The S100 Family Heterodimer, MRP-8/14, Binds with High Affinity to Heparin and Heparan Sulfate Glycosaminoglycans on Endothelial Cells

The Journal of Biological Chemistry. Feb, 2002  |  Pubmed ID: 11723110

The S100 family proteins MRP-8 (S100A8) and MRP-14 (S100A9) form a heterodimer that is abundantly expressed in neutrophils, monocytes, and some secretory epithelia. In inflamed tissues, the MRP-8/14 complex is deposited onto the endothelium of venules associated with extravasating leukocytes. To explore the receptor interactions of MRP-8/14, we use a model system in which the purified MRP-8/14 complex binds to the cell surface of an endothelial cell line, HMEC-1. This interaction is mediated by the MRP-14 subunit and is mirrored by recombinant MRP-14 alone. The cell surface binding of MRP-14 was blocked by heparin, heparan sulfate, and chondroitin sulfate B, and the binding sites were sensitive to heparinase I and trypsin treatment but not to chondroitinase ABC. Furthermore MRP-8/14 and MRP-14 did not bind to a glycosaminoglycan-minus cell line. MRP-14 has a high affinity for heparin (K(d) = 6.1 +/- 3.4 nm), and this interaction mimicked that with the endothelial cells. We therefore conclude that the MRP-8/14 complex binds to endothelial cells via the MRP-14 subunit interacting chiefly with heparan sulfate proteoglycans. CD36 and RAGE, two other putative receptors for MRP-8/14, were not expressed by HMEC-1 cells. This binding activity may explain the immobilization of the MRP-8/14 complex on endothelium that is observed in vivo.

Amorphous Iridium Complexes for Electrophosphorescent Light Emitting Devices

Chemical Communications (Cambridge, England). Apr, 2002  |  Pubmed ID: 12119722

Iridium complexes with fluorene-modified phenylpyridine ligands are resistant to crystallization and can be used in the fabrication of single layer light emitting diodes.

Myeloid Cell Function in MRP-14 (S100A9) Null Mice

Molecular and Cellular Biology. Apr, 2003  |  Pubmed ID: 12640137

Myeloid-related protein 14 (MRP-14) and its heterodimeric partner, MRP-8, are cytosolic calcium-binding proteins, highly expressed in neutrophils and monocytes. To understand the function of MRP-14, we performed targeted disruption of the MRP-14 gene in mice. MRP-14(-/-) mice showed no obvious phenotype and were fertile. MRP-8 mRNA but not protein is present in the myeloid cells of these mice, suggesting that the stability of MRP-8 protein is dependent on MRP-14 expression. A compensatory increase in other proteins was not detected in cells lacking MRP-8 and MRP-14. Although the morphology of MRP-14(-/-) myeloid cells was not altered, they were significantly less dense. When Ca(2+) responses were investigated, there was no change in the maximal response to the chemokine MIP-2. At lower concentrations, however, there was reduced responsiveness in MRP-14(-/-) compared with MRP-14(+/+) neutrophils. This alteration in the ability to flux Ca(2+) did not impair the ability of the MRP-14(-/-) neutrophils to respond chemotactically to MIP-2. In addition, the myeloid cell functions of phagocytosis, superoxide burst, and apoptosis were unaffected in MRP-14(-/-) cells. In an in vivo model of peritonitis, MRP-14(-/-) mice showed no difference from wild-type mice in induced inflammatory response. The data indicate that MRP-14 and MRP-8 are dispensable for many myeloid cell functions.

Conformation and Luminescence of Isolated Molecular Semiconductor Molecules

Journal of the American Chemical Society. Apr, 2003  |  Pubmed ID: 12708872

In this article, we describe, for the first time, direct comparisons of the detailed structures of two small molecule organic semiconductors, oligo(phenylenvinylene) (OPV) molecules with chains of five and six phenyl rings (5R-OC(8)H(17) and 6R-OC(8)H(17)), respectively, and their luminescence properties on a single molecule level. Our data originate from a combination of two powerful diagnostic tools in physical chemistry: ion mobility and single molecule fluorescence spectroscopy. These techniques enable us to precisely determine the shapes of isolated molecules in the gas phase and to correlate these structures to the emission from single molecules supported on bare glass substrates. The principal structural uncertainty in OPVs is the (possible) presence and location of cis-vinylene linkages (cis-defects) in the oligomer. The results show that the structures observed in the gas phase are strongly correlated to the categories of molecules observed in the single molecule polarization anisotropy measurements with nearly identical distributions for the two OPV molecules studied. Each category is also characterized by the luminescence efficiency of the molecules in each class, providing a direct correlation between the luminescence efficiency and the shape of the molecule. This combination of techniques provides a level of information far beyond that obtained via any other analytical technique.

UNC-71, a Disintegrin and Metalloprotease (ADAM) Protein, Regulates Motor Axon Guidance and Sex Myoblast Migration in C. Elegans

Development (Cambridge, England). Jul, 2003  |  Pubmed ID: 12783787

The migration of cells and growth cones is a process that is guided by extracellular cues and requires the controlled remodeling of the extracellular matrix along the migratory path. The ADAM proteins are important regulators of cellular adhesion and recognition because they can combine regulated proteolysis with modulation of cell adhesion. We report that the C. elegans gene unc-71 encodes a unique ADAM with an inactive metalloprotease domain. Loss-of-function mutations in unc-71 cause distinct defects in motor axon guidance and sex myoblast migration. Many unc-71 mutations affect the disintegrin and the cysteine-rich domains, supporting a major function of unc-71 in cell adhesion. UNC-71 appears to be expressed in a selected set of cells. Genetic mosaic analysis and tissue-specific expression studies indicate that unc-71 acts in a cell non-autonomous manner for both motor axon guidance and sex myoblast migration. Finally, double mutant analysis of unc-71 with other axon guidance signaling molecules suggests that UNC-71 probably functions in a combinatorial manner with integrins and UNC-6/netrin to provide distinct axon guidance cues at specific choice points for motoneurons.

Alternative Splicing Affecting a Novel Domain in the C. Elegans EGL-15 FGF Receptor Confers Functional Specificity

Development (Cambridge, England). Aug, 2003  |  Pubmed ID: 12835392

Fibroblast growth factor (FGF) receptors trigger a wide variety of cellular responses as diverse as cell migration, cell proliferation and cell differentiation. However, the molecular basis of the specificity of these responses is not well understood. The C. elegans FGF receptor EGL-15 similarly mediates a number of different responses, including transducing a chemoattractive signal and mediating an essential function. Analysis of the migration-specific alleles of egl-15 has identified a novel EGL-15 isoform that provides a molecular explanation for the different phenotypic effects of lesions at this locus. Alternative splicing yields two EGL-15 proteins containing different forms of a domain located within the extracellular region of the receptors immediately after the first IG domain. Neither of these two domain forms is found in any other FGF receptor. We have tested the roles of these EGL-15 receptor isoforms and their two FGF ligands for their signaling specificity. Our analyses demonstrate different physiological functions for the two receptor variants. EGL-15(5A) is required for the response to the FGF chemoattractant that guides the migrating sex myoblasts to their final positions. By contrast, EGL-15(5B) is both necessary and sufficient to elicit the essential function mediated by this receptor.

Abl Interactor 1 (Abi-1) Wave-binding and SNARE Domains Regulate Its Nucleocytoplasmic Shuttling, Lamellipodium Localization, and Wave-1 Levels

Molecular and Cellular Biology. Jun, 2004  |  Pubmed ID: 15143189

The Abl interactor 1 (Abi-1) protein has been implicated in the regulation of actin dynamics and localizes to the tips of lamellipodia and filopodia. Here, we show that Abi-1 binds the actin nucleator protein Wave-1 through an amino-terminal Wave-binding (WAB) domain and that disruption of the Abi-1-Wave-1 interaction prevents Abi-1 from reaching the tip of the lamellipodium. Abi-1 binds to the Wave homology domain of Wave-1, a region that is required for translocation of Wave-1 to the lamellipodium. Mouse embryo fibroblasts that lack one allele of Abi-1 and are homozygous null for the related Abi-2 protein exhibit decreased Wave-1 protein levels. This phenotype is rescued by Abi-1 proteins that retain Wave-1 binding but not by Abi-1 mutants that cannot bind to Wave-1. Moreover, we uncovered an overlapping SNARE domain in the amino terminus of Abi-1 that interacts with Syntaxin-1, a SNARE family member. Further, we demonstrated that Abi-1 shuttles in and out of the nucleus in a leptomycin B (LMB)-dependent manner and that complete nuclear translocation of Abi-1 in the absence of LMB requires the combined inactivation of the SNARE, WAB, and SH3 domains of Abi-1. Thus, Abi-1 undergoes nucleocytoplasmic shuttling and functions at the leading edge to regulate Wave-1 localization and protein levels.

