Phase sensitive Brillouin scattering measurements with a novel magneto-optic modulator

A recently reported phase sensitive Brillouin light scattering technique is improved by use of a magnetic modulator. This modulator is based on Brillouin light scattering in a thin ferrite film. Using this magnetic modulator in time- and space Brillouin light scattering measurements we have increased phase contrast and excluded influence of optical inhomogeneities in the sample. We also demonstrate that the quality of the resulting interference patterns can be improved by data postprocessing using the simultaneously recorded information about the reference light

Optical detection of spin transport in non-magnetic metals

We determine the dynamic magnetization induced in non-magnetic metal wedges composed of silver, copper and platinum by means of Brillouin light scattering (BLS) microscopy. The magnetization is transferred from a ferromagnetic Ni80Fe20 layer to the metal wedge via the spin pumping effect. The spin pumping efficiency can be controlled by adding an insulating but transparent interlayer between the magnetic and non-magnetic layer. By comparing the experimental results to a dynamical macroscopic spin-transport model we determine the transverse relaxation time of the pumped spin current which is much smaller than the longitudinal relaxation time

GTP and Ca2+ Modulate the Inositol 1,4,5-Trisphosphate-Dependent Ca2+ Release in Streptolysin O-Permeabilized Bovine Adrenal Chromaffin Cells

The inositol 1,4,5-trisphosphate (IP3)-induced Ca2+ release was studied using streptolysin O-permeabilized bovine adrenal chromaffin cells. The IP3-induced Ca2+ release was followed by Ca2+ reuptake into intracellular compartments. The IP3-induced Ca2+ release diminished after sequential applications of the same amount of IP3. Addition of 20 μM GTP fully restored the sensitivity to IP3. Guanosine 5'-O-(3-thio)triphosphate (GTPγS) could not replace GTP but prevented the action of GTP. The effects of GTP and GTPγS were reversible. Neither GTP nor GTPγS induced release of Ca2+ in the absence of IP3. The amount of Ca2+ whose release was induced by IP3 depended on the free Ca2+ concentration of the medium. At 0.3 μM free Ca2+, a half-maximal Ca2+ release was elicited with ∼0.1 μM IP3. At 1 μM free Ca2+, no Ca2+ release was observed with 0.1 μM IP3; at this Ca2+ concentration, higher concentrations of IP3 (0.25 μM) were required to evoke Ca2+ release. At 8 μM free Ca2+, even 0.25 μM IP3 failed to induce release of Ca2+ from the store. The IP3-induced Ca2+ release at constant low (0.2 μM) free Ca2+ concentrations correlated directly with the amount of stored Ca2+. Depending on the filling state of the intracellular compartment, 1 mol of IP3 induced release of between 5 and 30 mol of Ca2+

Association of 12 serum biochemical markers of angiogenesis, tumour invasion and bone turnover with bone metastases from breast cancer: a crossectional and longitudinal evaluation

Complex biological pathways including angiogenesis, invasion, osteoclastic activation and bone matrix degradation are involved in the formation of bone metastasis (BM). The aim of our study was to investigate the cross-sectional and longitudinal associations of a panel of 12 serum biochemical markers reflecting biological pathways underlying BM development. In a cross-sectional study, we investigated 29 patients with primary breast carcinoma without BM (BC/BM−), 28 patients with breast carcinoma and BM (BC/BM+) and 15 healthy women. In longitudinal analyses, we investigated 34 patients for whom serum was obtained a two different time points: at the time of primary BC diagnosis and after a median time of 3 years. During this follow-up, 15 patients developed BM, whereas the other 19 remained free of BM. In patients who developed BM, the second samples were obtained before BM was documented by bone scan. The cross-sectional analyses have shown all biochemical markers to be significantly elevated in patients with BM, when compared to the patients without BM and healthy controls, except TGFβ1 that was significantly decreased. Multivariable analyses showed that only the bone resorption markers TRACP 5b, CTX and ICTP, and the marker of angiogenesis VEGF were independently associated with BM. Those markers correctly distinguished 85% of BC patients with or without BM from normal individuals. Longitudinal analyses showed that patients with primary BC who developed BM during follow-up had higher levels of TRACP5b (+95%, P=0.08) at the time of primary diagnosis, those patients had also a higher increases of ICTP (P=0.006), MMP-7 (P=0.004) and TIMP-1 (P=0.017) during follow-up than patients who did not progress toward bone metastasis. This study provides evidence of increase and interrelationship of circulating markers of angiogenesis, invasion and bone resorption in patients with BC with and without BM. Markers of bone resorption have the highest independent diagnostic value for detecting and potentially predicting BM in breast carcinoma patients

Protein-Binding Microarray Analysis of Tumor Suppressor AP2α Target Gene Specificity

Cheap and massively parallel methods to assess the DNA-binding specificity of transcription factors are actively sought, given their prominent regulatory role in cellular processes and diseases. Here we evaluated the use of protein-binding microarrays (PBM) to probe the association of the tumor suppressor AP2α with 6000 human genomic DNA regulatory sequences. We show that the PBM provides accurate relative binding affinities when compared to quantitative surface plasmon resonance assays. A PBM-based study of human healthy and breast tumor tissue extracts allowed the identification of previously unknown AP2α target genes and it revealed genes whose direct or indirect interactions with AP2α are affected in the diseased tissues. AP2α binding and regulation was confirmed experimentally in human carcinoma cells for novel target genes involved in tumor progression and resistance to chemotherapeutics, providing a molecular interpretation of AP2α role in cancer chemoresistance. Overall, we conclude that this approach provides quantitative and accurate assays of the specificity and activity of tumor suppressor and oncogenic proteins in clinical samples, interfacing genomic and proteomic assays

Bone Sialoprotein Mediates Human Endothelial Cell Attachment and Migration and Promotes Angiogenesis

Bone sialoprotein (BSP) is a secreted glycoprotein primarily found in sites of biomineralization. Recently, we demonstrated that BSP is strongly upregulated in osteotropic cancers and particularly those that exhibit microcalcifications. BSP contains an Arg-Gly-Asp (RGD) motif found in other adhesive molecules that interact with cellular integrins. In bone, BSP has been shown to mediate the attachment of osteoblasts and osteoclasts via alpha(v)beta(3) integrin receptors. Ligands for alpha(v)beta(3) integrin are considered to play a central role during angiogenesis. Therefore, we used human umbilical vein endothelial cells (HUVECs) to study the potential role of BSP in angiogenesis. We found that purified eukaryotic recombinant human BSP (rhBSP) is able to promote both adhesion and chemotactic migration of HUVECs in a dose-dependent manner. These interactions involve HUVEC alpha(v)beta(3) integrin receptors and the RGD domain of BSP. Indeed, HUVECs attach to a recombinant BSP fragment containing the RGD domain, whereas this response is not observed with the same fragment in which RGD has been mutated to Lys-Ala-Glu (KAE). A cyclic RGD BSP peptide inhibits both adhesion and migration of HUVECs to rhBSP. Moreover, anti-alpha(v)beta(3) but not anti-alpha(v)beta(5) monoclonal antibodies also prevent BSP-mediated adhesion and migration of HUVECs. We observed that both rhBSP and the RGD BSP recombinant fragment stimulated ongoing angiogenesis on the chorioallantoic chick membrane assay. BSP angiogenic activity was inhibited by anti-alpha(v)beta(3) antibody, and the KAE BSP fragment was inactive. Our findings represent the first report implicating BSP in angiogenesis. BSP could play a critical role in angiogenesis associated with bone formation and with tumor growth and metastatic dissemination