Invasiveness-triggered state transition in malignant melanoma cells.

Cancer cells are considered to have high morphological heterogeneity in human melanoma tissue. Here, we report that epithelial cancer cells are dominant in different development stages of human melanoma tissues. The cellular and molecular mechanisms that maintain melanoma cells in the epithelial state are further investigated in the A2058 cell line. We find that micropore (8 µm) transwell invasion, but not superficial migration in the scratch assay, can induce remarkable morphological changes between epithelial and mesenchymal melanoma cells within 4 days. The morphological switch is associated with dynamic changes of epithelial-mesenchymal transition (EMT) hallmarks E-cadherin and vimentin. Further immunoflurencent staining and co-immunoprecipitation assay showed the uncoupling of the M3 muscarinic acetylcholine receptor (mAChR) and the p75 neurotrophin receptor (p75NTR) in epithelial melanoma cells. Specific knockdown of M3 mAChR by small interfering RNA (siRNA) significantly abrogates the transition of spindle-shaped mesenchymal cells to epithelial cells. Collectively, we report a cellular model of invasiveness-triggered state transition (ITST) in which melanoma cell invasion can induce morphological changes between epithelial and mesenchymal cells. ITST is one of the biological basis for maintaining metastatic melanoma cells in the epithelial state. Furthermore, M3 mAChR receptor-mediated ITST provides a novel therapeutic strategy to inhibit the development of malignant melanoma.

Modeled microgravity suppressed invasion and migration of human glioblastoma U87 cells through downregulating store-operated calcium entry.

Glioblastoma is the most common brain tumor and is characterized with robust invasion and migration potential resulting in poor prognosis. Previous investigations have demonstrated that modeled microgravity (MMG) could decline the cell proliferation and attenuate the metastasis potential in several cell lines. In this study, we studied the effects of MMG on the invasion and migration potentials of glioblastoma in human glioblastoma U87 cells. We found that MMG stimulation significantly attenuated the invasion and migration potentials, decreased thapsigargin (TG) induced store-operated calcium entry (SOCE) and downregulated the expression of Orai1 in U87 cells. Inhibition of SOCE by 2-APB or stromal interaction molecule 1 (STIM1) downregulation both mimicked the effects of MMG on the invasion and migration potentials in U87 cells. Furthermore, upregulation of Orai1 significantly weakened the effects of MMG on the invasion and migration potentials in U87 cells. Therefore, these findings indicated that MMG stimulation inhibited the invasion and migration potentials of U87 cells by downregulating the expression of Orai1 and sequentially decreasing the SOCE, suggesting that MMG might be a new potential therapeutic strategy in glioblastoma treatment in the future.

Notch signaling regulates CXCR4 expression and the migration of mesenchymal stem cells.

Mesenchymal stem cells (MSCs) have been used to repair injured tissues through immune-suppression and/or cell replace mechanisms. However, a significant barrier to MSC therapy is insufficient MSC engraftment in injured tissues after systemic administration. Here, we report that cell surface, total protein, and mRNA levels of CXCR4 were significantly increased in MSCs when Notch signaling was interrupted by γ-secretase inhibitor (GSI) or knockout of the transcription factor RBP-J, which mediates signaling from all four mammalian Notch receptors. The GSI-treated or RBP-J deficient MSCs showed stronger migration toward stromal cell-derived factor-1α (SDF-1α) than that of the control. In a mouse hepatic ischemia/reperfusion model, RBP-J deficient MSCs migrated into the injured liver tissues at a significantly higher efficiency than that of the control MSCs. Mice transfused with RBP-J deficient MSCs showed reduced liver damage. Therefore, Notch signaling regulates MSC migration and function, at least partially via the modulation of CXCR4 expression.

Differences in Non-suicidal Self-injury Behaviors between Unipolar Depression and Bipolar Depression in Adolescent Outpatients.