Quantitation of Small-animal (124)I Activity Distributions Using a Clinical PET/CT Scanner

Journal of Nuclear Medicine : Official Publication, Society of Nuclear Medicine. Jul, 2004  |  Pubmed ID: 15235072

Time-dependent PET imaging can be an important tool in the assessment of radiotracer performance in murine models. We have performed a quantitative analysis of PET images of (124)I, acquired on a clinical PET system using a small-animal phantom. We then compared the recovered activity concentrations with the known activity concentration in the phantom spheres. The recovery coefficients found from the phantom data were applied to in vivo (124)I anti-HER2/neu C6.5 diabody PET data and compared with necropsy biodistribution data from the same tumor-bearing immunodeficient mouse.

ABI2-deficient Mice Exhibit Defective Cell Migration, Aberrant Dendritic Spine Morphogenesis, and Deficits in Learning and Memory

Molecular and Cellular Biology. Dec, 2004  |  Pubmed ID: 15572692

The Abl-interactor (Abi) family of adaptor proteins has been linked to signaling pathways involving the Abl tyrosine kinases and the Rac GTPase. Abi proteins localize to sites of actin polymerization in protrusive membrane structures and regulate actin dynamics in vitro. Here we demonstrate that Abi2 modulates cell morphogenesis and migration in vivo. Homozygous deletion of murine abi2 produced abnormal phenotypes in the eye and brain, the tissues with the highest Abi2 expression. In the absence of Abi2, secondary lens fiber orientation and migration were defective in the eye, without detectable defects in proliferation, differentiation, or apoptosis. These phenotypes were consistent with the localization of Abi2 at adherens junctions in the developing lens and at nascent epithelial cell adherens junctions in vitro. Downregulation of Abi expression by RNA interference impaired adherens junction formation and correlated with downregulation of the Wave actin-nucleation promoting factor. Loss of Abi2 also resulted in cell migration defects in the neocortex and hippocampus, abnormal dendritic spine morphology and density, and severe deficits in short- and long-term memory. These findings support a role for Abi2 in the regulation of cytoskeletal dynamics at adherens junctions and dendritic spines, which is critical for intercellular connectivity, cell morphogenesis, and cognitive functions.

Bariatric and Associated Operations in Private and Academic Practices

Obesity Surgery. Nov-Dec, 2004  |  Pubmed ID: 15603660

The types of bariatric and the associated operations performed by academic and private surgeons were surveyed.

Quantitative Immuno-positron Emission Tomography Imaging of HER2-positive Tumor Xenografts with an Iodine-124 Labeled Anti-HER2 Diabody

Cancer Research. Feb, 2005  |  Pubmed ID: 15735035

Positron emission tomography (PET) provides an effective means of both diagnosing/staging several types of cancer and evaluating efficacy of treatment. To date, the only U.S. Food and Drug Administration-approved radiotracer for oncologic PET is (18)F-fluoro-deoxyglucose, which measures glucose accumulation as a surrogate for malignant activity. Engineered antibody fragments have been developed with the appropriate targeting specificity and systemic elimination properties predicted to allow for effective imaging of cancer based on expression of tumor associated antigens. We evaluated a small engineered antibody fragment specific for the HER2 receptor tyrosine kinase (C6.5 diabody) for its ability to function as a PET radiotracer when labeled with iodine-124. Our studies revealed HER2-dependent imaging of mouse tumor xenografts with a time-dependent increase in tumor-to-background signal over the course of the experiments. Radioiodination via an indirect method attenuated uptake of radioiodine in tissues that express the Na/I symporter without affecting the ability to image the tumor xenografts. In addition, we validated a method for using a clinical PET/computed tomography scanner to quantify tumor uptake in small-animal model systems; quantitation of the tumor targeting by PET correlated with traditional necropsy-based analysis at all time points analyzed. Thus, diabodies may represent an effective molecular structure for development of novel PET radiotracers.

Isolation of ScFvs to in Vitro Produced Extracellular Domains of EGFR Family Members

Cancer Biotherapy & Radiopharmaceuticals. Dec, 2005  |  Pubmed ID: 16398612

The members of the epidermal growth factor receptor (EGFR) family are over expressed in a variety of malignancies and are frequently linked to aggressive disease and a poor prognosis. Although clinically effective monoclonal antibodies (MAbs) have been developed to target HER2 and EGFR, the remaining two family members, HER3 and HER4, have not been the subject of significant efforts. In this paper, we have taken the initial steps required to generate antibodies with potential clinically utility that target the members of the EGFR family. The genes for the extracellular domains (ECDs) of all four members of the EGFR family were cloned and used to stably transfect 293 (HEK) cells. Milligram quantities of each ECD were produced and characterized. The HER3, HER4, and EGFR ECDs were then employed as targets for the selection of antibodies from naïve human scFv (single-chain Fv) phage display libraries. Six unique scFv clones were isolated that bound specifically to HER3, 13 unique clones were isolated with specificity for HER4 and 52 unique anti-EGFR clones were isolated. These scFvs provide a valuable and potentially clinically relevant panel of agents to target the members of the EGFR family.

Development of Engineered Antibodies Specific for the Müllerian Inhibiting Substance Type II Receptor: a Promising Candidate for Targeted Therapy of Ovarian Cancer

Molecular Cancer Therapeutics. Aug, 2006  |  Pubmed ID: 16928831

The Müllerian inhibiting substance type II receptor (MISIIR) is involved in Müllerian duct regression as part of the development of the male reproductive system. In adult females, MISIIR is present on ovarian surface epithelium and is frequently expressed on human epithelial ovarian cancer cells. Müllerian inhibiting substance has been found to be capable of inhibiting the growth of primary human ovarian cancer cells derived from ascites and ovarian cancer cell lines. This suggested to us that MISIIR could be an attractive target for antibody-based tumor targeting and growth inhibition strategies. Here, we describe the production of recombinant human MISIIR extracellular domain-human immunoglobulin Fc domain fusion proteins and their use as targets for the selection of MISIIR-specific human single-chain variable fragments (scFv) molecules from a human nonimmune scFv phage display library. The binding kinetics of the resulting anti-MISIIR scFv clones were characterized and two were employed as the basis for the construction of bivalent scFv:Fc antibody-based molecules. Both bound specifically to human ovarian carcinoma cells in flow cytometry assays and cross-reacted with mouse MISIIR. These results indicate that antibody-based constructs may provide a highly specific means of targeting MISIIR on human ovarian carcinoma cells for the purpose of diagnosing and treating this disease.

23-carboxy-24,25,26,27-tetranorvitamin D3 (calcioic Acid) and 24-carboxy-25,26,27-trinorvitamin D3 (cholacalcioic Acid): End Products of 25-hydroxyvitamin D3 Metabolism in Rat Kidney Through C-24 Oxidation Pathway

Archives of Biochemistry and Biophysics. Nov, 2006  |  Pubmed ID: 17027908

During the past two and half decades the elucidation of the metabolic pathways of 25OHD(3) and its active metabolite 1alpha,25(OH)(2)D(3) progressed in parallel. In spite of many advances in this area of vitamin D research, the unequivocal identification of the end products of 25OHD(3) metabolism through C-24 oxidation pathway has not been achieved. It is now well established that both 25OHD(3) and 1alpha,25(OH)(2)D(3) are metabolized through the same C-24 oxidation pathway initiated by the enzyme 24-hydroxylase (CYP24A1). Based on the information that the end product of 1alpha,25(OH)(2)D(3) metabolism through C-24 oxidation pathway is 1alpha-OH-23- COOH-24,25,26,27-tetranor D(3) or calcitroic acid; the metabolism of 25OHD(3) into 23-COOH-24,25,26,27-tetranor D(3) has been assumed. Furthermore, a previous study indicated 24-COOH-25,26,27-trinor D(3) as a water soluble metabolite of 24R,25(OH)(2)D(3) produced in rat kidney homogenates. Therefore, 24-COOH-25,26,27-trinor D(3) was also assumed as another end product of 25OHD(3) metabolism through C-24 oxidation pathway. We embarked on our present study to provide unequivocal proof for these assumptions. We first studied the metabolism of 25OHD(3) at low substrate concentration (3x10(-10)M) using [1,2-(3)H]25OHD(3) as the substrate in the perfused rat kidneys isolated from both normal and vitamin D(3) intoxicated rats. A highly polar water soluble metabolite, labeled as metabolite X was isolated from the kidney perfusate. The amount of metabolite X produced in the kidney of a vitamin D intoxicated rat was about seven times higher than that produced in the kidney of a normal rat. We then produced metabolite X in a quantity sufficient for its structure identification by perfusing kidneys isolated from vitamin D intoxicated rats with high substrate concentration of 25OHD(3) (5x10(-6)M). Using the techniques of electron impact and thermospray mass spectrometry, we established that the metabolite X contained both 23-COOH-24,25,26,27-tetranor D(3) and 24-COOH-25,26,27-trinor D(3) in a ratio of 4:1. The same metabolite X containing both acids in the same ratio of 4:1 was also produced when 24R,25(OH)(2)D(3) was used as the starting substrate. Previously, the trivial name of cholacalcioic acid was assigned to 24-COOH-25,26,27-trinorvitamin D(3). Using the same guidelines, we now assign the trivial name of calcioic acid to 23-COOH-24,25,26,27-tetranor D(3). In summary, for the first time our study provides unequivocal evidence to indicate that both calcioic and cholacalcioic acids as the end products of 25OHD(3) metabolism in rat kidney through C-24 oxidation pathway.

