175 results match your criteria: "Medical Biology Institute[Affiliation]"

Corneal scarring is a leading cause of worldwide blindness. Human mesenchymal stem cells (MSC) have been reported to promote corneal wound healing through secreted exosomes. This study investigated the wound healing and immunomodulatory effects of MSC-derived exosomes (MSC-exo) in corneal injury through an established rat model of corneal scarring.

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Gene fusion, as a prototypical pathognomonic mutation, contributes to genome complexity, and the -transcription-induced gene fusions generated by read-through transcription of adjacent genes have been found to be important for tumor development. We screened read-through transcription events from stomach adenocarcinoma RNA-seq data and selected three candidates , and , to assess their biological role in gastric cancer. The expression of all three read-through fusion transcripts was confirmed in gastric cancer cell lines and paired normal/tumor gastric cancer tissues by real-time quantitative reverse transcription polymerase chain reaction and their expression was found to be significantly higher in the tumor ( < 0.

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Naegleria fowleri, a free-living amoeba, is the causative pathogen of primary amoebic meningoencephalitis in humans and experimental mice. N. fowleri is capable of destroying tissues and host cells through lytic necrosis.

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Oligonucleotide N3'->P5' Phosphoramidates (PN) may confer advantages over unmodified phosphodiester compounds for therapeutic applications (1). Previous in vitro data demonstrated that PN Oligodeoxynucleotides (ODNs) possess several advantageous features, including RNase H-independence, an improved resistance to nuclease degradation, decreased protein binding, and high affinity sequence-specific binding to complementary RNAs (1, 2). Consequently, we undertook a study to investigate the effects of PN antisense (AS) oligos targeted against the p65 subunit of the Nuclear Factor Kappa beta (NF-kappaB) transcription factor in vivo, in mice.

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The aqueous phase monomers of fatty acids (FFA) appear in many steps of fat metabolism. Understanding metabolism requires that accurate measurements of FFA levels be determined in enzyme-mediated as well as in membrane and protein binding reactions. Measuring long chain FFA levels with sufficient sensitivity and temporal resolution is now possible using fluorescent probes constructed by ligating fluorescent groups and fatty acid binding proteins.

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The interactions of long chain fatty acids (FA) with wild type (WT) fatty acid binding proteins (FABP) and engineered FABP mutants have been monitored to determine the equilibrium binding constants as well as the rate constants for binding and dissociation. These measurements have been done using the fluorescent probes, ADIFAB and ADIFAB2, that allow the determination of the free fatty acid (FFA) concentration in the reaction of FA with proteins and membranes. The results of these studies indicate that forWT proteins from adipocyte, heart, intestine, and liver, Kd values are in the nM range and affinities decrease with increasing aqueous solubility of the FA.

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Engineered mutants of human complement component C3 were used to test the idea that sites of length polymorphisms in protein families (indels) can guide a search for protein:protein interaction sites. Sequence changes were introduced at each of the 27 indels in the C3/4/5 protein family, and mutants at 26 indels were expressed by transiently transfected COS cells. Expressed proteins were assayed 1) for concentration, by ELISA and by autoradiography of radiolabeled protein; 2) for classical pathway hemolytic activity; 3) for susceptibility to proteolytic activation by the alternative pathway and cobra venom factor C3 convertases; and 4) for susceptibility to complement factor I in the presence of factor H.

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We have developed a simple PCR strategy, termed vector-hexamer PCR, that is unique in its ability to easily recover every insert end from large insert clones in YAC and BAC vectors. We used this method to amplify and isolate all insert ends from a YAC contig covering the mouse Igh locus. Seventy-seven ends were amplified and sequenced from 36 YAC clones from four libraries in the pYAC4 vector.