OBJECTIVE: Non-suicidal self-injury (NSSI) has a higher prevalence in adolescents with depressive disorders than in community adolescents. This study examined the differences in NSSI behaviors between adolescents with unipolar depression (UD) and those with bipolar depression (BD). METHODS: Adolescents with UD or BD were recruited from 20 general or psychiatric hospitals across China. The methods, frequency, and function of NSSI were assessed by Functional Assessment of Self-Mutilation. The Beck Suicide Ideation Scale was used to evaluate adolescents' suicidal ideation, and the 10-item Kessler Psychological Distress Scale to estimate the anxiety and depression symptoms. RESULTS: The UD group had higher levels of depression (19.16 vs.17.37, F=15.23, P<0.001) and anxiety symptoms (17.73 vs.16.70, F=5.00, P=0.026) than the BD group. Adolescents with BD had a longer course of NSSI than those with UD (2.00 vs.1.00 year, Z=-3.39, P=0.001). There were no statistical differences in the frequency and the number of methods of NSSI between the UD and BD groups. Depression (r=0.408, P<0.01) and anxiety (r=0.391, P<0.01) were significantly and positively related to NSSI frequency. CONCLUSION: Adolescents with BD had a longer course of NSSI than those with UD. More importantly, NSSI frequency were positively and strongly correlated with depression and anxiety symptoms, indicating the importance of adequate treatment of depression and anxiety in preventing and intervening adolescents' NSSI behaviors.

Digital holographic reconstruction of a local object field using an adjustable magnification.

In the research of digital holography, this paper presents a numerical method using an adjustable magnification for local object field reconstruction together with experiment verification. The method first designs a spherical wave according to the given magnification to illuminate the digital hologram, then through a Fourier transform of diffraction, it calculates the reconstructed image plane. Afterward, a filtering window is set in the image plane to extract the image of the local object field, and then the object field reached hologram plane is formed using diffraction's inverse operation. Finally, the object field is reconstructed through diffraction's angular spectrum theory.

Real-time three-sensitivity measurements based on three-color digital Fresnel holographic interferometry.

This Letter presents a method for real-time 3D measurements based on three-color digital holographic interferometry. The optical setup is considerably simplified, since the reference beams are combined into a unique beam. A convolution algorithm allows the three monochrome images to be superposed to provide simultaneous full-field 3D measurements. Experimental results confirm the suitability of the proposed method.

Method of digital holographic recording and reconstruction using a stacked color image sensor.

We discuss a method to record and reconstruct color holograms by using a stack of photodiode sensors associated to a one-way reference beam. The reconstruction algorithm follows a convolution strategy in which a transverse magnification leads to the full reconstruction of the object in the reconstructed horizon. The transverse magnification of the object depends on the curvature of the reference wave. Analysis of the spatial resolution indicates that it is linked to the transversal magnification but that no extra information is gained or lost in the process. Experimental results confirm the validity of the proposed approach for two-color digital holography. The error due to spectral mixing is investigated and found to be quite irrelevant compared to the range of the phase measurement.

Spatial bandwidth extended reconstruction for digital color Fresnel holograms.

This paper presents a reconstruction algorithm based on the convolution formula of diffraction which uses the Fresnel impulse response of free space propagation. The bandwidth of the reconstructing convolution kernel is extended to the one of the object in order to allow the direct reconstruction of objects with size quite larger than the recording area. The spatial bandwidth extension is made possible by the use of a numerical spherical wave as a virtual reconstructing wave, thus modifying the virtual reconstruction distance and increasing the kernel bandwidth. Experimental results confirm the suitability of the proposed method in the case of the simultaneous recording of two-color digital holograms by using a spatial color multiplexing scheme.

Notch signaling regulates the FOXP3 promoter through RBP-J- and Hes1-dependent mechanisms.

