Ivo M B Francischetti

National Institute of Allergy and Infectious Disease, Maryland, United States

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Publications (110)470.3 Total impact

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    ABSTRACT: Ticks rely exclusively on vertebrate blood for their survival. During feeding ticks inject into their hosts a sophisticated salivary potion that overcomes host hemostasis and adverse inflammatory responses. These mediators may also enhance pathogen transmission. Knowledge of the tick salivary protein repertoire may lead to vaccine targets to disrupt feeding and/or parasite transmission as well as to the discovery of novel pharmacological agents. Male saliva may also assist reproduction because males use their mouthparts to lubricate and introduce their spermatophores into the females' genital pore. The analyses of the sialomes of male and female ticks independently allow us to understand the strategy used by each gender to feed successfully. We sequenced cDNA libraries from pools of salivary glands from adult male and female R. pulchellus feeding at different time points, using the Illumina HiSeq protocol. De novo assembly of a total of 241,229,128 paired-end reads lead to extraction of 50,460 coding sequences (CDS), 11,277 of which had more than 75% coverage to known transcripts, or represented novel sequences, and were submitted to GenBank. Additionally, we generated the proteome, from the salivary gland extracts of male and female R. pulchellus, yielding a total of 454 and 2,063 proteins respectively which were identified by one or more peptides with at least 95% confidence. The data set is presented as an annotated hyperlinked Excel spreadsheet, describing 121 putative secreted protein families. Female and male specific transcripts were identified. This annotated Rhipicephalus pulchellus database represents a mining field for future experiments involving the resolution of time-dependent transcript expression in this tick species, as well as to define novel vaccine targets and discover novel pharmaceuticals. Gender specific proteins may represent different repertoires of pharmacological reagents to assist feeding by each sex, and in males may represent proteins that assist reproduction similarly to seminal proteins in other animals. Copyright © 2015. Published by Elsevier B.V.
    Journal of proteomics. 01/2015;
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    Ivo M B Francischetti, Dongying Ma, John F Andersen, José M C Ribeiro
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    ABSTRACT: Salivary gland homogenate (SGH) from the female mosquitoes Anopheles gambiae, An. stephensi, An. freeborni, An. dirus and An. albimanus were found to exhibit hemagglutinating (lectin) activity. Lectin activity was not found for male An. gambiae, or female Ae aegypti, Culex quinquefasciatus, Phlebotomus duboscqi, and Lutzomyia longipalpis. With respect to species-specificity, An. gambiae SGH agglutinates red blood cells (RBC) from humans, horse, sheep, goat, pig, and cow; it is less active for rats RBC, and not detectable for guinea-pigs or chicken RBC. Notably, lectin activity was inhibited by low concentrations of dextran sulfate 50-500 K, fucoidan, heparin, laminin, heparin sulfate proteoglycan, sialyl-containing glycans (e.g. 3'-sialyl Lewis X, and 6'-sialyl lactose), and gangliosides (e.g. GM3, GD1, GD1b, GTB1, GM1, GQ1B), but not by simple sugars. These results imply that molecule(s) in the salivary gland target sulfated glycans. SGH from An. gambiae was also found to promote agglutination of HL-60 cells which are rich in sialyl Lewis X, a glycan that decorates PSGL-1, the neutrophils receptor that interacts with endothelial cell P-selectin. Accordingly, SGH interferes with HL-60 cells adhesion to immobilized P-selectin. Because An. gambiae SGH expresses galectins, one member of this family (herein named Agalectin) was expressed in E. coli. Recombinant Agalectin behaves as a non-covalent homodimer. It does not display lectin activity, and does not interact with 500 candidates tested in a Glycan microarray. Gel-filtration chromatography of the SGH of An. gambiae identified a fraction with hemagglutinating activity, which was analyzed by 1D PAGE followed by in-gel tryptic digestion, and nano-LC MS/MS. This approach identified several genes which emerge as candidates for a lectin targeting sulfated glycans, the first with this selectivity to be reported in the SGH of a blood-sucking arthropod. The role of salivary molecules (sialogenins) with lectin activity is discussed in the context of inflammation, and parasite-vector-host interactions.
