In-vivo longitudinal MRI study: an assessment of melanoma brain metastases in a clinically relevant mouse model.

Brain metastases are an important clinical problem. Few animal models exist for melanoma brain metastases; many of which are not clinically relevant. Longitudinal MRI was implemented to examine the development of tumors in a clinically relevant mouse model of melanoma brain metastases. Fifty thousand human metastatic melanoma (A2058) cells were injected intracardially into nude mice. Three Tesla MRI was performed using a custom-built gradient insert coil and a mouse solenoid head coil. Imaging was performed on consecutive days at four time points. Tumor burden and volumes of metastases were measured from balanced steady-state free precession image data. Metastases with a disrupted blood-tumor barrier were identified from T1-weighted spin echo images acquired after administration of gadopentetic acid (Gd-DTPA). Metastases permeable to Gd-DTPA showed signal enhancement. The number of enhancing metastases was determined by comparing balanced steady-state free precession images with T1-weighted spin echo images. After the final imaging session, ex-vivo permeability and histological analyses were carried out. Imaging showed that both enhancing and nonenhancing brain metastases coexist in the brain, and that most metastases switched from the nonenhancing to the enhancing phenotype. Small numbers of brain metastases were enhancing when first detected by MRI and remained enhancing, whereas other metastases remained nonenhancing to Gd-DTPA throughout the experiment. No clear relationship existed between the permeability of brain metastases and size, brain location and age. Longitudinal in-vivo MRI is key to studying the complex and dynamic processes of metastasis and changes in the blood-tumor barrier permeability, which may lead to a better understanding of the variable responses of brain metastases to treatments

Density functional theory study of the structural, electronic, lattice dynamical, and thermodynamic properties of Li

The structural, electronic, lattice dynamical, optical, thermodynamic, and CO{sub 2} capture properties of monoclinic and triclinic phases of Li{sub 4}SiO{sub 4} are investigated by combining density functional theory with phonon lattice dynamics calculations. We found that these two phases have some similarities in their bulk and thermodynamic properties. The calculated bulk modulus and the cohesive energies of these two phases are close to each other. Although both of them are insulators, the monoclinic phase of Li{sub 4}SiO{sub 4} has a direct band gap of 5.24 eV while the triclinic Li{sub 4}SiO{sub 4} phase has an indirect band gap of 4.98 eV. In both phases of Li{sub 4}SiO{sub 4}, the s orbital of O mainly contributes to the lower-energy second valence band (VB{sub 2}) and the p orbitals contribute to the fist valence band (VB{sub 1}) and the conduction bands (CBs). The s orbital of Si mainly contributes to the lower portions of the VB1 and VB{sub 2}, and Si p orbitals mainly contribute to the higher portions of the VB{sub 1} and VB{sub 2}. The s and p orbitals of Li contribute to both VBs and to CBs, and Li p orbitals have a higher contribution than the Li s orbital. There is possibly a phonon soft mode existing in triclinic {gamma}-Li{sub 4}SiO{sub 4}; in the monoclinic Li{sub 4}SiO{sub 4}, there are three phonon soft modes, which correspond to the one type of Li disordered over a few sites. Their LO-TO splitting indicates that both phases of Li{sub 4}SiO{sub 4} are polar anisotropic materials. The calculated infrared absorption spectra for LO and TO modes are different for these two phases of Li{sub 4}SiO{sub 4}. The calculated relationships of the chemical potential versus temperature and CO{sub 2} pressure for reaction of Li{sub 4}SiO{sub 4} with CO{sub 2} shows that Li{sub 4}SiO{sub 4} could be a good candidate for a high-temperature CO{sub 2} sorbent while used for postcombustion capture technology

