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Sample records for leukocyte lineage-specific genes

  1. SUMOylation of DRIL1 Directs Its Transcriptional Activity Towards Leukocyte Lineage-Specific Genes

    PubMed Central

    van Lohuizen, Maarten; Peeper, Daniel S.

    2009-01-01

    DRIL1 is an ARID family transcription factor that can immortalize primary mouse fibroblasts, bypass RASV12-induced cellular senescence and collaborate with RASV12 or MYC in mediating oncogenic transformation. It also activates immunoglobulin heavy chain transcription and engages in heterodimer formation with E2F to stimulate E2F-dependent transcription. Little, however, is known about the regulation of DRIL1 activity. Recently, DRIL1 was found to interact with the SUMO-conjugating enzyme Ubc9, but the functional relevance of this association has not been assessed. Here, we show that DRIL1 is sumoylated both in vitro and in vivo at lysine 398. Moreover, we provide evidence that PIASy functions as a specific SUMO E3-ligase for DRIL1 and promotes its sumoylation both in vitro and in vivo. Furthermore, consistent with the subnuclear localization of PIASy in the Matrix-Associated Region (MAR), SUMO-modified DRIL1 species are found exclusively in the MAR fraction. This post-translational modification interferes neither with the subcellular localization nor the DNA-binding activity of the protein. In contrast, DRIL1 sumoylation impairs its interaction with E2F1 in vitro and modifies its transcriptional activity in vivo, driving transcription of subset of genes regulating leukocyte fate. Taken together, these results identify sumoylation as a novel post-translational modification of DRIL1 that represents an important mechanism for targeting and modulating DRIL1 transcriptional activity. PMID:19436740

  2. Inferring Gene Family Histories in Yeast Identifies Lineage Specific Expansions

    PubMed Central

    Ames, Ryan M.; Money, Daniel; Lovell, Simon C.

    2014-01-01

    The complement of genes found in the genome is a balance between gene gain and gene loss. Knowledge of the specific genes that are gained and lost over evolutionary time allows an understanding of the evolution of biological functions. Here we use new evolutionary models to infer gene family histories across complete yeast genomes; these models allow us to estimate the relative genome-wide rates of gene birth, death, innovation and extinction (loss of an entire family) for the first time. We show that the rates of gene family evolution vary both between gene families and between species. We are also able to identify those families that have experienced rapid lineage specific expansion/contraction and show that these families are enriched for specific functions. Moreover, we find that families with specific functions are repeatedly expanded in multiple species, suggesting the presence of common adaptations and that these family expansions/contractions are not random. Additionally, we identify potential specialisations, unique to specific species, in the functions of lineage specific expanded families. These results suggest that an important mechanism in the evolution of genome content is the presence of lineage-specific gene family changes. PMID:24921666

  3. Multiple lineage specific expansions within the guanylyl cyclase gene family

    PubMed Central

    Fitzpatrick, David A; O'Halloran, Damien M; Burnell, Ann M

    2006-01-01

    Background Guanylyl cyclases (GCs) are responsible for the production of the secondary messenger cyclic guanosine monophosphate, which plays important roles in a variety of physiological responses such as vision, olfaction, muscle contraction, homeostatic regulation, cardiovascular and nervous function. There are two types of GCs in animals, soluble (sGCs) which are found ubiquitously in cell cytoplasm, and receptor (rGC) forms which span cell membranes. The complete genomes of several vertebrate and invertebrate species are now available. These data provide a platform to investigate the evolution of GCs across a diverse range of animal phyla. Results In this analysis we located GC genes from a broad spectrum of vertebrate and invertebrate animals and reconstructed molecular phylogenies for both sGC and rGC proteins. The most notable features of the resulting phylogenies are the number of lineage specific rGC and sGC expansions that have occurred during metazoan evolution. Among these expansions is a large nematode specific rGC clade comprising 21 genes in C. elegans alone; a vertebrate specific expansion in the natriuretic receptors GC-A and GC-B; a vertebrate specific expansion in the guanylyl GC-C receptors, an echinoderm specific expansion in the sperm rGC genes and a nematode specific sGC clade. Our phylogenetic reconstruction also shows the existence of a basal group of nitric oxide (NO) insensitive insect and nematode sGCs which are regulated by O2. This suggests that the primordial eukaryotes probably utilized sGC as an O2 sensor, with the ligand specificity of sGC later switching to NO which provides a very effective local cell-to-cell signalling system. Phylogenetic analysis of the sGC and bacterial heme nitric oxide/oxygen binding protein domain supports the hypothesis that this domain originated from a cyanobacterial source. Conclusion The most salient feature of our phylogenies is the number of lineage specific expansions, which have occurred within

  4. Lineage-Specific Expansion of IFIT Gene Family: An Insight into Coevolution with IFN Gene Family

    PubMed Central

    Liu, Ying; Zhang, Yi-Bing; Liu, Ting-Kai; Gui, Jian-Fang

    2013-01-01

    In mammals, IFIT (Interferon [IFN]-induced proteins with Tetratricopeptide Repeat [TPR] motifs) family genes are involved in many cellular and viral processes, which are tightly related to mammalian IFN response. However, little is known about non-mammalian IFIT genes. In the present study, IFIT genes are identified in the genome databases from the jawed vertebrates including the cartilaginous elephant shark but not from non-vertebrates such as lancelet, sea squirt and acorn worm, suggesting that IFIT gene family originates from a vertebrate ancestor about 450 million years ago. IFIT family genes show conserved gene structure and gene arrangements. Phylogenetic analyses reveal that this gene family has expanded through lineage-specific and species-specific gene duplication. Interestingly, IFN gene family seem to share a common ancestor and a similar evolutionary mechanism; the function link of IFIT genes to IFN response is present early since the origin of both gene families, as evidenced by the finding that zebrafish IFIT genes are upregulated by fish IFNs, poly(I:C) and two transcription factors IRF3/IRF7, likely via the IFN-stimulated response elements (ISRE) within the promoters of vertebrate IFIT family genes. These coevolution features creates functional association of both family genes to fulfill a common biological process, which is likely selected by viral infection during evolution of vertebrates. Our results are helpful for understanding of evolution of vertebrate IFN system. PMID:23818968

  5. Lineage-specific expansion of IFIT gene family: an insight into coevolution with IFN gene family.

    PubMed

    Liu, Ying; Zhang, Yi-Bing; Liu, Ting-Kai; Gui, Jian-Fang

    2013-01-01

    In mammals, IFIT (Interferon [IFN]-induced proteins with Tetratricopeptide Repeat [TPR] motifs) family genes are involved in many cellular and viral processes, which are tightly related to mammalian IFN response. However, little is known about non-mammalian IFIT genes. In the present study, IFIT genes are identified in the genome databases from the jawed vertebrates including the cartilaginous elephant shark but not from non-vertebrates such as lancelet, sea squirt and acorn worm, suggesting that IFIT gene family originates from a vertebrate ancestor about 450 million years ago. IFIT family genes show conserved gene structure and gene arrangements. Phylogenetic analyses reveal that this gene family has expanded through lineage-specific and species-specific gene duplication. Interestingly, IFN gene family seem to share a common ancestor and a similar evolutionary mechanism; the function link of IFIT genes to IFN response is present early since the origin of both gene families, as evidenced by the finding that zebrafish IFIT genes are upregulated by fish IFNs, poly(I:C) and two transcription factors IRF3/IRF7, likely via the IFN-stimulated response elements (ISRE) within the promoters of vertebrate IFIT family genes. These coevolution features creates functional association of both family genes to fulfill a common biological process, which is likely selected by viral infection during evolution of vertebrates. Our results are helpful for understanding of evolution of vertebrate IFN system. PMID:23818968

  6. In silico analysis of stomach lineage specific gene set expression pattern in gastric cancer

    SciTech Connect

    Pandi, Narayanan Sathiya Suganya, Sivagurunathan; Rajendran, Suriliyandi

    2013-10-04

    Highlights: •Identified stomach lineage specific gene set (SLSGS) was found to be under expressed in gastric tumors. •Elevated expression of SLSGS in gastric tumor is a molecular predictor of metabolic type gastric cancer. •In silico pathway scanning identified estrogen-α signaling is a putative regulator of SLSGS in gastric cancer. •Elevated expression of SLSGS in GC is associated with an overall increase in the survival of GC patients. -- Abstract: Stomach lineage specific gene products act as a protective barrier in the normal stomach and their expression maintains the normal physiological processes, cellular integrity and morphology of the gastric wall. However, the regulation of stomach lineage specific genes in gastric cancer (GC) is far less clear. In the present study, we sought to investigate the role and regulation of stomach lineage specific gene set (SLSGS) in GC. SLSGS was identified by comparing the mRNA expression profiles of normal stomach tissue with other organ tissue. The obtained SLSGS was found to be under expressed in gastric tumors. Functional annotation analysis revealed that the SLSGS was enriched for digestive function and gastric epithelial maintenance. Employing a single sample prediction method across GC mRNA expression profiles identified the under expression of SLSGS in proliferative type and invasive type gastric tumors compared to the metabolic type gastric tumors. Integrative pathway activation prediction analysis revealed a close association between estrogen-α signaling and SLSGS expression pattern in GC. Elevated expression of SLSGS in GC is associated with an overall increase in the survival of GC patients. In conclusion, our results highlight that estrogen mediated regulation of SLSGS in gastric tumor is a molecular predictor of metabolic type GC and prognostic factor in GC.

  7. Evolution of the globin gene family in deuterostomes: lineage-specific patterns of diversification and attrition.

    PubMed

    Hoffmann, Federico G; Opazo, Juan C; Hoogewijs, David; Hankeln, Thomas; Ebner, Bettina; Vinogradov, Serge N; Bailly, Xavier; Storz, Jay F

    2012-07-01

    In the Metazoa, globin proteins display an underlying unity in tertiary structure that belies an extraordinary diversity in primary structures, biochemical properties, and physiological functions. Phylogenetic reconstructions can reveal which of these functions represent novel, lineage-specific innovations, and which represent ancestral functions that are shared with homologous globin proteins in other eukaryotes and even prokaryotes. To date, our understanding of globin diversity in deuterostomes has been hindered by a dearth of genomic sequence data from the Ambulacraria (echinoderms + hemichordates), the sister group of chordates, and the phylum Xenacoelomorpha, which includes xenoturbellids, acoelomorphs, and nemertodermatids. Here, we report the results of a phylogenetic and comparative genomic analysis of the globin gene repertoire of deuterostomes. We first characterized the globin genes of the acorn worm, Saccoglossus kowalevskii, a representative of the phylum Hemichordata. We then integrated genomic sequence data from the acorn worm into a comprehensive analysis of conserved synteny and phylogenetic relationships among globin genes from representatives of the eight lineages that comprise the superphylum Deuterostomia. The primary aims were 1) to unravel the evolutionary history of the globin gene superfamily in deuterostomes and 2) to use the estimated phylogeny to gain insights into the functional evolution of deuterostome globins. Results of our analyses indicate that the deuterostome common ancestor possessed a repertoire of at least four distinct globin paralogs and that different subsets of these ancestral genes have been retained in each of the descendant organismal lineages. In each major deuterostome group, a different subset of ancestral precursor genes underwent lineage-specific expansions of functional diversity through repeated rounds of gene duplication and divergence. By integrating results of the phylogenetic analysis with available

  8. Lineage-specific enhancers activate self-renewal genes in macrophages and embryonic stem cells

    PubMed Central

    Soucie, Erinn L.; Weng, Ziming; Geirsdóttir, Laufey; Molawi, Kaaweh; Maurizio, Julien; Fenouil, Romain; Mossadegh-Keller, Noushine; Gimenez, Gregory; VanHille, Laurent; Beniazza, Meryam; Favret, Jeremy; Berruyer, Carole; Perrin, Pierre; Hacohen, Nir; Andrau, J.-C.; Ferrier, Pierre; Dubreuil, Patrice; Sidow, Arend; Sieweke, Michael H.

    2016-01-01

    Differentiated macrophages can self-renew in tissues and expand long-term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network controlling self-renewal. Single cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells. PMID:26797145

  9. Comparative genomics of rhizobia nodulating soybean suggests extensive recruitment of lineage-specific genes in adaptations

    PubMed Central

    Tian, Chang Fu; Zhou, Yuan Jie; Zhang, Yan Ming; Li, Qin Qin; Zhang, Yun Zeng; Li, Dong Fang; Wang, Shuang; Wang, Jun; Gilbert, Luz B.; Li, Ying Rui; Chen, Wen Xin

    2012-01-01

    The rhizobium–legume symbiosis has been widely studied as the model of mutualistic evolution and the essential component of sustainable agriculture. Extensive genetic and recent genomic studies have led to the hypothesis that many distinct strategies, regardless of rhizobial phylogeny, contributed to the varied rhizobium–legume symbiosis. We sequenced 26 genomes of Sinorhizobium and Bradyrhizobium nodulating soybean to test this hypothesis. The Bradyrhizobium core genome is disproportionally enriched in lipid and secondary metabolism, whereas several gene clusters known to be involved in osmoprotection and adaptation to alkaline pH are specific to the Sinorhizobium core genome. These features are consistent with biogeographic patterns of these bacteria. Surprisingly, no genes are specifically shared by these soybean microsymbionts compared with other legume microsymbionts. On the other hand, phyletic patterns of 561 known symbiosis genes of rhizobia reflected the species phylogeny of these soybean microsymbionts and other rhizobia. Similar analyses with 887 known functional genes or the whole pan genome of rhizobia revealed that only the phyletic distribution of functional genes was consistent with the species tree of rhizobia. Further evolutionary genetics revealed that recombination dominated the evolution of core genome. Taken together, our results suggested that faithfully vertical genes were rare compared with those with history of recombination including lateral gene transfer, although rhizobial adaptations to symbiotic interactions and other environmental conditions extensively recruited lineage-specific shell genes under direct or indirect control through the speciation process. PMID:22586130

  10. Lineage-Specific Expansion of the Chalcone Synthase Gene Family in Rosids

    PubMed Central

    Zavala, Kattina; Opazo, Juan C.

    2015-01-01

    Rosids are a monophyletic group that includes approximately 70,000 species in 140 families, and they are found in a variety of habitats and life forms. Many important crops such as fruit trees and legumes are rosids. The evolutionary success of this group may have been influenced by their ability to produce flavonoids, secondary metabolites that are synthetized through a branch of the phenylpropanoid pathway where chalcone synthase is a key enzyme. In this work, we studied the evolution of the chalcone synthase gene family in 12 species belonging to the rosid clade. Our results show that the last common ancestor of the rosid clade possessed six chalcone synthase gene lineages that were differentially retained during the evolutionary history of the group. In fact, of the six gene lineages that were present in the last common ancestor, 7 species retained 2 of them, whereas the other 5 only retained one gene lineage. We also show that one of the gene lineages was disproportionately expanded in species that belonged to the order Fabales (soybean, barrel medic and Lotus japonicas). Based on the available literature, we suggest that this gene lineage possesses stress-related biological functions (e.g., response to UV light, pathogen defense). We propose that the observed expansion of this clade was a result of a selective pressure to increase the amount of enzymes involved in the production of phenylpropanoid pathway-derived secondary metabolites, which is consistent with the hypothesis that suggested that lineage-specific expansions fuel plant adaptation. PMID:26181912

  11. Lineage-Specific Loss of Function of Bitter Taste Receptor Genes in Humans and Nonhuman Primates

    PubMed Central

    Go, Yasuhiro; Satta, Yoko; Takenaka, Osamu; Takahata, Naoyuki

    2005-01-01

    Since the process of becoming dead genes or pseudogenes (pseudogenization) is irreversible and can occur rather rapidly under certain environmental circumstances, it is one plausible determinant for characterizing species specificity. To test this evolutionary hypothesis, we analyzed the tempo and mode of duplication and pseudogenization of bitter taste receptor (T2R) genes in humans as well as in 12 nonhuman primates. The results show that primates have accumulated more pseudogenes than mice after their separation from the common ancestor and that lineage-specific pseudogenization becomes more conspicuous in humans than in nonhuman primates. Although positive selection has operated on some amino acids in extracellular domains, functional constraints against T2R genes are more relaxed in primates than in mice and this trend has culminated in the rapid deterioration of the bitter-tasting capability in humans. Since T2R molecules play an important role in avoiding generally bitter toxic and harmful substances, substantial modification of the T2R gene repertoire is likely to reflect different responses to changes in the environment and to result from species-specific food preference during primate evolution. PMID:15744053

  12. Genome-Wide Analysis Indicates Lineage-Specific Gene Loss during Papilionoideae Evolution

    PubMed Central

    Gu, Yongzhe; Xing, Shilai; He, Chaoying

    2016-01-01

    Gene loss is the driving force for changes in genome and morphology; however, this particular evolutionary event has been poorly investigated in leguminous plants. Legumes (Fabaceae) have some lineage-specific and diagnostic characteristics that are distinct from other angiosperms. To understand the potential role of gene loss in the evolution of legumes, we compared six genome-sequenced legume species of Papilionoideae, the largest representative clade of Fabaceae, such as Glycine max, with 34 nonlegume plant species, such as Arabidopsis thaliana. The results showed that the putative orthologs of the 34 Arabidopsis genes belonging to 29 gene families were absent in these legume species but these were conserved in the sequenced nonlegume angiosperm lineages. Further evolutionary analyses indicated that the orthologs of these genes were almost completely lost in the Papillionoideae ancestors, thus designated as the legume lost genes (LLGs), and these underwent purifying selection in nonlegume plants. Most LLGs were functionally unknown. In Arabidopsis, two LLGs were well-known genes that played a role in plant immunity such as HARMLESS TO OZONE LAYER 1 and HOPZ-ACTIVATED RESISTANCE 1, and 16 additional LLGs were predicted to participate in plant–pathogen interactions in in silico expression and protein–protein interaction network analyses. Most of these LLGs’ orthologs in various plants were also found to be associated with biotic stress response, indicating the conserved role of these genes in plant defense. The evolutionary implication of LLGs during the development of the ability of symbiotic nitrogen fixation involving plant and bacterial interactions, which is a well-known characteristic of most legumes, is also discussed. Our work sheds light on the evolutionary implication of gene loss events in Papilionoideae evolution, as well as provides new insights into crop design to improve nitrogen fixation capacity. PMID:26868598

  13. Genome-Wide Analysis Indicates Lineage-Specific Gene Loss during Papilionoideae Evolution.

    PubMed

    Gu, Yongzhe; Xing, Shilai; He, Chaoying

    2016-03-01

    Gene loss is the driving force for changes in genome and morphology; however, this particular evolutionary event has been poorly investigated in leguminous plants. Legumes (Fabaceae) have some lineage-specific and diagnostic characteristics that are distinct from other angiosperms. To understand the potential role of gene loss in the evolution of legumes, we compared six genome-sequenced legume species of Papilionoideae, the largest representative clade of Fabaceae, such as Glycine max, with 34 nonlegume plant species, such as Arabidopsis thaliana. The results showed that the putative orthologs of the 34 Arabidopsis genes belonging to 29 gene families were absent in these legume species but these were conserved in the sequenced nonlegume angiosperm lineages. Further evolutionary analyses indicated that the orthologs of these genes were almost completely lost in the Papillionoideae ancestors, thus designated as the legume lost genes (LLGs), and these underwent purifying selection in nonlegume plants. Most LLGs were functionally unknown. In Arabidopsis, two LLGs were well-known genes that played a role in plant immunity such as HARMLESS TO OZONE LAYER 1 and HOPZ-ACTIVATED RESISTANCE 1, and 16 additional LLGs were predicted to participate in plant-pathogen interactions in in silico expression and protein-protein interaction network analyses. Most of these LLGs' orthologs in various plants were also found to be associated with biotic stress response, indicating the conserved role of these genes in plant defense. The evolutionary implication of LLGs during the development of the ability of symbiotic nitrogen fixation involving plant and bacterial interactions, which is a well-known characteristic of most legumes, is also discussed. Our work sheds light on the evolutionary implication of gene loss events in Papilionoideae evolution, as well as provides new insights into crop design to improve nitrogen fixation capacity. PMID:26868598

  14. Lineage-specific evolution of the vertebrate Otopetrin gene family revealed by comparative genomic analyses

    PubMed Central

    2011-01-01

    Background Mutations in the Otopetrin 1 gene (Otop1) in mice and fish produce an unusual bilateral vestibular pathology that involves the absence of otoconia without hearing impairment. The encoded protein, Otop1, is the only functionally characterized member of the Otopetrin Domain Protein (ODP) family; the extended sequence and structural preservation of ODP proteins in metazoans suggest a conserved functional role. Here, we use the tools of sequence- and cytogenetic-based comparative genomics to study the Otop1 and the Otop2-Otop3 genes and to establish their genomic context in 25 vertebrates. We extend our evolutionary study to include the gene mutated in Usher syndrome (USH) subtype 1G (Ush1g), both because of the head-to-tail clustering of Ush1g with Otop2 and because Otop1 and Ush1g mutations result in inner ear phenotypes. Results We established that OTOP1 is the boundary gene of an inversion polymorphism on human chromosome 4p16 that originated in the common human-chimpanzee lineage more than 6 million years ago. Other lineage-specific evolutionary events included a three-fold expansion of the Otop genes in Xenopus tropicalis and of Ush1g in teleostei fish. The tight physical linkage between Otop2 and Ush1g is conserved in all vertebrates. To further understand the functional organization of the Ushg1-Otop2 locus, we deduced a putative map of binding sites for CCCTC-binding factor (CTCF), a mammalian insulator transcription factor, from genome-wide chromatin immunoprecipitation-sequencing (ChIP-seq) data in mouse and human embryonic stem (ES) cells combined with detection of CTCF-binding motifs. Conclusions The results presented here clarify the evolutionary history of the vertebrate Otop and Ush1g families, and establish a framework for studying the possible interaction(s) of Ush1g and Otop in developmental pathways. PMID:21261979

  15. Constraint of gene expression by the chromatin remodelling protein CHD4 facilitates lineage specification

    PubMed Central

    O'Shaughnessy-Kirwan, Aoife; Signolet, Jason; Costello, Ita; Gharbi, Sarah; Hendrich, Brian

    2015-01-01

    Chromatin remodelling proteins are essential for different aspects of metazoan biology, yet functional details of why these proteins are important are lacking. Although it is possible to describe the biochemistry of how they remodel chromatin, their chromatin-binding profiles in cell lines, and gene expression changes upon loss of a given protein, in very few cases can this easily translate into an understanding of how the function of that protein actually influences a developmental process. Here, we investigate how the chromatin remodelling protein CHD4 facilitates the first lineage decision in mammalian embryogenesis. Embryos lacking CHD4 can form a morphologically normal early blastocyst, but are unable to successfully complete the first lineage decision and form functional trophectoderm (TE). In the absence of a functional TE, Chd4 mutant blastocysts do not implant and are hence not viable. By measuring transcript levels in single cells from early embryos, we show that CHD4 influences the frequency at which unspecified cells in preimplantation stage embryos express lineage markers prior to the execution of this first lineage decision. In the absence of CHD4, this frequency is increased in 16-cell embryos, and by the blastocyst stage cells fail to properly adopt a TE gene expression programme. We propose that CHD4 allows cells to undertake lineage commitment in vivo by modulating the frequency with which lineage-specification genes are expressed. This provides novel insight into both how lineage decisions are made in mammalian cells, and how a chromatin remodelling protein functions to facilitate lineage commitment. PMID:26116663

  16. The Lineage-Specific Transcription Factor PU.1 Prevents Polycomb-Mediated Heterochromatin Formation at Macrophage-Specific Genes.