    PLoS ONE 09/2014; 9(9):e107295. · 3.53 Impact Factor
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    ABSTRACT: Invasion of mosquito salivary glands (SGs) by Plasmodium falciparum sporozoites is an essential step in the malaria life cycle. How infection modulates gene expression, and affects hematophagy remains unclear.
    PLoS Pathogens 09/2014; 10(9):e1004338. · 8.14 Impact Factor
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    ABSTRACT: The bilin-binding proteins (BBP) from lepidopteran insects are members of the lipocalin family of proteins and play a special role in pigmentation through the binding of biliverdin IXγ. Lopap, a BBP-like protein from the venom of the toxic caterpillar Lonomia obliqua has been reported to act as a serine protease that activates the coagulation proenzyme prothrombin. Here we show that BBPLo, a variant of lopap from the same organism binds biliverdin IXγ, forming a complex that is spectrally identical with previously described BBP proteins. Although BBPLo is nearly identical in sequence to lopap, no prothrombinase activity was detected in our recombinant preparations using reconstituted systems containing coagulation factors Xa and Va, as well as anionic phospholipids. In addition to biliverdin, BBPLo was found to form a 1∶1 complex with heme prompting us to examine whether the unusual biliverdin IXγ ligand of BBPs forms as a result of oxidation of bound heme in situ rather than by a conventional heme oxygenase. Using ascorbate or a NADPH+-ferredoxin reductase-ferredoxin system as a source of reducing equivalents, spectral changes are seen that suggest an initial reduction of heme to the Fe(II) state and formation of an oxyferrous complex. The complex then disappears and a product identified as a 5-coordinate carbonyl complex of verdoheme, an intermediate in the biosynthesis of biliverdin, is formed. However, further reaction to form biliverdin was not observed, making it unlikely that biliverdin IXγ is formed by this pathway.
    PLoS ONE 06/2014; 9(6):e95424. · 3.53 Impact Factor
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    ABSTRACT: The role of intracellular radical oxygen species (ROS) in pathogenesis of cerebral malaria (CM) remains incompletely understood. We undertook testing Tempol-a superoxide dismutase (SOD) mimetic and pleiotropic intracellular antioxidant-in cells relevant to malaria pathogenesis in the context of coagulation and inflammation. Tempol was also tested in a murine model of CM induced by Plasmodium berghei Anka infection. Tempol was found to prevent transcription and functional expression of procoagulant tissue factor in endothelial cells (ECs) stimulated by lipopolysaccharide (LPS). This effect was accompanied by inhibition of IL-6, IL-8, and monocyte chemoattractant protein (MCP-1) production. Tempol also attenuated platelet aggregation and human promyelocytic leukemia HL60 cells oxidative burst. In dendritic cells, Tempol inhibited LPS-induced production of TNF-α, IL-6, and IL-12p70, downregulated expression of co-stimulatory molecules, and prevented antigen-dependent lymphocyte proliferation. Notably, Tempol (20 mg/kg) partially increased the survival of mice with CM. Mechanistically, treated mice had lowered plasma levels of MCP-1, suggesting that Tempol downmodulates EC function and vascular inflammation. Tempol also diminished blood brain barrier permeability associated with CM when started at day 4 post infection but not at day 1, suggesting that ROS production is tightly regulated. Other antioxidants-such as α-phenyl N-tertiary-butyl nitrone (PBN; a spin trap), MnTe-2-PyP and MnTBAP (Mn-phorphyrin), Mitoquinone (MitoQ) and Mitotempo (mitochondrial antioxidants), M30 (an iron chelator), and epigallocatechin gallate (EGCG; polyphenol from green tea) did not improve survival. By contrast, these compounds (except PBN) inhibited Plasmodium falciparum growth in culture with different IC50s. Knockout mice for SOD1 or phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (gp91(phox-/-)) or mice treated with inhibitors of SOD (diethyldithiocarbamate) or NADPH oxidase (diphenyleneiodonium) did not show protection or exacerbation for CM. Results with Tempol suggest that intracellular ROS contribute, in part, to CM pathogenesis. Therapeutic targeting of intracellular ROS in CM is discussed.