pH-sensitive core-shell nanoparticles for intracellular drug delivery

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemical Engineering, 2008.This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.Vita.Includes bibliographical references (p. 193-208).Therapeutics such as proteins, DNA, or siRNA, can only exert their function in the cell cytosol or nucleus. However, most of them are cell membrane impermeable molecules that can only be taken up by cells via endocytosis or phagocytosis. Such drug molecules are thus confined in endolysosomes, where reduced pH and degradative enzymes may destroy them without therapeutic gain. Efficient escape of drug molecules to the cytosol before destruction in endolysosomes is a major challenge for intracellular drug delivery. To address this issue, we designed a pH-sensitive core-shell nanoparticle to segregate the functions of the particle into an endosome-disrupting pH-responsive core that would absorb protons at endolysosomal pH, and a shell whose composition could be tuned to facilitate particle targeting, cell binding, and drug binding. Two-stage surfactant-free emulsion polymerization of 2-diethylamino ethyl methacrylate (DEAEMA) (core) and 2-amino ethyl methacrylate (AEMA) (shell) in the presence of a crosslinker was used for the synthesis of monodisperse core-shell hydrogel nanoparticles of 200 nm in diameter. The protonation of tertiary amine groups on the polyDEAEMA core on moving from extracellular to endolysosomal pH resulted in reversible swelling of the nanoparticles with a 2.8-fold diameter change. With the aid of pH-sensitivity of these nanoparticles, efficient cytosolic delivery of calcein (with ~95% efficiency) was achieved by disrupting endolysosomes via proton sponge effect. The primary amine rich shell was found to facilitate cell and drug binding, and provided negligible cytotoxicity by sequestering the proton sponge component from any direct interactions with cells. These particles demonstrated a useful means to deliver therapeutic molecules to the cytosol of cells of interest efficiently.(cont.) The applications of nanoparticles showed significant improvement in delivering a model antigen vaccine protein ovalbumin (OVA) to primary dendritic cells for T cell activation, and promising knockdown of mRNA by delivering siRNA to epithelial cells for gene silencing. To extend this approach to a fully biodegradable system, nanoparticles with a cleavable crosslinker bis (acryloyl) cystamine (BAC) were synthesized. Preliminary explorations of this approach showed that such particles can degrade in the presence of glutathione in vitro, a reducing peptide present at mM concentrations in the cytosol of mammalian cells. This design could potentially serve as a drug releasing mechanism to further improve delivery efficiency.by Yuhua Hu.Ph.D

Multi-sensor large scale land surface data assimilation using ensemble approaches

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Civil and Environmental Engineering, 2006.Includes bibliographical references (p. 223-234).One of the ensemble Kalman filter's (EnKF) attractive features in land surface applications is its ability to provide distributional information. The EnKF relies on normality approximations that improve its efficiency but can also compromise the accuracy of its distributional estimates. The effects of these approximations are evaluated by comparing the conditional marginal distributions and moments estimated by the EnKF to those obtained from an SIR particle filter, which gives exact solutions for large ensemble sizes. The results show that overall the EnKF appears to provide a good approximation for nonlinear, non-normal land surface problems. A difficulty in land data assimilation problems results from the high dimensionality of states created by spatial discretization over large computational grids. The high dimensionality can be reduced by exploiting the fact that soil moisture field may have significant spatial correlation structure especially after extensive rainfall while it may have local structure determined by soil and vegetation variability after prolonged drydown. This is confirmed by SVD of the replicate matrix produced in an ensemble forecasting experiment. Local EnKF's are suitable for problems during dry periods but give less accurate results after rainfall.(cont.) The most promising option is to develop a generalized method that reflects structural changes in the ensemble. A highly efficient ensemble multiscale filter (EnMSF) is then proposed to solve large scale nonlinear estimation problems with arbitrary uncertainties. At each prediction step realizations of the state variables are propagated. At update times, joint Gaussian distribution of states and measurements are assumed and the Predictive Efficiency method is used to identify a multiscale tree to approximate statistics of the propagated ensemble. Then a two-sweep update is performed to estimate the state variables using all the data. By controlling the tree parameters, the EnMSF can reduce sampling error while keep long range correlation in the ensemble. Applications of the EnMSF to Navier-Stokes equation and a nonlinear diffusion problem are demonstrated. Finally, the EnMSF is successfully applied to soil moisture and surface fluxes estimation over the Great Plains using synthetic multiresolution L-band passive and active microwave soil moisture measurements following HYDROS specifications.by Yuhua Zhou.Ph.D

O-GlcNAc modifications regulate cell survival and epiboly during zebrafish development