    PubMed

    Tagore, Mohita; McAndrew, Michael J; Gjidoda, Alison; Floer, Monique

    2015-08-01

    Lineage-specific transcription factors (TFs) are important determinants of cellular identity, but their exact mode of action has remained unclear. Here we show using a macrophage differentiation system that the lineage-specific TF PU.1 keeps macrophage-specific genes accessible during differentiation by preventing Polycomb repressive complex 2 (PRC2) binding to transcriptional regulatory elements. We demonstrate that the distal enhancer of a gene becomes bound by PRC2 as cells differentiate in the absence of PU.1 binding and that the gene is wrapped into heterochromatin, which is characterized by increased nucleosome occupancy and H3K27 trimethylation. This renders the gene inaccessible to the transcriptional machinery and prevents induction of the gene in response to an external signal in mature cells. In contrast, if PU.1 is bound at the transcriptional regulatory region of a gene during differentiation, PRC2 is not recruited, nucleosome occupancy is kept low, and the gene can be induced in mature macrophages. Similar results were obtained at the enhancers of other macrophage-specific genes that fail to bind PU.1 as an estrogen receptor fusion (PUER) in this system. These results show that one role of PU.1 is to exclude PRC2 and to prevent heterochromatin formation at macrophage-specific genes. PMID:26012552

  17. Amoebozoa possess lineage-specific globin gene repertoires gained by individual horizontal gene transfers.

    PubMed

    Dröge, Jasmin; Buczek, Dorota; Suzuki, Yutaka; Makałowski, Wojciech

    2014-01-01

    The Amoebozoa represent a clade of unicellular amoeboid organisms that display a wide variety of lifestyles, including free-living and parasitic species. For example, the social amoeba Dictyostelium discoideum has the ability to aggregate into a multicellular fruiting body upon starvation, while the pathogenic amoeba Entamoeba histolytica is a parasite of humans. Globins are small heme proteins that are present in almost all extant organisms. Although several genomes of amoebozoan species have been sequenced, little is known about the phyletic distribution of globin genes within this phylum. Only two flavohemoglobins (FHbs) of D. discoideum have been reported and characterized previously while the genomes of Entamoeba species are apparently devoid of globin genes. We investigated eleven amoebozoan species for the presence of globin genes by genomic and phylogenetic in silico analyses. Additional FHb genes were identified in the genomes of four social amoebas and the true slime mold Physarum polycephalum. Moreover, a single-domain globin (SDFgb) of Hartmannella vermiformis, as well as two truncated hemoglobins (trHbs) of Acanthamoeba castellanii were identified. Phylogenetic evidence suggests that these globin genes were independently acquired via horizontal gene transfer from some ancestral bacteria. Furthermore, the phylogenetic tree of amoebozoan FHbs indicates that they do not share a common ancestry and that a transfer of FHbs from bacteria to amoeba occurred multiple times. PMID:25013378

  18. DLGP: A database for lineage-conserved and lineage-specific gene pairs in animal and plant genomes.

    PubMed

    Wang, Dapeng

    2016-01-15

    The conservation of gene organization in the genome with lineage-specificity is an invaluable resource to decipher their potential functionality with diverse selective constraints, especially in higher animals and plants. Gene pairs appear to be the minimal structure for such kind of gene clusters that tend to reside in their preferred locations, representing the distinctive genomic characteristics in single species or a given lineage. Despite gene families having been investigated in a widespread manner, the definition of gene pair families in various taxa still lacks adequate attention. To address this issue, we report DLGP (http://lcgbase.big.ac.cn/DLGP/) that stores the pre-calculated lineage-based gene pairs in currently available 134 animal and plant genomes and inspect them under the same analytical framework, bringing out a set of innovational features. First, the taxonomy or lineage has been classified into four levels such as Kingdom, Phylum, Class and Order. It adopts all-to-all comparison strategy to identify the possible conserved gene pairs in all species for each gene pair in certain species and reckon those that are conserved in over a significant proportion of species in a given lineage (e.g. Primates, Diptera or Poales) as the lineage-conserved gene pairs. Furthermore, it predicts the lineage-specific gene pairs by retaining the above-mentioned lineage-conserved gene pairs that are not conserved in any other lineages. Second, it carries out pairwise comparison for the gene pairs between two compared species and creates the table including all the conserved gene pairs and the image elucidating the conservation degree of gene pairs in chromosomal level. Third, it supplies gene order browser to extend gene pairs to gene clusters, allowing users to view the evolution dynamics in the gene context in an intuitive manner. This database will be able to facilitate the particular comparison between animals and plants, between vertebrates and arthropods, and

  19. Genome-wide identification of lineage-specific genes in Arabidopsis, Oryza and Populus

    SciTech Connect

    Yang, Xiaohan; Jawdy, Sara; Tschaplinski, Timothy J; Tuskan, Gerald A

    2009-01-01

    Protein sequences were compared among Arabidopsis, Oryza and Populus to identify differential gene (DG) sets that are in one but not the other two genomes. The DG sets were screened against a plant transcript database, the NR protein database and six newly-sequenced genomes (Carica, Glycine, Medicago, Sorghum, Vitis and Zea) to identify a set of species-specific genes (SS). Gene expression, protein motif and intron number were examined. 192, 641 and 109 SS genes were identified in Arabidopsis, Oryza and Populus, respectively. Some SS genes were preferentially expressed in flowers, roots, xylem and cambium or up-regulated by stress. Six conserved motifs in Arabidopsis and Oryza SS proteins were found in other distant lineages. The SS gene sets were enriched with intronless genes. The results reflect functional and/or anatomical differences between monocots and eudicots or between herbaceous and woody plants. The Populus-specific genes are candidates for carbon sequestration and biofuel research.

  20. The polyphenol oxidase gene family in land plants: Lineage-specific duplication and expansion

    PubMed Central

    2012-01-01

    Background Plant polyphenol oxidases (PPOs) are enzymes that typically use molecular oxygen to oxidize ortho-diphenols to ortho-quinones. These commonly cause browning reactions following tissue damage, and may be important in plant defense. Some PPOs function as hydroxylases or in cross-linking reactions, but in most plants their physiological roles are not known. To better understand the importance of PPOs in the plant kingdom, we surveyed PPO gene families in 25 sequenced genomes from chlorophytes, bryophytes, lycophytes, and flowering plants. The PPO genes were then analyzed in silico for gene structure, phylogenetic relationships, and targeting signals. Results Many previously uncharacterized PPO genes were uncovered. The moss, Physcomitrella patens, contained 13 PPO genes and Selaginella moellendorffii (spike moss) and Glycine max (soybean) each had 11 genes. Populus trichocarpa (poplar) contained a highly diversified gene family with 11 PPO genes, but several flowering plants had only a single PPO gene. By contrast, no PPO-like sequences were identified in several chlorophyte (green algae) genomes or Arabidopsis (A. lyrata and A. thaliana). We found that many PPOs contained one or two introns often near the 3’ terminus. Furthermore, N-terminal amino acid sequence analysis using ChloroP and TargetP 1.1 predicted that several putative PPOs are synthesized via the secretory pathway, a unique finding as most PPOs are predicted to be chloroplast proteins. Phylogenetic reconstruction of these sequences revealed that large PPO gene repertoires in some species are mostly a consequence of independent bursts of gene duplication, while the lineage leading to Arabidopsis must have lost all PPO genes. Conclusion Our survey identified PPOs in gene families of varying sizes in all land plants except in the genus Arabidopsis. While we found variation in intron numbers and positions, overall PPO gene structure is congruent with the phylogenetic relationships based on

  1. Topologically associated domains enriched for lineage-specific genes reveal expression-dependent nuclear topologies during myogenesis

    PubMed Central

    Neems, Daniel S.; Garza-Gongora, Arturo G.; Smith, Erica D.; Kosak, Steven T.

    2016-01-01

    The linear distribution of genes across chromosomes and the spatial localization of genes within the nucleus are related to their transcriptional regulation. The mechanistic consequences of linear gene order, and how it may relate to the functional output of genome organization, remain to be fully resolved, however. Here we tested the relationship between linear and 3D organization of gene regulation during myogenesis. Our analysis has identified a subset of topologically associated domains (TADs) that are significantly enriched for muscle-specific genes. These lineage-enriched TADs demonstrate an expression-dependent pattern of nuclear organization that influences the positioning of adjacent nonenriched TADs. Therefore, lineage-enriched TADs inform cell-specific genome organization during myogenesis. The reduction of allelic spatial distance of one of these domains, which contains Myogenin, correlates with reduced transcriptional variability, identifying a potential role for lineage-specific nuclear topology. Using a fusion-based strategy to decouple mitosis and myotube formation, we demonstrate that the cell-specific topology of syncytial nuclei is dependent on cell division. We propose that the effects of linear and spatial organization of gene loci on gene regulation are linked through TAD architecture, and that mitosis is critical for establishing nuclear topologies during cellular differentiation. PMID:26957603

  2. Lineage-specific gene radiations underlie the evolution of novel betalain pigmentation in Caryophyllales.

    PubMed

    Brockington, Samuel F; Yang, Ya; Gandia-Herrero, Fernando; Covshoff, Sarah; Hibberd, Julian M; Sage, Rowan F; Wong, Gane K S; Moore, Michael J; Smith, Stephen A

    2015-09-01

    Betalain pigments are unique to the Caryophyllales and structurally and biosynthetically distinct from anthocyanins. Two key enzymes within the betalain synthesis pathway have been identified: 4,5-dioxygenase (DODA) that catalyzes the formation of betalamic acid and CYP76AD1, a cytochrome P450 gene that catalyzes the formation of cyclo-DOPA. We performed phylogenetic analyses to reveal the evolutionary history of the DODA and CYP76AD1 lineages and in the context of an ancestral reconstruction of pigment states we explored the evolution of these genes in relation to the complex evolution of pigments in Caryophylalles. Duplications within the CYP76AD1 and DODA lineages arose just before the origin of betalain pigmentation in the core Caryophyllales. The duplications gave rise to DODA-α and CYP76AD1-α isoforms that appear specific to betalain synthesis. Both betalain-specific isoforms were then lost or downregulated in the anthocyanic Molluginaceae and Caryophyllaceae. Our findings suggest a single origin of the betalain synthesis pathway, with neofunctionalization following gene duplications in the CYP76AD1 and DODA lineages. Loss of DODA-α and CYP76AD1-α in anthocyanic taxa suggests that betalain pigmentation has been lost twice in Caryophyllales, and exclusion of betalain pigments from anthocyanic taxa is mediated through gene loss or downregulation. [Correction added after online publication 13 May 2015: in the last two paragraphs of the Summary the gene name CYP761A was changed to CYP76AD1.]. PMID:25966996

  3. Lineage-specific gene radiations underlie the evolution of novel betalain pigmentation in Caryophyllales

    PubMed Central

    Brockington, Samuel F; Yang, Ya; Gandia-Herrero, Fernando; Covshoff, Sarah; Hibberd, Julian M; Sage, Rowan F; Wong, Gane K S; Moore, Michael J; Smith, Stephen A

    2015-01-01

    Betalain pigments are unique to the Caryophyllales and structurally and biosynthetically distinct from anthocyanins. Two key enzymes within the betalain synthesis pathway have been identified: 4,5-dioxygenase (DODA) that catalyzes the formation of betalamic acid and CYP76AD1, a cytochrome P450 gene that catalyzes the formation of cyclo-DOPA. We performed phylogenetic analyses to reveal the evolutionary history of the DODA and CYP76AD1 lineages and in the context of an ancestral reconstruction of pigment states we explored the evolution of these genes in relation to the complex evolution of pigments in Caryophylalles. Duplications within the CYP76AD1 and DODA lineages arose just before the origin of betalain pigmentation in the core Caryophyllales. The duplications gave rise to DODA-α and CYP76AD1-α isoforms that appear specific to betalain synthesis. Both betalain-specific isoforms were then lost or downregulated in the anthocyanic Molluginaceae and Caryophyllaceae. Our findings suggest a single origin of the betalain synthesis pathway, with neofunctionalization following gene duplications in the CYP76AD1 and DODA lineages. Loss of DODA-α and CYP76AD1-α in anthocyanic taxa suggests that betalain pigmentation has been lost twice in Caryophyllales, and exclusion of betalain pigments from anthocyanic taxa is mediated through gene loss or downregulation. [Correction added after online publication 13 May 2015: in the last two paragraphs of the Summary the gene name CYP761A was changed to CYP76AD1.] PMID:25966996

  4. Helicobacter pylori Evolution: Lineage- Specific Adaptations in Homologs of Eukaryotic Sel1-Like Genes

    PubMed Central

    Mittl, Peer R. E; Lee, Hae-Kyung; Dailide, Geidrius; Tan, Shumin; Ito, Yoshiyuki; Secka, Ousman; Dailidiene, Daiva; Putty, Kalyani; Berg, Douglas E; Kalia, Awdhesh

    2007-01-01

    Geographic partitioning is postulated to foster divergence of Helicobacter pylori populations as an adaptive response to local differences in predominant host physiology. H. pylori's ability to establish persistent infection despite host inflammatory responses likely involves active management of host defenses using bacterial proteins that may themselves be targets for adaptive evolution. Sequenced H. pylori genomes encode a family of eight or nine secreted proteins containing repeat motifs that are characteristic of the eukaryotic Sel1 regulatory protein, whereas the related Campylobacter and Wolinella genomes each contain only one or two such “Sel1-like repeat” (SLR) genes (“slr genes”). Signatures of positive selection (ratio of nonsynonymous to synonymous mutations, dN/dS = ω > 1) were evident in the evolutionary history of H. pylori slr gene family expansion. Sequence analysis of six of these slr genes (hp0160, hp0211, hp0235, hp0519, hp0628, and hp1117) from representative East Asian, European, and African H. pylori strains revealed that all but hp0628 had undergone positive selection, with different amino acids often selected in different regions. Most striking was a divergence of Japanese and Korean alleles of hp0519, with Japanese alleles having undergone particularly strong positive selection (ωJ > 25), whereas alleles of other genes from these populations were intermingled. Homology-based structural modeling localized most residues under positive selection to SLR protein surfaces. Rapid evolution of certain slr genes in specific H. pylori lineages suggests a model of adaptive change driven by selection for fine-tuning of host responses, and facilitated by geographic isolation. Characterization of such local adaptations should help elucidate how H. pylori manages persistent infection, and potentially lead to interventions tailored to diverse human populations. PMID:17696605

  5. The Lineage-Specific Evolution of Aquaporin Gene Clusters Facilitated Tetrapod Terrestrial Adaptation

    PubMed Central

    Finn, Roderick Nigel; Chauvigné, François; Hlidberg, Jón Baldur; Cutler, Christopher P.; Cerdà, Joan

    2014-01-01

    A major physiological barrier for aquatic organisms adapting to terrestrial life is dessication in the aerial environment. This barrier was nevertheless overcome by the Devonian ancestors of extant Tetrapoda, but the origin of specific molecular mechanisms that solved this water problem remains largely unknown. Here we show that an ancient aquaporin gene cluster evolved specifically in the sarcopterygian lineage, and subsequently diverged into paralogous forms of AQP2, -5, or -6 to mediate water conservation in extant Tetrapoda. To determine the origin of these apomorphic genomic traits, we combined aquaporin sequencing from jawless and jawed vertebrates with broad taxon assembly of >2,000 transcripts amongst 131 deuterostome genomes and developed a model based upon Bayesian inference that traces their convergent roots to stem subfamilies in basal Metazoa and Prokaryota. This approach uncovered an unexpected diversity of aquaporins in every lineage investigated, and revealed that the vertebrate superfamily consists of 17 classes of aquaporins (Aqp0 - Aqp16). The oldest orthologs associated with water conservation in modern Tetrapoda are traced to a cluster of three aqp2-like genes in Actinistia that likely arose >500 Ma through duplication of an aqp0-like gene present in a jawless ancestor. In sea lamprey, we show that aqp0 first arose in a protocluster comprised of a novel aqp14 paralog and a fused aqp01 gene. To corroborate these findings, we conducted phylogenetic analyses of five syntenic nuclear receptor subfamilies, which, together with observations of extensive genome rearrangements, support the coincident loss of ancestral aqp2-like orthologs in Actinopterygii. We thus conclude that the divergence of sarcopterygian-specific aquaporin gene clusters was permissive for the evolution of water conservation mechanisms that facilitated tetrapod terrestrial adaptation. PMID:25426855

  6. Dynamic changes in replication timing and gene expression during lineage specification of human pluripotent stem cells.

    PubMed

    Rivera-Mulia, Juan Carlos; Buckley, Quinton; Sasaki, Takayo; Zimmerman, Jared; Didier, Ruth A; Nazor, Kristopher; Loring, Jeanne F; Lian, Zheng; Weissman, Sherman; Robins, Allan J; Schulz, Thomas C; Menendez, Laura; Kulik, Michael J; Dalton, Stephen; Gabr, Haitham; Kahveci, Tamer; Gilbert, David M

    2015-08-01

    Duplication of the genome in mammalian cells occurs in a defined temporal order referred to as its replication-timing (RT) program. RT changes dynamically during development, regulated in units of 400-800 kb referred to as replication domains (RDs). Changes in RT are generally coordinated with transcriptional competence and changes in subnuclear position. We generated genome-wide RT profiles for 26 distinct human cell types, including embryonic stem cell (hESC)-derived, primary cells and established cell lines representing intermediate stages of endoderm, mesoderm, ectoderm, and neural crest (NC) development. We identified clusters of RDs that replicate at unique times in each stage (RT signatures) and confirmed global consolidation of the genome into larger synchronously replicating segments during differentiation. Surprisingly, transcriptome data revealed that the well-accepted correlation between early replication and transcriptional activity was restricted to RT-constitutive genes, whereas two-thirds of the genes that switched RT during differentiation were strongly expressed when late replicating in one or more cell types. Closer inspection revealed that transcription of this class of genes was frequently restricted to the lineage in which the RT switch occurred, but was induced prior to a late-to-early RT switch and/or down-regulated after an early-to-late RT switch. Analysis of transcriptional regulatory networks showed that this class of genes contains strong regulators of genes that were only expressed when early replicating. These results provide intriguing new insight into the complex relationship between transcription and RT regulation during human development. PMID:26055160

  7. Lineage-Specific Regulation of Epigenetic Modifier Genes in Human Liver and Brain

    PubMed Central

    Weng, Matthias K.; Natarajan, Karthick; Scholz, Diana; Ivanova, Violeta N.; Sachinidis, Agapios; Hengstler, Jan G.; Waldmann, Tanja; Leist, Marcel

    2014-01-01

    Despite an abundance of studies on chromatin states and dynamics, there is an astonishing dearth of information on the expression of genes responsible for regulating histone and DNA modifications. We used here a set of 156 defined epigenetic modifier genes (EMG) and profiled their expression pattern in cells of different lineages. As reference value, expression data from human embryonic stem cells (hESC) were used. Hepatocyte-like cells were generated from hESC, and their EMG expression was compared to primary human liver cells. In parallel, we generated postmitotic human neurons (Lu d6), and compared their relative EMG expression to human cortex (Ctx). Clustering analysis of all cell types showed that neuronal lineage samples grouped together (94 similarly regulated EMG), as did liver cells (61 similarly-regulated), while the two lineages were clearly distinct. The general classification was followed by detailed comparison of the major EMG groups; genes that were higher expressed in differentiated cells than in hESC included the acetyltransferase KAT2B and the methyltransferase SETD7. Neuro-specific EMGs were the histone deacetylases HDAC5 and HDAC7, and the arginine-methyltransferase PRMT8. Comparison of young (Lu d6) and more aged (Ctx) neuronal samples suggested a maturation-dependent switch in the expression of functionally homologous proteins. For instance, the ratio of the histone H3 K27 methyltransfereases, EZH1 to EZH2, was high in Ctx and low in Lu d6. The same was observed for the polycomb repressive complex 1 (PRC1) subunits CBX7 and CBX8. A large proportion of EMGs in differentiated cells was very differently expressed than in hESC, and absolute levels were significantly higher in neuronal samples than in hepatic cells. Thus, there seem to be distinct qualitative and quantitative differences in EMG expression between cell lineages. PMID:25054330

  8. Origin, evolution and classification of type-3 copper proteins: lineage-specific gene expansions and losses across the Metazoa

    PubMed Central

    2013-01-01

    Background Tyrosinases, tyrosinase-related proteins, catechol oxidases and hemocyanins comprise the type-3 copper protein family and are involved in a variety of biological processes, including pigment formation, innate immunity and oxygen transport. Although this family is present in the three domains of life, its origin and early evolution are not well understood. Previous analyses of type-3 copper proteins largely have focussed on specific animal and plant phyla. Results Here, we combine genomic, phylogenetic and structural analyses to show that the original type-3 copper protein possessed a signal peptide and may have been secreted (we designate proteins of this type the α subclass). This ancestral type-3 copper protein gene underwent two duplication events, the first prior to the divergence of the unikont eukaryotic lineages and the second before the diversification of animals. The former duplication gave rise to a cytosolic form (β) and the latter to a membrane-bound form (γ). Structural comparisons reveal that the active site of α and γ forms are covered by aliphatic amino acids, and the β form has a highly conserved aromatic residue in this position. The subsequent evolution of this gene family in modern lineages of multicellular eukaryotes is typified by the loss of one or more of these three subclasses and the lineage-specific expansion of one or both of the remaining subclasses. Conclusions The diversity of type-3 copper proteins in animals and other eukaryotes is consistent with two ancient gene duplication events leading to α, β and γ subclasses, followed by the differential loss and expansion of one or more of these subclasses in specific kingdoms and phyla. This has led to many lineage-specific type-3 copper protein repertoires and in some cases the independent evolution of functionally-classified tyrosinases and hemocyanins. For example, the oxygen-carrying hemocyanins in arthropods evolved from a β-subclass tyrosinase, whilst hemocyanins

  9. An LTR Retrotransposon-Derived Gene Displays Lineage-Specific Structural and Putative Species-Specific Functional Variations in Eutherians.

    PubMed

    Irie, Masahito; Koga, Akihiko; Kaneko-Ishino, Tomoko; Ishino, Fumitoshi

    2016-01-01

    Amongst the 11 eutherian-specific genes acquired from a sushi-ichi retrotransposon is the CCHC type zinc-finger protein-encoding gene SIRH11/ZCCHC16. Its contribution to eutherian brain evolution is implied because of its involvement in cognitive function in mice, possibly via the noradrenergic system. Although, the possibility that Sirh11/Zcchc16 functions as a non-coding RNA still remains, dN/dS ratios in pairwise comparisons between its orthologs have provided supportive evidence that it acts as a protein. It became a pseudogene in armadillos (Cingulata) and sloths (Pilosa), the only two extant orders of xenarthra, which prompted us to examine the lineage-specific variations of SIRH11/ZCCHC16 in eutherians. We examined the predicted SIRH11/ZCCHC16 open reading frame (ORF) in 95 eutherian species based on the genomic DNA information in GenBank. A large variation in the SIRH11/ZCCHC16 ORF was detected in several lineages. These include a lack of a CCHC RNA-binding domain in its C-terminus, observed in gibbons (Hylobatidae: Primates) and megabats (Megachiroptera: Chiroptera). A lack of the N-terminal half, on the other hand, was observed in New World monkeys (Platyrrhini: Primates) and species belonging to New World and African Hystricognaths (Caviomorpha and Bathyergidae: Rodents) along with Cetacea and Ruminantia (Cetartiodactyla). Among the hominoids, interestingly, three out of four genera of gibbons have lost normal SIRH11/ZCCHC16 function by deletion or the lack of the CCHC RNA-binding domain. Our extensive dN/dS analysis suggests that such truncated SIRH11/ZCCHC16 ORFs are functionally diversified even within lineages. Combined, our results show that SIRH11/ZCCHC16 may contribute to the diversification of eutherians by lineage-specific structural changes after its domestication in the common eutherian ancestor, followed by putative species-specific functional changes that enhanced fitness and occurred as a consequence of complex natural selection events

  10. An LTR Retrotransposon-Derived Gene Displays Lineage-Specific Structural and Putative Species-Specific Functional Variations in Eutherians

    PubMed Central

    Irie, Masahito; Koga, Akihiko; Kaneko-Ishino, Tomoko; Ishino, Fumitoshi

    2016-01-01

    Amongst the 11 eutherian-specific genes acquired from a sushi-ichi retrotransposon is the CCHC type zinc-finger protein-encoding gene SIRH11/ZCCHC16. Its contribution to eutherian brain evolution is implied because of its involvement in cognitive function in mice, possibly via the noradrenergic system. Although, the possibility that Sirh11/Zcchc16 functions as a non-coding RNA still remains, dN/dS ratios in pairwise comparisons between its orthologs have provided supportive evidence that it acts as a protein. It became a pseudogene in armadillos (Cingulata) and sloths (Pilosa), the only two extant orders of xenarthra, which prompted us to examine the lineage-specific variations of SIRH11/ZCCHC16 in eutherians. We examined the predicted SIRH11/ZCCHC16 open reading frame (ORF) in 95 eutherian species based on the genomic DNA information in GenBank. A large variation in the SIRH11/ZCCHC16 ORF was detected in several lineages. These include a lack of a CCHC RNA-binding domain in its C-terminus, observed in gibbons (Hylobatidae: Primates) and megabats (Megachiroptera: Chiroptera). A lack of the N-terminal half, on the other hand, was observed in New World monkeys (Platyrrhini: Primates) and species belonging to New World and African Hystricognaths (Caviomorpha and Bathyergidae: Rodents) along with Cetacea and Ruminantia (Cetartiodactyla). Among the hominoids, interestingly, three out of four genera of gibbons have lost normal SIRH11/ZCCHC16 function by deletion or the lack of the CCHC RNA-binding domain. Our extensive dN/dS analysis suggests that such truncated SIRH11/ZCCHC16 ORFs are functionally diversified even within lineages. Combined, our results show that SIRH11/ZCCHC16 may contribute to the diversification of eutherians by lineage-specific structural changes after its domestication in the common eutherian ancestor, followed by putative species-specific functional changes that enhanced fitness and occurred as a consequence of complex natural selection events

  11. Highly Synchronized Expression of Lineage-Specific Genes during In Vitro Hepatic Differentiation of Human Pluripotent Stem Cell Lines.

    PubMed

    Ghosheh, Nidal; Olsson, Björn; Edsbagge, Josefina; Küppers-Munther, Barbara; Van Giezen, Mariska; Asplund, Annika; Andersson, Tommy B; Björquist, Petter; Carén, Helena; Simonsson, Stina; Sartipy, Peter; Synnergren, Jane

    2016-01-01

    Human pluripotent stem cells- (hPSCs-) derived hepatocytes have the potential to replace many hepatic models in drug discovery and provide a cell source for regenerative medicine applications. However, the generation of fully functional hPSC-derived hepatocytes is still a challenge. Towards gaining better understanding of the differentiation and maturation process, we employed a standardized protocol to differentiate six hPSC lines into hepatocytes and investigated the synchronicity of the hPSC lines by applying RT-qPCR to assess the expression of lineage-specific genes (OCT4, NANOG, T, SOX17, CXCR4, CER1, HHEX, TBX3, PROX1, HNF6, AFP, HNF4a, KRT18, ALB, AAT, and CYP3A4) which serve as markers for different stages during liver development. The data was evaluated using correlation and clustering analysis, demonstrating that the expression of these markers is highly synchronized and correlated well across all cell lines. The analysis also revealed a distribution of the markers in groups reflecting the developmental stages of hepatocytes. Functional analysis of the differentiated cells further confirmed their hepatic phenotype. Taken together, these results demonstrate, on the molecular level, the highly synchronized differentiation pattern across multiple hPSC lines. Moreover, this study provides additional understanding for future efforts to improve the functionality of hPSC-derived hepatocytes and thereby increase the value of related models. PMID:26949401

  12. Highly Synchronized Expression of Lineage-Specific Genes during In Vitro Hepatic Differentiation of Human Pluripotent Stem Cell Lines

    PubMed Central

    Ghosheh, Nidal; Olsson, Björn; Edsbagge, Josefina; Küppers-Munther, Barbara; Van Giezen, Mariska; Asplund, Annika; Andersson, Tommy B.; Björquist, Petter; Carén, Helena; Simonsson, Stina; Sartipy, Peter; Synnergren, Jane

    2016-01-01

    Human pluripotent stem cells- (hPSCs-) derived hepatocytes have the potential to replace many hepatic models in drug discovery and provide a cell source for regenerative medicine applications. However, the generation of fully functional hPSC-derived hepatocytes is still a challenge. Towards gaining better understanding of the differentiation and maturation process, we employed a standardized protocol to differentiate six hPSC lines into hepatocytes and investigated the synchronicity of the hPSC lines by applying RT-qPCR to assess the expression of lineage-specific genes (OCT4, NANOG, T, SOX17, CXCR4, CER1, HHEX, TBX3, PROX1, HNF6, AFP, HNF4a, KRT18, ALB, AAT, and CYP3A4) which serve as markers for different stages during liver development. The data was evaluated using correlation and clustering analysis, demonstrating that the expression of these markers is highly synchronized and correlated well across all cell lines. The analysis also revealed a distribution of the markers in groups reflecting the developmental stages of hepatocytes. Functional analysis of the differentiated cells further confirmed their hepatic phenotype. Taken together, these results demonstrate, on the molecular level, the highly synchronized differentiation pattern across multiple hPSC lines. Moreover, this study provides additional understanding for future efforts to improve the functionality of hPSC-derived hepatocytes and thereby increase the value of related models. PMID:26949401

  13. Lineage-specific STAT5 target gene activation in hematopoietic progenitor cells predicts the FLT3(+)-mediated leukemic phenotype.

