Two-photon imaging of cancer cell extravasation in live mice

Abstract MDA-MB-231 breast cancer cells were engineered to express cytoplasmic paxillin-GFP and nuclear H2B-mCherry. In order to image extravasation, the cancer cells were injected in the blood stream of nude mice. Using 2-photon excitation microscopy we can simultaneously excite the two probes and also visualize the autofluorescence of tissues. A skin flap was opened to visualize blood vessels and recognize the position of the cancer cells. Two-photon imaging showed that after an initial phase in which the cells are non-adherent, some cells spread on the internal surface of the capillaries. Days later some cells started to appear on the external side of the capillary. The extravasated cells extend very long protrusions into the tissue. The goal was to determine if at the end of the long protrusion, if it is possible to observe the formation of focal adhesions by imaging paxillin-GFP. Preliminary results show that when cells start to adhere to the blood vessel wall they form focal adhesions as determined by the characteristic elongated features observed in the paxillin-GFP channel. New approaches will allow the tracking of the tip of the protrusion to determine if focal adhesions are forming there as the cells extravasate. This is important in establishing the mechanism of cell extravasation and migration in tissues. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1412. doi:10.1158/1538-7445.AM2011-141

Magnetotransport in Sr3PbO antiperovskite with three-dimensional massive Dirac electrons

Novel topological phenomena are anticipated for three-dimensional (3D) Dirac electrons. The magnetotransport properties of cubic ${\rm Sr_{3}PbO}$ antiperovskite, theoretically proposed to be a 3D massive Dirac electron system, are studied. The measurements of Shubnikov-de Haas oscillations and Hall resistivity indicate the presence of a low density ($\sim 1 \times 10^{18}$ ${\rm cm^{-3}}$) of holes with an extremely small cyclotron mass of 0.01-0.06$m_{e}$. The magnetoresistance $\Delta\rho_{xx}(B)$ is linear in magnetic field $B$ with the magnitude independent of temperature. These results are fully consistent with the presence of 3D massive Dirac electrons in ${\rm Sr_{3}PbO}$. The chemical flexibility of the antiperovskites and our findings in the family member, ${\rm Sr_{3}PbO}$, point to their potential as a model system in which to explore exotic topological phases

Real-time imaging of 3-dimensional cancer cell movement in tissues

Abstract Our knowledge of how cells move in 3D in tissues is limited due to the lack of imaging methods that can produce 3D images fast enough and with sufficient resolution. Cancer cells migrate in 3D by forming adhesion points at the end of very long cellular protrusions. These protrusions are very thin and it is difficult to visualize adhesions along the protrusion surface. Conventional 3D stack reconstruction has relatively low resolution unless it is done using many frames. This results in a very slow acquisition in 3D confocal microscopy. Faster methods of 3D data acquisition (spinning disk microscopy) cannot be easily implemented since there is significant amount of scatter in tissues. A major obstacle in imaging adhesions is to find and track them so that they will not go out of focus. We are developing a new method which is based on orbiting imaging around cellular protrusions to visualize protein dynamics during extravasation. A feedback mechanism controls the center of the orbit to be at the center of the fluorescence distribution. A program reconstructs the shape of the protrusions in 3D. The fluorescence intensity in one or more channels is also simultaneously measured. The fluorescence intensity of one channel is used to paint the protrusion shape, which results in the 3D reconstruction of the protrusion. During the orbit, the second channel of the microscope measures the second harmonic generation (SHG) signal. We then correlated the appearance of bright fluorescence spots on the protrusion surface with the points of contact of the protrusion. This method will enable imaging of cancer cell invasion in 3-dimentions in live mice in real time. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4750. doi:10.1158/1538-7445.AM2011-475

Lens Regeneration in Axolotl: New Evidence of Developmental Plasticity

Background: Among vertebrates lens regeneration is most pronounced in newts, which have the ability to regenerate the entire lens throughout their lives. Regeneration occurs from the dorsal iris by transdifferentiation of the pigment epithelial cells. Interestingly, the ventral iris never contributes to regeneration. Frogs have limited lens regeneration capacity elicited from the cornea during pre-metamorphic stages. The axolotl is another salamander which, like the newt, regenerates its limbs or its tail with the spinal cord, but up until now all reports have shown that it does not regenerate the lens