Myeloid C-type Lectins in Innate Immunity

Nature Immunology. Dec, 2006  |  Pubmed ID: 17110942

C-type lectins expressed on myeloid cells comprise a family of proteins that share a common structural motif, and some act as receptors in pathogen recognition. But just as the presence of leucine-rich repeats alone is not sufficient to define a Toll-like receptor, the characterization of C-type lectin receptors in innate immunity requires the identification of accompanying signaling motifs. Here we focus on the known signaling pathways of myeloid C-type lectins and on their possible functions as autonomous activating or inhibitory receptors involved in innate responses to pathogens or self.

Live Fast, Die Young: Trade-offs Between Fitness Components and Sexually Antagonistic Selection on Weaponry in Soay Sheep

Evolution; International Journal of Organic Evolution. Oct, 2006  |  Pubmed ID: 17133873

Males are predicted to compete for reproductive opportunities, with sexual selection driving the evolution of large body size and weaponry through the advantage they confer for access to females. Few studies have explored potential trade-offs of investment in secondary sexual traits between different components of fitness or tested for sexually antagonistic selection pressures. These factors may provide explanations for observed polymorphisms in both form and quality of secondary sexual traits. We report here an analysis of selection on horn phenotype in a feral population of Soay sheep (Ovis aries) on the island of Hirta, St. Kilda, Scotland. Soay sheep display a phenotypic polymorphism for horn type with males growing either normal or reduced (scurred) horns, and females growing either normal, scurred, or no (polled) horns; further variation in size exists within horn morphs. We show that horn phenotype and the size of the trait displayed is subject to different selection pressures in males and females, generating sexually antagonistic selection. Furthermore, there was evidence of a trade-off between breeding success and longevity in normal-horned males, with both the normal horn type and larger horn size being associated with greater annual breeding success but reduced longevity. Therefore, selection through lifetime breeding success was not found to act upon horn phenotype in males. In females, a negative association of annual breeding success within the normal-horned phenotype did not result in a significant difference in lifetime fitness when compared to scurred individuals, as no significant difference in longevity was found. However, increased horn size within this group was negatively associated with breeding success and longevity. Females without horns (polled) suffered reduced longevity and thus reduced lifetime breeding success relative the other horn morphs. Our results therefore suggest that trade-offs between different components of fitness and antagonistic selection between the sexes may maintain genetic variation for secondary sexual traits within a population.

Method Modification (2004.08) to Field Testing of Visible Powders on a Variety of Nonporous Environmental Surfaces: Field Study

Journal of AOAC International. Nov-Dec, 2006  |  Pubmed ID: 17225611

The RAMP Anthrax Test Cartridge for detecting Bacillus anthracis was validated for use in the field for detection of B. anthracis spores in visible powder residues on 7 nonporous environmental surfaces. Six teams of trained first responders and civil support personnel in Class C personal protective equipment sampled visible powder residues on plastic, stainless steel, ceramic tile, wood, rubber, sealed concrete, and food-grade painted wood and analyzed the samples on the RAMP Anthrax Test System. The accuracy for each surface was at least 97% and the overall average was 98.8%. The overall average false-positive rate was 1.79% and false-negative rate was 1.07% for all surfaces. There were no significant differences between surfaces or between spore levels.

Standard Practice for Bulk Sample Collection and Swab Sample Collection of Visible Powders Suspected of Being Biological Agents from Nonporous Surfaces: Collaborative Study

Journal of AOAC International. Jan-Feb, 2007  |  Pubmed ID: 17373464

The draft ASTM Standard, "Standard Practice for Bulk Sample Collection and Swab Sample Collection of Visible Powders Suspected of Being Biological Agents from Nonporous Surfaces," was validated in a collaborative study consisting of 6 teams comprised of Civil Support personnel and First Responders, 2 levels of Bacillus anthracis Sterne and Bacillus thuringiensis Kurstaki spores, and 7 nonporous surfaces. The sample collection standard includes collection of the bulk sample (Method A) using a dry swab to push the sample onto a collection card and collection of residual sample (Method B) using an onsite test kit followed by a wet swab intended for additional onsite testing. Method A is to be performed prior to Method B in order to preserve unadulterated sample as potential criminal evidence. While statistical differences were observed between surfaces, between teams, and the interaction of surfaces and teams for the various sample types collected, these differences are due to the very low variability of the data and a much more narrow distribution than an ideal normal distribution, rather than to any practical differences. The data demonstrate that from both the 1.0 and 0.01 g powder samples, high levels of spores (mean >10(6) CFU) are recovered from the 7 surfaces by both the dry swab used in bulk sample collection (Method A) and the wet swab (Method B) sampling of the residual powder after bulk sample collection. Thus, after bulk sample collection, there is a high level of residual spores remaining for onsite biological testing and both Methods A and B should be performed in the field.

Syk- and CARD9-dependent Coupling of Innate Immunity to the Induction of T Helper Cells That Produce Interleukin 17

Nature Immunology. Jun, 2007  |  Pubmed ID: 17450144

The C-type lectin dectin-1 binds to yeast and signals through the kinase Syk and the adaptor CARD9 to induce production of interleukin 10 (IL-10) and IL-2 in dendritic cells (DCs). However, whether this pathway promotes full DC activation remains unclear. Here we show that dectin-1-Syk-CARD9 signaling induced DC maturation and the secretion of proinflammatory cytokines, including IL-6, tumor necrosis factor and IL-23, but little IL-12. Dectin-1-activated DCs 'instructed' the differentiation of CD4+ IL-17-producing effector T cells (T(H)-17 cells) in vitro, and a dectin-1 agonist acted as an adjuvant promoting the differentiation of T(H)-17 and T helper type 1 cells in vivo. Infection with Candida albicans induced CARD9-dependent T(H)-17 responses to the organism. Our data indicate that signaling through Syk and CARD9 can couple innate to adaptive immunity independently of Toll-like receptor signals and that CARD9 is required for the development of T(H)-17 responses to some pathogens.

Syk-dependent ERK Activation Regulates IL-2 and IL-10 Production by DC Stimulated with Zymosan

European Journal of Immunology. Jun, 2007  |  Pubmed ID: 17458858

Zymosan is a particulate yeast preparation that elicits high levels of IL-2 and IL-10 from dendritic cells (DC) and engages multiple innate receptors, including the Syk-coupled receptor dectin-1 and the MyD88-coupled receptor TLR2. Here, we show that induction of IL-2 and IL-10 by zymosan requires activation of ERK MAP kinase in murine DC. Surprisingly, ERK activation in response to zymosan is completely blocked in Syk-deficient DC and unaffected by MyD88 deficiency. Conversely, ERK activation in response to the TLR2 agonist Pam3Cys is completely MyD88 dependent and unaffected by Syk deficiency. The inability of TLR2 ligands in zymosan to couple to ERK may explain the Syk dependence of the IL-2 and IL-10 response in DC and emphasises the importance of Syk-coupled pattern recognition receptors such as dectin-1 in the detection of yeasts. Furthermore, the lack of receptor compensation observed here suggests that responses induced by complex innate stimuli cannot always be predicted by the signalling pathways downstream of individual receptors.