Evidence has shown that Notch signaling modulates CD4(+)CD25(+) regulatory T-cells (Tregs). As transcription factor Foxp3 acts as a master molecule governing the development and function of Tregs, we investigated whether Notch signaling might directly regulate Foxp3 expression. Here, we provide evidence that Notch signaling can modulate the FOXP3 promoter through RBP-J- and Hes1-dependent mechanisms. A conserved RBP-J-binding site and N-box sites were identified within the FOXP3 promoter. We show that the Notch intracellular domain (NIC), the active form of Notch receptors, activates a reporter driven by the FOXP3 promoter. Dissection of the FOXP3 promoter revealed bipartite effects of the RBP-J-binding site and the N-boxes: the RBP-J-binding site positively, while the N-boxes negatively regulated the FOXP3 promoter activity. Moreover, in freshly isolated Tregs, NIC-RBP-J complex is bound to the FOXP3 promoter in Tregs. Our results suggest that Notch signaling might be involved in the development and function of Tregs through regulating Foxp3 expression.

[Regulative effects of aquaporin 4 expression by rhein in rhubarb to intestinal epithelial cell line LoVo].

OBJECTIVE: To investigate the effects of rhein on regulating aquaporin4 (AQP4) to LoVo cells cultured with RPMI-1640 medium containing rhein. METHODS: LoVo cells were cultured with RPMI-1640 medium containing different concentration rhein for 24 hours and were cultured with RPMI-1640 containing rhein (20 mg/L) for different time. Four groups were assigned as LoVo cells were cultured respectively with RPMI-1640 medium containing different concentration rhein (40, 20, 10 mg/L and control group), while six groups were assigned as LoVo cells were cultured with RPMI-1640 medium containing rhein (20 mg/L) for different time (3, 6, 12, 24, 48 h and control group). The location of AQP4 protein in LoVo cells was definited by immuocytochemistry dying. Western blotting and semiquantitative RT-PCR were adopted to detect the relative expression of AQP4 protein and mRNA. RESULTS: AQP4 was located mainly in plasma membrane of LoVo cells while partly in cytoplasm. The relative expression of AQP4 protein and mRNA decreased with the increasing of rhein concentration; there was no significant difference of the relative expression of AQP4 in 10 mg/L group compared with that in control group, but it decreased significantly in 40 and 20 mg/L groups. The relative expression of AQP4 in 3 and 6 h groups was lower than that in control group but there was no statistical significance, however that in 12, 24, 48 h groups was lower significantly compared with that in control group. CONCLUSION: Rhein can inhibit the genetic transcription and the translation of AQP4 gene in LoVo cells, which demonstrates that the change of AQP4 expression regulated by rhein may be related to the cathartic effect of rhubarb.

Disruption of Notch signaling aggravates irradiation-induced bone marrow injury, which is ameliorated by a soluble Dll1 ligand through Csf2rb2 upregulation.

Physical and chemical insult-induced bone marrow (BM) damage often leads to lethality resulting from the depletion of hematopoietic stem and progenitor cells (HSPCs) and/or a deteriorated BM stroma. Notch signaling plays an important role in hematopoiesis, but whether it is involved in BM damage remains unclear. In this study, we found that conditional disruption of RBP-J, the transcription factor of canonical Notch signaling, increased irradiation sensitivity in mice. Activation of Notch signaling with the endothelial cell (EC)-targeted soluble Dll1 Notch ligand mD1R promoted BM recovery after irradiation. mD1R treatment resulted in a significant increase in myeloid progenitors and monocytes in the BM, spleen and peripheral blood after irradiation. mD1R also enhanced hematopoiesis in mice treated with cyclophosphamide, a chemotherapeutic drug that induces BM suppression. Mechanistically, mD1R increased the proliferation and reduced the apoptosis of myeloid cells in the BM after irradiation. The ß chain cytokine receptor Csf2rb2 was identified as a downstream molecule of Notch signaling in hematopoietic cells. mD1R improved hematopoietic recovery through up-regulation of the hematopoietic expression of Csf2rb2. Our findings reveal the role of Notch signaling in irradiation- and drug-induced BM suppression and establish a new potential therapy of BM- and myelo-suppression induced by radiotherapy and chemotherapy.

