High level of miR-196b at newly diagnosed pediatric acute myeloid leukemia predicts a poor outcome

Differential expression of microRNAs (miRNAs) has been implicated in leukemogenesis. We investigate the ex- pression pattern of miR-196b. Using quantitative real-time PCR (qRT-PCR), we detected the expression of miR- 196b and its correlated genes (SMC1A/MLH1) in initial pediatric AML. A significant association was observed between overexpression of miR-196b and inferior overall survival of pediatric AML (Log Rank P<0.0001). AML M4/5 subtype, high white blood cell (WBC) count at presentation, MLL rearrangement, or FLT3-ITD mutation at diagnosis and non-remission group after the first induction chemotherapy possessed higher miR-196b expression. Furthermore, a positive relationship was found between the expression of miR-196b and SMC1A/MLH1 (Spear- man’s r=0.37 and 0.44, P=0.001 and <0.0001, respectively). Taken together, these findings suggest that differen- tially high expression of miR-196b in diagnostic marrow samples of pediatric AML is associated with unfavorable outcome, and miR-196b potentially can be a novel biomarker for the diagnosis, prognosis and treatment in pediatric AML

Matrix Metalloproteinase-2 and -9 Secreted by Leukemic Cells Increase the Permeability of Blood-Brain Barrier by Disrupting Tight Junction Proteins

Central nervous system (CNS) involvement remains an important cause of morbidity and mortality in acute leukemia, the mechanisms of leukemic cell infiltration into the CNS have not yet been elucidated. The blood-brain barrier (BBB) makes CNS become a refugee to leukemic cells and serves as a resource of cells that seed extraneural sites. How can the leukemic cells disrupt this barrier and invasive the CNS, even if many of the currently available chemotherapies can not cross the BBB? Tight junction in endothelial cells occupies a central role in the function of the BBB. Except the well known role of degrading extracellular matrix in metastasis of cancer cells, here we show matrix metalloproteinase (MMP)-2 and -9, secreted by leukemic cells, mediate the BBB opening by disrupting tight junction proteins in the CNS leukemia. We demonstrated that leukemic cells impaired tight junction proteins ZO-1, claudin-5 and occludin resulting in increased permeability of the BBB. However, these alterations reduced when MMP-2 and -9 activities were inhibited by RNA interference strategy or by MMP inhibitor GM6001 in an in vitro BBB model. We also found that the disruption of the BBB in company with the down-regulation of ZO-1, claudin-5 and occludin and the up-regulation of MMP-2 and -9 in mouse brain tissues with leukemic cell infiltration by confocal imaging and the assay of in situ gelatin zymography. Besides, GM6001 protected all mice against CNS leukemia. Our findings suggest that the degradation of tight junction proteins ZO-1, claudin-5 and occludin by MMP-2 and -9 secreted by leukemic cells constitutes an important mechanism in the BBB breakdown which contributes to the invasion of leukemic cells to the CNS in acute leukemia

Localized surface plasmon-enhanced ultraviolet electroluminescence from n-ZnO/i-ZnO/p-GaN heterojunction light-emitting diodes via optimizing the thickness of MgO spacer layer

Localized surface plasmon (LSP)-enhanced ultraviolet light-emitting diodes were manufactured by introducing Ag nanoparticles and MgO spacer layer into n-ZnO/i-ZnO/p-GaN heterostructures. By optimizing the MgO thickness, which can suppress the undesired charge transfer and nonradiative Forster resonant energy transfer between Ag and ZnO, a 7-fold electroluminescence enhancement was achieved. Time-resolved and temperature-dependent photoluminescence measurements reveal that both spontaneous emission rate and internal quantum efficiency are increased as a result of coupling between ZnO excitons and Ag LSPs, and simple calculations, based on experimental data, also indicate that most of LSP's energy can be converted into the photon energy. (C) 2012 American Institute of Physics. [http://dx.doi.org/10.1063/1.4757127

Expression of MMP-2 and -9, gelatinase secretion and invasion rate in different leukemic cells.