Removal of C-ring from the CD-ring Skeleton of 1alpha,25-dihydroxyvitamin D3 Does Not Alter Its Target Tissue Metabolism Significantly

Archives of Biochemistry and Biophysics. Apr, 2007  |  Pubmed ID: 17196157

It is now well established that 1alpha,25(OH)2D3 is metabolized in its target tissues through the modifications of both side chain and A-ring. The C-24 oxidation pathway is the side chain modification pathway through which 1alpha,25(OH)2D3 is metabolized into calcitroic acid. The C-3 epimerization pathway is the A-ring modification pathway through which 1alpha,25(OH)2D3 is metabolized into 1alpha,25(OH)2-3-epi-D3. During the past two decades, a great number of vitamin D analogs were synthesized by altering the structure of both side chain and A-ring of 1alpha,25(OH)2D3 with the aim to generate novel vitamin D compounds that inhibit proliferation and induce differentiation of various types of normal and cancer cells without causing significant hypercalcemia. Previously, we used some of these analogs as molecular probes to examine how changes in 1alpha,25(OH)2D3 structure would affect its target tissue metabolism. Recently, several nonsteroidal analogs of 1alpha,25(OH)2D3 with unique biological activity profiles were synthesized. Two of the analogs, SL 117 and WU 515 lack the C-ring of the CD-ring skeleton of 1alpha,25(OH)2D3. SL 117 contains the same side chain as that of 1alpha,25(OH)2D3, while WU 515 contains an altered side chain with a 23-yne modification combined with hexafluorination at C-26 and C-27. Presently, it is unknown how the removal of C-ring from the CD-ring skeleton of 1alpha,25(OH)2D3 would affect its target tissue metabolism. In the present study, we compared the metabolic fate of SL 117 and WU 515 with that of 1alpha,25(OH)2D3 in both the isolated perfused rat kidney, which expresses only the C-24 oxidation pathway and rat osteosarcoma cells (UMR 106), which express both the C-24 oxidation and C-3 epimerization pathways. The results of our present study indicate that SL 117 is metabolized like 1alpha,25(OH)2D3, into polar metabolites via the C-24 oxidation pathway in both rat kidney and UMR 106 cells. As expected, WU 515 with altered side chain structure is not metabolized via the C-24 oxidation pathway. Unlike in rat kidney, both SL 117 and WU 515 are also metabolized into less polar metabolites in UMR 106 cells. These metabolites displayed GC and MS characteristics consistent with A-ring epimerization and were putatively assigned as C-3 epimers of SL 117 and WU 515. In summary, we report that removal of the C-ring from the CD-ring skeleton of 1alpha,25(OH)2D3 does not alter its target tissue metabolism significantly.

Regulation of Antibody-dependent Cellular Cytotoxicity by IgG Intrinsic and Apparent Affinity for Target Antigen

Journal of Immunology (Baltimore, Md. : 1950). Sep, 2007  |  Pubmed ID: 17709495

Unconjugated mAbs have emerged as useful cancer therapeutics. Ab-dependent cellular cytotoxicity (ADCC) is believed to be a major antitumor mechanism of some anticancer Abs. However, the factors that regulate the magnitude of ADCC are incompletely understood. In this study, we described the relationship between Ab affinity and ADCC. A series of human IgG1 isotype Abs was created from the anti-HER2/neu (also named c-erbB2) C6.5 single-chain Fv (scFv) and its affinity mutants. The scFv affinities range from 10(-7) to 10(-11) M, and the IgG Abs retain the affinities of the scFv from which they were derived. The apparent affinity of the Abs ranged from nearly 10(-10) M (the lowest affinity variant) to almost 10(-11) M (the other variants). The IgG molecules were tested for their ability to elicit ADCC in vitro against three tumor cell lines with differing levels of HER2/neu expression using unactivated human PBMC from healthy donors as the effector cells. The results demonstrated that both the apparent affinity and intrinsic affinity of the Abs studied regulate ADCC. High-affinity tumor Ag binding by the IgGs led to the most efficient and powerful ADCC. Tumor cells expressing high levels of HER2/neu are more susceptible to the ADCC triggered by Abs than the cells expressing lower amounts of HER2/neu. These findings justify the examination of high affinity Abs for ADCC promotion. Because high affinity may impair in vivo tumor targeting, a careful examination of Ab structure to function relationships is required to develop optimized therapeutic unconjugated Abs.

Function of Weaponry in Females: the Use of Horns in Intrasexual Competition for Resources in Female Soay Sheep

Biology Letters. Dec, 2007  |  Pubmed ID: 17711817

In many species, females show reduced expression of a trait that is under sexual selection in males, and this expression is thought to be maintained through genetic associations with the male phenotype. However, there is also the potential for the female trait to convey an advantage in intrasexual conflicts over resources. We tested this hypothesis in a feral population of Soay sheep, in which males and females have a polymorphism for horn development, producing either full (normal horned), reduced (scurred) or no (polled, females only) horns. During the lambing period, females who possessed horns were more likely to initiate and win aggressive interactions, independent of age, weight and birthing status. The occurrence of aggression was also context dependent, decreasing over the lambing period and associated with local density. Our results demonstrate that a trait that confers benefits to males during intrasexual competition for mates may also be used by females in intrasexual competition over resources: males use weaponry to gain mates, whereas females use weaponry to gain food.

Effective Treatment of Established Human Breast Tumor Xenografts in Immunodeficient Mice with a Single Dose of the Alpha-emitting Radioisotope Astatine-211 Conjugated to Anti-HER2/neu Diabodies

Clinical Cancer Research : an Official Journal of the American Association for Cancer Research. Feb, 2008  |  Pubmed ID: 18245551

Successful radioimmunotherapy strategies depend on selecting radioisotopes with physical properties complementary to the biological properties of the targeting vehicle. Small, engineered antitumor antibody fragments are capable of rapid, highly specific tumor targeting in immunodeficient mouse models. We hypothesized that the C6.5 diabody, a noncovalent anti-HER2 single-chain Fv dimer, would be an ideal radioisotope carrier for the radioimmunotherapy of established tumors using the short-lived alpha-emitting radioisotope (211)At.

Spectacular Impalement Through the Face and Neck: a Case Report and Literature Review

The Journal of Trauma. Dec, 2008  |  Pubmed ID: 18277292

Selective Inhibition of Cyclooxygenase-2 (COX-2) by 1alpha,25-dihydroxy-16-ene-23-yne-vitamin D3, a Less Calcemic Vitamin D Analog

Journal of Cellular Biochemistry. Aug, 2008  |  Pubmed ID: 18348265

Inducible cyclooxygenase-2 (COX-2) has been implicated to play a role in inflammation and carcinogenesis and selective COX-2 inhibitors have been considered as anti-inflammatory and cancer chemopreventive agents. 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3), the active hormonal form of vitamin D3 also has been considered to be a cancer chemopreventive agent in addition to its important role in maintaining calcium homeostasis. Based on these observations, we studied the direct effect of 1alpha,25(OH)2D3 and one of its less calcemic synthetic analogs, 1alpha,25(OH)2-16-ene-23-yne-D3 on the activity of both COX-1 and COX-2 in an in vitro enzyme assay. Preliminary data indicated that both 1alpha,25(OH)2D3 and 1alpha,25(OH)2-16-ene-23-yne-D3 inhibited selectively the activity of COX-2 with no effect on the activity of COX-1. Out of the two compounds, 1alpha,25(OH)2-16-ene-23-yne-D3 was found to be more effective with an IC50 of 5.8 nM. Therefore, the rest of the experiments were performed using 1alpha,25(OH)2-16-ene-23-yne-D3 only. 1alpha,25(OH)2-16-ene-23-yne-D3 inhibited the proliferation of lipopolysaccharide (LPS) stimulated mouse macrophage cells (RAW 264.7) with a reduction in the expression of COX-2 along with other inflammatory mediators like inducible nitric oxide synthase (iNOS) and interleukin-2 (IL-2). Furthermore, 1alpha,25(OH)2-16-ene-23-yne-D3 also inhibited carrageenan induced inflammation in an air pouch of a rat and effectively reduced the expression of COX-2, iNOS, and IL-2 in the tissues of the same air pouch. In both cases, 1alpha,25(OH)2-16-ene-23-yne-D3 did not show any effect on the expression of COX-1. In summary, our results indicate that 1alpha,25(OH)2-16-ene-23-yne-D3, a less calcemic vitamin D analog, exhibits potent anti-inflammatory effects and is a selective COX-2 inhibitor.

Environmental Heterogeneity Generates Fluctuating Selection on a Secondary Sexual Trait

Current Biology : CB. May, 2008  |  Pubmed ID: 18485708

In any population in which resources are limiting, the allocation of resources toward increased reproductive success may generate costs to survival [1-8]. The relationship between a sexually selected trait and fitness will therefore represent a balance between its relative associations with fecundity versus viability [3, 6, 7]. Because the risk of mortality in a population is likely to be heavily determined by ecological conditions, survival costs may vary as a function of the prevailing environment [7]. As a result, for populations experiencing heterogeneous ecological conditions, there may not be a single optimal level of allocation toward reproduction versus survival [9]. Here, we show that early viability and fecundity selection act in opposing directions on a secondary sexual trait and that their relative magnitude depends upon ecological conditions, generating fluctuating selection. In a wild population of Soay sheep (Ovis aries), phenotypic and genetic associations between male horn growth and lifetime reproductive success were positive under good environmental conditions (because of increased breeding success) and negative under poor environmental conditions (because of reduced survival). In an unpredictable environment, high allocation to early horn growth is a gamble that will only pay off if ensuing conditions are favorable. Such fluctuating selection may play an important role in preventing the erosion of genetic variance in secondary sexual traits.