Hif-1α and Hif-2α differentially regulate Notch signaling through competitive interaction with the intracellular domain of Notch receptors in glioma stem cells.

Hypoxia contributes to GSC expansion principally through Hif-1α and Hif-2α, but how these two factors work together has not been completely understood. We show that hypoxia promoted proliferation, self-renewal and inhibited the conversion of GSCs into INP-like cells through activating Notch signaling. Further data suggested that Hif-2α interacted with NICD and repressed the activity of Notch signaling, in contrast to the role of Hif-1α in Notch signaling. Together, our findings suggest that Hif-1α and Hif-2α competitively bind to NICD and dynamically regulate the activation of Notch signaling in GSCs likely depending on different oxygen tensions, providing improved therapeutic opportunities for malignant gliomas.

Notch-RBP-J signaling is required by bone marrow stromal cells for the treatment of acute graft versus host disease.

Recent evidence has shown that bone marrow stromal cells (BMSCs) may exhibit immuno-suppression activities through soluble mediators and direct cell-cell contact, but how these processes are modulated has been poorly understood. In this study, we show that the Notch signaling pathway participates in the modulation of BMSCs to elicit their immuno-suppressive roles. In a murine lethal acute graft versus host disease (aGvHD) model, BMSCs deficient for RBP-J, the critical transcription factor mediating signaling from all four mammalian Notch receptors, failed to delay the development of the disease. RBP-J deficient BMSCs were not able to inhibit the proliferation and activation of allogenic T-cells. Moreover, RBP-J deficient BMSCs could not down-regulate the expression of MHC II and co-stimulation molecules CD80 and CD86 on dendritic cells (DCs). The antigen presentation capacity of DCs co-cultured with RBP-J deficient BMSCs was not impaired in contrast to wild type BMSCs. Furthermore, we showed that the productions of IL-6 and PGE2, two critical molecules mediating the immuno-suppressive activities of BMSCs, were reduced significantly in RBP-J deficient BMSCs. Both of the two molecules were importantly involved in the regulation of BMSCs by Notch signaling. In conclusion, our data suggests that the immuno-suppressive effects of BMSCs in aGvHD are dependent on Notch-RBP-J signaling, which regulates the productions of IL-6 and PGE2.

Differential regulation of bone marrow-derived endothelial progenitor cells and endothelial outgrowth cells by the Notch signaling pathway.

Endothelial progenitor cells (EPCs) are heterogeneous populations of cells that participate in vasculogenesis and promote tissue regeneration. However the different roles of EPC populations in vasculogenesis and tissue regeneration, as well as their regulation and mechanisms remain elusive. In the present study, we cultured bone marrow (BM)-derived early EPCs (EEPCs) and endothelial outgrowth cells (EOCs), and investigated their roles in liver regeneration and their regulation by the Notch signaling pathway. We found that Notch signaling exhibited different effects on the proliferation and migration of EEPCs and EOCs. Our results also showed that while EEPCs failed to form vessel-like structures in a three dimensional sprouting model in vitro, EOCs could sprout and form endothelial cords, and this was regulated by the Notch signaling. We further showed that, by using a conditional knockout model of RBP-J (the critical transcription factor mediating Notch signaling), Notch signaling differentially regulates EEPCs and EOCs. In a partial hepatectomy (PHx) model, EEPCs Notch-dependently benefitted liver regeneration with respect to liver function and hepatocyte proliferation and apoptosis. In contrast, EOCs appeared not directly involved in the recovery of liver function and the increase of hepatocytes. These data suggested that the RBP-J-mediated Notch signaling differentially regulated the two types of EPCs, which showed different roles in liver regeneration.

Digital holographic reconstruction of large objects using a convolution approach and adjustable magnification.

We present a numerical method for reconstructing large objects using a convolution method with an adjustable magnification. The method is based on the image locations and magnification relations of holography when the illuminating beam is a spherical wavefront. A modified version of the angular spectrum transfer function is proposed that allows the filtering in the spatial frequency spectrum. Experimental results confirm the suitability of the proposed method.