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Evidence from a number of laboratories suggests that membrane proteins may meditate the transport of physiologic fatty acids (FA) across cell membranes. However, actual transport of unbound free fatty acids (unbound FFA) from the aqueous phase on one side of a cell membrane to the aqueous phase on the other side has not been measured previously. In this study, we have used the fluorescent probe of unbound FFA, ADIFAB, to monitor the time course of FA movement from the outer to the inner aqueous compartments, and from the lipid membrane to the outer aqueous compartment of red cell ghosts.

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We constructed 18 single amino acid mutants of the adipocyte fatty acid-binding protein (A-FABP) and 17 of the intestinal fatty acid-binding protein (I-FABP), at locations in the fatty acid (FA) binding sites. For each mutant protein, we measured thermodynamic parameters that characterize FA binding. Binding affinities ranged from about 200-fold smaller to 30-fold larger than the wild type (WT) proteins.

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Evidence from a number of laboratories suggests that membrane proteins may meditate the transport of physiologic fatty acids (FA) across cell membranes. However, studies using lipid membranes indicate that FA are capable of spontaneous flip-flip, raising the possibility that rapid transport through the lipid phase obviates the need for a transport protein. Determining the rate-limiting steps for transport of FA across lipid membranes, therefore, is central to understanding FA transport across cell membranes.

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Site-specific variants of rat intestinal fatty acid-binding protein were constructed to identify the molecular interactions that are important for binding to fatty acids (FAs). Several variants displayed affinities that appeared incompatible with the crystal structure of the protein-FA complex. Thermodynamic measurements provided an explanation for these apparent inconsistencies and revealed that binding affinities often inaccurately reported changes in protein-FA interactions because changes in the binding entropy and enthalpy were usually compensatory.

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Stimulation of cloned cytotoxic T lymphocytes (CTL) with anti-T-cell receptor (TCR) monoclonal antibody (mAb) in solution resulted in rapid and sustained activation of adhesion to immobilized fibronectin (FN) but did not initiate degranulation. Addition of a second antibody (Ab) to further cross-link the TCR substantially increased the level of adhesion and also activated degranulation, as measured by release of serine esterase, in the presence of immobilized FN but not in its absence. Thus, binding to FN can provide a costimulatory signal to activate degranulation.

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An issue that is central to understanding cellular fatty acid (FA) metabolism is whether physiologic transport of FA across cell membranes requires protein mediation or can be satisfied by the rate of spontaneous movement through the lipid phase. For this reason, considerable effort has been devoted to determining the rate-limiting steps for transport of FA across pure lipid bilayer membranes. Previously, we found that transbilayer flip-flop was the rate-limiting step for transport of long chain anthroyloxy FA (AOFA) across lipid bilayers and that the times for long chain AOFA flip-flop were > or = 100 s, yielding rate constants for flip-flop (k(ff)) that were < or = 0.

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To find protein-protein interactive sites in complement component C3, we examined regions of C3 that are proximal to sites of length polymorphism or indels in the C345 protein family. We reasoned that indels probably mark protein interactive sites because they usually involve residues at protein surfaces. To test for the involvement of individual indels, we examined the effects on complement function of synthetic peptides corresponding to indel-proximal segments of C3.

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Murine acquired immune deficiency syndrome (MAIDS) is an immunosuppressive disease of mice induced by infection with the LP-BM5 murine leukemia virus (MuLV) retrovirus isolate. Certain inbred strains of mice are resistant to disease, but F1 crosses between sensitive and resistant strains are predominantly sensitive to MAIDS. One inbred strain, BDP, demonstrates a novel disease phenotype, recovery of immune function after a period of profound immune suppression.

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IL-4 and CD40 ligand stimulate transcription of CD23 (Fc epsilonRII) in B cells and are necessary for the expression of germline epsilon mRNA and production of IgE. Because in vivo studies have shown that the Fc epsilonRII is involved in the regulation of IgE, a study was initiated to compare how IL-4 and engagement of CD40 up-regulate the Fc epsilonRII and epsilon genes. Herein, we describe the preparation of a series of linker-scanning mutants that cover the IL-4 response region in the murine Fc epsilonRII promoter, and their function when transfected into M12.