Antibody Engineering Principles and Applications

Cancer Journal (Sudbury, Mass.). May-Jun, 2008  |  Pubmed ID: 18536554

Antibodies have emerged as significant agents for the treatment of a number of diseases including cancer and autoimmunity. However, most of the antibodies currently used in clinical practice were developed from humanized or chimeric molecules based on mouse monoclonal antibodies. Recent advances in antibody selection and engineering techniques have led to the development of antibodies specific for highly conserved targets, the creation of novel antibody-based structures, significant improvements in affinity for target antigens, enhanced ability to engage immune effector functions, and the creation of fusion proteins with direct cytotoxic properties. This review provides an overview of the techniques that we expect will have the greatest impact on the field of antibody engineering.

Diabetic Ketoacidosis in Toddler with a Diaper Rash

The American Journal of Emergency Medicine. Sep, 2008  |  Pubmed ID: 18774050

The term diabetes does not denote a single disease entity but rather a clinical syndrome. Fundamental to all types of diabetes is impairment of insulin secretion by the pancreatic beta cells. Diabetes is divided into (1) diabetes associated with certain syndromes or conditions, (2) gestational diabetes, (3) non-insulin-dependent diabetes or type 2 diabetes, and (4) insulin-dependent diabetes (IDDM) or type 1 diabetes. The impairment of insulin secretion seen in diabetes is due to progressive loss of pancreatic beta-cell function secondary to an autoimmune-mediated process. Diabetes mellitus is the most common metabolic disorder of childhood (2). We present a patient with a common finding in children, diaper candidiasis. Surprisingly, our patient was found to have IDDM and be in diabetic ketoacidosis.

Costs and Clinical Outcomes After Coronary Multidetector CT Angiography in Patients Without Known Coronary Artery Disease: Comparison to Myocardial Perfusion SPECT

Radiology. Oct, 2008  |  Pubmed ID: 18796668

To assess costs and clinical outcomes in individuals without known coronary artery disease (CAD) who underwent multidetector computed tomographic (CT) angiography compared with those in matched patients who underwent myocardial perfusion single photon emission computed tomography (SPECT).

Factors Influencing the Natural History of HIV-1 Infection

Chinese Medical Journal. Dec, 2008  |  Pubmed ID: 19187605

Differences in Episode-based Care Costs for Multidetector Computed Tomographic Coronary Angiography Versus Myocardial Perfusion Imaging for the Diagnosis of Coronary Artery Disease

Journal of Medical Economics. 2008  |  Pubmed ID: 19450089

Multidetector computed tomography (MDCT) is a novel method for diagnosis and prognosis of coronary artery disease (CAD). The opportunity costs that favour MDCT over other CAD diagnostic methods is currently unknown.

Isolation of Anti-MISIIR ScFv Molecules from a Phage Display Library by Cell Sorter Biopanning

Cancer Immunology, Immunotherapy : CII. Mar, 2008  |  Pubmed ID: 17676323

While cell surface antigens represent the most common targets for antibody-based cancer therapy, isolation of new antibodies specific for these targets from single-chain Fv phage display libraries has been hindered by limitations associated with traditional selection techniques. Solid phase panning is often associated with conformational changes to the target protein due to its immobilization on plastic tubes that can limit the ability of the isolated scFv to bind to conformational epitopes and solution panning methods require the use of secondary tags that often mask desired sequences and create unintended epitopes. Commonly utilized cell-based panning methods typically yield a panel of single-chain Fv (scFv) molecules that are specific for numerous cell surface antigens, often obscuring the desired clones. Here, we describe a novel cell sorter-based system to isolate single-chain Fv molecules specific for defined antigen targets expressed on stably-transformed mammalian cells. We employed these methods to isolate promising scFv clones that bind specifically to the Müllerian inhibiting substance type II receptor, a cell surface ovarian cancer antigen that has proven to be a difficult target for selection strategies.

Deamidation of Alpha-synuclein

Protein Science : a Publication of the Protein Society. Aug, 2009  |  Pubmed ID: 19521992

The rates of deamidation of alpha-synuclein and single Asn residues in 13 Asn-sequence mutants have been measured for 5 x 10(-5)M protein in both the absence and presence of 10(-2)M sodium dodecyl sulfate (SDS). In the course of these experiments, 370 quantitative protein deamidation measurements were performed and 37 deamidation rates were determined by ion cyclotron resonance Fourier transform mass spectrometry, using an improved whole protein isotopic envelope method and a mass defect method with both enzymatic and collision-induced fragmentation. The measured deamidation index of alpha-synuclein was found to be 0.23 for an overall deamidation half-time of 23 days, without or with SDS micelles, owing primarily to the deamidation of Asn(103) and Asn(122). Deamidation rates of 15 Asn residues in the wild-type and mutant proteins were found to be primary sequence controlled without SDS. However, the presence of SDS micelles slowed the deamidation rates of nine N-terminal region Asn residues, caused by the known three-dimensional structures induced through protein binding to SDS micelles.

Impact of Improved Cookstoves on Indoor Air Pollution and Adverse Health Effects Among Honduran Women

International Journal of Environmental Health Research. Oct, 2009  |  Pubmed ID: 19626518

Elevated indoor air pollution levels due to the burning of biomass in developing countries are well established. Few studies have quantitatively assessed air pollution levels of improved cookstoves and examined these measures in relation to health effects. We conducted a cross-sectional survey among 79 Honduran women cooking with traditional or improved cookstoves. Carbon monoxide and fine particulate matter (PM(2.5)) levels were assessed via indoor and personal monitoring. Pulmonary function and respiratory symptoms were ascertained. Finger-stick blood spot samples were collected to measure C-reactive protein (CRP) concentrations. The use of improved stoves was associated with 63% lower levels of personal PM(2.5), 73% lower levels of indoor PM(2.5), and 87% lower levels of indoor carbon monoxide as compared to traditional stoves. Women using traditional stoves reported symptoms more frequently than those using improved stoves. There was no evidence of associations between cookstove type or air quality measures with lung function or CRP.

Dectin-2 is a Syk-coupled Pattern Recognition Receptor Crucial for Th17 Responses to Fungal Infection

The Journal of Experimental Medicine. Aug, 2009  |  Pubmed ID: 19703985

Innate immune cells detect pathogens via pattern recognition receptors (PRRs), which signal for initiation of immune responses to infection. Studies with Dectin-1, a PRR for fungi, have defined a novel innate signaling pathway involving Syk kinase and the adaptor CARD9, which is critical for inducing Th17 responses to fungal infection. We show that another C-type lectin, Dectin-2, also signals via Syk and CARD9, and contributes to dendritic cell (DC) activation by fungal particles. Unlike Dectin-1, Dectin-2 couples to Syk indirectly, through association with the FcRgamma chain. In a model of Candida albicans infection, blockade of Dectin-2 did not affect innate immune resistance but abrogated Candida-specific T cell production of IL-17 and, in combination with the absence of Dectin-1, decreased Th1 responses to the organism. Thus, Dectin-2 constitutes a major fungal PRR that can couple to the Syk-CARD9 innate signaling pathway to activate DCs and regulate adaptive immune responses to fungal infection.

Human Immunodeficiency Virus and Hepatitis C Virus Co-infection: Epidemiology, Natural History and the Situation in China

Chinese Medical Journal. Jan, 2009  |  Pubmed ID: 19187624

The Impact of Environmental Heterogeneity on Genetic Architecture in a Wild Population of Soay Sheep

Genetics. Apr, 2009  |  Pubmed ID: 19204380

This work demonstrates that environmental conditions experienced by individuals can shape their development and affect the stability of genetic associations. The implication of this observation is that the environmental response may influence the evolution of traits in the wild. Here, we examined how the genetic architecture of a suite of sexually dimorphic traits changed as a function of environmental conditions in an unmanaged population of Soay sheep (Ovis aries) on the island of Hirta, St. Kilda, northwest Scotland. We examined the stability of phenotypic, genetic, and environmental (residual) covariance in males during the first year of life between horn length, body weight, and parasite load in environments of different quality. We then examined the same covariance structures across environments within and between the adult sexes. We found significant genotype-by-environment interactions for lamb male body weight and parasite load, leading to a change in the genetic correlation among environments. Horn length was genetically correlated with body weight in males but not females and the genetic correlation among traits within and between the sexes was dependent upon the environmental conditions experienced during adulthood. Genetic correlations were smaller in more favorable environmental conditions, suggesting that in good environments, loci are expressed that have sex-specific effects. The reduction in genetic correlation between the sexes may allow independent evolutionary trajectories for each sex. This study demonstrates that the genetic architecture of traits is not stable under temporally varying environments and highlights the fact that evolutionary processes may depend largely upon ecological conditions.