    PubMed

    Müller, T A; Grundler, R; Istvanffy, R; Rudelius, M; Hennighausen, L; Illert, A L; Duyster, J

    2016-08-01

    Mutations that activate FMS-like tyrosine kinase 3 (FLT3) are frequent occurrences in acute myeloid leukemia. Two distinct types of mutations have been described: internal duplication of the juxtamembranous domain (ITD) and point mutations of the tyrosine kinase domain (TKD). Although both mutations lead to constitutive FLT3 signaling, only FLT3-ITD strongly activates signal transducer and activator of transcription 5 (STAT5). In a murine transplantation model, FLT3-ITD induces a myeloproliferative neoplasm, whereas FLT3-TKD leads to a lymphoid malignancy with significantly longer latency. Here we report that the presence of STAT5 is critical for the development of a myeloproliferative disease by FLT3-ITD in mice. Deletion of Stat5 in FLT3-ITD-induced leukemogenesis leads not only to a significantly longer survival (82 vs 27 days) of the diseased mice, but also to an immunophenotype switch with expansion of the lymphoid cell compartment. Interestingly, we were able to show differential STAT5 activation in FLT3-ITD(+) myeloid and lymphoid murine progenitors. STAT5 target genes such as Oncostatin M were highly expressed in FLT3-ITD(+) myeloid but not in FLT3-ITD(+) lymphoid progenitor cells. Strikingly, FLT3-TKD expression in combination with Oncostatin M is sufficient to reverse the phenotype to a myeloproliferative disease in FLT3-TKD mice. Thus, lineage-specific STAT5 activation in hematopoietic progenitor cells predicts the FLT3(+)-mediated leukemic phenotype in mice. PMID:27046463

  14. Neuroblast lineage identification and lineage-specific Hox gene action during postembryonic development of the subesophageal ganglion in the Drosophila central brain.

    PubMed

    Kuert, Philipp A; Hartenstein, Volker; Bello, Bruno C; Lovick, Jennifer K; Reichert, Heinrich

    2014-06-15

    The central brain of Drosophila consists of the supraesophageal ganglion (SPG) and the subesophageal ganglion (SEG), both of which are generated by neural stem cell-like neuroblasts during embryonic and postembryonic development. Considerable information has been obtained on postembryonic development of the neuroblasts and their lineages in the SPG. In contrast, very little is known about neuroblasts, neural lineages, or any other aspect of the postembryonic development in the SEG. Here we characterize the neuroanatomy of the larval SEG in terms of tracts, commissures, and other landmark features as compared to a thoracic ganglion. We then use clonal MARCM labeling to identify all adult-specific neuroblast lineages in the late larval SEG and find a surprisingly small number of neuroblast lineages, 13 paired and one unpaired. The Hox genes Dfd, Scr, and Antp are expressed in a lineage-specific manner in these lineages during postembryonic development. Hox gene loss-of-function causes lineage-specific defects in axonal targeting and reduction in neural cell numbers. Moreover, it results in the formation of novel ectopic neuroblast lineages. Apoptosis block also results in ectopic lineages suggesting that Hox genes are required for lineage-specific termination of proliferation through programmed cell death. Taken together, our findings show that postembryonic development in the SEG is mediated by a surprisingly small set of identified lineages and requires lineage-specific Hox gene action to ensure the correct formation of adult-specific neurons in the Drosophila brain. PMID:24713419

  15. Characterization of MAT gene functions in the life cycle of Sclerotinia sclerotiorum reveals a lineage-specific MAT gene functioning in apothecium morphogenesis.

    PubMed

    Doughan, Benjamin; Rollins, Jeffrey A

    2016-09-01

    Sclerotinia sclerotiorum (Lib.) de Bary is a phytopathogenic fungus that relies on the completion of the sexual cycle to initiate aerial infections. The sexual cycle produces apothecia required for inoculum dispersal. In this study, insight into the regulation of apothecial multicellular development was pursued through functional characterization of mating-type genes. These genes are hypothesized to encode master regulatory proteins required for aspects of sexual development ranging from fertilization through fertile fruiting body development. Experimentally, loss-of-function mutants were created for the conserved core mating-type genes (MAT1-1-1, and MAT1-2-1), and the lineage-specific genes found only in S. sclerotiorum and closely related fungi (MAT1-1-5, and MAT1-2-4). The MAT1-1-1, MAT1-1-5, and MAT1-2-1 mutants are able to form ascogonia but are blocked in all aspects of apothecium development. These mutants also exhibit defects in secondary sexual characters including lower numbers of spermatia. The MAT1-2-4 mutants are delayed in carpogenic germination accompanied with altered disc morphogenesis and ascospore production. They too produce lower numbers of spermatia. All four MAT gene mutants showed alterations in the expression of putative pheromone precursor (Ppg-1) and pheromone receptor (PreA, PreB) genes. Our findings support the involvement of MAT genes in sexual fertility, gene regulation, meiosis, and morphogenesis in S. sclerotiorum. PMID:27567717

  16. An analysis of the gene complement of a marsupial, Monodelphis domestica: Evolution of lineage-specific genes and giant chromosomes

    PubMed Central

    Goodstadt, Leo; Heger, Andreas; Webber, Caleb; Ponting, Chris P.

    2007-01-01

    The newly sequenced genome of Monodelphis domestica not only provides the out-group necessary to better understand our own eutherian lineage, but it enables insights into the innovative biology of metatherians. Here, we compare Monodelphis with Homo sequences from alignments of single nucleotides, genes, and whole chromosomes. Using PhyOP, we have established orthologs in Homo for 82% (15,250) of Monodelphis gene predictions. Those with single orthologs in each species exhibited a high median synonymous substitution rate (dS = 1.02), thereby explaining the relative paucity of aligned regions outside of coding sequences. Orthology assignments were used to construct a synteny map that illustrates the considerable fragmentation of Monodelphis and Homo karyotypes since their therian last common ancestor. Fifteen percent of Monodelphis genes are predicted, from their low divergence at synonymous sites, to have been duplicated in the metatherian lineage. The majority of Monodelphis-specific genes possess predicted roles in chemosensation, reproduction, adaptation to specific diets, and immunity. Using alignments of Monodelphis genes to sequences from either Homo or Trichosurus vulpecula (an Australian marsupial), we show that metatherian X chromosomes have elevated silent substitution rates and high G+C contents in comparison with both metatherian autosomes and eutherian chromosomes. Each of these elevations is also a feature of subtelomeric chromosomal regions. We attribute these observations to high rates of female-specific recombination near the chromosomal ends and within the X chromosome, which act to sustain or increase G+C levels by biased gene conversion. In particular, we propose that the higher G+C content of the Monodelphis X chromosome is a direct consequence of its small size relative to the giant autosomes. PMID:17495010

  17. Gain of a New Exon by a Lineage-Specific Alu Element-Integration Event in the BCS1L Gene during Primate Evolution

    PubMed Central

    Park, Sang-Je; Kim, Young-Hyun; Lee, Sang-Rae; Choe, Se-Hee; Kim, Myung-Jin; Kim, Sun-Uk; Kim, Ji-Su; Sim, Bo-Woong; Song, Bong-Seok; Jeong, Kang-Jin; Jin, Yeung-Bae; Lee, Youngjeon; Park, Young-Ho; Park, Young Il; Huh, Jae-Won; Chang, Kyu-Tae

    2015-01-01

    BCS1L gene encodes mitochondrial protein and is a member of conserved AAA protein family. This gene is involved in the incorporation of Rieske FeS and Qcr10p into complex III of respiratory chain. In our previous study, AluYRa2-derived alternative transcript in rhesus monkey genome was identified. However, this transcript has not been reported in human genome. In present study, we conducted evolutionary analysis of AluYRa2-exonized transcript with various primate genomic DNAs and cDNAs from humans, rhesus monkeys, and crab-eating monkeys. Remarkably, our results show that AluYRa2 element has only been integrated into genomes of Macaca species. This Macaca lineage-specific integration of AluYRa2 element led to exonization event in the first intron region of BCS1L gene by producing a conserved 3′ splice site. Intriguingly, in rhesus and crab-eating monkeys, more diverse transcript variants by alternative splicing (AS) events, including exon skipping and different 5′ splice sites from humans, were identified. Alignment of amino acid sequences revealed that AluYRa2-exonized transcript has short N-terminal peptides. Therefore, AS events play a major role in the generation of various transcripts and proteins during primate evolution. In particular, lineage-specific integration of Alu elements and species-specific Alu-derived exonization events could be important sources of gene diversification in primates. PMID:26537194

  18. Expansion of banana (Musa acuminata) gene families involved in ethylene biosynthesis and signalling after lineage-specific whole-genome duplications.

    PubMed

    Jourda, Cyril; Cardi, Céline; Mbéguié-A-Mbéguié, Didier; Bocs, Stéphanie; Garsmeur, Olivier; D'Hont, Angélique; Yahiaoui, Nabila

    2014-05-01

    Whole-genome duplications (WGDs) are widespread in plants, and three lineage-specific WGDs occurred in the banana (Musa acuminata) genome. Here, we analysed the impact of WGDs on the evolution of banana gene families involved in ethylene biosynthesis and signalling, a key pathway for banana fruit ripening. Banana ethylene pathway genes were identified using comparative genomics approaches and their duplication modes and expression profiles were analysed. Seven out of 10 banana ethylene gene families evolved through WGD and four of them (1-aminocyclopropane-1-carboxylate synthase (ACS), ethylene-insensitive 3-like (EIL), ethylene-insensitive 3-binding F-box (EBF) and ethylene response factor (ERF)) were preferentially retained. Banana orthologues of AtEIN3 and AtEIL1, two major genes for ethylene signalling in Arabidopsis, were particularly expanded. This expansion was paralleled by that of EBF genes which are responsible for control of EIL protein levels. Gene expression profiles in banana fruits suggested functional redundancy for several MaEBF and MaEIL genes derived from WGD and subfunctionalization for some of them. We propose that EIL and EBF genes were co-retained after WGD in banana to maintain balanced control of EIL protein levels and thus avoid detrimental effects of constitutive ethylene signalling. In the course of evolution, subfunctionalization was favoured to promote finer control of ethylene signalling. PMID:24716518

  19. Phylogenetic Analysis, Lineage-Specific Expansion and Functional Divergence of seed dormancy 4-Like Genes in Plants

    PubMed Central

    Subburaj, Saminathan; Cao, Shuanghe; Xia, Xianchun; He, Zhonghu

    2016-01-01

    The rice gene seed dormancy 4 (OsSdr4) functions in seed dormancy and is a major factor associated with pre-harvest sprouting (PHS). Although previous studies of this protein family were reported for rice and other species, knowledge of the evolution of genes homologous to OsSdr4 in plants remains inadequate. Fifty four Sdr4-like (hereafter designated Sdr4L) genes were identified in nine plant lineages including 36 species. Phylogenetic analysis placed these genes in eight subfamilies (I-VIII). Genes from the same lineage clustered together, supported by analysis of conserved motifs and exon-intron patterns. Segmental duplications were present in both dicot and monocot clusters, while tandemly duplicated genes occurred only in monocot clusters indicating that both tandem and segmental duplications contributed to expansion of the grass I and II subfamilies. Estimation of the approximate ages of the duplication events indicated that ancestral Sdr4 genes evolved from a common angiosperm ancestor, about 160 million years ago (MYA). Moreover, diversification of Sdr4L genes in mono and dicot plants was mainly associated with genome-wide duplication and speciation events. Functional divergence was observed in all subfamily pairs, except IV/VIIIa. Further analysis indicated that functional constraints between subfamily pairs I/II, I/VIIIb, II/VI, II/VIIIb, II/IV, and VI/VIIIb were statistically significant. Site and branch-site model analyses of positive selection suggested that these genes were under strong adaptive selection pressure. Critical amino acids detected for both functional divergence and positive selection were mostly located in the loops, pointing to functional importance of these regions in this protein family. In addition, differential expression studies by transcriptome atlas of 11 Sdr4L genes showed that the duplicated genes may have undergone divergence in expression between plant species. Our findings showed that Sdr4L genes are functionally divergent

  20. Analysis of expression in the Anopheles gambiae developing testes reveals rapidly evolving lineage-specific genes in mosquitoes

    PubMed Central

    Krzywinska, Elzbieta; Krzywinski, Jaroslaw

    2009-01-01

    Background Male mosquitoes do not feed on blood and are not involved in delivery of pathogens to humans. Consequently, they are seldom the subjects of research, which results in a very poor understanding of their biology. To gain insights into male developmental processes we sought to identify genes transcribed exclusively in the reproductive tissues of male Anopheles gambiae pupae. Results Using a cDNA subtraction strategy, five male-specifically or highly male-biased expressed genes were isolated, four of which remain unannotated in the An. gambiae genome. Spatial and temporal expression patterns suggest that each of these genes is involved in the mid-late stages of spermatogenesis. Their sequences are rapidly evolving; however, two genes possess clear homologs in a wide range of taxa and one of these probably acts in a sperm motility control mechanism conserved in many organisms, including humans. The other three genes have no match to sequences from non-mosquito taxa, thus can be regarded as orphans. RNA in situ hybridization demonstrated that one of the orphans is transcribed in spermatids, which suggests its involvement in sperm maturation. Two other orphans have unknown functions. Expression analysis of orthologs of all five genes indicated that male-biased transcription was not conserved in the majority of cases in Aedes and Culex. Conclusion Discovery of testis-expressed orphan genes in mosquitoes opens new prospects for the development of innovative control methods. The orphan encoded proteins may represent unique targets of selective anti-mosquito sterilizing agents that will not affect non-target organisms. PMID:19580678

  1. Gene Duplication, Lineage-Specific Expansion, and Subfunctionalization in the MADF-BESS Family Patterns the Drosophila Wing Hinge

    PubMed Central

    Shukla, Vallari; Habib, Farhat; Kulkarni, Apurv; Ratnaparkhi, Girish S.

    2014-01-01

    Gene duplication, expansion, and subsequent diversification are features of the evolutionary process. Duplicated genes can be lost, modified, or altered to generate novel functions over evolutionary timescales. These features make gene duplication a powerful engine of evolutionary change. In this study, we explore these features in the MADF-BESS family of transcriptional regulators. In Drosophila melanogaster, the family contains 16 similar members, each containing an N-terminal, DNA-binding MADF domain and a C-terminal, protein-interacting, BESS domain. Phylogenetic analysis shows that members of the MADF-BESS family are expanded in the Drosophila lineage. Three members, which we name hinge1, hinge2, and hinge3 are required for wing development, with a critical role in the wing hinge. hinge1 is a negative regulator of Winglesss expression and interacts with core wing-hinge patterning genes such as teashirt, homothorax, and jing. Double knockdowns along with heterologous rescue experiments are used to demonstrate that members of the MADF-BESS family retain function in the wing hinge, in spite of expansion and diversification for over 40 million years. The wing hinge connects the blade to the thorax and has critical roles in fluttering during flight. MADF-BESS family genes appear to retain redundant functions to shape and form elements of the wing hinge in a robust and fail-safe manner. PMID:24336749

  2. The Evaluation of Nerve Growth Factor Over Expression on Neural Lineage Specific Genes in Human Mesenchymal Stem Cells

    PubMed Central

    Mortazavi, Yousef; Sheikhsaran, Fatemeh; Khamisipour, Gholamreza Khamisipour; Soleimani, Masoud; Teimuri, Ali; Shokri, Somayeh

    2016-01-01

    Objective Treatment and repair of neurodegenerative diseases such as brain tumors, spinal cord injuries, and functional disorders, including Alzheimer’s disease, are challenging problems. A common treatment approach for such disorders involves the use of mesenchymal stem cells (MSCs) as an alternative cell source to replace injured cells. However, use of these cells in hosts may potentially cause adverse outcomes such as tumorigenesis and uncontrolled differentiation. In attempt to generate mesenchymal derived neural cells, we have infected MSCs with recombinant lentiviruses that expressed nerve growth factor (NGF) and assessed their neural lineage genes. Materials and Methods In this experimental study, we cloned the NGF gene sequence into a helper dependent lentiviral vector that contained the green fluorescent protein (GFP) gene. The recombinant vector was amplified in DH5 bacterial cells. Recombinant viruses were generated in the human embryonic kidney 293 (HEK-293) packaging cell line with the helper vectors and analyzed under fluorescent microscopy. Bone marrow mesenchymal cells were infected by recombinant viruses for three days followed by assessment of neural differentiation. We evaluated expression of NGF through measurement of the NGF protein in culture medium by ELISA; neural specific genes were quantified by real-time polymerase chain reaction (PCR). Results We observed neural morphological changes after three days. Quantitative PCR showed that expressions of NESTIN, glial derived neurotrophic factor (GDNF), glial fibrillary acidic protein (GFAP) and Microtubule-associated protein 2 (MAP2) genes increased following induction of NGF overexpression, whereas expressions of endogenous NGF and brain derived neural growth factor (BDNF) genes reduced. Conclusion Ectopic expression of NGF can induce neurogenesis in MSCs. Direct injection of MSCs may cause tumorigenesis and an undesirable outcome. Therefore an alternative choice to overcome this obstacle may

  3. Gene Loss and Lineage-Specific Restriction-Modification Systems Associated with Niche Differentiation in the Campylobacter jejuni Sequence Type 403 Clonal Complex

    PubMed Central

    Morley, Laura; McNally, Alan; Paszkiewicz, Konrad; Corander, Jukka; Méric, Guillaume; Sheppard, Samuel K.; Blom, Jochen

    2015-01-01

    Campylobacter jejuni is a highly diverse species of bacteria commonly associated with infectious intestinal disease of humans and zoonotic carriage in poultry, cattle, pigs, and other animals. The species contains a large number of distinct clonal complexes that vary from host generalist lineages commonly found in poultry, livestock, and human disease cases to host-adapted specialized lineages primarily associated with livestock or poultry. Here, we present novel data on the ST403 clonal complex of C. jejuni, a lineage that has not been reported in avian hosts. Our data show that the lineage exhibits a distinctive pattern of intralineage recombination that is accompanied by the presence of lineage-specific restriction-modification systems. Furthermore, we show that the ST403 complex has undergone gene decay at a number of loci. Our data provide a putative link between the lack of association with avian hosts of C. jejuni ST403 and both gene gain and gene loss through nonsense mutations in coding sequences of genes, resulting in pseudogene formation. PMID:25795671

  4. Lentiviral Delivery of RNAi for In Vivo Lineage-Specific Modulation of Gene Expression in Mouse Lung Macrophages

    PubMed Central

    Wilson, Andrew A; Kwok, Letty W; Porter, Emily L; Payne, Julie G; McElroy, Gregory S; Ohle, Sarah J; Greenhill, Sara R; Blahna, Matthew T; Yamamoto, Kazuko; Jean, Jyh C; Mizgerd, Joseph P; Kotton, Darrell N

    2013-01-01

    Although RNA interference (RNAi) has become a ubiquitous laboratory tool since its discovery 12 years ago, in vivo delivery to selected cell types remains a major technical challenge. Here, we report the use of lentiviral vectors for long-term in vivo delivery of RNAi selectively to resident alveolar macrophages (AMs), key immune effector cells in the lung. We demonstrate the therapeutic potential of this approach by RNAi-based downregulation of p65 (RelA), a component of the pro-inflammatory transcriptional regulator, nuclear factor κB (NF-κB) and a key participant in lung disease pathogenesis. In vivo RNAi delivery results in decreased induction of NF-κB and downstream neutrophilic chemokines in transduced AMs as well as attenuated lung neutrophilia following stimulation with lipopolysaccharide (LPS). Through concurrent delivery of a novel lentiviral reporter vector (lenti-NF-κB-luc-GFP) we track in vivo expression of NF-κB target genes in real time, a critical step towards extending RNAi-based therapy to longstanding lung diseases. Application of this system reveals that resident AMs persist in the airspaces of mice following the resolution of LPS-induced inflammation, thus allowing these localized cells to be used as effective vehicles for prolonged RNAi delivery in disease settings. PMID:23403494

  5. Multi-species sequence comparison reveals dynamic evolution of the elastin gene that has involved purifying selection and lineage-specific insertions/deletions

    PubMed Central

    Piontkivska, Helen; Zhang, Yi; Green, Eric D; Elnitski, Laura

    2004-01-01

    Background The elastin gene (ELN) is implicated as a factor in both supravalvular aortic stenosis (SVAS) and Williams Beuren Syndrome (WBS), two diseases involving pronounced complications in mental or physical development. Although the complete spectrum of functional roles of the processed gene product remains to be established, these roles are inferred to be analogous in human and mouse. This view is supported by genomic sequence comparison, in which there are no large-scale differences in the ~1.8 Mb sequence block encompassing the common region deleted in WBS, with the exception of an overall reversed physical orientation between human and mouse. Results Conserved synteny around ELN does not translate to a high level of conservation in the gene itself. In fact, ELN orthologs in mammals show more sequence divergence than expected for a gene with a critical role in development. The pattern of divergence is non-conventional due to an unusually high ratio of gaps to substitutions. Specifically, multi-sequence alignments of eight mammalian sequences reveal numerous non-aligning regions caused by species-specific insertions and deletions, in spite of the fact that the vast majority of aligning sites appear to be conserved and undergoing purifying selection. Conclusions The pattern of lineage-specific, in-frame insertions/deletions in the coding exons of ELN orthologous genes is unusual and has led to unique features of the gene in each lineage. These differences may indicate that the gene has a slightly different functional mechanism in mammalian lineages, or that the corresponding regions are functionally inert. Identified regions that undergo purifying selection reflect a functional importance associated with evolutionary pressure to retain those features. PMID:15149554

  6. Step-wise and lineage-specific diversification of plant RNA polymerase genes and origin of the largest plant-specific subunits.

    PubMed

    Wang, Yaqiong; Ma, Hong

    2015-09-01

    Proteins often function as complexes, yet little is known about the evolution of dissimilar subunits of complexes. DNA-directed RNA polymerases (RNAPs) are multisubunit complexes, with distinct eukaryotic types for different classes of transcripts. In addition to Pol I-III, common in eukaryotes, plants have Pol IV and V for epigenetic regulation. Some RNAP subunits are specific to one type, whereas other subunits are shared by multiple types. We have conducted extensive phylogenetic and sequence analyses, and have placed RNAP gene duplication events in land plant history, thereby reconstructing the subunit compositions of the novel RNAPs during land plant evolution. We found that Pol IV/V have experienced step-wise duplication and diversification of various subunits, with increasingly distinctive subunit compositions. Also, lineage-specific duplications have further increased RNAP complexity with distinct copies in different plant families and varying divergence for subunits of different RNAPs. Further, the largest subunits of Pol IV/V probably originated from a gene fusion in the ancestral land plants. We propose a framework of plant RNAP evolution, providing an excellent model for protein complex evolution. PMID:25921392

  7. Evolutionary Genomics Reveals Lineage-Specific Gene Loss and Rapid Evolution of a Sperm-Specific Ion Channel Complex: CatSpers and CatSperβ

    PubMed Central

    Cai, Xinjiang; Clapham, David E.

    2008-01-01

    The mammalian CatSper ion channel family consists of four sperm-specific voltage-gated Ca2+ channels that are crucial for sperm hyperactivation and male fertility. All four CatSper subunits are believed to assemble into a heteromultimeric channel complex, together with an auxiliary subunit, CatSperβ. Here, we report a comprehensive comparative genomics study and evolutionary analysis of CatSpers and CatSperβ, with important correlation to physiological significance of molecular evolution of the CatSper channel complex. The development of the CatSper channel complex with four CatSpers and CatSperβ originated as early as primitive metazoans such as the Cnidarian Nematostella vectensis. Comparative genomics revealed extensive lineage-specific gene loss of all four CatSpers and CatSperβ through metazoan evolution, especially in vertebrates. The CatSper channel complex underwent rapid evolution and functional divergence, while distinct evolutionary constraints appear to have acted on different domains and specific sites of the four CatSper genes. These results reveal unique evolutionary characteristics of sperm-specific Ca2+ channels and their adaptation to sperm biology through metazoan evolution. PMID:18974790

  8. Lineage-specific expansions of TET/JBP genes and a new class of DNA transposons shape fungal genomic and epigenetic landscapes

    PubMed Central

    Iyer, Lakshminarayan M.; Zhang, Dapeng; de Souza, Robson F.; Pukkila, Patricia J.; Rao, Anjana; Aravind, L.