    PLoS ONE 02/2014; 9(2):e87140. · 3.53 Impact Factor
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    ABSTRACT: The saliva of ticks is critical to their survival as parasites and hematophagous animals. In this study, we have purified an enzyme with trypsin-like activity from the saliva of the tick vector of Lyme Disease, Ixodes scapularis. This enzyme, named as IXOSP (Ixodes scapularis salivary serine protease), is a 29.9 kDa molecule with N-terminus FPxMVxLRIKxR. A BLAST search identified IXOSP as a secreted serine protease (AAY66740) with a conserved catalytic triad His, Asp, and Ser. In vitro studies demonstrated that IXOSP cleaves chromogenic substrates with arginine in the P1 position, by a mechanism inhibited by PMSF or aprotinin. Gene expression studies revealed that IXOSP is expressed at different tick developmental stages, including eggs, and unfed or fed adult tick salivary glands, but not in nymphs or in the midgut. While the physiological substrate for IXOSP remains to be identified, we demonstrated that I. scapularis saliva activate protein C (PC) resulting in the production of activated PC, a potent anticoagulant that also regulates a myriad of inflammatory responses through protease activated receptors. In contrast, the salivary glands of Anopheles gambiae, An. stephensi, An. albimanus, Aedes aegypti, Lutzomyia longipalpis, and Phlebotomus ariasi did not activate protein C. These discoveries are discussed in the context of blood coagulation, inflammation and vector-host interactions.
    Toxicon 10/2013; · 2.58 Impact Factor
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    ABSTRACT: The identity of the vampire bat saliva anticoagulant remained elusive for almost a century. Sequencing the salivary gland genes from the vampire bat Desmodus rotundus identified Desmolaris as a novel 21.5-kDa naturally deleted (Kunitz 1-domainless) form of tissue factor pathway inhibitor (TFPI). Recombinant Desmolaris was expressed in HEK293 cells and characterized as a slow, tight, and noncompetitive inhibitor of FXIa by a mechanism modulated by heparin (KD in pM range). Desmolaris also inhibits FXa with lower affinity, independently of protein S. In addition, Desmolaris binds kallikrein and reduces bradykinin generation in plasma activated with kaolin. Truncated- and mutated (R32L)-forms of Desmolaris determined that Arg32 in the Kunitz-1 domain is critical for protease inhibition. Moreover, Kunitz-2 and the C-terminus domains mediate interaction of Desmolaris with heparin and are required for optimal inhibition of FXIa and FXa. Notably, Desmolaris (100 µg/kg) inhibited FeCl3-induced carotid artery thrombus without impairing hemostasis. These results imply that FXIa is the primary in vivo target for Desmolaris, at antithrombotic concentrations. Desmolaris also reduces the polyphosphate (polyP)-induced increase in vascular permeability and collagen/epinephrine-mediated thromboembolism in mice. Desmolaris emerges as a novel anticoagulant targeting FXIa under conditions where the coagulation activation, particularly the contact pathway, plays a major pathological role.