Trace element and stable isotope analyses of deep sea fish from the Sulu sea, Philippines

Thirty-five deep sea fishes belonging to 22 species and one unidentified specimen obtained from the Sulu Sea, located in the southwestern area of the Philippines were analyzed in the late 2002, for 23 trace elements using ICP-MS, HGAAS and CV-AAS. Predominant accumulation of strontium (Sr) was observed in all the samples. This stems from the fact that the whole body of fish was homogenized since Sr is known to accumulate in bones and hard tissues. Mercury concentrations in all the 36 samples were below the detection limit. Cadmium concentrations were generally below 1ìg/g dry weight (dw) except in Pterygotrigla spp. (4.29 ìg/g dw) and Sternoptyx pseudodiaphana (2.89 ìg/g dw). Concentrations of Pb were predominantly low with about 90% of the specimens having less than 1 ìg/g dw. In general, concentrations of Sr, Zn, Cu, Se and Cd appeared to increase with increasing depth of occurrence of the species.Manganese, Tl, Pb, Bi, In, Cs and As showed significant positive correlation (p < 0.05) with d15N, suggesting that these elements were biomagnified. To our knowledge, this is the first study reporting Tl biomagnification in fish. Rubidium and Cs showed significant positive correlation with d13C, implying that Rb and Cs would originate from offshore waters as oceanic plankton has high d13C. Comparing results from this study to the dietary standards and guidelines for Hg, Pb, Cu and Zn in fish and shellfish of the Ministry of Agriculture, Fisheries and Food of the United Kingdom,these levels were not high to warrant concern if they were to be consumed by humans. However, 16.7% of the fish samples had high Cr levels when compared with the Hong Kong’s safe limit of 4 ìg/g dw for Cr in sea food. This constitutes a health risk to humans, as Cr is potentially toxic

Inverse-perovskites A3BO (A = Sr, Ca, Eu/B = Pb, Sn) : a platform for control of Dirac and Weyl fermions

This work was partly supported by the Japan Society for the Promotion of Science (JSPS) KAKENHI (Grant Nos. 24224010, 15K13523, JP15H05852, JP15K21717, and 17H01140), EPSRC (Grant No. EP/P024564/1), and the Alexander von Humboldt FoundationBulk Dirac electron systems have attracted strong interest for their unique magnetoelectric properties as well as their close relation to topological (crystalline) insulators. Recently, the focus has been shifting toward the role of magnetism in stabilizing Weyl fermions as well as chiral surface states in such materials. While a number of nonmagnetic systems are well known, experimental realizations of magnetic analogs are a key focus of current studies. Here, we report on the physical properties of a large family of inverse perovskites A3BO (A = Sr, Ca, Eu/B = Pb, Sn) in which we are able to not only stabilize 3D Dirac electrons at the Fermi energy but also chemically control their properties. In particular, it is possible to introduce a controllable Dirac gap, change the Fermi velocity, tune the anisotropy of the Dirac dispersion, and—crucially—introduce complex magnetism into the system. This family of compounds therefore opens up unique possibilities for the chemical control and systematic investigation of the fascinating properties of such topological semimetals.Publisher PDFPeer reviewe

Transcultural adaptation to the Brazilian Portuguese of the Postpartum Bonding Questionnaire for assessing the postpartum bond between mother and baby

The establishment of the bond between mother and baby in the postpartum period is important for ensuring the physical and psychological health of both. This short communication reports the first phase of the cross-cultural translation and adaptation to the Brazilian context of the Postpartum Bonding Questionnaire (PBQ). Four aspects of equivalence between the original scale and the Portuguese version were evaluated: the conceptual, semantic, operational and item equivalences. Literature review, the study of PBQ history, translation, expert evaluation, back-translation and pretests involving 30 mothers with children aging up to 7 months using a primary healthcare unit were conducted. Each step demonstrated the need for adjustments, which were made during the adaptation process. At the end of the study, a version of PBQ in Brazilian Portuguese equivalent to the original one was obtained, offering promise for national studies on the mother-baby bond, and its influence on health, and for use in health services