    2014-01-01

    TET/JBP dioxygenases oxidize methylpyrimidines in nucleic acids and are implicated in generation of epigenetic marks and potential intermediates for DNA demethylation. We show that TET/JBP genes are lineage-specifically expanded in all major clades of basidiomycete fungi, with the majority of copies predicted to encode catalytically active proteins. This pattern differs starkly from the situation in most other organisms that possess just a single or a few copies of the TET/JBP family. In most basidiomycetes, TET/JBP genes are frequently linked to a unique class of transposons, KDZ (Kyakuja, Dileera, and Zisupton) and appear to have dispersed across chromosomes along with them. Several of these elements typically encode additional proteins, including a divergent version of the HMG domain. Analysis of their transposases shows that they contain a previously uncharacterized version of the RNase H fold with multiple distinctive Zn-chelating motifs and a unique insert, which are predicted to play roles in structural stabilization and target sequence recognition, respectively. We reconstruct the complex evolutionary history of TET/JBPs and associated transposons as involving multiple rounds of expansion with concomitant lineage sorting and loss, along with several capture events of TET/JBP genes by different transposon clades. On a few occasions, these TET/JBP genes were also laterally transferred to certain Ascomycota, Glomeromycota, Viridiplantae, and Amoebozoa. One such is an inactive version, calnexin-independence factor 1 (Cif1), from Schizosaccharomyces pombe, which has been implicated in inducing an epigenetically transmitted prion state. We argue that this unique transposon-TET/JBP association is likely to play important roles in speciation during evolution and epigenetic regulation. PMID:24398522

  9. Histone Demethylase Jumonji AT-rich Interactive Domain 1B (JARID1B) Controls Mammary Gland Development by Regulating Key Developmental and Lineage Specification Genes*

    PubMed Central

    Zou, Mike Ran; Cao, Jian; Liu, Zongzhi; Huh, Sung Jin; Polyak, Kornelia; Yan, Qin

    2014-01-01

    The JmjC domain-containing H3K4 histone demethylase jumonji AT-rich interactive domain 1B (JARID1B) (also known as KDM5B and PLU1) is overexpressed in breast cancer and is a potential target for breast cancer treatment. To investigate the in vivo function of JARID1B, we developed Jarid1b−/− mice and characterized their phenotypes in detail. Unlike previously reported Jarid1b−/− strains, the majority of these Jarid1b−/− mice were viable beyond embryonic and neonatal stages. This allowed us to further examine phenotypes associated with the loss of JARID1B in pubertal development and pregnancy. These Jarid1b−/− mice exhibited decreased body weight, premature mortality, decreased female fertility, and delayed mammary gland development. Related to these phenotypes, JARID1B loss decreased serum estrogen level and reduced mammary epithelial cell proliferation in early puberty. In mammary epithelial cells, JARID1B loss diminished the expression of key regulators for mammary morphogenesis and luminal lineage specification, including FOXA1 and estrogen receptor α. Mechanistically, JARID1B was required for GATA3 recruitment to the Foxa1 promoter to activate Foxa1 expression. These results indicate that JARID1B positively regulates mammary ductal development through both extrinsic and cell-autonomous mechanisms. PMID:24802759

  10. Two C or not two C: recurrent disruption of Zn-ribbons, gene duplication, lineage-specific gene loss, and horizontal gene transfer in evolution of bacterial ribosomal proteins

    PubMed Central

    Makarova, Kira S; Ponomarev, Vladimir A; Koonin, Eugene V

    2001-01-01

    + and C- forms in different lineages. Additionally, evidence was obtained for a role of horizontal gene transfer in the evolution of these ribosomal proteins, with multiple cases of gene displacement 'in situ', that is, without a change of the gene order in the recipient genome. Conclusions A more complex picture of evolution of bacterial ribosomal proteins than previously suspected is emerging from these results, with major contributions of lineage-specific gene loss and horizontal gene transfer. The recurrent theme of emergence and disruption of Zn-ribbons in bacterial ribosomal proteins awaits a functional interpretation. PMID:11574053

  11. Sequencing analysis of 20,000 full-length cDNA clones from cassava reveals lineage specific expansions in gene families related to stress response

    PubMed Central

    Sakurai, Tetsuya; Plata, Germán; Rodríguez-Zapata, Fausto; Seki, Motoaki; Salcedo, Andrés; Toyoda, Atsushi; Ishiwata, Atsushi; Tohme, Joe; Sakaki, Yoshiyuki; Shinozaki, Kazuo; Ishitani, Manabu

    2007-01-01

    Background Cassava, an allotetraploid known for its remarkable tolerance to abiotic stresses is an important source of energy for humans and animals and a raw material for many industrial processes. A full-length cDNA library of cassava plants under normal, heat, drought, aluminum and post harvest physiological deterioration conditions was built; 19968 clones were sequence-characterized using expressed sequence tags (ESTs). Results The ESTs were assembled into 6355 contigs and 9026 singletons that were further grouped into 10577 scaffolds; we found 4621 new cassava sequences and 1521 sequences with no significant similarity to plant protein databases. Transcripts of 7796 distinct genes were captured and we were able to assign a functional classification to 78% of them while finding more than half of the enzymes annotated in metabolic pathways in Arabidopsis. The annotation of sequences that were not paired to transcripts of other species included many stress-related functional categories showing that our library is enriched with stress-induced genes. Finally, we detected 230 putative gene duplications that include key enzymes in reactive oxygen species signaling pathways and could play a role in cassava stress response features. Conclusion The cassava full-length cDNA library here presented contains transcripts of genes involved in stress response as well as genes important for different areas of cassava research. This library will be an important resource for gene discovery, characterization and cloning; in the near future it will aid the annotation of the cassava genome. PMID:18096061

  12. CopyRighter: a rapid tool for improving the accuracy of microbial community profiles through lineage-specific gene copy number correction

    PubMed Central

    2014-01-01

    Background Culture-independent molecular surveys targeting conserved marker genes, most notably 16S rRNA, to assess microbial diversity remain semi-quantitative due to variations in the number of gene copies between species. Results Based on 2,900 sequenced reference genomes, we show that 16S rRNA gene copy number (GCN) is strongly linked to microbial phylogenetic taxonomy, potentially under-representing Archaea in amplicon microbial profiles. Using this relationship, we inferred the GCN of all bacterial and archaeal lineages in the Greengenes database within a phylogenetic framework. We created CopyRighter, new software which uses these estimates to correct 16S rRNA amplicon microbial profiles and associated quantitative (q)PCR total abundance. CopyRighter parses microbial profiles and, because GCN estimates are pre-computed for all taxa in the reference taxonomy, rapidly corrects GCN bias. Software validation with in silico and in vitro mock communities indicated that GCN correction results in more accurate estimates of microbial relative abundance and improves the agreement between metagenomic and amplicon profiles. Analyses of human-associated and anaerobic digester microbiomes illustrate that correction makes tangible changes to estimates of qPCR total abundance, α and β diversity, and can significantly change biological interpretation. For example, human gut microbiomes from twins were reclassified into three rather than two enterotypes after GCN correction. Conclusions The CopyRighter bioinformatic tools permits rapid correction of GCN in microbial surveys, resulting in improved estimates of microbial abundance, α and β diversity. PMID:24708850

  13. Matrix elasticity directs stem cell lineage specification

    NASA Astrophysics Data System (ADS)

    Discher, Dennis

    2010-03-01

    Adhesion of stem cells - like most cells - is not just a membrane phenomenon. Most tissue cells need to adhere to a ``solid'' for viability, and over the last decade it has become increasingly clear that the physical ``elasticity'' of that solid is literally ``felt'' by cells. Here we show that Mesenchymal Stem Cells (MSCs) specify lineage and commit to phenotypes with extreme sensitivity to the elasticity typical of tissues [1]. In serum only media, soft matrices that mimic brain appear neurogenic, stiffer matrices that mimic muscle are myogenic, and comparatively rigid matrices that mimic collagenous bone prove osteogenic. Inhibition of nonmuscle myosin II activity blocks all elasticity directed lineage specification, which indicates that the cytoskeleton pulls on matrix through adhesive attachments. Results have significant implications for `therapeutic' stem cells and have motivated development of a proteomic-scale method to identify mechano-responsive protein structures [2] as well as deeper physical studies of matrix physics [3] and growth factor pathways [4]. [4pt] [1] A. Engler, et al. Matrix elasticity directs stem cell lineage specification. Cell (2006).[0pt] [2] C.P. Johnson, et al. Forced unfolding of proteins within cells. Science (2007).[0pt] [3] A.E.X. Brown, et al. Multiscale mechanics of fibrin polymer: Gel stretching with protein unfolding and loss of water. Science (2009).[0pt] [4] D.E. Discher, et al. Growth factors, matrices, and forces combine and control stem cells. Science (2009).

  14. Lineage-Specific Conserved Noncoding Sequences of Plant Genomes: Their Possible Role in Nucleosome Positioning

    PubMed Central

    Hettiarachchi, Nilmini; Kryukov, Kirill; Sumiyama, Kenta; Saitou, Naruya

    2014-01-01

    Many studies on conserved noncoding sequences (CNSs) have found that CNSs are enriched significantly in regulatory sequence elements. We conducted whole-genome analysis on plant CNSs to identify lineage-specific CNSs in eudicots, monocots, angiosperms, and vascular plants based on the premise that lineage-specific CNSs define lineage-specific characters and functions in groups of organisms. We identified 27 eudicot, 204 monocot, 6,536 grass, 19 angiosperm, and 2 vascular plant lineage-specific CNSs (lengths range from 16 to 1,517 bp) that presumably originated in their respective common ancestors. A stronger constraint on the CNSs located in the untranslated regions was observed. The CNSs were often flanked by genes involved in transcription regulation. A drop of A+T content near the border of CNSs was observed and CNS regions showed a higher nucleosome occupancy probability. These CNSs are candidate regulatory elements, which are expected to define lineage-specific features of various plant groups. PMID:25364802

  15. Genetic and epigenetic variation in the lineage specification of regulatory T cells

    PubMed Central

    Arvey, Aaron; van der Veeken, Joris; Plitas, George; Rich, Stephen S; Concannon, Patrick; Rudensky, Alexander Y

    2015-01-01

    Regulatory T (Treg) cells, which suppress autoimmunity and other inflammatory states, are characterized by a distinct set of genetic elements controlling their gene expression. However, the extent of genetic and associated epigenetic variation in the Treg cell lineage and its possible relation to disease states in humans remain unknown. We explored evolutionary conservation of regulatory elements and natural human inter-individual epigenetic variation in Treg cells to identify the core transcriptional control program of lineage specification. Analysis of single nucleotide polymorphisms in core lineage-specific enhancers revealed disease associations, which were further corroborated by high-resolution genotyping to fine map causal polymorphisms in lineage-specific enhancers. Our findings suggest that a small set of regulatory elements specify the Treg lineage and that genetic variation in Treg cell-specific enhancers may alter Treg cell function contributing to polygenic disease. DOI: http://dx.doi.org/10.7554/eLife.07571.001 PMID:26510014

  16. Social regulation of gene expression in human leukocytes

    PubMed Central

    Cole, Steve W; Hawkley, Louise C; Arevalo, Jesusa M; Sung, Caroline Y; Rose, Robert M; Cacioppo, John T

    2007-01-01

    Background Social environmental influences on human health are well established in the epidemiology literature, but their functional genomic mechanisms are unclear. The present study analyzed genome-wide transcriptional activity in people who chronically experienced high versus low levels of subjective social isolation (loneliness) to assess alterations in the activity of transcription control pathways that might contribute to increased adverse health outcomes in social isolates. Results DNA microarray analysis identified 209 genes that were differentially expressed in circulating leukocytes from 14 high- versus low-lonely individuals, including up-regulation of genes involved in immune activation, transcription control, and cell proliferation, and down-regulation of genes supporting mature B lymphocyte function and type I interferon response. Promoter-based bioinformatic analyses showed under-expression of genes bearing anti-inflammatory glucocorticoid response elements (GREs; p = 0.032) and over-expression of genes bearing response elements for pro-inflammatory NF-κB/Rel transcription factors (p = 0.011). This reciprocal shift in pro- and anti-inflammatory signaling was not attributable to differences in circulating cortisol levels, or to other demographic, psychological, or medical characteristics. Additional transcription control pathways showing differential activity in bioinformatic analyses included the CREB/ATF, JAK/STAT, IRF1, C/EBP, Oct, and GATA pathways. Conclusion These data provide the first indication that human genome-wide transcriptional activity is altered in association with a social epidemiological risk factor. Impaired transcription of glucocorticoid response genes and increased activity of pro-inflammatory transcription control pathways provide a functional genomic explanation for elevated risk of inflammatory disease in individuals who experience chronically high levels of subjective social isolation. PMID:17854483

  17. Nitric Oxide-cGMP Signaling Stimulates Erythropoiesis through Multiple Lineage-Specific Transcription Factors: Clinical Implications and a Novel Target for Erythropoiesis

    PubMed Central

    Ikuta, Tohru; Sellak, Hassan; Odo, Nadine; Adekile, Adekunle D.; Gaensler, Karin M. L.

    2016-01-01

    Much attention has been directed to the physiological effects of nitric oxide (NO)-cGMP signaling, but virtually nothing is known about its hematologic effects. We reported for the first time that cGMP signaling induces human γ-globin gene expression. Aiming at developing novel therapeutics for anemia, we examined here the hematologic effects of NO-cGMP signaling in vivo and in vitro. We treated wild-type mice with NO to activate soluble guanylate cyclase (sGC), a key enzyme of cGMP signaling. Compared to untreated mice, NO-treated mice had higher red blood cell counts and total hemoglobin but reduced leukocyte counts, demonstrating that when activated, NO-cGMP signaling exerts hematopoietic effects on multiple types of blood cells in vivo. We next generated mice which overexpressed rat sGC in erythroid and myeloid cells. The forced expression of sGCs activated cGMP signaling in both lineage cells. Compared with non-transgenic littermates, sGC mice exhibited hematologic changes similar to those of NO-treated mice. Consistently, a membrane-permeable cGMP enhanced the differentiation of hematopoietic progenitors toward erythroid-lineage cells but inhibited them toward myeloid-lineage cells by controlling multiple lineage-specific transcription factors. Human γ-globin gene expression was induced at low but appreciable levels in sGC mice carrying the human β-globin locus. Together, these results demonstrate that NO-cGMP signaling is capable of stimulating erythropoiesis in both in vitro and vivo settings by controlling the expression of multiple lineage-specific transcription factors, suggesting that cGMP signaling upregulates erythropoiesis at the level of gene transcription. The NO-cGMP signaling axis may constitute a novel target to stimulate erythropoiesis in vivo. PMID:26727002

  18. Mapping the route from naive pluripotency to lineage specification.

    PubMed

    Kalkan, Tüzer; Smith, Austin

    2014-12-01

    In the mouse blastocyst, epiblast cells are newly formed shortly before implantation. They possess a unique developmental plasticity, termed naive pluripotency. For development to proceed, this naive state must be subsumed by multi-lineage differentiation within 72 h following implantation. In vitro differentiation of naive embryonic stem cells (ESCs) cultured in controlled conditions provides a tractable system to dissect and understand the process of exit from naive pluripotency and entry into lineage specification. Exploitation of this system in recent large-scale RNAi and mutagenesis screens has uncovered multiple new factors and modules that drive or facilitate progression out of the naive state. Notably, these studies show that the transcription factor network that governs the naive state is rapidly dismantled prior to upregulation of lineage specification markers, creating an intermediate state that we term formative pluripotency. Here, we summarize these findings and propose a road map for state transitions in ESC differentiation that reflects the orderly dynamics of epiblast progression in the embryo. PMID:25349449

  19. ATTED-II in 2016: A Plant Coexpression Database Towards Lineage-Specific Coexpression

    PubMed Central

    Aoki, Yuichi; Okamura, Yasunobu; Tadaka, Shu; Kinoshita, Kengo; Obayashi, Takeshi

    2016-01-01

    ATTED-II (http://atted.jp) is a coexpression database for plant species with parallel views of multiple coexpression data sets and network analysis tools. The user can efficiently find functional gene relationships and design experiments to identify gene functions by reverse genetics and general molecular biology techniques. Here, we report updates to ATTED-II (version 8.0), including new and updated coexpression data and analysis tools. ATTED-II now includes eight microarray- and six RNA sequencing-based coexpression data sets for seven dicot species (Arabidopsis, field mustard, soybean, barrel medick, poplar, tomato and grape) and two monocot species (rice and maize). Stand-alone coexpression analyses tend to have low reliability. Therefore, examining evolutionarily conserved coexpression is a more effective approach from the viewpoints of reliability and evolutionary importance. In contrast, the reliability of species-specific coexpression data remains poor. Our assessment scores for individual coexpression data sets indicated that the quality of the new coexpression data sets in ATTED-II is higher than for any previous coexpression data set. In addition, five species (Arabidopsis, soybean, tomato, rice and maize) in ATTED-II are now supported by both microarray- and RNA sequencing-based coexpression data, which has increased the reliability. Consequently, ATTED-II can now provide lineage-specific coexpression information. As an example of the use of ATTED-II to explore lineage-specific coexpression, we demonstrate monocot- and dicot-specific coexpression of cell wall genes. With the expanded coexpression data for multilevel evaluation, ATTED-II provides new opportunities to investigate lineage-specific evolution in plants. PMID:26546318

  20. Tracing the Evolution of Lineage-Specific Transcription Factor Binding Sites in a Birth-Death Framework

    PubMed Central

    Ma, Jian

    2014-01-01

    Changes in cis-regulatory element composition that result in novel patterns of gene expression are thought to be a major contributor to the evolution of lineage-specific traits. Although transcription factor binding events show substantial variation across species, most computational approaches to study regulatory elements focus primarily upon highly conserved sites, and rely heavily upon multiple sequence alignments. However, sequence conservation based approaches have limited ability to detect lineage-specific elements that could contribute to species-specific traits. In this paper, we describe a novel framework that utilizes a birth-death model to trace the evolution of lineage-specific binding sites without relying on detailed base-by-base cross-species alignments. Our model was applied to analyze the evolution of binding sites based on the ChIP-seq data for six transcription factors (GATA1, SOX2, CTCF, MYC, MAX, ETS1) along the lineage toward human after human-mouse common ancestor. We estimate that a substantial fraction of binding sites (∼58–79% for each factor) in humans have origins since the divergence with mouse. Over 15% of all binding sites are unique to hominids. Such elements are often enriched near genes associated with specific pathways, and harbor more common SNPs than older binding sites in the human genome. These results support the ability of our method to identify lineage-specific regulatory elements and help understand their roles in shaping variation in gene regulation across species. PMID:25144359

  1. Molecular characterization of swine leukocyte antigen (SLA) class II genes in outbred pig populations

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The highly polymorphic swine leukocyte antigen (SLA) genes are one of the most important determinants in swine immune, disease and vaccine responses. Thus, understanding how SLA gene polymorphism affects immunity, especially in outbred pig populations with a diverse genetic background, requires accu...

  2. Phylogenetic analysis of the swine leukocyte antigen - 2 gene for Korean native pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to investigate genetic distances of the SLA-2 gene, to characterize SLA-2 alleles, and to provide basic genetic information of Korean pigs. The swine leukocyte antigen - 2 (SLA-2) gene in the MHC classical region was cloned with spleen tissues from Korean native pigs ...

  3. Microfluidic Leukocyte Isolation for Gene Expression Analysis in Critically Ill Hospitalized Patients

    PubMed Central

    Russom, Aman; Sethu, Palaniappan; Irimia, Daniel; Mindrinos, Michael N.; Calvano, Steve E.; Garcia, Iris; Finnerty, Celeste; Tannahill, Cynthia; Abouhamze, Amer; Wilhelmy, Julie; López, M. Cecilia; Baker, Henry V.; Herndon, David N.; Lowry, Stephen F.; Maier, Ronald V.; Davis, Ronald W.; Moldawer, Lyle L.; Tompkins, Ronald G.; Toner, Mehmet

    2014-01-01

    BACKGROUND Microarray technology is becoming a powerful tool for diagnostic, therapeutic, and prognostic applications. There is at present no consensus regarding the optimal technique to isolate nucleic acids from blood leukocyte populations for subsequent expression analyses. Current collection and processing techniques pose significant challenges in the clinical setting. Here, we report the clinical validation of a novel microfluidic leukocyte nucleic acid isolation technique for gene expression analysis from critically ill, hospitalized patients that can be readily used on small volumes of blood. METHODS We processed whole blood from hospitalized patients after burn injury and severe blunt trauma according to the microfluidic and standard macroscale leukocyte isolation protocol. Side-by-side comparison of RNA quantity, quality, and genome-wide expression patterns was used to clinically validate the microfluidic technique. RESULTS When the microfluidic protocol was used for processing, sufficient amounts of total RNA were obtained for genome-wide expression analysis from 0.5 mL whole blood. We found that the leukocyte expression patterns from samples processed using the 2 protocols were concordant, and there was less variability introduced as a result of harvesting method than there existed between individuals. CONCLUSIONS The novel microfluidic approach achieves leukocyte isolation in <25 min, and the quality of nucleic acids and genome expression analysis is equivalent to or surpasses that obtained from macroscale approaches. Microfluidics can significantly improve the isolation of blood leukocytes for genomic analyses in the clinical setting. PMID:18375483

  4. Phylogenetic plant community structure along elevation is lineage specific

    PubMed Central

    Ndiribe, Charlotte; Pellissier, Loïc; Antonelli, Silvia; Dubuis, Anne; Pottier, Julien; Vittoz, Pascal; Guisan, Antoine; Salamin, Nicolas

    2013-01-01

    The trend of closely related taxa to retain similar environmental preferences mediated by inherited traits suggests that several patterns observed at the community scale originate from longer evolutionary processes. While the effects of phylogenetic relatedness have been previously studied within a single genus or family, lineage-specific effects on the ecological processes governing community assembly have rarely been studied for entire communities or flora. Here, we measured how community phylogenetic structure varies across a wide elevation gradient for plant lineages represented by 35 families, using a co-occurrence index and net relatedness index (NRI). We propose a framework that analyses each lineage separately and reveals the trend of ecological assembly at tree nodes. We found prevailing phylogenetic clustering for more ancient nodes and overdispersion in more recent tree nodes. Closely related species may thus rapidly evolve new environmental tolerances to radiate into distinct communities, while older lineages likely retain inherent environmental tolerances to occupy communities in similar environments, either through efficient dispersal mechanisms or the exclusion of older lineages with more divergent environmental tolerances. Our study illustrates the importance of disentangling the patterns of community assembly among lineages to better interpret the ecological role of traits. It also sheds light on studies reporting absence of phylogenetic signal, and opens new perspectives on the analysis of niche and trait conservatism across lineages. PMID:24455126

  5. KCNE gene expression is dependent on the proliferation and mode of activation of leukocytes

    PubMed Central

    Solé, Laura; Vallejo-Gracia, Albert; Roig, Sara R.; Serrano-Albarrás, Antonio; Marruecos, Laura; Manils, Joan; Gómez, Diana; Soler, Concepció; Felipe, Antonio

    2013-01-01

    Voltage-dependent K+ (Kv) channels are tightly regulated during the immune system response. Leukocytes have a limited repertoire of Kv channels, whose physiological role is under intense investigation. A functional Kv channel is an oligomeric complex composed of pore-forming and ancillary subunits. The KCNE gene family is a novel group of modulatory Kv channel elements in leukocytes. Here, we characterized the gene expression of KCNEs (1–5) in leukocytes and investigated their regulation during leukocyte proliferation and mode of activation. Murine bone-marrow-derived macrophages, human Jurkat T-lymphocytes and human Raji B-cells were analyzed. KCNEs (1–5) are expressed in all leukocytes lineages. Most KCNE mRNAs show cell cycle-dependent regulation and are differentially regulated under specific insults. Our results further suggest a new and yet undefined physiological role for KCNE subunits in the immune system. Putative associations of these ancillary proteins with Kv channels would yield a wide variety of biophysically and pharmacologically distinct channels that fine-tune the immunological response. PMID:23327879

  6. Lineage-Specific Modulation of Interleukin 4 Signaling by Interferon Regulatory Factor 4

    PubMed Central

    Gupta, Sanjay; Jiang, Man; Anthony, Alissa; Pernis, Alessandra B.

    1999-01-01

    Interleukin (IL)-4 is an immunoregulatory cytokine that exerts distinct biological activities on different cell types. Our studies indicate that interferon regulatory factor (IRF)-4 is both a target and a modulator of the IL-4 signaling cascade. IRF-4 expression is strongly upregulated upon costimulation of B cells with CD40 and IL-4. Furthermore, we find that IRF-4 can interact with signal transducer and activator of transcription (Stat)6 and drive the expression of IL-4–inducible genes. The transactivating ability of IRF-4 is blocked by the repressor factor BCL-6. Since expression of IRF-4 is mostly confined to lymphoid cells, these data provide a potential mechanism by which IL-4–inducible genes can be regulated in a lineage-specific manner. PMID:10601358

  7. MOLECULAR CHARACTERIZATION OF SWINE LEUKOCYTE ANTIGEN (SLA) CLASS I GENES IN OUTBRED PIG POPULATIONS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The highly polymorphic swine leukocyte antigen (SLA) genes are one of the most important determinants in swine immune responses to infectious diseases, vaccines, and in transplantation. Study of SLA influence requires accurate and effective typing methods. We developed a simple and rapid method to t...