    Blood 10/2013; · 9.78 Impact Factor
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    ABSTRACT: Polyphosphate and heparin are anionic polymers released by activated mast cells and platelets that are known to stimulate the contact pathway of coagulation. These polymers promote both the autoactivation of factor XII and the assembly of complexes containing factor XI, prekallikrein, and high-molecular-weight kininogen. We are searching for salivary proteins from blood-feeding insects that counteract the effect of procoagulant and proinflammatory factors in the host, including elements of the contact pathway. Here, we evaluate the ability of the sand fly salivary proteins, PdSP15a and PdSP15b, to inhibit the contact pathway by disrupting binding of its components to anionic polymers. We attempt to demonstrate binding of the proteins to polyphosphate, heparin, and dextran sulfate. We also evaluate the effect of this binding on contact pathway reactions. We also set out to determine the x-ray crystal structure of PdSP15b and examine the determinants of relevant molecular interactions. Both proteins bind polyphosphate, heparin, and dextran sulfate with high affinity. Through this mechanism they inhibit the autoactivation of factor XII and factor XI, the reciprocal activation of factor XII and prekallikrein, the activation of factor XI by thrombin and factor XIIa, the cleavage of high-molecular-weight kininogen in plasma, and plasma extravasation induced by polyphosphate. The crystal structure of PdSP15b contains an amphipathic helix studded with basic side chains that forms the likely interaction surface. The results of these studies indicate that the binding of anionic polymers by salivary proteins is used by blood feeders as an antihemostatic/anti-inflammatory mechanism.
    Arteriosclerosis Thrombosis and Vascular Biology 10/2013; · 6.34 Impact Factor
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    ABSTRACT: One dimensional gel electrophoresis was used to separate proteins from the saliva of Rhipicephalus sanguineus female ticks fed on rabbits. Gel slices were subjected to tryptic digestion and analyzed by reversed-phase HPLC followed by MS/MS analysis. The data were compared to a database of salivary proteins of the same tick and to the predicted proteins of the host. Saliva was obtained by either pilocarpine or dopamine stimulation of partially fed ticks. Electrophoretic separations of both yielded products that were identified by mass spectrometry, although the pilocarpine-derived sample was of much better quality. The majority of identified proteins were of rabbit origin, indicating the recycling of the host proteins in the tick saliva, including hemoglobin, albumin, haptoglobin, transferring, and a plasma serpin. The few proteins found that were previously associated with parasitism and blood feeding include 2 glycine-rich, cement-like proteins, 2 lipocalins, and a thyropin protease inhibitor. Among other of the 19 tick proteins identified, albeit with undefined roles, were SPARC and cyclophilin A. This catalog provides a resource that can be mined for secreted molecules that play a role in tick-host interactions.
    Ticks and Tick-borne Diseases 09/2013; · 2.35 Impact Factor
  • Daniella M. Mizurini, Ivo M.B. Francischetti, Robson Q. Monteiro
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    ABSTRACT: Aegyptin is a mosquito salivary gland protein and potent inhibitor of platelet aggregation. Aegyptin binds to the von Willebrand factor-binding site on collagen and prevents its interaction with platelets. Because collagen also induces plasma clotting by activation of factor XII, we evaluated the effects of aegyptin on collagen-induced coagulation activation and how it interferes with thrombosis in three different in vivo models. Our results demonstrate that aegyptin abolishes collagen-induced clot formation and thrombin generation in platelet-free plasma. Aegyptin has no antithrombotic activity in the arteriovenous shunt model (collagen-independent) but it prevents laser-induced collagen-mediated thrombus formation in rats. Furthermore, aegyptin protects mice from collagen and epinephrine-induced thromboembolism. Therefore, aegyptin has a dual antithrombotic mechanism: inhibition of platelet-collagen interaction and collagen’s pro-coagulant activity. This is the first description of a collagen-binding protein that also inhibits collagen-mediated coagulant activity.