Synthesis and Anti-inflammatory Properties of 1alpha,25-dihydroxy-16-ene-20-cyclopropyl-24-oxo-vitamin D3, a Hypocalcemic, Stable Metabolite of 1alpha,25-dihydroxy-16-ene-20-cyclopropyl-vitamin D3

Journal of Medicinal Chemistry. Apr, 2009  |  Pubmed ID: 19309155

1alpha,25(OH)(2)-16-ene-20-cyclopropyl-vitamin D(3) (13) is several fold more potent than the natural hormone 1alpha,25-dihydroxyvitamin D(3) (1) as an anti-inflammatory agent. Here, we have further analyzed the anti-inflammatory properties of 13, confirming it as the most potent analogue tested within this family. We then determined the structures of all the natural metabolites of 13, including the 24-oxo metabolite 14, and carried out its synthesis. A comparison of 13 with 14 showed a similar induction of the primary VDR target genes CYP24A1 and CAMP and comparable anti-inflammatory properties as revealed by a similar inhibition of TNF-alpha, IL-12/23p40, IL-6, and IFN-gamma production. Interestingly, 14 displays a 3-fold lower calcemic activity in vivo compared to 13. Collectively, these findings indicate that the strong potency of 13 can be explained by the accumulation of its stable 24-oxo metabolite, which shows immunoregulatory and anti-inflammatory properties superimposable to those exerted by 13 itself.

Investigation of Individuals Exposed to a Healthcare Worker with Cavitary Pulmonary Tuberculosis

Infection Control and Hospital Epidemiology : the Official Journal of the Society of Hospital Epidemiologists of America. May, 2009  |  Pubmed ID: 19344274

The Interactions of Some Commonly Consumed Drugs with Mitochondrial Adaptations to Exercise

Journal of Applied Physiology (Bethesda, Md. : 1985). Jul, 2009  |  Pubmed ID: 19423832

The importance of mitochondrial dysfunctions in the progression of chronic disease has been well established. Patients with chronic diseases are often prescribed a variety of medications, many of which have been shown to induce mitochondrial dysfunction. Exercise is a known stimulus for mitochondrial biogenesis and also recommended to patients as a lifestyle modification to supplement drug therapy. However, the potential interference of those drugs with mitochondrial adaptations to exercise has not been thoroughly investigated. This review provides a summary and discussion of known and potential interactions of commonly consumed drugs with exercise-induced mitochondrial adaptations.

Acute {beta}-adrenergic Stimulation Does Not Alter Mitochondrial Protein Synthesis or Markers of Mitochondrial Biogenesis in Adult Men

American Journal of Physiology. Regulatory, Integrative and Comparative Physiology. Jan, 2010  |  Pubmed ID: 19907002

Exercise-induced expression of peroxisome proliferator-activated receptor gamma coactivator-1alpha (PGC-1alpha) is dramatically inhibited in mice pretreated with a beta-adrenergic receptor (beta-AR) antagonist, suggesting that beta-ARs play an important role in the regulation of skeletal muscle PGC-1alpha expression, and potentially, mitochondrial biogenesis. Accordingly, we hypothesized that acute beta-AR stimulation would induce transcriptional pathways involved in skeletal muscle mitochondrial biogenesis in humans. Whole body protein turnover (WBPT), myofibrillar protein synthesis (MyPS), skeletal muscle mitochondrial protein synthesis (MiPS), and mitochondrial biogenic signaling were determined in samples of vastus lateralis obtained on two separate occasions in 10 young adult males following 1 h of continuous intravenous administration of saline (CON) or a nonspecific beta-AR agonist [isoproterenol (ISO): 12 ng.kg fat free mass(-1).min(-1)], combined with coinfusion of [1,2](13)C-leucine. beta-AR stimulation induced appreciable increases in heart rate and systolic blood pressure (both P < 0.001) but did not affect mitochondrial biogenic signaling (no change in PGC-1alpha, TFAM, NRF-1, NRF-2, COX, or NADHox expression via RT-PCR; P > 0.05). Additionally, MiPS [CON: 0.099 +/- 0.028, ISO: 0.074 +/- 0.046 (mean +/- SD); P > 0.05] and MyPS (CON: 0.059 +/- 0.008, ISO: 0.055 +/- 0.009; P > 0.05), as well as measures of WBPT were unaffected. On the basis of this investigation, we conclude that acute intravenous beta-AR stimulation does not increase mitochondrial protein synthesis or biogenesis signals in skeletal muscle.

Predictors of Virologic Failure in HIV-1-infected Adults Receiving First-line Antiretroviral Therapy in 8 Provinces in China

Clinical Infectious Diseases : an Official Publication of the Infectious Diseases Society of America. Jan, 2010  |  Pubmed ID: 20017637

Despite poor primary health care systems, free antiretroviral therapy (ART) has been available in China for >5 years. Virologic outcomes in Chinese patients receiving ART have not been described on a national level.

Hepatitis B and Hepatitis C Seroprevalence in Children Receiving Antiretroviral Therapy for Human Immunodeficiency Virus-1 Infection in China, 2005-2009

Journal of Acquired Immune Deficiency Syndromes (1999). Jun, 2010  |  Pubmed ID: 20032784

Coinfection of hepatitis B virus (HBV) or hepatitis C virus (HCV) may compromise pediatric antiretroviral therapy (ART) in China. In this study, we evaluated the seroprevalence of HBV and HCV in children receiving ART and associated factors.

Naptumomab Estafenatox: a New Immunoconjugate

Expert Opinion on Biological Therapy. Feb, 2010  |  Pubmed ID: 20053143

New agents that specifically engage the immune system are being tested in a variety of malignancies. This review provides an overview of naptumomab, an immunotoxin, with encouraging clinical activity in Phase I trials.

CBL is Frequently Altered in Lung Cancers: Its Relationship to Mutations in MET and EGFR Tyrosine Kinases

PloS One. 2010  |  Pubmed ID: 20126411

Non-small cell lung cancer (NSCLC) is a heterogeneous group of disorders with a number of genetic and proteomic alterations. c-CBL is an E3 ubiquitin ligase and adaptor molecule important in normal homeostasis and cancer. We determined the genetic variations of c-CBL, relationship to receptor tyrosine kinases (EGFR and MET), and functionality in NSCLC.

Blastomycosis of the Central Nervous System: a Multicenter Review of Diagnosis and Treatment in the Modern Era

Clinical Infectious Diseases : an Official Publication of the Infectious Diseases Society of America. Mar, 2010  |  Pubmed ID: 20166817

Central nervous system (CNS) involvement with Blastomyces dermatitidis is an uncommon and potentially fatal complication of blastomycosis.

Immuno-positron Emission Tomography in Cancer Models

Seminars in Nuclear Medicine. May, 2010  |  Pubmed ID: 20350627

Positron emission tomography (PET) is playing an increasingly important role in the diagnosis, staging, and monitoring response to treatment in a variety of cancers. Recent efforts have focused on immuno-PET, which uses antibody-based radiotracers, to image tumors based on expression of tumor-associated antigens. It is postulated that the specificity afforded by antibody targeting should both improve tumor detection and provide phenotypic information related to primary and metastatic lesions that will guide therapy decisions. Advances in antibody-engineering are providing the tools to develop antibody-based molecules with pharmacokinetic properties optimized for use as immuno-PET radiotracers. Coupled with technical advances in the design of PET scanners, immuno-PET holds promise to improve diagnostic imaging and to guide the use of targeted therapies. An overview of the preclinical immuno-PET studies in cancer models is reviewed here.

Hoxb8 Conditionally Immortalised Macrophage Lines Model Inflammatory Monocytic Cells with Important Similarity to Dendritic Cells

European Journal of Immunology. Dec, 2010  |  Pubmed ID: 21225953

We have examined the potential to generate bona fide macrophages (MØ) from conditionally immortalised murine bone marrow precursors. MØ can be derived from Hoxb8 conditionally immortalised macrophage precursor cell lines (MØP) using either M-CSF or GM-CSF. When differentiated in GM-CSF (GM-MØP) the resultant cells resemble GM-CSF bone marrow-derived dendritic cells (BMDC) in morphological phenotype, antigen phenotype and functional responses to microbial stimuli. In spite of this high similarity between the two cell types and the ability of GM-MØP to effectively present antigen to a T-cell hybridoma, these cells are comparatively poor at priming the expansion of IFN-γ responses from naïve CD4(+) T cells. The generation of MØP from transgenic or genetically aberrant mice provides an excellent opportunity to study the inflammatory role of GM-MØP, and reduces the need for mouse colonies in many studies. Hence differentiation of conditionally immortalised MØPs in GM-CSF represents a unique in vitro model of inflammatory monocyte-like cells, with important differences from bone marrow-derived dendritic cells, which will facilitate functional studies relating to the many 'sub-phenotypes' of inflammatory monocytes.