  8. Phenotyping of Leukocytes and Leukocyte-Derived Extracellular Vesicles

    PubMed Central

    Pugholm, Lotte Hatting; Bæk, Rikke; Søndergaard, Evo Kristina Lindersson; Revenfeld, Anne Louise Schacht; Jørgensen, Malene Møller; Varming, Kim

    2016-01-01

    Extracellular vesicles (EVs) have a demonstrated involvement in modulating the immune system. It has been proposed that EVs could be used as biomarkers for detection of inflammatory and immunological disorders. Consequently, it is of great interest to investigate EVs in more detail with focus on immunological markers. In this study, five major leukocyte subpopulations and the corresponding leukocyte-derived EVs were phenotyped with focus on selected immunological lineage-specific markers and selected vesicle-related markers. The leukocyte-derived EVs displayed phenotypic differences in the 34 markers investigated. The majority of the lineage-specific markers used for identification of the parent cell types could not be detected on EVs released from monocultures of the associated cell types. In contrast, the vesicular presentation of CD9, CD63, and CD81 correlated to the cell surface expression of these markers, however, with few exceptions. Furthermore, the cellular expression of CD9, CD63, and CD81 varied between leukocytes present in whole blood and cultured leukocytes. In summary, these data demonstrate that the cellular and vesicular presentation of selected lineage-specific and vesicle-related markers may differ, supporting the accumulating observations that sorting of molecular cargo into EVs is tightly controlled. PMID:27195303

  9. Lineage-specific evolution of Methylthioalkylmalate synthases (MAMs) involved in glucosinolates biosynthesis.

    PubMed

    Zhang, Jifang; Wang, Xiaobo; Cheng, Feng; Wu, Jian; Liang, Jianli; Yang, Wencai; Wang, Xiaowu

    2015-01-01

    Methylthioalkylmalate synthases (MAMs) encoded by MAM genes are central to the diversification of the glucosinolates, which are important secondary metabolites in Brassicaceae species. However, the evolutionary pathway of MAM genes is poorly understood. We analyzed the phylogenetic and synteny relationships of MAM genes from 13 sequenced Brassicaceae species. Based on these analyses, we propose that the syntenic loci of MAM genes, which underwent frequent tandem duplications, divided into two independent lineage-specific evolution routes and were driven by positive selection after the divergence from Aethionema arabicum. In the lineage I species Capsella rubella, Camelina sativa, Arabidopsis lyrata, and A. thaliana, the MAM loci evolved three tandem genes encoding enzymes responsible for the biosynthesis of aliphatic glucosinolates with different carbon chain-lengths. In lineage II species, the MAM loci encode enzymes responsible for the biosynthesis of short-chain aliphatic glucosinolates. Our proposed model of the evolutionary pathway of MAM genes will be useful for understanding the specific function of these genes in Brassicaceae species. PMID:25691886

  10. Lineage-specific evolution of Methylthioalkylmalate synthases (MAMs) involved in glucosinolates biosynthesis

    PubMed Central

    Zhang, Jifang; Wang, Xiaobo; Cheng, Feng; Wu, Jian; Liang, Jianli; Yang, Wencai; Wang, Xiaowu

    2015-01-01

    Methylthioalkylmalate synthases (MAMs) encoded by MAM genes are central to the diversification of the glucosinolates, which are important secondary metabolites in Brassicaceae species. However, the evolutionary pathway of MAM genes is poorly understood. We analyzed the phylogenetic and synteny relationships of MAM genes from 13 sequenced Brassicaceae species. Based on these analyses, we propose that the syntenic loci of MAM genes, which underwent frequent tandem duplications, divided into two independent lineage-specific evolution routes and were driven by positive selection after the divergence from Aethionema arabicum. In the lineage I species Capsella rubella, Camelina sativa, Arabidopsis lyrata, and A. thaliana, the MAM loci evolved three tandem genes encoding enzymes responsible for the biosynthesis of aliphatic glucosinolates with different carbon chain-lengths. In lineage II species, the MAM loci encode enzymes responsible for the biosynthesis of short-chain aliphatic glucosinolates. Our proposed model of the evolutionary pathway of MAM genes will be useful for understanding the specific function of these genes in Brassicaceae species. PMID:25691886

  11. Increased expression of telomere-regulating genes in endurance athletes with long leukocyte telomeres.

    PubMed

    Denham, Joshua; O'Brien, Brendan J; Prestes, Priscilla R; Brown, Nicholas J; Charchar, Fadi J

    2016-01-15

    Leukocyte telomeres shorten with age, and excessive shortening is associated with age-related cardiometabolic diseases. Exercise training may prevent disease through telomere length maintenance although the optimal amount of exercise that attenuates telomere attrition is unknown. Furthermore, the underlying molecular mechanisms responsible for the enhanced telomere maintenance observed in endurance athletes is poorly understood. We quantified the leukocyte telomere length and analyzed the expression of telomere-regulating genes in endurance athletes and healthy controls (both n = 61), using quantitative PCR. We found endurance athletes have significantly longer (7.1%, 208-416 nt) leukocyte telomeres and upregulated TERT (2.0-fold) and TPP1 (1.3-fold) mRNA expression compared with controls in age-adjusted analysis. The telomere length and telomere-regulating gene expression differences were no longer statistically significant after adjustment for resting heart rate and relative V̇O(2 max) (all P > 0.05). Resting heart rate emerged as an independent predictor of leukocyte telomere length and TERT and TPP1 mRNA expression in stepwise regression models. To gauge whether volume of exercise was associated with leukocyte telomere length, we divided subjects into running and cycling tertiles (distance covered per week) and found individuals in the middle and highest tertiles had longer telomeres than individuals in the lowest tertile. These data emphasize the importance of cardiorespiratory fitness and exercise training in the prevention of biological aging. They also support the concept that moderate amounts of exercise training protects against biological aging, while higher amounts may not elicit additional benefits. PMID:26586905

  12. Prospects for T. cruzi lineage-specific serological surveillance of wild mammals.

    PubMed

    Bhattacharyya, Tapan; Mills, Emily A; Jansen, Ana Maria; Miles, Michael A

    2015-11-01

    Sequence diversity in the Trypanosoma cruzi small surface molecule TSSA has yielded antigens for serology to investigate the T. cruzi lineage-specific infection history of patients with Chagas disease. Synthetic peptides can be used as the lineage-specific antigens. Here we consider the rationale, feasibility and potential of applying peptide-based lineage-specific serology to naturally infected wild mammals. The commercial availability of appropriate secondary antibodies encourages this further development, for discovery of new reservoir host species and to reveal the wider ecological distribution of T. cruzi lineages, currently hindered by the need to recover live isolates or to attempt genotyping of DNA extracted from blood samples. PMID:26116784

  13. Age-dependent expression of osteochondrosis-related genes in equine leukocytes

    PubMed Central

    Mendoza, L.; Piquemal, D.; Lejeune, J. P.; Vander Heyden, L.; Noguier, F.; Bruno, R.; Sandersen, C.; Serteyn, D.

    2015-01-01

    Introduction:  Osteochondrosis (OC) is a developmental disease in horses which has a significant impact on the horse's welfare and performance. The early disturbance in the process of endochondral ossification progresses to inflammatory and repair processes in older horses. Previously, differentially expressed genes in leukocytes of OC-affected horses have been identified. The aim of the present study is to detect age-related changes in these differentially expressed genes. Materials and Methods:  The expression of OC-related genes was analysed by real-time PCR and subsequent statistical analysis (ΔΔCT) in the leukocytes of 135 Belgian Warmblood horses divided into three different age groups: <12 months (n=47), 18–24 months (n=50) >30 months (n=38). Results:  Relative expression of genes of horses less than 12 months of age showed significant induction of the genes MGAT4A, PRKCG, MHCI, ApoB, ApoB3G, B4GALT6 and a significantly lower expression of the genes OAS3. Horses of 18–24 months of age, showed a significantly higher expression of the genes TBC1D9, MGAT4A, IFIH1, MHCIIa and MMP1. Horses of more than 30 months of age showed a significantly higher expression of the genes MGAT4A, HP, SECTM1 compared with their age-matched control groups. Conclusions:  The study demonstrates that OC-related genes are differentially expressed in horses of different ages compared with their age-matched controls. Some of the genes may be implicated in cell signalling and differentiation as well as carbohydrate and lipid metabolism and inflammation. However, the causal relationship between the differentially expressed genes and the development and progression of the OC lesions needs to be determined. PMID:26392886

  14. Evolution of lineage-specific functions in ancient cis-regulatory modules.

    PubMed

    Pauls, Stefan; Goode, Debbie K; Petrone, Libero; Oliveri, Paola; Elgar, Greg

    2015-11-01

    Morphological evolution is driven both by coding sequence variation and by changes in regulatory sequences. However, how cis-regulatory modules (CRMs) evolve to generate entirely novel expression domains is largely unknown. Here, we reconstruct the evolutionary history of a lens enhancer located within a CRM that not only predates the lens, a vertebrate innovation, but bilaterian animals in general. Alignments of orthologous sequences from different deuterostomes sub-divide the CRM into a deeply conserved core and a more divergent flanking region. We demonstrate that all deuterostome flanking regions, including invertebrate sequences, activate gene expression in the zebrafish lens through the same ancient cluster of activator sites. However, levels of gene expression vary between species due to the presence of repressor motifs in flanking region and core. These repressor motifs are responsible for the relatively weak enhancer activity of tetrapod flanking regions. Ray-finned fish, however, have gained two additional lineage-specific activator motifs which in combination with the ancient cluster of activators and the core constitute a potent lens enhancer. The exploitation and modification of existing regulatory potential in flanking regions but not in the highly conserved core might represent a more general model for the emergence of novel regulatory functions in complex CRMs. PMID:26538567

  15. Lower Methylation of the ANGPTL2 Gene in Leukocytes from Post-Acute Coronary Syndrome Patients

    PubMed Central

    Nguyen, Albert; Mamarbachi, Maya; Turcot, Valérie; Lessard, Samuel; Yu, Carol; Luo, Xiaoyan; Lalongé, Julie; Hayami, Doug; Gayda, Mathieu; Juneau, Martin; Thorin-Trescases, Nathalie; Lettre, Guillaume; Nigam, Anil; Thorin, Eric

    2016-01-01

    DNA methylation is believed to regulate gene expression during adulthood in response to the constant changes in environment. The methylome is therefore proposed to be a biomarker of health through age. ANGPTL2 is a circulating pro-inflammatory protein that increases with age and prematurely in patients with coronary artery diseases; integrating the methylation pattern of the promoter may help differentiate age- vs. disease-related change in its expression. We believe that in a pro-inflammatory environment, ANGPTL2 is differentially methylated, regulating ANGPTL2 expression. To test this hypothesis we investigated the changes in promoter methylation of ANGPTL2 gene in leukocytes from patients suffering from post-acute coronary syndrome (ACS). DNA was extracted from circulating leukocytes of post-ACS patients with cardiovascular risk factors and from healthy young and age-matched controls. Methylation sites (CpGs) found in the ANGPTL2 gene were targeted for specific DNA methylation quantification. The functionality of ANGPTL2 methylation was assessed by an in vitro luciferase assay. In post-ACS patients, C-reactive protein and ANGPTL2 circulating levels increased significantly when compared to healthy controls. Decreased methylation of specific CpGs were found in the promoter of ANGPTL2 and allowed to discriminate age vs. disease associated methylation. In vitro DNA methylation of specific CpG lead to inhibition of ANGPTL2 promoter activity. Reduced leukocyte DNA methylation in the promoter region of ANGPTL2 is associated with the pro-inflammatory environment that characterizes patients with post-ACS differently from age-matched healthy controls. Methylation of different CpGs in ANGPTL2 gene may prove to be a reliable biomarker of coronary disease. PMID:27101308

  16. Geographical Genomics of Human Leukocyte Gene Expression Variation in Southern Morocco

    PubMed Central

    Idaghdour, Youssef; Czika, Wendy; Shianna, Kevin V.; Lee, S. Hong; Visscher, Peter M.; Martin, Hilary C.; Miclaus, Kelci; Jadallah, Sami J.; Goldstein, David B.; Wolfinger, Russell D.; Gibson, Greg

    2009-01-01

    Studies of the genetics of gene expression reveal expression SNPs that explain variation in transcript abundance. Here we address the robustness of eSNP associations to environmental geography and population structure in a comparison of 194 Arab and Amazigh individuals from a city and two villages in southern Morocco. Gene expression differed between pairs of locations for up to a third of all transcripts, with notable enrichment for ribosomal biosynthesis and oxidative phosphorylation. Robust associations were observed in the leukocyte samples with cis-eSNPs (P < 10−08) for 346 genes, and trans-eSNPs (P < 10−11) with 10 genes. All of these were consistent across the three sample locations and after controlling for ethnicity and relatedness. No evidence for large-effect trans-acting mediators of the pervasive environmental influence was found and instead genetic and environmental factors acted in a largely additive manner. PMID:19966804

  17. Concordant Gene Expression in Leukemia Cells and Normal Leukocytes Is Associated with Germline cis-SNPs

    PubMed Central

    French, Deborah; Yang, Wenjian; Hamilton, Leo H.; Neale, Geoffrey; Fan, Yiping; Downing, James R.; Cox, Nancy J.; Pui, Ching-Hon; Evans, William E.; Relling, Mary V.

    2008-01-01

    The degree to which gene expression covaries between different primary tissues within an individual is not well defined. We hypothesized that expression that is concordant across tissues is more likely influenced by genetic variability than gene expression which is discordant between tissues. We quantified expression of 11,873 genes in paired samples of primary leukemia cells and normal leukocytes from 92 patients with acute lymphoblastic leukemia (ALL). Genetic variation at >500,000 single nucleotide polymorphisms (SNPs) was also assessed. The expression of only 176/11,783 (1.5%) genes was correlated (p<0.008, FDR = 25%) in the two tissue types, but expression of a high proportion (20 of these 176 genes) was significantly related to cis-SNP genotypes (adjusted p<0.05). In an independent set of 134 patients with ALL, 14 of these 20 genes were validated as having expression related to cis-SNPs, as were 9 of 20 genes in a second validation set of HapMap cell lines. Genes whose expression was concordant among tissue types were more likely to be associated with germline cis-SNPs than genes with discordant expression in these tissues; genes affected were involved in housekeeping functions (GSTM2, GAPDH and NCOR1) and purine metabolism. PMID:18478092

  18. Adipose lineage specification of bone marrow-derived myeloid cells

    PubMed Central

    Majka, Susan M.; Miller, Heidi L.; Sullivan, Timothy; Erickson, Paul F.; Kong, Raymond; Weiser-Evans, Mary; Nemenoff, Raphael; Moldovan, Radu; Morandi, Shelley A.; Davis, James A.; Klemm, Dwight J.

    2012-01-01

    We have reported the production of white adipocytes in adipose tissue from hematopoietic progenitors arising from bone marrow. However, technical challenges have hindered detection of this adipocyte population by certain other laboratories. These disparate results highlight the need for sensitive and definitive techniques to identify bone marrow progenitor (BMP)-derived adipocytes. In these studies we exploited new models and methods to enhance detection of this adipocyte population. Here we showed that confocal microscopy with spectrum acquisition could effectively identify green fluorescent protein (GFP) positive BMP-derived adipocytes by matching their fluorescence spectrum to that of native GFP. Likewise, imaging flow cytometry made it possible to visualize intact unilocular and multilocular GFP-positive BMP-derived adipocytes and distinguished them from non-fluorescent adipocytes and cell debris in the cytometer flow stream. We also devised a strategy to detect marker genes in flow-enriched adipocytes from which stromal cells were excluded. This technique also proved to be an efficient means for detecting genetically labeled adipocytes and should be applicable to models in which marker gene expression is low or absent. Finally, in vivo imaging of mice transplanted with BM from adipocyte-targeted luciferase donors showed a time-dependent increase in luciferase activity, with the bulk of luciferase activity confined to adipocytes rather than stromal cells. These results confirmed and extended our previous reports and provided proof-of-principle for sensitive techniques and models for detection and study of these unique cells. PMID:23700536

  19. Characterisation of monotreme caseins reveals lineage-specific expansion of an ancestral casein locus in mammals.

    PubMed

    Lefèvre, Christophe M; Sharp, Julie A; Nicholas, Kevin R

    2009-01-01

    Using a milk-cell cDNA sequencing approach we characterised milk-protein sequences from two monotreme species, platypus (Ornithorhynchus anatinus) and echidna (Tachyglossus aculeatus) and found a full set of caseins and casein variants. The genomic organisation of the platypus casein locus is compared with other mammalian genomes, including the marsupial opossum and several eutherians. Physical linkage of casein genes has been seen in the casein loci of all mammalian genomes examined and we confirm that this is also observed in platypus. However, we show that a recent duplication of beta-casein occurred in the monotreme lineage, as opposed to more ancient duplications of alpha-casein in the eutherian lineage, while marsupials possess only single copies of alpha- and beta-caseins. Despite this variability, the close proximity of the main alpha- and beta-casein genes in an inverted tail-tail orientation and the relative orientation of the more distant kappa-casein genes are similar in all mammalian genome sequences so far available. Overall, the conservation of the genomic organisation of the caseins indicates the early, pre-monotreme development of the fundamental role of caseins during lactation. In contrast, the lineage-specific gene duplications that have occurred within the casein locus of monotremes and eutherians but not marsupials, which may have lost part of the ancestral casein locus, emphasises the independent selection on milk provision strategies to the young, most likely linked to different developmental strategies. The monotremes therefore provide insight into the ancestral drivers for lactation and how these have adapted in different lineages. PMID:19874726

  20. miRNA-embedded shRNAs for Lineage-specific BCL11A Knockdown and Hemoglobin F Induction

    PubMed Central

    Guda, Swaroopa; Brendel, Christian; Renella, Raffaele; Du, Peng; Bauer, Daniel E; Canver, Matthew C; Grenier, Jennifer K; Grimson, Andrew W; Kamran, Sophia C; Thornton, James; de Boer, Helen; Root, David E; Milsom, Michael D; Orkin, Stuart H; Gregory, Richard I; Williams, David A

    2015-01-01

    RNA interference (RNAi) technology using short hairpin RNAs (shRNAs) expressed via RNA polymerase (pol) III promoters has been widely exploited to modulate gene expression in a variety of mammalian cell types. For certain applications, such as lineage-specific knockdown, embedding targeting sequences into pol II-driven microRNA (miRNA) architecture is required. Here, using the potential therapeutic target BCL11A, we demonstrate that pol III-driven shRNAs lead to significantly increased knockdown but also increased cytotoxcity in comparison to pol II-driven miRNA adapted shRNAs (shRNAmiR) in multiple hematopoietic cell lines. We show that the two expression systems yield mature guide strand sequences that differ by a 4 bp shift. This results in alternate seed sequences and consequently influences the efficacy of target gene knockdown. Incorporating a corresponding 4 bp shift into the guide strand of shRNAmiRs resulted in improved knockdown efficiency of BCL11A. This was associated with a significant de-repression of the hemoglobin target of BCL11A, human γ-globin or the murine homolog Hbb-y. Our results suggest the requirement for optimization of shRNA sequences upon incorporation into a miRNA backbone. These findings have important implications in future design of shRNAmiRs for RNAi-based therapy in hemoglobinopathies and other diseases requiring lineage-specific expression of gene silencing sequences. PMID:26080908

  1. Environmental Physical Cues Determine the Lineage Specification of Mesenchymal Stem Cells

    PubMed Central

    Huang, Chao; Dai, Jingxing; Zhang, Xin A.

    2015-01-01

    Background Physical cues of cellular environment affect cell fate and differentiation. For example, an environment with high stiffness drives mesenchymal stem cells (MSCs) to undergo osteogenic differentiation, while low stiffness leads to lipogenic differentiation. Such effects could be independent of chemical/biochemical inducers. Scope of review Stiffness and/or topography of cellular environment can control MSC differentiation and fate determination. In addition, physical factors such as tension, resulted from profound cytoskeleton reorganization during MSC differentiation, affect the gene expression essential for the differentiation. Although physical cues control MSC lineage specification probably by reorganizing and tuning cytoskeleton, the full mechanism is largely unclear. It also remains elusive how physical signals are sensed by cells and transformed into biochemical and biological signals. More importantly, it becomes pivotal to define explicitly the physical cue(s) essential for cell differentiation and fate decision. With a focus on MSC, we present herein current understanding of the interplay between i) physical cue and factors and ii) MSC differentiation and fate determination. Major conclusions Biophysical cues can initiate or strengthen the biochemical signaling for MSC fate determination and differentiation. Physical properties of cellular environment direct the structural adaptation and functional coupling of the cells to their environment. General significance These observations not only open a simple avenue to engineer cell fate in vitro, but also start to reveal the physical elements that regulate and determine cell fate. PMID:25727396

  2. Cluster Analysis of Tumor Suppressor Genes in Canine Leukocytes Identifies Activation State

    PubMed Central

    Daly, Julie-Anne; Mortlock, Sally-Anne; Taylor, Rosanne M.; Williamson, Peter

    2015-01-01

    Cells of the immune system undergo activation and subsequent proliferation in the normal course of an immune response. Infrequently, the molecular and cellular events that underlie the mechanisms of proliferation are dysregulated and may lead to oncogenesis, leading to tumor formation. The most common forms of immunological cancers are lymphomas, which in dogs account for 8%–20% of all cancers, affecting up to 1.2% of the dog population. Key genes involved in negatively regulating proliferation of lymphocytes include a group classified as tumor suppressor genes (TSGs). These genes are also known to be associated with progression of lymphoma in humans, mice, and dogs and are potential candidates for pathological grading and diagnosis. The aim of the present study was to analyze TSG profiles in stimulated leukocytes from dogs to identify genes that discriminate an activated phenotype. A total of 554 TSGs and three gene set collections were analyzed from microarray data. Cluster analysis of three subsets of genes discriminated between stimulated and unstimulated cells. These included 20 most upregulated and downregulated TSGs, TSG in hallmark gene sets significantly enriched in active cells, and a selection of candidate TSGs, p15 (CDKN2B), p18 (CDKN2C), p19 (CDKN1A), p21 (CDKN2A), p27 (CDKN1B), and p53 (TP53) in the third set. Analysis of two subsets suggested that these genes or a subset of these genes may be used as a specialized PCR set for additional analysis. PMID:27478369

  3. Lineage-specific Virulence Determinants of Haemophilus influenzae Biogroup aegyptius

    PubMed Central

    Strouts, Fiona R.; Power, Peter; Croucher, Nicholas J.; Corton, Nicola; van Tonder, Andries; Quail, Michael A.; Langford, Paul R.; Hudson, Michael J.; Parkhill, Julian; Bentley, Stephen D.

    2012-01-01

    An emergent clone of Haemophilus influenzae biogroup aegyptius (Hae) is responsible for outbreaks of Brazilian purpuric fever (BPF). First recorded in Brazil in 1984, the so-called BPF clone of Hae caused a fulminant disease that started with conjunctivitis but developed into septicemic shock; mortality rates were as high as 70%. To identify virulence determinants, we conducted a pan-genomic analysis. Sequencing of the genomes of the BPF clone strain F3031 and a noninvasive conjunctivitis strain, F3047, and comparison of these sequences with 5 other complete H. influenzae genomes showed that >77% of the F3031 genome is shared among all H. influenzae strains. Delineation of the Hae accessory genome enabled characterization of 163 predicted protein-coding genes; identified differences in established autotransporter adhesins; and revealed a suite of novel adhesins unique to Hae, including novel trimeric autotransporter adhesins and 4 new fimbrial operons. These novel adhesins might play a critical role in host–pathogen interactions. PMID:22377449

  4. Lineage-specific genomics: Frequent birth and death in the human genome: The human genome contains many lineage-specific elements created by both sequence and functional turnover.

    PubMed

    Young, Robert S

    2016-07-01

    Frequent evolutionary birth and death events have created a large quantity of biologically important, lineage-specific DNA within mammalian genomes. The birth and death of DNA sequences is so frequent that the total number of these insertions and deletions in the human population remains unknown, although there are differences between these groups, e.g. transposable elements contribute predominantly to sequence insertion. Functional turnover - where the activity of a locus is specific to one lineage, but the underlying DNA remains conserved - can also drive birth and death. However, this does not appear to be a major driver of divergent transcriptional regulation. Both sequence and functional turnover have contributed to the birth and death of thousands of functional promoters in the human and mouse genomes. These findings reveal the pervasive nature of evolutionary birth and death and suggest that lineage-specific regions may play an important but previously underappreciated role in human biology and disease. PMID:27231054

  5. Lineage-Specific Patterns of Genome Deterioration in Obligate Symbionts of Sharpshooter Leafhoppers

    PubMed Central

    Bennett, Gordon M.; McCutcheon, John P.; McDonald, Bradon R.; Moran, Nancy A.