    Biochemical and Biophysical Research Communications 06/2013; 436(2):235–239. · 2.28 Impact Factor
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    ABSTRACT: The function of the antigen-5/CAP family of proteins ubiquitously found in the salivary gland (SG) of bloodsucking animals has remained elusive for decades. Antigen 5 members from the hematophagous insects Dipetalogaster maxima (DMAV) and Triatoma infestans (TIAV) were expressed and discovered to attenuate platelet aggregation, ATP secretion and TXA2 generation by low doses of collagen (<1ug/mL) but no other agonists. DMAV did not interact with collagen, GPVI or integrin a2β1. This inhibitory profile resembles the effects of antioxidants (e.g., Cu,Zn superoxide dismutase [Cu,Zn SOD]) in platelet function. Accordingly, Cu2+DMAV was found to inhibit cytochrome c reduction by O2 generated by the xanthine/xanthine oxidase implying that it exhibits antioxidant activity. Moreover, our results demonstrate that DMAV blunts the luminescence signal of superoxide generated by phorbol myristate acetate (PMA) stimulated neutrophils. Mechanistically, inductively coupled plasma mass spectrometry (ICP MS) and fluorescence spectroscopy revealed that DMAV, like Cu,Zn-SOD, interacts with Cu2 + which provides redox potential for catalytic removal of superoxide. Notably, fractions of the SG of D. maxima with native DMAV contain Cu2+ and display metal-dependent antioxidant properties. Finally, surface plasmon resonance experiments (BIAcore) determined that DMAV binds sulfated glycosaminoglycans (e.g. heparin, KD100 nmol/L), as reported for extracellular SOD. Antigen-5/CAP emerges as novel family of Cu2+ dependent antioxidant enzymes that inhibit neutrophil oxidative burst and negatively modulate platelet aggregation by a unique salivary mechanism.
    Journal of Biological Chemistry 04/2013; · 4.60 Impact Factor
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    ABSTRACT: Vampire bats are notorious for being the sole mammals that strictly feed on fresh blood for their survival. While their saliva has been historically associated with anticoagulants, only one antihemostatic (plasminogen activator) has been molecularly and functionally characterized. Here, RNAs from both principal and accessory submaxillary (submandibular) salivary glands of Desmodus rotundus were extracted, and ~ 200 million reads were sequenced by Illumina. The principal gland was enriched with plasminogen activators with fibrinolytic properties, members of lipocalin and secretoglobin families, which bind prohemostatic prostaglandins, and endonucleases, which cleave neutrophil-derived procoagulant NETs. Anticoagulant (tissue factor pathway inhibitor, TFPI), vasodilators (PACAP and C-natriuretic peptide), and metalloproteases (ADAMTS-1) were also abundantly expressed. Members of the TSG-6 (anti-inflammatory), antigen 5/CRISP, and CCL28-like (antimicrobial) protein families were also sequenced. Apyrases (which remove platelet agonist ADP), phosphatases (which degrade procoagulant polyphosphates), and sphingomyelinase were found at lower transcriptional levels. Accessory glands were enriched with antimicrobials (lysozyme, defensin, lactotransferrin) and protease inhibitors (TIL-domain, cystatin, Kazal). Mucins, heme-oxygenase, and IgG chains were present in both glands. Proteome analysis by nano LC–MS/MS confirmed that several transcripts are expressed in the glands. The database presented herein is accessible online at http://exon.niaid.nih.gov/transcriptome/D_rotundus/Supplemental-web.xlsx. These results reveal that bat saliva emerges as a novel source of modulators of vascular biology.Biological significanceVampire bat saliva emerges as a novel source of antihemostatics which modulate several aspects of vascular biology.
    Journal of proteomics 02/2013; 82:288–319. · 5.07 Impact Factor
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    ABSTRACT: Bornavirus, a non-segmented, negative-strand RNA viruses, is currently classified into several genetically distinct genotypes, such as Borna disease virus (BDV) and avian bornaviruses (ABVs). Recent studies revealed that bornavirus genotypes show unique sequence variability in the putative 5' untranslated region (5' UTR) of X/P mRNA, a bicistronic mRNA for the X protein and phosphoprotein (P). In this study, to understand the evolutionary relationship among the bornavirus genotypes, we investigated the functional interaction between the X and P proteins of four bornavirus genotypes, BDV, ABV genotype 4 and 5 and reptile bornavirus (RBV), the putative 5' UTRs of which exhibit variation in the length. Immunofluorescence and immunoprecipitation analyses using mammalian and avian cell lines revealed that the X proteins of bornaviruses conserve the ability to facilitate the export of P from the nucleus to the cytoplasm via interaction with P. Furthermore, we showed that inter-genotypic interactions may occur between X and P among the genotypes, except for X of RBV. In addition, a BDV minireplicon assay demonstrated that the X and P proteins of ABVs, but not RBV, can affect the polymerase activity of BDV. This study demonstrates that bornaviruses may have conserved the fundamental function of a regulatory protein during their evolution, whereas RBV has evolved distinctly from the other bornavirus genotypes.
