the glycan and substitution of 1803–1810 has little or no effect on the apparent FIXa binding. FVIIIa-FIXa assembly on membranes containing 15% PS was studied using lipid- coated glass beads (lipospheres). Lipospheres were incubated with fluorescein-labeled FIXa and different FVIIIa concentrations. FIXa did only display liposphere binding in the presence of FVIIIa. Therefore, the mean fluorescent intensity on the lipospheres at increasing FVIIIa concentration could be used as a measure for FVIIIa-FIXa assembly on lipids. Results from these experiments showed that the 1811–1818 variant fails to promote FVIIIa-FIXa assembly, whereas the other variants behave like WT. To investigate FVIIIa stability, and thereby the role of the mutations on A2 domain dissociation, the activity of the variants was followed over time. Results showed that the 1811–1818 variant has a decreased half life of 1.5 min, compared to 6.9 min for WT. Also substitution of region 1803–1810 results in a lower half life, although to a lesser extent (2.8 min), whereas the glycan lacking variant behaves like WT (6.8 min). Incubation of FVIIIa variants with FIXa is known to stabilize FVIIIa and leads to an increased half life for all variants. However, the 1803–1810 variant is most efficiently stabilized by FIXa, shown by a 3-fold increase in half life, instead of 1.6-fold as seen for both WT and N1810C. The 1811–1818 variant is stabilized by 2.2-fold and therefore remains significantly less stable than WT. Together these results show that the 1811–1818 region is not only involved in FIXa binding, but additionally plays a major role in A2 domain retention. Region 1803–1810 also plays a role in FVIIIa stability, although to a lesser extent. Remarkably, the glycan at position Asn1810 does not influence neither FIXa binding nor FVIIIa stability, and apparently serves some other function. Disclosures: No relevant conflicts of interest to declare.

scholarly journals IGFBP7 As a Potential Therapeutic Target for AML

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5134-5134
Author(s):  
Yina Niu ◽  
Shuiyan Wu ◽  
Shaoyan Hu
Keyword(s):  
Cell Lines ◽  
Conflicts Of Interest ◽  
Fold Increase ◽  
Dependent Manner ◽  
Mice Model ◽  
Cell Counts ◽  
And Migration ◽  
Proliferation And Migration

Abstract Insulin-like growth factor binding proteins (IGFBPs) are secretory factors that play essential roles in regulation of insulin-like growth factors (IGFs) in tissue as well as in modulating IGF binding to its receptors. IGFBP7, known as IGFBP-related protein 1 (IGFBP-rP1), mac25/angiomodulin, function as a potential tumor suppressor in various human solid cancers, including breast, prostate, gastric and liver cancer. We have reported the overexpression of IGFBP7 in the context of acute myeloid leukemia (AML), showing that IGFBP7 expression level in AML patients is significantly increased compared with controls (P<0.001). IGFBP7 expression was obviously decreased in AML patients achieving complete remission (P<0.01), and was significantly increased in relapsed AML patients (P<0.01). In addition, AML patients with high expression of IGFBP7 had shorter overall survival. Here, we investigate the role and mechanism of IGFBP7 in the development and progression of AML. In order to study the role of IGFBP7 in AML, stable cell lines expressing IGFBP7 and control in AML cells were constructed using lentiviral packaging system. Expression microarray assay was carried out to analyze the global gene level changes driven by IGFBP7. MTT and transwell assays were performed to evaluate the effect of IGFBP7 on cell proliferation and migration. Bioinformatics results found that IGFBP7 appeared to utilize multiple cellular processes for its oncogenic roles, including adhesion, migration, and proliferation. Experimental data showed overexpression of IGFBP7 in K562 cells resulted in a 2-3 fold increase in migration in contrast to control cells. Moreover, enforced expression of IGFBP7 also led to phosphorylation of Akt and Erk, whose activities inactivation by pharmacologically inhibitors resulted in the loss of ability to migrating. Finally, knockdown of IGFBP7 in cells with high IGFBP7 level, their migration abilities were significantly decreased. To assess the role of IGFBP7 in leukemogenesis in vivo, the same numbers of K562/IGFBP7 and K562-Vector cells, U937-shIGFBP7 and U937-shNEG cells were injected into NOD-SCID mice by tail vein injection, respectively. About two weeks later, it was showed that mice of K562/IGFBP7 and U937 groups displayed higher white blood cell counts compared with mice of K562-Vector and U937-shIGFBP7 groups, respectively. Mice of K562/IGFBP7 and U937 groups had more severe splenomegaly and hepatomagaly compared with its corresponding control groups. We further characterized the molecular mechanism underlying leukemogenesis driven by IGFBP7 in AML cell lines. The global expression profiling and molecular biological experiments showed PI3K/AKT signaling was activated by overexpression of IGFBP7, and knockdown of IGFBP7 in AML cells led to a decrease of PI3K/AKT activity in PTEN-dependent manner. IGFBP7 promotes proliferation and migration of AML cells, the promotion could be suppressed by both RNA interference and pharmacological inhibition of PI3K/AKT pathway. Immuno-precipitation assay showed that IGFBP7 associated with AXAN2 and induced PTEN degradation. The expression of ANXA2 was significantly positive correlated with the expression ANXA2 in AML patients. The expression of IGFBP7 in AML, overexpression as well as knockdown of IGFBP7 in leukemia cells and in mice model, all suggest that IGFBP7 is a potential proto-oncogene. Collectively this work suggests that targeting IGFBP7 activity may be an effective therapeutic strategy for AML. Disclosures No relevant conflicts of interest to declare.

scholarly journals Suppression of Mir-708 Promotes DKK3 to Inhibit Wnt/β-Catenin Signaling Pathway in Adult B-ALL

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5090-5090
Author(s):  
Yingjie Zhang ◽  
Huibo Li ◽  
Rongyi Cao ◽  
Lili Sun ◽  
Yan Wang ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Solid Tumors ◽  
Conflicts Of Interest ◽  
Hematologic Malignancies ◽  
Leukemia Cells ◽  
Childhood All ◽  
Apoptosis Inhibition ◽  
Low Levels ◽  
Matched Samples

Abstract Dickkopf-3 (DKK-3) is a Wnt signaling antagonist, and DKK3 inactivation is associated with poor prognosis in various solid tumors and hematologic malignancies. Epigenetic hypermethylation has been implicated in downregulation of DKK3, but other regulatory mechanisms remain to be investigated. In this study, we examined DKK3 expression and the role of microRNA in the regulation of DKK3 in adult B-ALL. Our results showed low levels of DKK3 expression in Nalm-6 and BALL-1 cell lines, and leukemia cells from adult B-ALL patients. DKK3 expression was remarkably lower in the initial diagnosis and relapse samples than in the matched samples during remission. In addition to hypermethylation, low levels of DKK3 were associated with high expression of miR-708. This finding was similar to that in childhood ALL. The miR-708 was predicted to bind to the 3'-UTR of DKK3. By using miR-708 mimics, we found that miR-708 targets DKK3 to promote B-ALL cell proliferation through cell cycle promotion and apoptosis inhibition. The treatment with 5-aza-2'-deoxycytidine significantly increased DKK3 and decreased p-GSK3β, cyclin D1 and nuclear and cytoplasmic β-catenin protein expression. A synergistic effect was seen when the miR-708 inhibitor and 5-aza were used simultaneously. Although miR-708 was reported to either promote or suppress tumorigenecity in some solid tumors, our findings indicate that suppression of miR-708 increases DKK3 expression to inhibit the Wnt/β- catenin signaling pathway in B-ALL. Disclosures No relevant conflicts of interest to declare.

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