    2016-01-01

    Plant sap-feeding insects (Hemiptera) rely on obligate bacterial symbionts that provision nutrients. Some of these symbionts are ancient and have evolved tiny genomes, whereas others are younger and retain larger, dynamic genomes. Baumannia cicadellinicola, an obligate symbiont of sharpshooter leafhoppers, is derived from a relatively recent symbiont replacement. To better understand evolutionary decay of genomes, we compared Baumannia from three host species. A newly sequenced genome for Baumannia from the green sharpshooter (B-GSS) was compared with genomes of Baumannia from the blue-green sharpshooter (B-BGSS, 759 kilobases [kb]) and from the glassy-winged sharpshooter (B-GWSS, 680 kb). B-GSS has the smallest Baumannia genome sequenced to date (633 kb), with only three unique genes, all involved in membrane function. It has lost nearly all pathways involved in vitamin and cofactor synthesis, as well as amino acid biosynthetic pathways that are redundant with pathways of the host or the symbiotic partner, Sulcia muelleri. The entire biosynthetic pathway for methionine is eliminated, suggesting that methionine has become a dietary requirement for hosts. B-GSS and B-BGSS share 33 genes involved in bacterial functions (e.g., cell division, membrane synthesis, metabolite transport, etc.) that are lost from the more distantly related B-GWSS and most other tiny genome symbionts. Finally, pairwise divergence estimates indicate that B-GSS has experienced a lineage-specific increase in substitution rates. This increase correlates with accelerated protein-level changes and widespread gene loss. Thus, the mode and tempo of genome reduction vary widely among symbiont lineages and result in wide variation in metabolic capabilities across hosts. PMID:26260652

  6. Lineage-Specific Patterns of Genome Deterioration in Obligate Symbionts of Sharpshooter Leafhoppers.

    PubMed

    Bennett, Gordon M; McCutcheon, John P; McDonald, Bradon R; Moran, Nancy A

    2016-01-01

    Plant sap-feeding insects (Hemiptera) rely on obligate bacterial symbionts that provision nutrients. Some of these symbionts are ancient and have evolved tiny genomes, whereas others are younger and retain larger, dynamic genomes. Baumannia cicadellinicola, an obligate symbiont of sharpshooter leafhoppers, is derived from a relatively recent symbiont replacement. To better understand evolutionary decay of genomes, we compared Baumannia from three host species. A newly sequenced genome for Baumannia from the green sharpshooter (B-GSS) was compared with genomes of Baumannia from the blue-green sharpshooter (B-BGSS, 759 kilobases [kb]) and from the glassy-winged sharpshooter (B-GWSS, 680 kb). B-GSS has the smallest Baumannia genome sequenced to date (633 kb), with only three unique genes, all involved in membrane function. It has lost nearly all pathways involved in vitamin and cofactor synthesis, as well as amino acid biosynthetic pathways that are redundant with pathways of the host or the symbiotic partner, Sulcia muelleri. The entire biosynthetic pathway for methionine is eliminated, suggesting that methionine has become a dietary requirement for hosts. B-GSS and B-BGSS share 33 genes involved in bacterial functions (e.g., cell division, membrane synthesis, metabolite transport, etc.) that are lost from the more distantly related B-GWSS and most other tiny genome symbionts. Finally, pairwise divergence estimates indicate that B-GSS has experienced a lineage-specific increase in substitution rates. This increase correlates with accelerated protein-level changes and widespread gene loss. Thus, the mode and tempo of genome reduction vary widely among symbiont lineages and result in wide variation in metabolic capabilities across hosts. PMID:26260652

  7. The BAF chromatin remodelling complex is an epigenetic regulator of lineage specification in the early mouse embryo

    PubMed Central

    Panamarova, Maryna; Cox, Andy; Wicher, Krzysztof B.; Butler, Richard; Bulgakova, Natalia; Jeon, Shin; Rosen, Barry; Seong, Rho H.; Skarnes, William; Crabtree, Gerald; Zernicka-Goetz, Magdalena

    2016-01-01

    Dynamic control of gene expression is essential for the development of a totipotent zygote into an embryo with defined cell lineages. The accessibility of genes responsible for cell specification to transcriptional machinery is dependent on chromatin remodelling complexes such as the SWI\\SNF (BAF) complex. However, the role of the BAF complex in early mouse development has remained unclear. Here, we demonstrate that BAF155, a major BAF complex subunit, regulates the assembly of the BAF complex in vivo and regulates lineage specification of the mouse blastocyst. We find that associations of BAF155 with other BAF complex subunits become enriched in extra-embryonic lineages just prior to implantation. This enrichment is attributed to decreased mobility of BAF155 in extra-embryonic compared with embryonic lineages. Downregulation of BAF155 leads to increased expression of the pluripotency marker Nanog and its ectopic expression in extra-embryonic lineages, whereas upregulation of BAF155 leads to the upregulation of differentiation markers. Finally, we show that the arginine methyltransferase CARM1 methylates BAF155, which differentially influences assembly of the BAF complex between the lineages and the expression of pluripotency markers. Together, our results indicate a novel role of BAF-dependent chromatin remodelling in mouse development via regulation of lineage specification. PMID:26952987

  8. The BAF chromatin remodelling complex is an epigenetic regulator of lineage specification in the early mouse embryo.

    PubMed

    Panamarova, Maryna; Cox, Andy; Wicher, Krzysztof B; Butler, Richard; Bulgakova, Natalia; Jeon, Shin; Rosen, Barry; Seong, Rho H; Skarnes, William; Crabtree, Gerald; Zernicka-Goetz, Magdalena

    2016-04-15

    Dynamic control of gene expression is essential for the development of a totipotent zygote into an embryo with defined cell lineages. The accessibility of genes responsible for cell specification to transcriptional machinery is dependent on chromatin remodelling complexes such as the SWI\\SNF (BAF) complex. However, the role of the BAF complex in early mouse development has remained unclear. Here, we demonstrate that BAF155, a major BAF complex subunit, regulates the assembly of the BAF complexin vivoand regulates lineage specification of the mouse blastocyst. We find that associations of BAF155 with other BAF complex subunits become enriched in extra-embryonic lineages just prior to implantation. This enrichment is attributed to decreased mobility of BAF155 in extra-embryonic compared with embryonic lineages. Downregulation of BAF155 leads to increased expression of the pluripotency markerNanogand its ectopic expression in extra-embryonic lineages, whereas upregulation of BAF155 leads to the upregulation of differentiation markers. Finally, we show that the arginine methyltransferase CARM1 methylates BAF155, which differentially influences assembly of the BAF complex between the lineages and the expression of pluripotency markers. Together, our results indicate a novel role of BAF-dependent chromatin remodelling in mouse development via regulation of lineage specification. PMID:26952987

  9. Effects of Smoking Cessation on Gene Expression in Human Leukocytes of Chronic Smoker

    PubMed Central

    Kim, Soo-Jeong; Kim, Su Young; Kim, Jae Hwa

    2014-01-01

    Objective The risks of cigarette smoking concerning higher systemic disease mortality are lessened by smoking cessation. Methods Microarray analysis compared the expression profiles of smokers who were successful and not successful at smoking cessation, with the goal of identifying genes that might serve as potential biomarkers or that might be valuable in elucidating distinct biological mechanisms. The mRNAs were isolated and compared from peripheral leukocytes of six smokers who were successful in cessation and six smokers who failed in smoking cessation. Results Two hundred ninety nine genes displayed significantly different expression; 196 genes were up-regulated and 103 genes were down-regulated in the success group compared to the failure group. Twenty four of these genes were identified with biological processes including immunity, cytoskeleton and cell growth/cycle. Real-time PCR confirmed the differential gene expression. The mRNA levels of HEPACAM family member 2 (HEPACAM2) and tropomodulin 1 (TMOD1) were significantly more expressed in the success group, while the mRNA ubiquitin specific peptides 18 (USP18) were significantly less expressed in the success group compared to the failure group. Conclusion The results suggest that smoking cessation can modulate cell adhesion and immune response by regulating expression levels of genes, especially HEPACAM2, TMOD1 and USP18, which have an important relationship with smoking cessation. PMID:25110502

  10. Understanding the molecular circuitry of cell lineage specification in the early mouse embryo.

    PubMed

    Bergsmedh, Anna; Donohoe, Mary E; Hughes, Rebecca-Ayme; Hadjantonakis, Anna-Katerina

    2011-01-01

    Pluripotent stem cells hold great promise for cell-based therapies in regenerative medicine. However, critical to understanding and exploiting mechanisms of cell lineage specification, epigenetic reprogramming, and the optimal environment for maintaining and differentiating pluripotent stem cells is a fundamental knowledge of how these events occur in normal embryogenesis. The early mouse embryo has provided an excellent model to interrogate events crucial in cell lineage commitment and plasticity, as well as for embryo-derived lineage-specific stem cells and induced pluripotent stem (iPS) cells. Here we provide an overview of cell lineage specification in the early (preimplantation) mouse embryo focusing on the transcriptional circuitry and epigenetic marks necessary for successive differentiation events leading to the formation of the blastocyst. PMID:24710206

  11. Age gene expression and coexpression progressive signatures in peripheral blood leukocytes.

    PubMed

    Irizar, Haritz; Goñi, Joaquín; Alzualde, Ainhoa; Castillo-Triviño, Tamara; Olascoaga, Javier; Lopez de Munain, Adolfo; Otaegui, David

    2015-12-01

    Both cellular senescence and organismic aging are known to be dynamic processes that start early in life and progress constantly during the whole life of the individual. In this work, with the objective of identifying signatures of age-related progressive change at the transcriptomic level, we have performed a whole-genome gene expression analysis of peripheral blood leukocytes in a group of healthy individuals with ages ranging from 14 to 93 years. A set of genes with progressively changing gene expression (either increase or decrease with age) has been identified and contextualized in a coexpression network. A modularity analysis has been performed on this network and biological-term and pathway enrichment analyses have been used for biological interpretation of each module. In summary, the results of the present work reveal the existence of a transcriptomic component that shows progressive expression changes associated to age in peripheral blood leukocytes, highlighting both the dynamic nature of the process and the need to complement young vs. elder studies with longitudinal studies that include middle aged individuals. From the transcriptional point of view, immunosenescence seems to be occurring from a relatively early age, at least from the late 20s/early 30s, and the 49-56 year old age-range appears to be critical. In general, the genes that, according to our results, show progressive expression changes with aging are involved in pathogenic/cellular processes that have classically been linked to aging in humans: cancer, immune processes and cellular growth vs. maintenance. PMID:26362218

  12. Methylation of a panel of genes in peripheral blood leukocytes is associated with colorectal cancer.

    PubMed

    Luo, Xiang; Huang, Rong; Sun, Hongru; Liu, Yupeng; Bi, Haoran; Li, Jing; Yu, Hongyuan; Sun, Jiamei; Lin, Shangqun; Cui, Binbin; Zhao, Yashuang

    2016-01-01

    The relationship between the DNA methylation status of the CpG islands of multiple genes in blood leukocytes in CRC susceptibility and prognosis, as well as possible interactions with dietary factors on CRC risk are unclear. We carried out a case-control study including 421 CRC patients and 506 controls to examine the associations between six genes (AOX-1, RARB2, RERG, ADAMTS9, IRF4, and FOXE-1), multiple CpG site methylation (MCSM) and susceptibility to CRC. High-level MCSM (MCSM-H) was defined as methylation of greater than or equal to 2 of 5 candidate genes (except for RARB2); low-level MCSM (MCSM-L) was when 1 candidate gene was methylated; non-MCSM was when none of the candidate genes were methylated. Blood cell-derived DNA methylation status was detected using methylation-sensitive high-resolution melting analysis. The hypermethylation status of each individual gene was statistically significantly associated with CRC. MCSM status was also associated with CRC (OR = 1.54, 95% CI: 1.15-2.05, P = 0.004). We observed interactions between a high level of dietary intake of cereals, pungent food, and stewed fish with brown sauce, age (older than 60 yrs), smoking and hypermethylation on risk of CRC. MCSM in peripheral blood DNA may be an important biomarker for susceptibility to CRC. PMID:27453436

  13. Methylation of a panel of genes in peripheral blood leukocytes is associated with colorectal cancer

    PubMed Central

    Luo, Xiang; Huang, Rong; Sun, Hongru; Liu, Yupeng; Bi, Haoran; Li, Jing; Yu, Hongyuan; Sun, Jiamei; Lin, Shangqun; Cui, Binbin; Zhao, Yashuang

    2016-01-01

    The relationship between the DNA methylation status of the CpG islands of multiple genes in blood leukocytes in CRC susceptibility and prognosis, as well as possible interactions with dietary factors on CRC risk are unclear. We carried out a case-control study including 421 CRC patients and 506 controls to examine the associations between six genes (AOX-1, RARB2, RERG, ADAMTS9, IRF4, and FOXE-1), multiple CpG site methylation (MCSM) and susceptibility to CRC. High-level MCSM (MCSM-H) was defined as methylation of greater than or equal to 2 of 5 candidate genes (except for RARB2); low-level MCSM (MCSM-L) was when 1 candidate gene was methylated; non-MCSM was when none of the candidate genes were methylated. Blood cell-derived DNA methylation status was detected using methylation-sensitive high-resolution melting analysis. The hypermethylation status of each individual gene was statistically significantly associated with CRC. MCSM status was also associated with CRC (OR = 1.54, 95% CI: 1.15–2.05, P = 0.004). We observed interactions between a high level of dietary intake of cereals, pungent food, and stewed fish with brown sauce, age (older than 60 yrs), smoking and hypermethylation on risk of CRC. MCSM in peripheral blood DNA may be an important biomarker for susceptibility to CRC. PMID:27453436

  14. Transient accumulation of 5-carboxylcytosine indicates involvement of active demethylation in lineage specification of neural stem cells.

    PubMed

    Wheldon, Lee M; Abakir, Abdulkadir; Ferjentsik, Zoltan; Dudnakova, Tatiana; Strohbuecker, Stephanie; Christie, Denise; Dai, Nan; Guan, Shengxi; Foster, Jeremy M; Corrêa, Ivan R; Loose, Matthew; Dixon, James E; Sottile, Virginie; Johnson, Andrew D; Ruzov, Alexey

    2014-06-12

    5-Methylcytosine (5mC) is an epigenetic modification involved in regulation of gene activity during differentiation. Tet dioxygenases oxidize 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). Both 5fC and 5caC can be excised from DNA by thymine-DNA glycosylase (TDG) followed by regeneration of unmodified cytosine via the base excision repair pathway. Despite evidence that this mechanism is operative in embryonic stem cells, the role of TDG-dependent demethylation in differentiation and development is currently unclear. Here, we demonstrate that widespread oxidation of 5hmC to 5caC occurs in postimplantation mouse embryos. We show that 5fC and 5caC are transiently accumulated during lineage specification of neural stem cells (NSCs) in culture and in vivo. Moreover, 5caC is enriched at the cell-type-specific promoters during differentiation of NSCs, and TDG knockdown leads to increased 5fC/5caC levels in differentiating NSCs. Our data suggest that active demethylation contributes to epigenetic reprogramming determining lineage specification in embryonic brain. PMID:24882006

  15. A missense mutation in the beta-2 integrin gene (ITGB2) causes canine leukocyte adhesion deficiency.

    PubMed

    Kijas, J M; Bauer, T R; Gäfvert, S; Marklund, S; Trowald-Wigh, G; Johannisson, A; Hedhammar, A; Binns, M; Juneja, R K; Hickstein, D D; Andersson, L

    1999-10-01

    Canine leukocyte adhesion deficiency (CLAD) is a fatal immunodeficiency disease found in Irish setters. The clinical manifestations of CLAD are very similar to LAD in humans and BLAD in cattle, which are both caused by mutations in ITGB2 encoding the leukocyte integrin beta-2 subunit (CD18). Sequence analysis of the ITGB2 coding sequence from a CLAD dog and a healthy control revealed a single missense mutation, Cys36Ser. This cysteine residue is conserved among all beta integrins, and the mutation most likely disrupts a disulfide bond. The mutation showed a complete association with CLAD in Irish setters and was not found in a sample of dogs from other breeds. The causative nature of this mutation was confirmed by transduction experiments using retroviral vectors and human LAD EBV B-cells. The normal canine CD18 formed heterodimers with the human CD11 subunit, whereas gene transfer of the mutant CD18 resulted in very low levels of CD11/CD18 expression. The identification of the causative mutation for CLAD now makes it possible to identify carrier animals with a simple diagnostic DNA test, and it forms the basis for using CLAD as a large animal model for the development and evaluation of clinical treatments for human LAD. PMID:10512685

  16. The effect of cell subset isolation method on gene expression in leukocytes

    PubMed Central

    White, Cory; Lada, Steven; Du, Pinyi; Vaida, Florin; Blanco, Julià; Spina, Celsa A.; Woelk, Christopher H.

    2014-01-01

    Multiple scientific disciplines require the isolation of specific subsets of blood cells from patient samples for gene expression analysis by microarray or RNA-sequencing, preserving disease- or treatment-related signatures. However, little is known with respect to the impact of different cell isolation methods on gene expression and the effects of positive selection, negative selection and fluorescence activated cell sorting (FACS) have not previously been assessed in parallel. To address this knowledge gap, CD4+ T cells, CD8+ T cells, B cells and monocytes were isolated from blood samples from 5 independent donors using positive immunomagnetic selection, negative immunomagnetic selection and FACS. We hypothesized that positive selection and FACS would yield higher purity but may have an impact on gene expression since both methods utilize antibodies that bind surface receptors of the cell type of interest. Moreover, FACS might upregulate stress response genes due to passage of the cells through the sorter. Microarray gene expression data was generated and subjected to unsupervised clustering and differential gene expression analysis. Surprisingly, these analyses revealed that gene expression signatures were more similar between cells isolated by negative selection and FACS compared to cells isolated by positive selection. Moreover, genes that are involved in the response to stress generally had the highest expression in cells isolated by negative or positive selection and not FACS. Thus, FACS is the recommended method for isolation of leukocyte subsets for gene expression studies since this method results in the purest subset populations and does not appear to induce a stress response. PMID:24115734

  17. Lineage-specific and ubiquitous biological roles of the mammalian transcription factor LSF

    PubMed Central

    Veljkovic, Jelena; Hansen, Ulla

    2012-01-01

    Transcriptional regulation in mammalian cells is driven by a complex interplay of multiple transcription factors that respond to signals from either external or internal stimuli. A single transcription factor can control expression of distinct sets of target genes, dependent on its state of post-translational modifications, interacting partner proteins, and the chromatin environment of the cellular genome. Furthermore, many transcription factors can act as either transcriptional repressors or activators, depending on promoter and cellular contexts (Alvarez, et al., 2003). Even in this light, the versatility of LSF (Late SV40 Factor) is remarkable. A hallmark of LSF is its unusual DNA binding domain, as evidenced both by lack of homology to any other established DNA-binding domains and by its DNA recognition sequence. Although a dimer in solution, LSF requires additional multimerization with itself or partner proteins in order to interact with DNA. Transcriptionally, LSF can function as an activator or a repressor. It is a direct target of an increasing number of signal transduction pathways. Biologically, LSF plays roles in cell cycle progression and cell survival, as well as in cell lineage-specific functions, shown most strikingly to date in hematopoietic lineages. This review discusses how the unique aspects of LSF DNA-binding activity may make it particularly susceptible to regulation by signal transduction pathways and may relate to its distinct biological roles. We present current progress in elucidation of both tissue-specific and more universal cellular roles of LSF. Finally, we discuss suggestive data linking LSF to signaling by the amyloid precursor protein and to Alzheimer's disease, as well as to the regulation of latency of the human immunodeficiency virus (HIV). PMID:15563829

  18. Early cell lineage specification in a marsupial: a case for diverse mechanisms among mammals.

    PubMed

    Frankenberg, Stephen; Shaw, Geoff; Freyer, Claudia; Pask, Andrew J; Renfree, Marilyn B

    2013-03-01

    Early cell lineage specification in eutherian mammals results in the formation of a pluripotent inner cell mass (ICM) and trophoblast. By contrast, marsupials have no ICM. Here, we present the first molecular analysis of mechanisms of early cell lineage specification in a marsupial, the tammar wallaby. There was no overt differential localisation of key lineage-specific transcription factors in cleavage and early unilaminar blastocyst stages. Pluriblast cells (equivalent to the ICM) became distinguishable from trophoblast cells by differential expression of POU5F1 and, to a greater extent, POU2, a paralogue of POU5F1. Unlike in the mouse, pluriblast-trophoblast differentiation coincided with a global nuclear-to-cytoplasmic transition of CDX2 localisation. Also unlike in the mouse, Hippo pathway factors YAP and WWTR1 showed mutually distinct localisation patterns that suggest non-redundant roles. NANOG and GATA6 were conserved as markers of epiblast and hypoblast, respectively, but some differences to the mouse were found in their mode of differentiation. Our results suggest that there is considerable evolutionary plasticity in the mechanisms regulating early lineage specification in mammals. PMID:23344710

  19. Effect of Genetic Diversity in Swine Leukocyte Antigen-DRA Gene on Piglet Diarrhea

    PubMed Central

    Huang, Xiaoyu; Yang, Qiaoli; Yuan, Junhu; Liu, Lixia; Sun, Wenyang; Jiang, Yingdi; Zhao, Shengguo; Zhang, Shengwei; Huang, Wangzhou; Gun, Shuangbao

    2016-01-01

    The swine leukocyte antigens (SLAs) are the multigene families related to immune responses. Little is known about the effect of the DRA gene on diarrheal disease. This study reported the genetic diversity of the DRA gene in exons 1, 3 and 4 in 290 Chinese Yantai black pigs. No variation was identified in exon 3. In exon 1, three genotypes and two alleles were identified, generated by two single nucleotide polymorphisms (SNPs). In exon 4, there were eight genotypes and five alleles containing seven SNPs were detected with four SNPs being novel SNPs. The low polymorphism found in swine DRA is consistent with the concept that the DRA gene is highly conserved among all mammalian species. Statistical analyses indicated that the genotypes of exon 1 were not significantly associated with piglet diarrhea (p > 0.05); however, genotypes C4C4 (1.80 ± 0.33) and A4E4 (1.66 ± 0.25) of exon 4 were significantly susceptible to diarrhea (p < 0.01). These indicate that the particular genotypes of the DRA gene are susceptible to diarrheal disease, which provides valuable information for disease-resistance breeding in swine. PMID:27429004

  20. Effect of Genetic Diversity in Swine Leukocyte Antigen-DRA Gene on Piglet Diarrhea.

    PubMed

    Huang, Xiaoyu; Yang, Qiaoli; Yuan, Junhu; Liu, Lixia; Sun, Wenyang; Jiang, Yingdi; Zhao, Shengguo; Zhang, Shengwei; Huang, Wangzhou; Gun, Shuangbao

    2016-01-01

    The swine leukocyte antigens (SLAs) are the multigene families related to immune responses. Little is known about the effect of the DRA gene on diarrheal disease. This study reported the genetic diversity of the DRA gene in exons 1, 3 and 4 in 290 Chinese Yantai black pigs. No variation was identified in exon 3. In exon 1, three genotypes and two alleles were identified, generated by two single nucleotide polymorphisms (SNPs). In exon 4, there were eight genotypes and five alleles containing seven SNPs were detected with four SNPs being novel SNPs. The low polymorphism found in swine DRA is consistent with the concept that the DRA gene is highly conserved among all mammalian species. Statistical analyses indicated that the genotypes of exon 1 were not significantly associated with piglet diarrhea (p > 0.05); however, genotypes C₄C₄ (1.80 ± 0.33) and A₄E₄ (1.66 ± 0.25) of exon 4 were significantly susceptible to diarrhea (p < 0.01). These indicate that the particular genotypes of the DRA gene are susceptible to diarrheal disease, which provides valuable information for disease-resistance breeding in swine. PMID:27429004

  1. Ovine leukocyte profiles do not associate with variation in the prion gene, but are breed-dependent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Prion genotype in sheep confer resistance to scrapie. In cattle, lymphocyte profile has been found to be associated with prion genotype. Therefore, the aim of this study was to determine if variations in the sheep prion gene were associated with leukocyte populations as measured by complete blood ce...

  2. Hypermethylation of gene promoters in peripheral blood leukocytes in humans long term after radiation exposure.

    PubMed

    Kuzmina, Nina S; Lapteva, Nellya Sh; Rubanovich, Alexander V

    2016-04-01

    Some human genes known to undergo age-related promoter hypermethylation. These epigenetic modifications are similar to those occurring in the course of certain diseases, e.g. some types of cancer, which in turn may also associate with age. Given external genotoxic factors may additionally contribute to hypermethylation, this study was designed to analyzes, using methylation-sensitive polymerase chain reaction (PCR), the CpG island hypermethylation in RASSF1A, CDKN2A (including p16/INK4A and p14/ARF) and GSTP1 promoters in peripheral blood leukocytes of individuals exposed to ionizing radiation long time ago. One hundred and twenty-four irradiated subjects (24-77 years old at sampling: 83 Chernobyl Nuclear Power Plant clean-up workers, 21 nuclear workers, 20 residents of territories with radioactive contamination) and 208 unirradiated volunteers (19-77 years old at sampling) were enrolled. In addition, 74 non-exposed offspring (2-51 years old at sampling) born to irradiated parents were examined. The frequency of individuals displaying promoter methylation of at least one gene in exposed group was significantly higher as compared to the control group (OR=5.44, 95% CI=2.62-11.76, p=3.9×10(-7)). No significant difference was found between the frequency of subjects with the revealed promoter methylation in the group of offspring born to irradiated parents and in the control group. The increase in the number of methylated loci of RASSF1A and p14/ARF was associated with age (β=0.242; p=1.7×10(-5)). In contrast, hypermethylation of p16/INK4A and GSTP1 genes correlated with the fact of radiation exposure only (β=0.290; p=1.7×10(-7)). The latter finding demonstrates that methylation changes in blood leukocytes of healthy subjects exposed to radiation resemble those reported in human malignancies. Additional studies are required to identify the dose-response of epigenetic markers specifically associating with radiation-induced premature aging and/or with the development

  3. Rewiring mesenchymal stem cell lineage specification by switching the biophysical microenvironment

    NASA Astrophysics Data System (ADS)

    Lee, Junmin; Abdeen, Amr A.; Kilian, Kristopher A.

    2014-06-01

    The propensity of stem cells to specify and commit to a particular lineage program is guided by dynamic biophysical and biochemical signals that are temporally regulated. However, most in vitro studies rely on ``snapshots'' of cell state under static conditions. Here we asked whether changing the biophysical aspects of the substrate could modulate the degree of mesenchymal stem cell (MSC) lineage specification. We chose to explore two diverse differentiation outcomes: MSC osteogenesis and trans-differentiation to neuron-like cells. MSCs were cultured on soft (~0.5 kPa) or stiff (~40 kPa) hydrogels followed by transfer to gels of the opposite stiffness. MSCs on soft gels express elevated neurogenesis markers while MSCs on stiff substrates express elevated osteogenesis markers. Transfer of MSCs from soft to stiff or stiff to soft substrates led to a switch in lineage specification. However, MSCs transferred from stiff to soft substrates maintained elevated osteogenesis markers, suggesting a degree of irreversible activation. Transferring MSCs to micropatterned substrates reveal geometric cues that further modulate lineage reversal. Taken together, this study demonstrates that MSCs remain susceptible to the biophysical properties of the extracellular matrix--even after several weeks of culture--and can redirect lineage specification in response to changes in the microenvironment.

  4. Lineage-specific detection of influenza B virus using real-time polymerase chain reaction with melting curve analysis.