Hoxb8 Conditionally Immortalised Macrophage Lines Model Inflammatory Monocytic Cells with Important Similarity to Dendritic Cells

European Journal of Immunology. Feb, 2011  |  Pubmed ID: 21268006

We have examined the potential to generate bona fide macrophages (MØ) from conditionally immortalised murine bone marrow precursors. MØ can be derived from Hoxb8 conditionally immortalised macrophage precursor cell lines (MØP) using either M-CSF or GM-CSF. When differentiated in GM-CSF (GM-MØP) the resultant cells resemble GM-CSF bone marrow-derived dendritic cells (BMDC) in morphological phenotype, antigen phenotype and functional responses to microbial stimuli. In spite of this high similarity between the two cell types and the ability of GM-MØP to effectively present antigen to a T-cell hybridoma, these cells are comparatively poor at priming the expansion of IFN-γ responses from naïve CD4(+) T cells. The generation of MØP from transgenic or genetically aberrant mice provides an excellent opportunity to study the inflammatory role of GM-MØP, and reduces the need for mouse colonies in many studies. Hence differentiation of conditionally immortalised MØPs in GM-CSF represents a unique in vitro model of inflammatory monocyte-like cells, with important differences from bone marrow-derived dendritic cells, which will facilitate functional studies relating to the many 'sub-phenotypes' of inflammatory monocytes.

Highly Sensitive Detection of HER2 Extracellular Domain in the Serum of Breast Cancer Patients by Piezoelectric Microcantilevers

Analytical Chemistry. May, 2011  |  Pubmed ID: 21449604

Rapid and sensitive detection of serum tumor biomarkers are needed to monitor cancer patients for disease progression. Highly sensitive piezoelectric microcantilever sensors (PEMS) offer an attractive tool for biomarker detection; however, their utility in the complex environment encountered in serum has yet to be determined. As a proof of concept, we have functionalized PEMS with antibodies that specifically bind to HER2, a biomarker (antigen) that is commonly overexpressed in the blood of breast cancer patients. The function and sensitivity of these anti-HER2 PEMS biosensors was initially assessed using recombinant HER2 spiked into human serum. Their ability to detect native HER2 present in the serum of breast cancer patients was then determined. We have found that the anti-HER2 PEMS were able to accurately detect both recombinant and naturally occurring HER2 at clinically relevant levels (>2 ng/mL). This indicates that PEMS-based biosensors provide a potentially effective tool for biomarker detection.

Improved Detection of Canine Angiostrongylus Vasorum Infection Using Real-time PCR and Indirect ELISA

Parasitology Research. Dec, 2011  |  Pubmed ID: 21537986

This study reports the development of a real-time PCR assay and an indirect ELISA to improve on current detection of canine Angiostrongylus vasorum infection. A highly specific fluorescent probe-based, real-time PCR assay was developed to target the A. vasorum second internal transcribed spacer region and detected DNA in EDTA blood, lung tissue, broncho-alveolar larvage fluid, endotracheal mucus, pharyngeal swabs and faecal samples. PCR was fast (∼1 h), highly efficient when using EDTA blood samples, consistently detected a single molecule of parasite DNA and did not amplify DNA from other parasitic nematodes or definitive host species. An indirect ELISA was also developed using the soluble protein fraction from adult A. vasorum worms. Some cross-reactive antigen recognition was observed when tested against sera from dogs infected with Crenosoma vulpis (n = 8), Toxocara canis (n = 5) and Dirofilaria immitis (n = 5). This was largely overcome by setting the cut-off for a positive result at an appropriately high level. Field evaluation of the real-time PCR and ELISA was conducted by testing sera and EDTA blood from dogs with suspected A. vasorum infection (n = 148) and compared with the Baermann's larval migration test in faeces. Thirty-one dogs were positive by at least one test. Of these, 20 (65%) were detected by the Baermann method, 18 (58%) by blood PCR, 24 (77%) by ELISA and 28 (90%) by blood PCR and ELISA together. Combined testing using real-time PCR and ELISA therefore improved the detection rate of A. vasorum infection and holds promise for improved clinical diagnosis and epidemiological investigation.

Proteomic Characterization of Non-small Cell Lung Cancer in a Comprehensive Translational Thoracic Oncology Database

Journal of Clinical Bioinformatics. Feb, 2011  |  Pubmed ID: 21603121

In recent years, there has been tremendous growth and interest in translational research, particularly in cancer biology. This area of study clearly establishes the connection between laboratory experimentation and practical human application. Though it is common for laboratory and clinical data regarding patient specimens to be maintained separately, the storage of such heterogeneous data in one database offers many benefits as it may facilitate more rapid accession of data and provide researchers access to greater numbers of tissue samples.

Long-term Synthesis Rates of Skeletal Muscle DNA and Protein Are Higher During Aerobic Training in Older Humans Than in Sedentary Young Subjects but Are Not Altered by Protein Supplementation

FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology. Sep, 2011  |  Pubmed ID: 21613572

Consuming protein following exercise has been shown to stimulate protein synthesis acutely in skeletal muscle and has been recommended to prevent sarcopenia. It is not known, however, whether acute stimulation persists long term or includes muscle cell division. We asked here whether consuming protein following exercise during aerobic training increases long-term protein and DNA synthesis rates in skeletal muscle of adult humans. Sixteen previously untrained participants (50 ± 8 yr) consumed either a carbohydrate or carbohydrate and protein drink following each session during 6 wk of treadmill training. A younger untrained group provided a nonexercising comparison. Participants were administered heavy water (²H₂O; deuterium oxide) continuously for 6 wk to isotopically label newly synthesized skeletal muscle proteins and DNA. Muscle biopsies were performed after 6 wk of training. Contrary to acute studies, consuming protein after exercise did not increase skeletal muscle protein synthesis rates. In contrast, muscle protein synthesis, DNA, and phospholipid synthesis were significantly higher in the older exercise groups than the younger sedentary group. The higher DNA replication rate could not be attributed to mitochondrial DNA and may be due to satellite cell activation. We conclude that postexercise protein supplementation does not increase rates of mixed protein synthesis over 6 wk and that aerobic exercise may stimulate long-term cell division (DNA synthesis) in skeletal muscle of humans. Measurements of long-term synthesis rates provide important insights into aging and exercise adaptations.

β-Adrenergic Receptor Blockade Blunts Postexercise Skeletal Muscle Mitochondrial Protein Synthesis Rates in Humans

American Journal of Physiology. Regulatory, Integrative and Comparative Physiology. Aug, 2011  |  Pubmed ID: 21613574

β-Adrenergic receptor (AR) signaling is a regulator of skeletal muscle protein synthesis and mitochondrial biogenesis in mice. We hypothesized that β-AR blockade blunts postexercise skeletal muscle mitochondrial protein synthesis rates in adult humans. Six healthy men (mean ± SD: 26 ± 6 yr old, 39.9 ± 4.9 ml·kg(-1)·min(-1) peak O(2) uptake, 26.7 ± 2.0 kg/m(2) body mass index) performed 1 h of stationary cycle ergometer exercise (60% peak O(2) uptake) during 1) β-AR blockade (intravenous propranolol) and 2) administration of saline (control). Skeletal muscle mitochondrial, myofibrillar, and sarcoplasmic protein synthesis rates were assessed using [(2)H(5)]phenylalanine incorporation into skeletal muscle proteins after exercise. The mRNA content of signals for mitochondrial biogenesis was determined using real-time PCR. β-AR blockade decreased mitochondrial (from 0.217 ± 0.076 to 0.135 ± 0.031%/h, P < 0.05), but not myofibrillar or sarcoplasmic, protein synthesis rates. Peroxisome proliferator-activated receptor-γ coactivator-1α mRNA was increased ∼2.5-fold (P < 0.05) at 5 h compared with 1 h postexercise but was not influenced by β-AR blockade. We conclude that decreased β-AR signaling during cycling can blunt the postexercise increase in mitochondrial protein synthesis rates without affecting mRNA content.

Acute Effects of an Avena Sativa Herb Extract on Responses to the Stroop Color-Word Test

Journal of Alternative and Complementary Medicine (New York, N.Y.). Jul, 2011  |  Pubmed ID: 21711204

Extracts from oat (Avena sativa) herb may benefit cognitive performance. This study investigated whether Neuravena(®), an oat herb extract, could acutely improve responses to the Stroop Color-Word test, a measure of attention and concentration and the ability to maintain task focus.