    PLoS ONE 12/2012; 7(12):e51161. · 3.53 Impact Factor
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    ABSTRACT: BACKGROUND: Saliva of hematophagous arthropods contains a diverse mixture of compounds that counteracts host hemostasis. Immunomodulatory and antiinflammatory components are also found in these organisms' saliva. Blood feeding evolved at least ten times within arthropods, providing a scenario of convergent evolution for the solution of the salivary potion. Perhaps because of immune pressure from hosts, the salivary proteins of related organisms have considerable divergence, and new protein families are often found within different genera of the same family or even among subgenera. Fleas radiated with their vertebrate hosts, including within the mammal expansion initiated 65 million years ago. Currently, only one flea species-the rat flea Xenopsylla cheopis-has been investigated by means of salivary transcriptome analysis to reveal salivary constituents, or sialome. We present the analysis of the sialome of cat flea Ctenocephaides felis. METHODOLOGY AND CRITICAL FINDINGS: A salivary gland cDNA library from adult fleas was randomly sequenced, assembled, and annotated. Sialomes of cat and rat fleas have in common the enzyme families of phosphatases (inactive), CD-39-type apyrase, adenosine deaminases, and esterases. Antigen-5 members are also common to both sialomes, as are defensins. FS-I/Cys7 and the 8-Cys families of peptides are also shared by both fleas and are unique to these organisms. The Gly-His-rich peptide similar to holotricin was found only in the cat flea, as were the abundantly expressed Cys-less peptide and a novel short peptide family. CONCLUSIONS/SIGNIFICANCE: Fleas, in contrast to bloodsucking Nematocera (mosquitoes, sand flies, and black flies), appear to concentrate a good portion of their sialome in small polypeptides, none of which have a known function but could act as inhibitors of hemostasis or inflammation. They are also unique in expansion of a phosphatase family that appears to be deficient of enzyme activity and has an unknown function.
    PLoS ONE 09/2012; 7(9):e44612. · 3.53 Impact Factor
  • Toxicon 08/2012; 60(2):130–131. · 2.58 Impact Factor
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    ABSTRACT: Tissue factor (TF) is frequently overexpressed in cancer cells and correlated with more aggressive tumor phenotypes and poor prognosis. In addition to promoting coagulation-dependent metastasis and cancer-associated thrombosis, tumor cell-expressed TF mediates direct cell signaling involving the protease-activated receptor (PAR) 2. Ixolaris is a tick-derived inhibitor of the TF-factor (F)VIIa-Xa coagulation initiation complex which blocks primary tumor growth and angiogenesis in glioblastoma and melanoma models. In this study we address the anti-tumor effects of Ixolaris in TF-VIIa-PAR2 signaling-dependent breast cancer models, a xenograft model of highly aggressive human MDA-MB-231 mfp cells and a syngeneic model of PAR2-deficient and replete PyMT mouse mammary carcinoma cells. Ixolaris potently inhibited the procoagulant activity of human MDA-MB-231mfp or murine PyMT breast cancer cells. Ixolaris blocked signaling by the ternary TF-FVIIa-FXa complex, and, surprisingly, at higher concentrations also the binary TF-FVIIa complex on MDA-MB-231 cells. We show that Ixolaris interacts with certain residues in the human VIIa protease domain that are involved in PAR2 cleavage. In contrast to human VIIa, Ixolaris was a poor inhibitor of murine TF-FVIIa signaling and did not attenuate PAR2-dependent tumor growth in a syngeneic mouse model of breast cancer progression. These data show that Ixolaris inhibits PAR2 cleavage specifically by human TF signaling complexes and suggest that Ixolaris may block tumor growth of human cell models with ectopic FVIIa expression through inhibition of direct TF-FVIIa-PAR2 signaling as well as its anticoagulant activity.