<p>Abstract</p> <p>Background</p> <p>The post-translational addition of the monosaccharide O-linked β-<it>N</it>-acetylglucosamine (O-GlcNAc) regulates the activity of a wide variety of nuclear and cytoplasmic proteins. The enzymes O-GlcNAc Transferase (Ogt) and O-GlcNAcase (Oga) catalyze, respectively, the attachment and removal of O-GlcNAc to target proteins. In adult mice, Ogt and Oga attenuate the response to insulin by modifying several components of the signal transduction pathway. Complete loss of <it>ogt </it>function, however, is lethal to mouse embryonic stem cells, suggesting that the enzyme has additional, unstudied roles in development. We have utilized zebrafish as a model to determine role of O-GlcNAc modifications in development. Zebrafish has two <it>ogt </it>genes, encoding six different enzymatic isoforms that are expressed maternally and zygotically.</p> <p>Results</p> <p>We manipulated O-GlcNAc levels in zebrafish embryos by overexpressing zebrafish <it>ogt</it>, human <it>oga </it>or by injecting morpholinos against <it>ogt </it>transcripts. Each of these treatments results in embryos with shortened body axes and reduced brains at 24 hpf. The embryos had 23% fewer cells than controls, and displayed increased rates of cell death as early as the mid-gastrula stages. An extensive marker analysis indicates that derivatives of three germ layers are reduced to variable extents, and the embryos are severely disorganized after gastrulation. Overexpression of Ogt and Oga delayed epiboly and caused a severe disorganization of the microtubule and actin based cytoskeleton in the extra-embryonic yolk syncytial layer (YSL). The cytoskeletal defects resemble those previously reported for embryos lacking function of the Pou5f1/Oct4 transcription factor <it>spiel ohne grenzen</it>. Consistent with this, Pou5f1/Oct4 is modified by O-GlcNAc in human embryonic stem cells.</p> <p>Conclusion</p> <p>We conclude that O-GlcNAc modifications control the activity of proteins that regulate apoptosis and epiboly movements, but do not seem to regulate germ layer specification. O-GlcNAc modifies the transcription factor Spiel ohne grenzen/Pou5f1 and may regulate its activity.</p

Human immunodeficiency virus type 1 specific cytotoxic T lymphocyte responses in Chinese infected with HIV-1 B'/C Recombinant (CRF07_BC)

<p>Abstract</p> <p>Background</p> <p>The characterization of HIV-1-specific T cell responses in people infected with locally circulating HIV-1 strain will facilitate the development of HIV-1 vaccine. Sixty intravenous drug users infected with HIV-1 circulating recombinant form 07_BC (CRF07_BC), which has been spreading rapidly in western China from north to south, were recruited from Xinjiang, China to assess the HIV-1-specific T cell responses at single peptide level with overlapping peptides (OLP) covering the whole concensus clades B and C proteome.</p> <p>Results</p> <p>The median of the total magnitude and total number of OLPs recognized by CTL responses were 10925 SFC/million PBMC and 25 OLPs, respectively, when tested by clade C peptides, which was significantly higher than when tested by clade B peptides. The immunodominant regions, which cover 14% (58/413) of the HIV-1 proteome, are widely distributed throughout the HIV-1 proteome except in Tat, Vpu and Pol-PR, with Gag, Pol-RT, Pol-Int and Nef being most frequently targeted. The subdominant epitopes are mostly located in p24, Nef, integrase, Vpr and Vif. Of the responses directed to clade C OLPs, 61.75% (972/1574) can be observed when tested with corresponding clade B OLPs. However, Pol-PR and Vpu tend to be targeted in the clade B sequence rather than the clade C sequence, which is in line with the recombinant pattern of CRF07_BC. Stronger and broader CTL responses in subjects with CD4 cell counts ranging from 200 to 400/mm³were observed when compared to those with less than 200/mm³or more than 400/mm³, though there have been no significant correlations identified between the accumulative CTL responses or overall breadth and CD4 cell count or plasma viral load.</p> <p>Conclusion</p> <p>This is the first study conducted to comprehensively address T cell responses in Chinese subjects infected with HIV-1 CRF07_BC in which subtle differences in cross-reactivity were observed, though similar patterns of overall immune responses were demonstrated with clade B infected populations. The immunodominant regions identified in this population can facilitate future HIV-1 vaccine development in China.</p

Benzoate Catabolite Repression of the Phenol Degradation in Acinetobacter calcoaceticus PHEA-2

Acinetobacter calcoaceticus PHEA-2 exhibited a delayed utilization of phenol in the presence of benzoate. Benzoate supplementation completely inhibited phenol degradation in a benzoate 1,2-dioxygenase knockout mutant. The mphR encoding the transcriptional activator and mphN encoding the largest subunit of multi-component phenol hydroxylase in the benA mutant were significantly downregulated (about 7- and 70-fold) on the basis of mRNA levels when benzoate was added to the medium. The co-transformant assay of E. coli JM109 with mphK::lacZ fusion and the plasmid pETR carrying mphR gene showed that MphR did not activate the mph promoter in the presence of benzoate. These results suggest that catabolite repression of phenol degradation by benzoate in A. calcoaceticus PHEA-2 is mediated by the inhibition of the activator protein MphR