    PubMed

    Tewawong, Nipaporn; Chansaenroj, Jira; Klinfueng, Sirapa; Vichiwattana, Preeyaporn; Korkong, Sumeth; Thongmee, Thanunrat; Theamboonlers, Apiradee; Payungporn, Sunchai; Vongpunsawad, Sompong; Poovorawan, Yong

    2016-06-01

    Influenza B viruses comprise two lineages, Victoria (B/Vic) and Yamagata (B/Yam), which co-circulate globally. The surveillance data on influenza B virus lineages in many countries often underestimate the true prevalence due to the lack of a rapid, accurate, and cost-effective method for virus detection. We have developed a real-time PCR with melting curve analysis for lineage-specific differential detection of influenza B virus. By amplifying a region of the hemagglutinin gene using real-time PCR with SYBR Green I dye, B/Vic and B/Yam could be differentiated based on their melting temperature peaks. This method was efficient (B/Vic = 93.2 %; B/Yam 97.7 %), sensitive (B/Vic, 94.6 %; B/Yam, 96.3 %), and specific (B/Vic, 97.7 %; B/Yam, 97.1 %). The lower detection limit was 10(2) copies per microliter. The assay was evaluated using 756 respiratory specimens that were positive for influenza B virus, obtained between 2010 and 2015. The incidence of influenza B virus was approximately 18.9 % of all influenza cases, and the percentage was highest among children aged 6-17 years (7.57 %). The overall percentage of mismatched influenza B vaccine was 21.1 %. Our findings suggest that real-time PCR with melting curve analysis can provide a rapid, simple, and sensitive lineage-specific influenza B virus screening method to facilitate influenza surveillance. PMID:26923928

  5. Recombination and lineage-specific gene loss in the aflatoxin gene cluster of Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aflatoxins produced by Aspergillus flavus are potent carcinogens that contaminate agricultural crops. Recent efforts to reduce aflatoxin concentrations in crops have focused on biological control using nonaflatoxigenic A. flavus strains AF36 (= NRRL 18543) and NRRL 21882 (the active component of af...

  6. Lineage-Specific Reductions of Plastid Genomes in an Orchid Tribe with Partially and Fully Mycoheterotrophic Species.

    PubMed

    Feng, Yan-Lei; Wicke, Susann; Li, Jian-Wu; Han, Yu; Lin, Choun-Sea; Li, De-Zhu; Zhou, Ting-Ting; Huang, Wei-Chang; Huang, Lu-Qi; Jin, Xiao-Hua

    2016-01-01

    The plastid genome (plastome) of heterotrophic plants like mycoheterotrophs and parasites shows massive gene losses in consequence to the relaxation of functional constraints on photosynthesis. To understand the patterns of this convergent plastome reduction syndrome in heterotrophic plants, we studied 12 closely related orchids of three different lifeforms from the tribe Neottieae (Orchidaceae). We employ a comparative genomics approach to examine structural and selectional changes in plastomes within Neottieae. Both leafy and leafless heterotrophic species have functionally reduced plastid genome. Our analyses show that genes for the NAD(P)H dehydrogenase complex, the photosystems, and the RNA polymerase have been lost functionally multiple times independently. The physical reduction proceeds in a highly lineage-specific manner, accompanied by structural reconfigurations such as inversions or modifications of the large inverted repeats. Despite significant but minor selectional changes, all retained genes continue to evolve under purifying selection. All leafless Neottia species, including both visibly green and nongreen members, are fully mycoheterotrophic, likely evolved from leafy and partially mycoheterotrophic species. The plastomes of Neottieae span many stages of plastome degradation, including the longest plastome of a mycoheterotroph, providing invaluable insights into the mechanisms of plastome evolution along the transition from autotrophy to full mycoheterotrophy. PMID:27412609

  7. Flagellated Algae Protein Evolution Suggests the Prevalence of Lineage-Specific Rules Governing Evolutionary Rates of Eukaryotic Proteins

    PubMed Central

    Chang, Ting-Yan; Liao, Ben-Yang

    2013-01-01

    Understanding the general rules governing the rate of protein evolution is fundamental to evolutionary biology. However, attempts to address this issue in yeasts and mammals have revealed considerable differences in the relative importance of determinants for protein evolutionary rates. This phenomenon was previously explained by the fact that yeasts and mammals are different in many cellular and genomic properties. Flagellated algae species have several cellular and genomic characteristics that are intermediate between yeasts and mammals. Using partial correlation analyses on the evolution of 6,921 orthologous proteins from Chlamydomonas reinhardtii and Volvox carteri, we examined factors influencing evolutionary rates of proteins in flagellated algae. Previous studies have shown that mRNA abundance and gene compactness are strong determinants for protein evolutionary rates in yeasts and mammals, respectively. We show that both factors also influence algae protein evolution with mRNA abundance having a larger impact than gene compactness on the rates of algae protein evolution. More importantly, among all the factors examined, coding sequence (CDS) length has the strongest (positive) correlation with protein evolutionary rates. This correlation between CDS length and the rates of protein evolution is not due to alignment-related issues or domain density. These results suggest no simple and universal rules governing protein evolutionary rates across different eukaryotic lineages. Instead, gene properties influence the rate of protein evolution in a lineage-specific manner. PMID:23563973

  8. Lineage-Specific Reductions of Plastid Genomes in an Orchid Tribe with Partially and Fully Mycoheterotrophic Species

    PubMed Central

    Feng, Yan-Lei; Wicke, Susann; Li, Jian-Wu; Han, Yu; Lin, Choun-Sea; Li, De-Zhu; Zhou, Ting-Ting; Huang, Wei-Chang; Huang, Lu-Qi; Jin, Xiao-Hua

    2016-01-01

    The plastid genome (plastome) of heterotrophic plants like mycoheterotrophs and parasites shows massive gene losses in consequence to the relaxation of functional constraints on photosynthesis. To understand the patterns of this convergent plastome reduction syndrome in heterotrophic plants, we studied 12 closely related orchids of three different lifeforms from the tribe Neottieae (Orchidaceae). We employ a comparative genomics approach to examine structural and selectional changes in plastomes within Neottieae. Both leafy and leafless heterotrophic species have functionally reduced plastid genome. Our analyses show that genes for the NAD(P)H dehydrogenase complex, the photosystems, and the RNA polymerase have been lost functionally multiple times independently. The physical reduction proceeds in a highly lineage-specific manner, accompanied by structural reconfigurations such as inversions or modifications of the large inverted repeats. Despite significant but minor selectional changes, all retained genes continue to evolve under purifying selection. All leafless Neottia species, including both visibly green and nongreen members, are fully mycoheterotrophic, likely evolved from leafy and partially mycoheterotrophic species. The plastomes of Neottieae span many stages of plastome degradation, including the longest plastome of a mycoheterotroph, providing invaluable insights into the mechanisms of plastome evolution along the transition from autotrophy to full mycoheterotrophy. PMID:27412609

  9. Flagellated algae protein evolution suggests the prevalence of lineage-specific rules governing evolutionary rates of eukaryotic proteins.

    PubMed

    Chang, Ting-Yan; Liao, Ben-Yang

    2013-01-01

    Understanding the general rules governing the rate of protein evolution is fundamental to evolutionary biology. However, attempts to address this issue in yeasts and mammals have revealed considerable differences in the relative importance of determinants for protein evolutionary rates. This phenomenon was previously explained by the fact that yeasts and mammals are different in many cellular and genomic properties. Flagellated algae species have several cellular and genomic characteristics that are intermediate between yeasts and mammals. Using partial correlation analyses on the evolution of 6,921 orthologous proteins from Chlamydomonas reinhardtii and Volvox carteri, we examined factors influencing evolutionary rates of proteins in flagellated algae. Previous studies have shown that mRNA abundance and gene compactness are strong determinants for protein evolutionary rates in yeasts and mammals, respectively. We show that both factors also influence algae protein evolution with mRNA abundance having a larger impact than gene compactness on the rates of algae protein evolution. More importantly, among all the factors examined, coding sequence (CDS) length has the strongest (positive) correlation with protein evolutionary rates. This correlation between CDS length and the rates of protein evolution is not due to alignment-related issues or domain density. These results suggest no simple and universal rules governing protein evolutionary rates across different eukaryotic lineages. Instead, gene properties influence the rate of protein evolution in a lineage-specific manner. PMID:23563973

  10. AMPK governs lineage specification through Tfeb-dependent regulation of lysosomes

    PubMed Central

    Young, Nathan P.; Kamireddy, Anwesh; Van Nostrand, Jeanine L.; Eichner, Lillian J.; Shokhirev, Maxim Nikolaievich; Dayn, Yelena; Shaw, Reuben J.

    2016-01-01

    Faithful execution of developmental programs relies on the acquisition of unique cell identities from pluripotent progenitors, a process governed by combinatorial inputs from numerous signaling cascades that ultimately dictate lineage-specific transcriptional outputs. Despite growing evidence that metabolism is integrated with many molecular networks, how pathways that control energy homeostasis may affect cell fate decisions is largely unknown. Here, we show that AMP-activated protein kinase (AMPK), a central metabolic regulator, plays critical roles in lineage specification. Although AMPK-deficient embryonic stem cells (ESCs) were normal in the pluripotent state, these cells displayed profound defects upon differentiation, failing to generate chimeric embryos and preferentially adopting an ectodermal fate at the expense of the endoderm during embryoid body (EB) formation. AMPK−/− EBs exhibited reduced levels of Tfeb, a master transcriptional regulator of lysosomes, leading to diminished endolysosomal function. Remarkably, genetic loss of Tfeb also yielded endodermal defects, while AMPK-null ESCs overexpressing this transcription factor normalized their differential potential, revealing an intimate connection between Tfeb/lysosomes and germ layer specification. The compromised endolysosomal system resulting from AMPK or Tfeb inactivation blunted Wnt signaling, while up-regulating this pathway restored expression of endodermal markers. Collectively, these results uncover the AMPK pathway as a novel regulator of cell fate determination during differentiation. PMID:26944679

  11. Lineage-specific splicing of a brain-enriched alternative exon promotes glioblastoma progression

    PubMed Central

    Ferrarese, Roberto; Harsh, Griffith R.; Yadav, Ajay K.; Bug, Eva; Maticzka, Daniel; Reichardt, Wilfried; Dombrowski, Stephen M.; Miller, Tyler E.; Masilamani, Anie P.; Dai, Fangping; Kim, Hyunsoo; Hadler, Michael; Scholtens, Denise M.; Yu, Irene L.Y.; Beck, Jürgen; Srinivasasainagendra, Vinodh; Costa, Fabrizio; Baxan, Nicoleta; Pfeifer, Dietmar; von Elverfeldt, Dominik; Backofen, Rolf; Weyerbrock, Astrid; Duarte, Christine W.; He, Xiaolin; Prinz, Marco; Chandler, James P.; Vogel, Hannes; Chakravarti, Arnab; Rich, Jeremy N.; Carro, Maria S.; Bredel, Markus

    2014-01-01

    Tissue-specific alternative splicing is critical for the emergence of tissue identity during development, yet the role of this process in malignant transformation is undefined. Tissue-specific splicing involves evolutionarily conserved, alternative exons that represent only a minority of the total alternative exons identified. Many of these conserved exons have functional features that influence signaling pathways to profound biological effect. Here, we determined that lineage-specific splicing of a brain-enriched cassette exon in the membrane-binding tumor suppressor annexin A7 (ANXA7) diminishes endosomal targeting of the EGFR oncoprotein, consequently enhancing EGFR signaling during brain tumor progression. ANXA7 exon splicing was mediated by the ribonucleoprotein PTBP1, which is normally repressed during neuronal development. PTBP1 was highly expressed in glioblastomas due to loss of a brain-enriched microRNA (miR-124) and to PTBP1 amplification. The alternative ANXA7 splicing trait was present in precursor cells, suggesting that glioblastoma cells inherit the trait from a potential tumor-initiating ancestor and that these cells exploit this trait through accumulation of mutations that enhance EGFR signaling. Our data illustrate that lineage-specific splicing of a tissue-regulated alternative exon in a constituent of an oncogenic pathway eliminates tumor suppressor functions and promotes glioblastoma progression. This paradigm may offer a general model as to how tissue-specific regulatory mechanisms can reprogram normal developmental processes into oncogenic ones. PMID:24865424

  12. Lineage-specific splicing of a brain-enriched alternative exon promotes glioblastoma progression.

    PubMed

    Ferrarese, Roberto; Harsh, Griffith R; Yadav, Ajay K; Bug, Eva; Maticzka, Daniel; Reichardt, Wilfried; Dombrowski, Stephen M; Miller, Tyler E; Masilamani, Anie P; Dai, Fangping; Kim, Hyunsoo; Hadler, Michael; Scholtens, Denise M; Yu, Irene L Y; Beck, Jürgen; Srinivasasainagendra, Vinodh; Costa, Fabrizio; Baxan, Nicoleta; Pfeifer, Dietmar; von Elverfeldt, Dominik; Backofen, Rolf; Weyerbrock, Astrid; Duarte, Christine W; He, Xiaolin; Prinz, Marco; Chandler, James P; Vogel, Hannes; Chakravarti, Arnab; Rich, Jeremy N; Carro, Maria S; Bredel, Markus

    2014-07-01

    Tissue-specific alternative splicing is critical for the emergence of tissue identity during development, yet the role of this process in malignant transformation is undefined. Tissue-specific splicing involves evolutionarily conserved, alternative exons that represent only a minority of the total alternative exons identified. Many of these conserved exons have functional features that influence signaling pathways to profound biological effect. Here, we determined that lineage-specific splicing of a brain-enriched cassette exon in the membrane-binding tumor suppressor annexin A7 (ANXA7) diminishes endosomal targeting of the EGFR oncoprotein, consequently enhancing EGFR signaling during brain tumor progression. ANXA7 exon splicing was mediated by the ribonucleoprotein PTBP1, which is normally repressed during neuronal development. PTBP1 was highly expressed in glioblastomas due to loss of a brain-enriched microRNA (miR-124) and to PTBP1 amplification. The alternative ANXA7 splicing trait was present in precursor cells, suggesting that glioblastoma cells inherit the trait from a potential tumor-initiating ancestor and that these cells exploit this trait through accumulation of mutations that enhance EGFR signaling. Our data illustrate that lineage-specific splicing of a tissue-regulated alternative exon in a constituent of an oncogenic pathway eliminates tumor suppressor functions and promotes glioblastoma progression. This paradigm may offer a general model as to how tissue-specific regulatory mechanisms can reprogram normal developmental processes into oncogenic ones. PMID:24865424

  13. Lineage-specific function of Engrailed-2 in the progression of chronic myelogenous leukemia to T-cell blast crisis

    PubMed Central

    Abollo-Jiménez, Fernando; Campos-Sánchez, Elena; Toboso-Navasa, Amparo; Vicente-Dueñas, Carolina; González-Herrero, Inés; Alonso-Escudero, Esther; González, Marcos; Segura, Víctor; Blanco, Óscar; Martínez-Climent, José Ángel; Sánchez-García, Isidro; Cobaleda, César

    2014-01-01

    In hematopoietic malignancies, oncogenic alterations interfere with cellular differentiation and lead to tumoral development. Identification of the proteins regulating differentiation is essential to understand how they are altered in malignancies. Chronic myelogenous leukemia (CML) is a biphasic disease initiated by an alteration taking place in hematopoietic stem cells. CML progresses to a blast crisis (BC) due to a secondary differentiation block in any of the hematopoietic lineages. However, the molecular mechanisms of CML evolution to T-cell BC remain unclear. Here, we have profiled the changes in DNA methylation patterns in human samples from BC-CML, in order to identify genes whose expression is epigenetically silenced during progression to T-cell lineage-specific BC. We have found that the CpG-island of the ENGRAILED-2 (EN2) gene becomes methylated in this progression. Afterwards, we demonstrate that En2 is expressed during T-cell development in mice and humans. Finally, we further show that genetic deletion of En2 in a CML transgenic mouse model induces a T-cell lineage BC that recapitulates human disease. These results identify En2 as a new regulator of T-cell differentiation whose disruption induces a malignant T-cell fate in CML progression, and validate the strategy used to identify new developmental regulators of hematopoiesis. PMID:24675889

  14. A Role for RE-1-Silencing Transcription Factor in Embryonic Stem Cells Cardiac Lineage Specification.

    PubMed

    Aksoy, Irene; Marcy, Guillaume; Chen, Jiaxuan; Divakar, Ushashree; Kumar, Vibhor; John-Sanchez, Daniel; Rahmani, Mehran; Buckley, Noel J; Stanton, Lawrence W

    2016-04-01

    During development, lineage specification is controlled by several signaling pathways involving various transcription factors (TFs). Here, we studied the RE-1-silencing transcription factor (REST) and identified an important role of this TF in cardiac differentiation. Using mouse embryonic stem cells (ESC) to model development, we found that REST knockout cells lost the ability to differentiate into the cardiac lineage. Detailed analysis of specific lineage markers expression showed selective downregulation of endoderm markers in REST-null cells, thus contributing to a loss of cardiogenic signals. REST regulates cardiac differentiation of ESCs by negatively regulating the Wnt/β-catenin signaling pathway and positively regulating the cardiogenic TF Gata4. We propose here a new role for REST in cell fate specification besides its well-known repressive role of neuronal differentiation. PMID:26864965

  15. Gene copy number variations in the leukocyte genome of hepatocellular carcinoma patients with integrated hepatitis B virus DNA

    PubMed Central

    Xu, Guixia; Cheng, Kai; Cao, Guangwen; Wu, Mengchao; Cheng, Shuqun; Liu, Shanrong

    2016-01-01

    Integration of hepatitis B virus (HBV) DNA into the human liver cell genome is believed to promote HBV-related carcinogenesis. This study aimed to quantify the integration of HBV DNA into the leukocyte genome in hepatocellular carcinoma (HCC) patients in order to identify potential biomarkers for HBV-related diseases. Whole-genome comparative genomic hybridization (CGH) chip array analyses were performed to screen gene copy number variations (CNV) in the leukocyte genome, and the results were confirmed by quantitative polymerase chain reaction (qPCR). The commonly detected regions included chromosome arms 19p, 5q, 1q and 15p, where 200 copy number gain events and 270 copy number loss events were noted. In particular, gains were observed in 5q35.3 (OR4F3) and 19p13.3 (OR4F17) in 90% of the samples. Successful homologous recombination of OR4F3 and the HBV P gene was demonstrated, and the amplification at 5q35.3 is potentially associated with the integration of HBV P gene into natural killer cells isolated from peripheral blood mononuclear cells (PBMCs). Receiver operating characteristic (ROC) curve analysis indicated that the combination of OR4F3 and OR4F17 a novel potential biomarker of HBV-related diseases. PMID:26769853

  16. Effect of lineage-specific metabolic traits of Lactobacillus reuteri on sourdough microbial ecology.

    PubMed

    Lin, Xiaoxi B; Gänzle, Michael G

    2014-09-01

    This study determined the effects of specific metabolic traits of Lactobacillus reuteri on its competitiveness in sourdoughs. The competitiveness of lactobacilli in sourdough generally depends on their growth rate; acid resistance additionally contributes to competitiveness in sourdoughs with long fermentation times. Glycerol metabolism via glycerol dehydratase (gupCDE) accelerates growth by the regeneration of reduced cofactors; glutamate metabolism via glutamate decarboxylase (gadB) increases acid resistance by generating a proton motive force. Glycerol and glutamate metabolisms are lineage-specific traits in L. reuteri; therefore, this study employed glycerol dehydratase-positive sourdough isolates of human-adapted L. reuteri lineage I, glutamate decarboxylase-positive strains of rodent-adapted L. reuteri lineage II, as well as mutants with deletions in gadB or gupCDE. The competitivenesses of the strains were quantified by inoculation of wheat and sorghum sourdoughs with defined strains, followed by propagation of doughs with a 10% inoculum and 12-h or 72-h fermentation cycles. Lineage I L. reuteri strains dominated sourdoughs propagated with 12-h fermentation cycles; lineage II L. reuteri strains dominated sourdoughs propagated with 72-h fermentation cycles. L. reuteri 100-23ΔgadB was outcompeted by its wild-type strain in sourdoughs fermented with 72-h fermentation cycles; L. reuteri FUA3400ΔgupCDE was outcompeted by its wild-type strain in sourdoughs fermented with both 12-h and 72-h fermentation cycles. Competition experiments with isogenic pairs of strains resulted in a constant rate of strain displacement of the less competitive mutant strain. In conclusion, lineage-specific traits of L. reuteri determine the competitiveness of this species in sourdough fermentations. PMID:25015888

  17. Lineage-Specific Changes in Biomarkers in Great Apes and Humans.

    PubMed

    Ronke, Claudius; Dannemann, Michael; Halbwax, Michel; Fischer, Anne; Helmschrodt, Christin; Brügel, Mathias; André, Claudine; Atencia, Rebeca; Mugisha, Lawrence; Scholz, Markus; Ceglarek, Uta; Thiery, Joachim; Pääbo, Svante; Prüfer, Kay; Kelso, Janet

    2015-01-01

    Although human biomedical and physiological information is readily available, such information for great apes is limited. We analyzed clinical chemical biomarkers in serum samples from 277 wild- and captive-born great apes and from 312 healthy human volunteers as well as from 20 rhesus macaques. For each individual, we determined a maximum of 33 markers of heart, liver, kidney, thyroid and pancreas function, hemoglobin and lipid metabolism and one marker of inflammation. We identified biomarkers that show differences between humans and the great apes in their average level or activity. Using the rhesus macaques as an outgroup, we identified human-specific differences in the levels of bilirubin, cholinesterase and lactate dehydrogenase, and bonobo-specific differences in the level of apolipoprotein A-I. For the remaining twenty-nine biomarkers there was no evidence for lineage-specific differences. In fact, we find that many biomarkers show differences between individuals of the same species in different environments. Of the four lineage-specific biomarkers, only bilirubin showed no differences between wild- and captive-born great apes. We show that the major factor explaining the human-specific difference in bilirubin levels may be genetic. There are human-specific changes in the sequence of the promoter and the protein-coding sequence of uridine diphosphoglucuronosyltransferase 1 (UGT1A1), the enzyme that transforms bilirubin and toxic plant compounds into water-soluble, excretable metabolites. Experimental evidence that UGT1A1 is down-regulated in the human liver suggests that changes in the promoter may be responsible for the human-specific increase in bilirubin. We speculate that since cooking reduces toxic plant compounds, consumption of cooked foods, which is specific to humans, may have resulted in relaxed constraint on UGT1A1 which has in turn led to higher serum levels of bilirubin in humans. PMID:26247603

  18. Effect of Lineage-Specific Metabolic Traits of Lactobacillus reuteri on Sourdough Microbial Ecology

    PubMed Central

    Lin, Xiaoxi B.

    2014-01-01

    This study determined the effects of specific metabolic traits of Lactobacillus reuteri on its competitiveness in sourdoughs. The competitiveness of lactobacilli in sourdough generally depends on their growth rate; acid resistance additionally contributes to competitiveness in sourdoughs with long fermentation times. Glycerol metabolism via glycerol dehydratase (gupCDE) accelerates growth by the regeneration of reduced cofactors; glutamate metabolism via glutamate decarboxylase (gadB) increases acid resistance by generating a proton motive force. Glycerol and glutamate metabolisms are lineage-specific traits in L. reuteri; therefore, this study employed glycerol dehydratase-positive sourdough isolates of human-adapted L. reuteri lineage I, glutamate decarboxylase-positive strains of rodent-adapted L. reuteri lineage II, as well as mutants with deletions in gadB or gupCDE. The competitivenesses of the strains were quantified by inoculation of wheat and sorghum sourdoughs with defined strains, followed by propagation of doughs with a 10% inoculum and 12-h or 72-h fermentation cycles. Lineage I L. reuteri strains dominated sourdoughs propagated with 12-h fermentation cycles; lineage II L. reuteri strains dominated sourdoughs propagated with 72-h fermentation cycles. L. reuteri 100-23ΔgadB was outcompeted by its wild-type strain in sourdoughs fermented with 72-h fermentation cycles; L. reuteri FUA3400ΔgupCDE was outcompeted by its wild-type strain in sourdoughs fermented with both 12-h and 72-h fermentation cycles. Competition experiments with isogenic pairs of strains resulted in a constant rate of strain displacement of the less competitive mutant strain. In conclusion, lineage-specific traits of L. reuteri determine the competitiveness of this species in sourdough fermentations. PMID:25015888

  19. Lineage-Specific Changes in Biomarkers in Great Apes and Humans

    PubMed Central

    Ronke, Claudius; Dannemann, Michael; Halbwax, Michel; Fischer, Anne; Helmschrodt, Christin; Brügel, Mathias; André, Claudine; Atencia, Rebeca; Mugisha, Lawrence; Scholz, Markus; Ceglarek, Uta; Thiery, Joachim; Pääbo, Svante; Prüfer, Kay; Kelso, Janet

    2015-01-01

    Although human biomedical and physiological information is readily available, such information for great apes is limited. We analyzed clinical chemical biomarkers in serum samples from 277 wild- and captive-born great apes and from 312 healthy human volunteers as well as from 20 rhesus macaques. For each individual, we determined a maximum of 33 markers of heart, liver, kidney, thyroid and pancreas function, hemoglobin and lipid metabolism and one marker of inflammation. We identified biomarkers that show differences between humans and the great apes in their average level or activity. Using the rhesus macaques as an outgroup, we identified human-specific differences in the levels of bilirubin, cholinesterase and lactate dehydrogenase, and bonobo-specific differences in the level of apolipoprotein A-I. For the remaining twenty-nine biomarkers there was no evidence for lineage-specific differences. In fact, we find that many biomarkers show differences between individuals of the same species in different environments. Of the four lineage-specific biomarkers, only bilirubin showed no differences between wild- and captive-born great apes. We show that the major factor explaining the human-specific difference in bilirubin levels may be genetic. There are human-specific changes in the sequence of the promoter and the protein-coding sequence of uridine diphosphoglucuronosyltransferase 1 (UGT1A1), the enzyme that transforms bilirubin and toxic plant compounds into water-soluble, excretable metabolites. Experimental evidence that UGT1A1 is down-regulated in the human liver suggests that changes in the promoter may be responsible for the human-specific increase in bilirubin. We speculate that since cooking reduces toxic plant compounds, consumption of cooked foods, which is specific to humans, may have resulted in relaxed constraint on UGT1A1 which has in turn led to higher serum levels of bilirubin in humans. PMID:26247603

  20. H3K27me3 Does Not Orchestrate the Expression of Lineage-Specific Markers in hESC-Derived Hepatocytes In Vitro.

    PubMed

    Vanhove, Jolien; Pistoni, Mariaelena; Welters, Marc; Eggermont, Kristel; Vanslembrouck, Veerle; Helsen, Nicky; Boon, Ruben; Najimi, Mustapha; Sokal, Etienne; Collas, Philippe; Voncken, J Willem; Verfaillie, Catherine M

    2016-08-01

    Although pluripotent stem cells can be differentiated into the hepatocyte lineages, such cells retain an immature phenotype. As the chromatin state of regulatory regions controls spatiotemporal gene expression during development, we evaluated changes in epigenetic histone marks in lineage-specific genes throughout in vitro hepatocyte differentiation from human embryonic stem cells (hESCs). Active acetylation and methylation marks at promoters and enhancers correlated with progressive changes in gene expression. However, repression-associated H3K27me3 marks at these control regions showed an inverse correlation with gene repression during transition from hepatic endoderm to a hepatocyte-like state. Inhibitor of Enhancer of Zeste Homolog 2 (EZH2) reduced H3K27me3 decoration but did not improve hepatocyte maturation. Thus, H3K27me3 at regulatory regions does not regulate transcription and appears dispensable for hepatocyte lineage differentiation of hESCs in vitro. PMID:27477635

  1. The effects of short term dietary restriction on haematological responses and leukocyte gene expression of anovulatory and ovulatory beef heifers.