    Journal of Thrombosis and Haemostasis 07/2012; 10(9):1849-58. · 6.08 Impact Factor
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    ABSTRACT: Blood-sucking arthropods' salivary glands contain a remarkable diversity of antihemostatics. The aim of the present study was to identify the unique salivary anticoagulant of the sand fly Lutzomyia longipalpis, which remained elusive for decades. Several L. longipalpis salivary proteins were expressed in human embryonic kidney 293 cells and screened for inhibition of blood coagulation. A novel 32.4-kDa molecule, named Lufaxin, was identified as a slow, tight, noncompetitive, and reversible inhibitor of factor Xa (FXa). Notably, Lufaxin's primary sequence does not share similarity to any physiological or salivary inhibitors of coagulation reported to date. Lufaxin is specific for FXa and does not interact with FX, Dansyl-Glu-Gly-Arg-FXa, or 15 other enzymes. In addition, Lufaxin blocks prothrombinase and increases both prothrombin time and activated partial thromboplastin time. Surface plasmon resonance experiments revealed that FXa binds Lufaxin with an equilibrium constant ≈3 nM, and isothermal titration calorimetry determined a stoichiometry of 1:1. Lufaxin also prevents protease-activated receptor 2 activation by FXa in the MDA-MB-231 cell line and abrogates edema formation triggered by injection of FXa in the paw of mice. Moreover, Lufaxin prevents FeCl(3)-induced carotid artery thrombus formation and prolongs activated partial thromboplastin time ex vivo, implying that it works as an anticoagulant in vivo. Finally, salivary gland of sand flies was found to inhibit FXa and to interact with the enzyme. Lufaxin belongs to a novel family of slow-tight FXa inhibitors, which display antithrombotic and anti-inflammatory activities. It is a useful tool to understand FXa structural features and its role in prohemostatic and proinflammatory events.
    Arteriosclerosis Thrombosis and Vascular Biology 07/2012; 32(9):2185-98. · 6.34 Impact Factor
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    ABSTRACT: Melanoma is a highly metastatic cancer and there is strong evidence that the clotting initiator protein, tissue factor (TF), contributes to its aggressive pattern. TF inhibitors may attenuate primary tumor growth and metastasis. In this study, we evaluated the effect of ixolaris, a TF inhibitor, on a murine model of melanoma B16F10 cells. Enzymatic assays performed with B16F10 and human U87-MG tumor cells as the TF source showed that ixolaris inhibits the generation of FX in either murine, human or hybrid FVIIa/TF complexes. The effect of ixolaris on the metastatic potential was further estimated by intravenous injection of B16F10 cells in C57BL/6 mice. Ixolaris (250 μg/kg) dramatically decreased the number of pulmonary tumor nodules (4 ± 1 compared to 47 ± 10 in the control group). Furthermore, a significant decrease in tumor weights was observed in primary tumor growth assays in animals treated with ixolaris (250 μg/kg) from days 3 to 18 after a subcutaneous inoculation of melanoma cells. Remarkably, immunohistochemical analyses showed that inhibition of melanoma growth by ixolaris is accompanied by a significant downregulation of both vascular endothelial growth factor (VEGF) expression and microvascular density in the tumor mass. Our data demonstrate that ixolaris targets B16F10 cell-derived TF, resulting in the reduction of both the primary tumor growth and the metastatic potential of melanoma, as well as the inhibition of tumor angiogenesis. Therefore TF may be a potential target for the treatment of this aggressive malignancy.