PACSIN2 accelerates nephrin trafficking and is up-regulated in diabetic kidney disease

Nephrin is a core component of podocyte (glomerular epithelial cell) slit diaphragm and is required for kidney ultrafiltration. Down-regulation or mislocalization of nephrin has been observed in diabetic kidney disease (DKD), characterized by albuminuria. Here, we investigate the role of protein kinase C and casein kinase 2 substrate in neurons 2 (PACSIN2), a regulator of endocytosis and recycling, in the trafficking of nephrin and development of DKD. We observe that PACSIN2 is up-regulated and nephrin mislocalized in podocytes of obese Zucker Diabetic Fatty (ZDF) rats that have altered renal function. In cultured podocytes, PACSIN2 and nephrin colocalize and interact. We show that nephrin is endocytosed in PACSIN2-positive membrane regions and that PACSIN2 overexpression increases both nephrin endocytosis and recycling. We identify rabenosyn-5, which is involved in early endosome maturation and endosomal sorting, as a novel interaction partner of PACSIN2. Interestingly, rabenosyn-5 expression is increased in podocytes in obese ZDF rats, and, in vitro, its overexpression enhances the association of PACSIN2 and nephrin. We also show that palmitate, which is elevated in diabetes, enhances this association. Collectively, PACSIN2 is up-regulated and nephrin is abnormally localized in podocytes of diabetic ZDF rats. In vitro, PACSIN2 enhances nephrin turnover apparently via a mechanism involving rabenosyn-5. The data suggest that elevated PACSIN2 expression accelerates nephrin trafficking and associates with albuminuria.Peer reviewe

A Complement Receptor C5a Antagonist Regulates Epithelial to Mesenchymal Transition and Crystallin Expression After Lens Cataract Surgery in Mice

Purpose: To evaluate the effects of complement employing a mouse model for secondary cataract. Methods: The role of complement receptor C5a (CD88) was evaluated after cataract surgery in mice. An antagonist specific to C5a receptor was administered intraperitoneally to mice. Epithelial to mesenchymal transition (EMT) was evaluated by alpha-smooth muscle actin (α-SMA) staining and proliferation by bromodeoxyuridine (5-bromo-2\u27- deoxyuridine, BrdU) incorporation. Gene expression patterns was examined by microarray analysis and quantitative polymerase chain reaction (QPCR). Results: We found that administration of a C5aR antagonist in C57BL/6J mice decreases EMT, as evidenced by α-SMA expression, and cell proliferation. Gene expression by microarray analysis reveals discreet steps of gene regulation in the two major stages that of EMT and lens fiber differentiation in vivo. A hallmark of the microarray analysis is that the antagonist seems to be a novel stage-specific regulator of crystallin genes. At week two, which is marked by lens fiber differentiation genes encoding 12 crystallins and 3 lens-specific structural proteins were severely down-regulated. Conclusions: These results suggest a possible therapeutic role of an antagonist to C5aR in preventing secondary cataracts after surgery. Also these results suggest that crystallin gene expression can be regulated by pro-inflammatory events in the eye

Mtss1 promotes cell-cell junction assembly and stability through the small GTPase Rac1

Cell-cell junctions are an integral part of epithelia and are often disrupted in cancer cells during epithelial-to-mesenchymal transition (EMT), which is a main driver of metastatic spread. We show here that Metastasis suppressor-1 (Mtss1; Missing in Metastasis, MIM), a member of the IMD-family of proteins, inhibits cell-cell junction disassembly in wound healing or HGF-induced scatter assays by enhancing cell-cell junction strength. Mtss1 not only makes cells more resistant to cell-cell junction disassembly, but also accelerates the kinetics of adherens junction assembly. Mtss1 drives enhanced junction formation specifically by elevating Rac-GTP. Lastly, we show that Mtss1 depletion reduces recruitment of F-actin at cell-cell junctions. We thus propose that Mtss1 promotes Rac1 activation and actin recruitment driving junction maintenance. We suggest that the observed loss of Mtss1 in cancers may compromise junction stability and thus promote EMT and metastasis