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Serum unbound free fatty acid levels (FFAu) were measured in patients undergoing percutaneous transluminal coronary angioplasty (PTCA) using the fluorescent probe acrylodan intestinal fatty acid binding protein (ADIFAB). These are the first measurements of FFAu, under nonphysiologic conditions. In these studies, FFAu, levels were determined in 22 patients 5 minutes before and 30 minutes after the procedure.

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Fatty acid-binding protein from rat liver (L-FABP) binds 2 fatty acids (FA) per protein, in contrast to FABPs from adipocyte, heart, and intestine, for which binding and structural studies are consistent with a single FA binding site. To understand better the unique characteristics of L-FABP, we have carried out equilibrium binding and kinetic measurements of long chain FA using the fluorescent probes of free fatty acids (FFA), ADIFAB and ADIFAB2, to monitor the concentration of FFA in the reaction of FA with L-FABP. We found that the dissociation constants (Kd) ranged from about 1 nM to 4 microM, being largest for myristate at 45 degrees C and smallest for oleate at 10 degrees C, and that 2 FA were bound per L-FABP for all temperatures and FA.

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Tyrosine phosphorylation of proteins plays a central role in T cell activation. Mitogens or anti-receptor antibodies have been employed to study these signaling events, but the extent to which these mimic receptor interactions with native ligands is unclear. Cytotoxic T lymphocytes can be activated for functional responses using purified, native class I ligands presented on a surface.

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It has been hypothesized that distinct stromal cells from niches within the microenvironment that selectively regulate stem cell functions. To test this hypothesis, we derived a panel of matched stromal cell lines from murine fetal liver. The lines were immortalized with a retroviral vector encoding a temperature sensitive SV40 T antigen, to provide a snapshot of potential heterogeneity of the in vivo stroma compartment.

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Rate constants for the interaction of fatty acids (FA) with fatty acid binding proteins (FABP) from adipocyte (A-FABP), heart (H-FABP), and intestine (I-FABP) were determined by using stopped-flow fluorometry and ADIFAB, the fluorescent probe of free fatty acids (FFA), or a new FFA probe, ADIFAB2, constructed by derivatizing with acrylodan the Leu72 --> Ala mutant of I-FABP. ADIFAB2, because its binding affinities are about 10-fold greater than ADIFAB, was found to be more accurate for monitoring the kinetics of the higher affinity reactions. On- (kappa on) and off- (kappa off) rate constants were determined as a function of temperature.

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The impact of intrinsic B lymphocyte heterogeneity and of microenvironmental influences on serum immunoglobulin production by B cells was examined by intravenous (i.v.) and intraperitoneal (i.

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Adoptive transfer of resistance to murine retrovirus-induced immune suppression.

Cell Immunol

April 1996

Department of Immunology, Medical Biology Institute, La Jolla, California, USA.

Infection of certain strains of mice, such as C57BL/6 and C57BL/10 [B10], with LP-BM5 murine leukemia virus (MuLV) rapidly causes a profound and lethal immune suppression. The H2d congenic strain of B10, B10.D2, is resistant to disease, but B10 x B10.

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Immunotherapy of established murine tumors with large multivalent immunogen and cyclophosphamide.

J Immunother Emphasis Tumor Immunol

March 1996

Division of Membrane Biology, Medical Biology Institute, La Jolla, California, USA.

Plasma membrane vesicles isolated from tumor cells can be incorporated onto 5-microns diameter microspheres and antigen in this form, termed large multivalent immunogen (LMI), augments generation of tumor-specific cytotoxic T lymphocyte (CTL) responses in vivo. Treatment of mice with LMI at the time of challenge with tumor significantly reduced growth of several tumors in their syngeneic hosts. Our report describes the effects of LMI on established progressing tumors, including P815 solid tumor and two fibrosarcomas in a lung-metastasis model.

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