    Thrombosis Research 06/2012; 130(3):e163-70. · 2.43 Impact Factor
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    ABSTRACT: Triatoma matogrossensis is a Hemiptera that belongs to the oliveirai complex, a vector of Chagas' disease that feeds on vertebrate blood in all life stages. Hematophagous insects' salivary glands (SGs) produce potent pharmacologic compounds that counteract host hemostasis, including anticlotting, antiplatelet, and vasodilatory molecules. Exposure to T. matogrossensis was also found to be a risk factor associated with the endemic form of the autoimmune skin disease pemphigus foliaceus, which is described in the same regions where Chagas' disease is observed in Brazil. To obtain a further insight into the salivary biochemical and pharmacologic diversity of this kissing bug and to identify possible allergens that might be associated with this autoimmune disease, a cDNA library from its SGs was randomly sequenced. We present the analysis of a set of 2,230 (SG) cDNA sequences, 1,182 of which coded for proteins of a putative secretory nature.
    The American journal of tropical medicine and hygiene 06/2012; 86(6):1005-14. · 2.53 Impact Factor
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    ABSTRACT: The kissing bug Triatoma rubida (Uhler, 1894) is found in southwestern United States and parts of Mexico where it is found infected with Trypanosoma cruzi, invades human dwellings and causes allergies from their bites. Although the protein salivary composition of several triatomine species is known, not a single salivary protein sequence is known from T. rubida. Furthermore, the salivary diversity of related hematophagous arthropods is very large probably because of the immune pressure from their hosts. Here we report the sialotranscriptome analysis of T. rubida based on the assembly of 1,820 high-quality expressed sequence tags, 51% of which code for putative secreted peptides, including lipocalins, members of the antigen five family, apyrase, hemolysin, and trialysin families. Interestingly, T. rubida lipocalins are at best 40% identical in primary sequence to those of T. protracta, a kissing bug that overlaps its range with T. rubida, indicating the diversity of the salivary lipocalins among species of the same hematophagous genus. We additionally found several expressed sequence tags coding for proteins of clear Trypanosoma spp. origin. This work contributes to the future development of markers of human and pet exposure to T. rubida and to the possible development of desensitization therapies. Supp. Data 1 and 2 (online only) of the transcriptome and deducted protein sequences can be obtained from http://exon.niaid.nih.gov/transcriptome/Trubida/Triru-S1-web.xlsx and http://exon.niaid.nih.gov/transcriptome/Trubida/Triru-S2-web.xlsx.
    Journal of Medical Entomology 05/2012; 49(3):563-72. · 1.82 Impact Factor

Publication Stats

4k Citations
470.30 Total Impact Points


  • 2005–2014
    • National Institute of Allergy and Infectious Disease
      Maryland, United States
  • 1999–2014
    • National Institute of Allergy and Infectious Diseases
      • Laboratory of Parasitic Diseases (LPD)
      Maryland, United States
  • 2013
    • Universidade Federal do Triangulo Mineiro (UFTM)
      Убераба, Minas Gerais, Brazil
    • University of Rhode Island
      • Department of Biomedical and Pharmaceutical Sciences
      Kingston, Rhode Island, United States
  • 1988–2013
    • Federal University of Rio de Janeiro
      • • Institute of Medical Biochemistry
      • • Departamento de Bioquímica
      Rio de Janeiro, Rio de Janeiro, Brazil
  • 2000–2012
    • National Institutes of Health
      • • Section on Organelle Biology
      • • Laboratory of Malaria and Vector Research Molecular Immunology
      • • Laboratory of Parasitic Diseases
      Bethesda, MD, United States
  • 2008
    • University of Brasília
      Brasília, Federal District, Brazil
  • 2005–2007
    • University of Naples Federico II
      • Department of Biology
      Napoli, Campania, Italy