CLEC-2 Signaling Via Syk in Myeloid Cells Can Regulate Inflammatory Responses

European Journal of Immunology. Oct, 2011  |  Pubmed ID: 21728173

Myeloid cells express a plethora of C-type lectin receptors (CLRs) that can regulate immune responses. CLEC-2 belongs to the Dectin-1 sub-family of CLRs that possess an extracellular C-type lectin-like domain and a single intracellular hemITAM motif. CLEC-2 is highly expressed on mouse and human platelets where it signals via Syk to promote aggregation. We generated a monoclonal antibody (mAb) against mouse CLEC-2 and found that CLEC-2 is additionally widely expressed on leukocytes and that its expression is upregulated during inflammation. MAb-mediated crosslinking of CLEC-2 leads to hemITAM-dependent signaling via Syk, Ca(2+) and NFAT and, in myeloid cells, modulates the effect of toll-like receptor (TLR) agonists to selectively potentiate production of IL-10. A macrophage/dendritic cell-dependent increase in IL-10 is also observed in mice given anti-CLEC-2 mAb together with LPS. Collectively, these data indicate that CLEC-2 is expressed in myeloid cells and acts as a Syk-coupled CLR able to modulate TLR signaling and inflammatory responses.

Protein-intrinsic and Signaling Network-based Sources of Resistance to EGFR- and ErbB Family-targeted Therapies in Head and Neck Cancer

Drug Resistance Updates : Reviews and Commentaries in Antimicrobial and Anticancer Chemotherapy. Dec, 2011  |  Pubmed ID: 21920801

Agents targeting EGFR and related ErbB family proteins are valuable therapies for the treatment of many cancers. For some tumor types, including squamous cell carcinomas of the head and neck (SCCHN), antibodies targeting EGFR were the first protein-directed agents to show clinical benefit, and remain a standard component of clinical strategies for management of the disease. Nevertheless, many patients display either intrinsic or acquired resistance to these drugs; hence, major research goals are to better understand the underlying causes of resistance, and to develop new therapeutic strategies that boost the impact of EGFR/ErbB inhibitors. In this review, we first summarize current standard use of EGFR inhibitors in the context of SCCHN, and described new agents targeting EGFR currently moving through pre-clinical and clinical development. We then discuss how changes in other transmembrane receptors, including IGF1R, c-Met, and TGF-β, can confer resistance to EGFR-targeted inhibitors, and discuss new agents targeting these proteins. Moving downstream, we discuss critical EGFR-dependent effectors, including PLC-γ; PI3K and PTEN; SHC, GRB2, and RAS and the STAT proteins, as factors in resistance to EGFR-directed inhibitors and as alternative targets of therapeutic inhibition. We summarize alternative sources of resistance among cellular changes that target EGFR itself, through regulation of ligand availability, post-translational modification of EGFR, availability of EGFR partners for hetero-dimerization and control of EGFR intracellular trafficking for recycling versus degradation. Finally, we discuss new strategies to identify effective therapeutic combinations involving EGFR-targeted inhibitors, in the context of new system level data becoming available for analysis of individual tumors.

Local Entropy and Structure in a Two-dimensional Frustrated System

Chaos (Woodbury, N.Y.). Sep, 2011  |  Pubmed ID: 21974677

We calculate the local contributions to the Shannon entropy and excess entropy and use these information theoretic measures as quantitative probes of the order arising from quenched disorder in the diluted Ising antiferromagnet on a triangular lattice. When one sublattice is sufficiently diluted, the system undergoes a temperature-driven phase transition, with the other two sublattices developing magnetizations of equal magnitude and opposite sign as the system is cooled.(1) The diluted sublattice has no net magnetization but exhibits spin glass ordering. The distribution of local entropies shows a dramatic broadening at low temperatures; this indicates that the system's total entropy is not shared equally across the lattice. The entropy contributions from some regions exhibit local reentrance, although the entropy of the system decreases monotonically as expected. The average excess entropy shows a sharp peak at the critical temperature, showing that the excess entropy is sensitive to the structural changes that occur as a result of the spin glass ordering.

Age Effect on Myocellular Remodeling: Response to Exercise and Nutrition in Humans

Ageing Research Reviews. Nov, 2011  |  Pubmed ID: 22085885

Aging is associated with decline in muscle mass and muscle functions. Muscle strength declines disproportionate to the decline in muscle mass indicating that muscle quality or protein quality also declines with age. Human studies have shown a progressive decline in muscle protein synthesis including proteins in the contractile apparatus and mitochondria with age. However, the decline in muscle protein synthesis is disproportionate to the decline in muscle mass that occurs with age prompting to hypothesize that muscle protein degradation also declines with age. A decline in mitochondrial capacity to synthesize ATP is likely a limiting factor of both synthesis and degradation, which are ATP dependent processes. In support of the above hypothesis, several studies have shown a decline in whole body protein turnover (synthesis and degradation). The timely and efficient degradation of irreversibly damaged or modified proteins is critical to maintain the quality of protein. It is proposed that a failure to degrade the damaged proteins and replacing them with newly synthesized proteins contribute to age related decline in muscle mass and quality of muscle proteins. The underlying molecular mechanism of these age related changes in human muscle needs further investigation.

Efficient and Cost-effective Estimation of the Influence of Respiratory Variables on Respiratory Sinus Arrhythmia

Psychophysiology. Apr, 2011  |  Pubmed ID: 20718933

Researchers are interested in respiratory sinus arrhythmia (RSA) as an index of cardiac vagal activity. Yet, debate exists about how to account for respiratory influences on quantitative indices of RSA. T. Ritz, M. Thons, and B. Dahme (2001) developed a within-individual correction procedure by which the effects of respiration on RSA may be estimated using regression models. We replicated their procedure substituting a spectral high-frequency measure of RSA for a time-domain statistic and a respiratory belt's relative measure of tidal volume for the direct assessment provided by a pneumotachograph. The standardized slopes from the respiratory belt and pneumotachography-derived regression equations (estimated across a 6-min paced breathing protocol) were positively correlated (r=0.93, p<.001); correlations were similar across 2- and 4-min time courses parsed from the 6-min protocol. Our results offer methodological alternatives to the research community.

Evaluation of the Anti-HER2 C6.5 Diabody As a PET Radiotracer to Monitor HER2 Status and Predict Response to Trastuzumab Treatment

Clinical Cancer Research : an Official Journal of the American Association for Cancer Research. Mar, 2011  |  Pubmed ID: 21177408

The rapid tumor targeting and pharmacokinetic properties of engineered antibodies make them potentially suitable for use in imaging strategies to predict and monitor response to targeted therapies. This study aims to evaluate C6.5 diabody (C6.5 db), a noncovalent anti-HER2 single-chain Fv dimer, as a radiotracer for predicting response to HER2-targeted therapies such as trastuzumab.

Senescence and Age-specific Trade-offs Between Reproduction and Survival in Female Asian Elephants

Ecology Letters. Mar, 2012  |  Pubmed ID: 22248152

Although studies on laboratory species and natural populations of vertebrates have shown reproduction to impair later performance, little is known of the age-specific associations between reproduction and survival, and how such findings apply to the ageing of large, long-lived species. Herein we develop a framework to examine population-level patterns of reproduction and survival across lifespan in long-lived organisms, and decompose those changes into individual-level effects, and the effects of age-specific trade-offs between fitness components. We apply this to an extensive longitudinal dataset on female semi-captive Asian timber elephants (Elephas maximus) and report the first evidence of age-specific fitness declines that are driven by age-specific associations between fitness components in a long-lived mammal. Associations between reproduction and survival are positive in early life, but negative in later life with up to 71% of later-life survival declines associated with investing in the production of offspring within this population of this critically endangered species.

A Comprehensive Assessment of Mitochondrial Protein Synthesis and Cellular Proliferation with Age and Caloric Restriction

Aging Cell. Feb, 2012  |  Pubmed ID: 22081942

It is proposed that caloric restriction (CR) increases mitochondrial biogenesis. However, it is not clear why CR increases an energetically costly biosynthetic process. We hypothesized that 40% CR would decrease mitochondrial protein synthesis and would be regulated by translational rather than transcriptional mechanisms. We assessed cumulative mitochondrial protein synthesis over 6 weeks and its transcriptional and translational regulation in the liver, heart, and skeletal muscle of young (6 month), middle (12 month), and old (24 month) male B6D2F1 mice that were lifelong CR or ad lib (AL) controls. Mitochondrial protein synthesis was not different between AL and CR (fractional synthesis over 6 weeks (range): liver, 91-100%; heart, 74-85%; skeletal muscle, 53-72%) despite a decreased cellular proliferation in liver and heart with CR. With CR, there was an increase in AMP-activated protein kinase phosphorylation/total (P:T) in heart and liver, and an increase in peroxisome proliferator-activated receptor gamma coactivator 1-α mRNA in all tissues, but not protein. Ribosomal protein S6 was decreased with CR. In conclusion, CR maintained mitochondrial protein synthesis while decreasing cellular proliferation during a time of energetic stress, which is consistent with the concept that CR increases somatic maintenance. Alternative mechanisms to global translation initiation may be responsible for selective translation of mitochondrial proteins.