<p>(A) qRT-PCR analysis of MMP-2 (A1) and MMP-9 (A2) gene transcription in SHI-1, HL-60 and U937 leukemic cells. (B) Gelatinase secretion from culture supernatants of three leukemic cells. (C) Cell invasion rates of SHI-1, HL-60 and U937 leukemic cells. Cell invasion rates were determined by the ratios relative to SHI-1 cells. Data represent the mean ± SD of triplicate measurement representative for three independent experiments relative to SHI-1 cells (set as 100%). Compare with SHI-1 cells *<i>P</i><.001.</p

Effects of MMP-2/MMP-9 knock-down on activity of gelatinase and rates of cell invasion by SHI-1 in the <i>in vitro</i> BBB model.

<p>(A) Transcription of specific mRNAs quantified by qRT-PCR 48 h after siRNA transfection. SHI-1 cells transfected with siRNAs targeting the expression of MMP-2 (KD-MMP-2) or (B) MMP-9 (KD- MMP-9). Control SHI-1 cells transfected with non-target-directed siRNA (set as 100%). (C) Gelatinase secretion from SHI-1 cells transfected with siRNAs. (D) Rates of cell invasion of SHI-1 cells 48 h after transfection with siRNA in in vitro model of BBB. Data represent the mean ± SD of triplicate measurement representative for three independent experiments relative to SHI-1 cells (set as 100%). *<i>P</i><.001.</p

The SCI-nu/nu mice model of CNS leukemia.

<p>(A) Clinical signs of CNS leukemia in mice. (A1, A2, arrow) The mouse developed paralysis in its left forelimb and cranial nerve with light loss. (A3, arrow) The mice developed paralysis in both fore limbs and hind limbs. (B, arrow) Neoplasms of different parts with or without intracranial hemorrhage in removed brains. (C) Infiltrations of SHI-1 examined with histopathology assay. (C1, C2, C3) Brain parenchyma pathobiology stained with H&E. (C4) Examination of bone marrow smears stained with Wright's-Giemsa. (D) Organs with infiltrations of SHI-1 examined with RT-PCR analysis of human MLL/AF6 fusion gene transcription in mice with or without GM6001 treatment. (D1) RT-PCR analysis of MLL/AF6 fusion gene of brains and bone marrow from SCI nu/nu-mice. Lanes 1–5: Human MLL/AF6 fusion genegene amplified in SHI-1 cell and in brain, bone marrow, stomach and liver of mice with CNS leukemia. lane 6–9: Human MLL/AF6 fusion gene amplified in stomach, liver, brain and bone marrow of mice with GM6001 treatment. lane 10: negative control. (D2) The number of organs with infiltrations of SHI-1 examined with RT-PCR analysis of human MLL/AF6 fusion gene transcription in mice with or without GM6001 treatment. Original magnification: (C1), ×40; (C2), ×200; (C3), ×1000; (C4), ×1000.</p

Increased <i>in vitro</i> BBB permeability due to MMP-2 and -9 secreted by SHI-1, HL-60 and U937 cells.

<p>(A) Morphological changes of BMVECs affected by SHI-1, HL-60 and U937 cells treated with or without GM6001. (B) Rates of leukemic cell invasion of co-culture with BMEVCs compared with those of monoculture (without BMEVCs) (<i>P</i>>.05) and those treated with GM6001 (*<i>P</i><.001). Results are shown as the mean ± SD of three independent experiments. (C) Zymography gelatinase secretion in SHI-1, HL-60 and U937 cells treated with or without GM6001. Original magnification: (A)×600.</p

Activities of MMP-2 and -9 secreted by SHI-1 with or without GM6001 treatment in CNS leukemia mice.

<p>(A) RT-PCR analysis of human MMP-2 and -9 gene transcription in normal brain tissues, brain tissues infiltrated with SHI-1 cells with or without GM6001 treatment. (B, C and D) <i>In situ</i> zymographic analysis of MMP-2 and -9 activities mouse tissues. (B) Normal brain tissues; (C) Brain tissues infiltrated with SHI-1 cells; (D) Brain tissues after treatment with GM6001. Results shown are from individual animals and are representative of findings from 3 experiments with 6 animals per condition per experiment. (Scale bars: 10 µm).</p