    PubMed

    Matthews, Daragh; Waters, Sinéad M; Diskin, Michael G; Kenny, David A; Morris, Dermot G; Earley, Bernadette

    2015-02-01

    The study objective was to characterise the impact of negative energy balance (NEB) on immune-stress responsiveness in beef heifers. A short term (18-day) dietary restriction model was used. Dietary restriction (0.4 maintenance (Mn) energy requirements) induced abrupt onset of anoestrus in nine heifers (Restricted Anovulatory; RA) while nineteen heifers maintained oestrous cyclicity (Restricted Ovulatory; RO). In addition a control (C) group of 12 heifers received a higher level of feeding (1.2 Mn). Haematological related biomarkers of husbandry stress, leukocyte gene expression of seven cytokine genes and five immunological biomarkers were investigated. After 18 days of differential feeding of the heifers alterations in eosinophil and monocyte numbers and altered expression of CXCL8, IL2 and TNFα could be attributed to diet restriction. More specifically, changes in these five variables were found in heifers that became anovulatory (RA) and are therefore considered to be more sensitive biomarkers to an energy deficit. PMID:25496833

  2. The Lineage Specification of Mesenchymal Stem Cells Is Directed by the Rate of Fluid Shear Stress.

    PubMed

    Lu, Juan; Fan, Yijuan; Gong, Xiaoyuan; Zhou, Xin; Yi, Caixia; Zhang, Yinxing; Pan, Jun

    2016-08-01

    The effective regulation of fluid shear stress (FSS) on the lineage specification of mesenchymal stem cells (MSCs) remains to be addressed. We hypothesized that when MSCs are recruited to musculoskeletal system following stimulation, their differentiation into osteogenic or chondrogenic cells is directed by the rate of FSS (ΔSS) through modulation of the mechanosensitive, cation-selective channels (MSCCs), intracellular calcium levels, and F-actin. To this end, MSCs were exposed to laminar FSS linearly increased from 0 to 10 dyn/cm(2) in 0, 2, or 20 min and maintained at 10 dyn/cm(2) for a total of 20 min (termed as ΔSS 0-0', 0-2', and 0-20', respectively, representing more physiological (0-0') and non-physiological (0-2' and 0-20') ΔSS treatments). Our results showed 0-0' facilitated MSC differentiation towards chondrogenic and not osteogenic phenotype, by promoting moderate intracellular calcium concentration ([Ca(2+) ]i ) increase from the calcium channels with the exception of MSCCs or intracellular calcium stores, and F-actin organization. In contrast, 0-2' promoted MSCs towards osteogenic and not chondrogenic phenotype, by inducing significant [Ca(2+) ]i increase mainly from the MSCCs, and F-actin assembly. However, 0-20' elicited the modest osteogenic and chondrogenic phenotypes, as it induced the lowest [Ca(2+) ]i increase mainly from MSCCs, and F-actin assembly. Our results suggest that compared to the more physiological ΔSS, the non-physiological ΔSS favors [Ca(2+) ]i influx from MSCCs. An appropriate non-physiological ΔSS (0-2') even elicits a large [Ca(2+) ]i influx from the MSCCs that reverses the lineage specification of MSCs, providing validation for the high mechanosensitivity of MSCs and guidance for training osteoporosis and osteoarthritis patients. J. Cell. Physiol. 231: 1752-1760, 2016. © 2015 Wiley Periodicals, Inc. PMID:26636289

  3. Leukocyte Gene Expression and Plasma Concentration in Multiple Sclerosis: Alteration of Transforming Growth Factor-βs, Claudin-11, and Matrix Metalloproteinase-2.

    PubMed

    Hassanzadeh, Gholamreza; Hosseini Quchani, Samaneh; Sahraian, Mohammad Ali; Abolhassani, Farid; Sadighi Gilani, Mohammad Ali; Dehghan Tarzjani, Masoomeh; Atoof, Fatemeh

    2016-08-01

    Multiple sclerosis is a neurodegenerative disease characterized by the present of leukocytes in the brain tissue and subsequently the formation of sclerotic plaques. Leukocytes penetration into the blood-brain barrier is related to several factors, such as, the conversion of leukocyte gene expression or plasma characteristics. In this frame, we explore alteration of matrix metalloproteinase-2 (MMP-2), transforming growth factor beta (TGF-β) family, and Claudin-11 (as a main myelin structural protein) in leukocytes and blood plasma of multiple sclerosis patients compared to the normal group. Blood samples were collected from thirteen men affected by MS and fifteen healthy men. Leukocyte gene expression was measured using real-time PCR and plasma parameters were examined by ELISA. The results of this study showed that the gene expression of Claudin-11 was significantly higher in MS group compared with normal. Interestingly, the MMP-2 pattern was similar to Claudin-11 and correlated positively with it. It was observed that, although the expressions of TGF-β1 and TGF-β2 are down-regulated in the leukocytes of subjects with MS, they showed higher levels of these cytokines in blood plasma. The plasma level of TGF-β3 in MS patients was higher than normal and correlated with Claudin-11 concentration. In conclusion, the aberrant pattern of Claudin-11, TGF-βs family, and MMP-2 expression in leukocytes of the MS patients was observed in this study. Moreover, the plasma levels of TGF-βs family increased in the MS group. The findings of this study provide clues for further investigations to assay MS pathogenesis. PMID:26768647

  4. Lineage-specific evolutionary rate in plants: Contributions of a screening for Cereus (Cactaceae)1

    PubMed Central

    Romeiro-Brito, Monique; Moraes, Evandro M.; Taylor, Nigel P.; Zappi, Daniela C.; Franco, Fernando F.

    2016-01-01

    Premise of the study: Predictable chloroplast DNA (cpDNA) sequences have been listed for the shallowest taxonomic studies in plants. We investigated whether plastid regions that vary between closely allied species could be applied for intraspecific studies and compared the variation of these plastid segments with two nuclear regions. Methods: We screened 16 plastid and two nuclear intronic regions for species of the genus Cereus (Cactaceae) at three hierarchical levels (species from different clades, species of the same clade, and allopatric populations). Results: Ten plastid regions presented interspecific variation, and six of them showed variation at the intraspecific level. The two nuclear regions showed both inter- and intraspecific variation, and in general they showed higher levels of variability in almost all hierarchical levels than the plastid segments. Discussion: Our data suggest no correspondence between variation of plastid regions at the interspecific and intraspecific level, probably due to lineage-specific variation in cpDNA, which appears to have less effect in nuclear data. Despite the heterogeneity in evolutionary rates of cpDNA, we highlight three plastid segments that may be considered in initial screenings in plant phylogeographic studies. PMID:26819857

  5. Lineage-specific transcriptional regulation of DICER by MITF in melanocytes.

    PubMed

    Levy, Carmit; Khaled, Mehdi; Robinson, Kathleen C; Veguilla, Rosa A; Chen, Po-Hao; Yokoyama, Satoru; Makino, Eiichi; Lu, Jun; Larue, Lionel; Beermann, Friedrich; Chin, Lynda; Bosenberg, Marcus; Song, Jun S; Fisher, David E

    2010-06-11

    DICER is a central regulator of microRNA maturation. However, little is known about mechanisms regulating its expression in development or disease. While profiling miRNA expression in differentiating melanocytes, two populations were observed: some upregulated at the pre-miRNA stage, and others upregulated as mature miRNAs (with stable pre-miRNA levels). Conversion of pre-miRNAs to fully processed miRNAs appeared to be dependent upon stimulation of DICER expression--an event found to occur via direct transcriptional targeting of DICER by the melanocyte master transcriptional regulator MITF. MITF binds and activates a conserved regulatory element upstream of DICER's transcriptional start site upon melanocyte differentiation. Targeted KO of DICER is lethal to melanocytes, at least partly via DICER-dependent processing of the pre-miRNA-17 approximately 92 cluster thus targeting BIM, a known proapoptotic regulator of melanocyte survival. These observations highlight a central mechanism underlying lineage-specific miRNA regulation which could exist for other cell types during development. PMID:20550935

  6. Lineage-specific molecular probing reveals novel diversity and ecological partitioning of haplosporidians

    PubMed Central

    Hartikainen, Hanna; Ashford, Oliver S; Berney, Cédric; Okamura, Beth; Feist, Stephen W; Baker-Austin, Craig; Stentiford, Grant D; Bass, David

    2014-01-01

    Haplosporidians are rhizarian parasites of mostly marine invertebrates. They include the causative agents of diseases of commercially important molluscs, including MSX disease in oysters. Despite their importance for food security, their diversity and distributions are poorly known. We used a combination of group-specific PCR primers to probe environmental DNA samples from planktonic and benthic environments in Europe, South Africa and Panama. This revealed several highly distinct novel clades, novel lineages within known clades and seasonal (spring vs autumn) and habitat-related (brackish vs littoral) variation in assemblage composition. High frequencies of haplosporidian lineages in the water column provide the first evidence for life cycles involving planktonic hosts, host-free stages or both. The general absence of haplosporidian lineages from all large online sequence data sets emphasises the importance of lineage-specific approaches for studying these highly divergent and diverse lineages. Combined with host-based field surveys, environmental sampling for pathogens will enhance future detection of known and novel pathogens and the assessment of disease risk. PMID:23966100

  7. FoxA1 as a lineage-specific oncogene in luminal type breast cancer

    SciTech Connect

    Yamaguchi, Noritaka; Ito, Emi; Azuma, Sakura; Honma, Reiko; Yanagisawa, Yuka; Nishikawa, Akira; Kawamura, Mika; Imai, Jun-ichi

    2008-01-25

    The forkhead transcription factor FoxA1 is thought to be involved in mammary tumorigenesis. However, the precise role of FoxA1 in breast cancer development is controversial. We examined expression of FoxA1 in 35 human breast cancer cell lines and compared it with that of ErbB2, a marker of poor prognosis in breast cancer. We found that FoxA1 is expressed at high levels in all ErbB2-positive cell lines and a subset of ErbB2-negative cell lines. Down-regulation of FoxA1 by RNA interference significantly suppressed proliferation of ErbB2-negative and FoxA1-positive breast cancer cell lines. Down-regulation of FoxA1 also enhanced the toxic effect of Herceptin on ErbB2-positive cell lines through induction of apoptosis. Taken together with previous data that FoxA1 is a marker of luminal cells in mammary gland, our present results suggest that FoxA1 plays an important role as a lineage-specific oncogene in proliferation of cancer cells derived from mammary luminal cells.

  8. Telomerase Protects Werner Syndrome Lineage-Specific Stem Cells from Premature Aging

    PubMed Central

    Cheung, Hoi-Hung; Liu, Xiaozhuo; Canterel-Thouennon, Lucile; Li, Lu; Edmonson, Catherine; Rennert, Owen M.

    2014-01-01

    Summary Werner syndrome (WS) patients exhibit premature aging predominantly in mesenchyme-derived tissues, but not in neural lineages, a consequence of telomere dysfunction and accelerated senescence. The cause of this lineage-specific aging remains unknown. Here, we document that reprogramming of WS fibroblasts to pluripotency elongated telomere length and prevented telomere dysfunction. To obtain mechanistic insight into the origin of tissue-specific aging, we differentiated iPSCs to mesenchymal stem cells (MSCs) and neural stem/progenitor cells (NPCs). We observed recurrence of premature senescence associated with accelerated telomere attrition and defective synthesis of the lagging strand telomeres in MSCs, but not in NPCs. We postulate this “aging” discrepancy is regulated by telomerase. Expression of hTERT or p53 knockdown ameliorated the accelerated aging phenotypein MSC, whereas inhibition of telomerase sensitized NPCs to DNA damage. Our findings unveil a role for telomerase in the protection of accelerated aging in a specific lineage of stem cells. PMID:24749076

  9. Therapeutic transdifferentiation of human fibroblasts into endothelial cells using forced expression of lineage-specific transcription factors

    PubMed Central

    Wong, Wing Tak; Cooke, John P

    2016-01-01

    Transdifferentiation is the direct conversion from one somatic cell type into another desired somatic cell type. This reprogramming method offers an attractive approach for regenerative medicine. Here, we demonstrate that neonatal fibroblasts can be transdifferentiated into endothelial cells using only four endothelial transcription factors, namely, ETV2, FLI1, GATA2, and KLF4. We observed a significant up-regulation of endothelial genes including KDR, CD31, CD144, and vWF in human neonatal foreskin (BJ) fibroblasts infected with the lentiviral construct encoding the open reading frame of the four transcription factors. We observed morphological changes in BJ fibroblasts from the fibroblastic spindle shape into a more endothelial-like cobblestone structures. Fluorescence-activated cell sorting analysis revealed that ~16% of the infected cells with the lentiviral constructs encoding 4F expressed CD31. The sorted cells were allowed to expand for 2 weeks and these cells were immunostained and found to express endothelial markers CD31. The induced endothelial cells also incorporated fluorescence-labeled acetylated low-density lipoprotein and efficiently formed capillary-like networks when seeded on Matrigel. These results suggested that the induced endothelial cells were functional in vitro. Taken together, we successfully demonstrated the direct conversion of human neonatal fibroblasts into endothelial cells by transduction of lentiviral constructs encoding endothelial lineage-specific transcription factors ETV2, FLI1, GATA2, and KLF4. The directed differentiation of fibroblasts into endothelial cells may have significant utility in diseases characterized by fibrosis and loss of microvasculature. PMID:27081470

  10. Integrative Genomic Analyses Identify BRF2 as a Novel Lineage-Specific Oncogene in Lung Squamous Cell Carcinoma

    PubMed Central

    Lockwood, William W.; Chari, Raj; Coe, Bradley P.; Thu, Kelsie L.; Garnis, Cathie; Malloff, Chad A.; Campbell, Jennifer; Williams, Ariane C.; Hwang, Dorothy; Zhu, Chang-Qi; Buys, Timon P. H.; Yee, John; English, John C.; MacAulay, Calum; Tsao, Ming-Sound; Gazdar, Adi F.; Minna, John D.; Lam, Stephen; Lam, Wan L.

    2010-01-01

    a key role in squamous cell lineage specificity of the disease. Our data suggest that genetic activation of BRF2 represents a unique mechanism of SqCC lung tumorigenesis through the increase of Pol III-mediated transcription. It can serve as a marker for lung SqCC and may provide a novel target for therapy. Please see later in the article for the Editors' Summary PMID:20668658

  11. Variability among the Most Rapidly Evolving Plastid Genomic Regions is Lineage-Specific: Implications of Pairwise Genome Comparisons in Pyrus (Rosaceae) and Other Angiosperms for Marker Choice

    PubMed Central

    Ter-Voskanyan, Hasmik; Allgaier, Martin; Borsch, Thomas

    2014-01-01

    Plastid genomes exhibit different levels of variability in their sequences, depending on the respective kinds of genomic regions. Genes are usually more conserved while noncoding introns and spacers evolve at a faster pace. While a set of about thirty maximum variable noncoding genomic regions has been suggested to provide universally promising phylogenetic markers throughout angiosperms, applications often require several regions to be sequenced for many individuals. Our project aims to illuminate evolutionary relationships and species-limits in the genus Pyrus (Rosaceae)—a typical case with very low genetic distances between taxa. In this study, we have sequenced the plastid genome of Pyrus spinosa and aligned it to the already available P. pyrifolia sequence. The overall p-distance of the two Pyrus genomes was 0.00145. The intergenic spacers between ndhC–trnV, trnR–atpA, ndhF–rpl32, psbM–trnD, and trnQ–rps16 were the most variable regions, also comprising the highest total numbers of substitutions, indels and inversions (potentially informative characters). Our comparative analysis of further plastid genome pairs with similar low p-distances from Oenothera (representing another rosid), Olea (asterids) and Cymbidium (monocots) showed in each case a different ranking of genomic regions in terms of variability and potentially informative characters. Only two intergenic spacers (ndhF–rpl32 and trnK–rps16) were consistently found among the 30 top-ranked regions. We have mapped the occurrence of substitutions and microstructural mutations in the four genome pairs. High AT content in specific sequence elements seems to foster frequent mutations. We conclude that the variability among the fastest evolving plastid genomic regions is lineage-specific and thus cannot be precisely predicted across angiosperms. The often lineage-specific occurrence of stem-loop elements in the sequences of introns and spacers also governs lineage-specific mutations

  12. Transcription analysis, physical mapping, and molecular characterization of a nonclassical human leukocyte antigen class I gene.

    PubMed Central

    Chorney, M J; Sawada, I; Gillespie, G A; Srivastava, R; Pan, J; Weissman, S M

    1990-01-01

    The human major histocompatibility complex contains approximately 20 class I genes, pseudogenes, and gene fragments. These include the genes for the three major transplantation antigens, HLA-A, HLA-B, and HLA-C, as well as a number of other genes or pseudogenes of unknown biological significance. Most of the latter have C + G-rich sequences in their 5' ends that are unmethylated in the B-lymphoblastoid cell line 3.1.0. We investigated one of these genes, HLA-H, in more detail. The gene is, overall, strongly homologous in sequence to HLA-A but differs in several potentially significant ways, including changes in conserved promoter sequences, a single-base deletion producing a translation termination codon in exon 4, and a region of sequence divergence downstream of the transcribed portion of the gene. Nevertheless, mouse L cells transfected with the gene accumulated small amounts of apparently full-length polyadenylated RNA. A portion of this RNA begins at the transcription site predicted by analogy to certain class I cDNA clones, while another portion appears to begin shortly upstream. L cells transfected with a hybrid gene containing the first three exons of HLA-H and the last five exons of HLA-B27 accumulated full-length HLA transcripts at the same level as cells transfected with an HLA-B27 gene; both levels are at least 15- to 20-fold higher than that directed by HLA-H alone. In addition, we isolated a cDNA clone for HLA-H that contains a portion of intron 3 attached to a normally spliced sequence comprising exons 4 through 8. These results suggest that low levels of translatable mRNA for the truncated class I heavy chain encoded by HLA-H are produced under physiologic circumstances and that sequences 3' of intron 3 decrease the levels of stable transcripts. Images PMID:2294403

  13. Lineage-specific late pleistocene expansion of an endemic subtropical gossamer-wing damselfly, Euphaea formosa, in Taiwan

    PubMed Central

    2011-01-01

    subtropical damselfly's historical demography is lineage-specific, depending predominantly on its colonization history and geography. It is proposed that the Riss and Würm glaciations in the late Pleistocene period had a greater impact on the evolutionary diversification of subtropical insular species than the last glacial maximum (LGM). PMID:21486452

  14. P01.08LINEAGE-SPECIFIC SPLICING OF AN ALTERNATIVE EXON OF ANXA7 PROMOTES EGFR SIGNALING ACTIVATION AND TUMOR PROGRESSION IN GLIOBLASTOMA

    PubMed Central

    Ferrarese, R.; Bug, E.; Maticzka, D.; Reichardt, W.; Masilamani, A.P.; Dai, F.; Weyerbrock, A.; Prinz, M.; Bredel, M.; Carro, M.S.

    2014-01-01

    Tissue-specific alternative splicing is critical to the emergence of tissue identity during development, yet its role in malignant transformation is undefined. Tissue-specific splicing involves evolutionarily-conserved, alternative exons, which represent only a minority of total alternative exons. Many, however, have functional features that influence signaling pathways to profound biological effect. In the brain, Annexin A7 isoform 1 (ANXA7-I1) is exclusively expressed in mature neurons, while isoform 2 (ANXA7-I2) in which exon 6 is skipped, is expressed in glial and progenitor cells. We show that lineage-specific splicing of the cassette exon 6 in the membrane-binding tumor suppressor ANXA7diminishes endosomal targeting and consequent signal termination of the EGFR oncoprotein during brain tumor progression. Splicing of this exon is mediated by Polypyrimidine Tract-Binding Protein 1 (PTBP1), a ribonucleoprotein normally repressed during neuronal development but which we found to be highly expressed also in glioblastomas through loss of a brain-enriched microRNA, miR-124, and gene amplification. Here, we show that the PTBP1-ANXA7 splicing-EGFR signal activation axis promotes in vitro cell migration and invasion, and tumor angiogenesis in vivo. In glioblastoma, ANXA7 splicing is likely inherited from a potential tumor-initiating ancestor but this trait is further exploited through accumulation of mutations that enhance EGFR signaling. Our data illustrate how lineage-specific splicing of a tissue-regulated alternative exon in a constituent of an oncogenic pathway eliminates its tumor suppressor function and promotes glioblastoma progression. This paradigm may offer a general model as to how tissue-specific regulatory mechanisms can contribute to reprogramming normal development to oncogenesis.

  15. Alcohol Consumption Modulates Host Defense in Rhesus Macaques by Altering Gene Expression in Circulating Leukocytes.

    PubMed

    Barr, Tasha; Girke, Thomas; Sureshchandra, Suhas; Nguyen, Christina; Grant, Kathleen; Messaoudi, Ilhem

    2016-01-01

    Several lines of evidence indicate that chronic alcohol use disorder leads to increased susceptibility to several viral and bacterial infections, whereas moderate alcohol consumption decreases the incidence of colds and improves immune responses to some pathogens. In line with these observations, we recently showed that heavy ethanol intake (average blood ethanol concentrations > 80 mg/dl) suppressed, whereas moderate alcohol consumption (blood ethanol concentrations < 50 mg/dl) enhanced, T and B cell responses to modified vaccinia Ankara vaccination in a nonhuman primate model of voluntary ethanol consumption. To uncover the molecular basis for impaired immunity with heavy alcohol consumption and enhanced immune response with moderate alcohol consumption, we performed a transcriptome analysis using PBMCs isolated on day 7 post-modified vaccinia Ankara vaccination, the earliest time point at which we detected differences in T cell and Ab responses. Overall, chronic heavy alcohol consumption reduced the expression of immune genes involved in response to infection and wound healing and increased the expression of genes associated with the development of lung inflammatory disease and cancer. In contrast, chronic moderate alcohol consumption upregulated the expression of genes involved in immune response and reduced the expression of genes involved in cancer. To uncover mechanisms underlying the alterations in PBMC transcriptomes, we profiled the expression of microRNAs within the same samples. Chronic heavy ethanol consumption altered the levels of several microRNAs involved in cancer and immunity and known to regulate the expression of mRNAs differentially expressed in our data set. PMID:26621857

  16. Risk assessment of relapse by lineage-specific monitoring of chimerism in children undergoing allogeneic stem cell transplantation for acute lymphoblastic leukemia

    PubMed Central

    Preuner, Sandra; Peters, Christina; Pötschger, Ulrike; Daxberger, Helga; Fritsch, Gerhard; Geyeregger, Rene; Schrauder, André; von Stackelberg, Arend; Schrappe, Martin; Bader, Peter; Ebell, Wolfram; Eckert, Cornelia; Lang, Peter; Sykora, Karl-Walter; Schrum, Johanna; Kremens, Bernhard; Ehlert, Karoline; Albert, Michael H.; Meisel, Roland; Lawitschka, Anita; Mann, Georg; Panzer-Grümayer, Renate; Güngör, Tayfun; Holter, Wolfgang; Strahm, Brigitte; Gruhn, Bernd; Schulz, Ansgar; Woessmann, Wilhelm; Lion, Thomas

    2016-01-01

    Allogeneic hematopoietic stem cell transplantation is required as rescue therapy in about 20% of pediatric patients with acute lymphoblastic leukemia. However, the relapse rates are considerable, and relapse confers a poor outcome. Early assessment of the risk of relapse is therefore of paramount importance for the development of appropriate measures. We used the EuroChimerism approach to investigate the potential impact of lineage-specific chimerism testing for relapse-risk analysis in 162 pediatric patients with acute lymphoblastic leukemia after allogeneic stem cell transplantation in a multicenter study based on standardized transplantation protocols. Within a median observation time of 4.5 years, relapses have occurred in 41/162 patients at a median of 0.6 years after transplantation (range, 0.13–5.7 years). Prospective screening at defined consecutive time points revealed that reappearance of recipient-derived cells within the CD34+ and CD8+ cell subsets display the most significant association with the occurrence of relapses with hazard ratios of 5.2 (P=0.003) and 2.8 (P=0.008), respectively. The appearance of recipient cells after a period of pure donor chimerism in the CD34+ and CD8+ leukocyte subsets revealed dynamics indicative of a significantly elevated risk of relapse or imminent disease recurrence. Assessment of chimerism within these lineages can therefore provide complementary information for further diagnostic and, potentially, therapeutic purposes aiming at the prevention of overt relapse. This study was registered at clinical.trials.gov with the number NC01423747. PMID:26869631

  17. H3K4/H3K9me3 Bivalent Chromatin Domains Targeted by Lineage-Specific DNA Methylation Pauses Adipocyte Differentiation.

    PubMed

    Matsumura, Yoshihiro; Nakaki, Ryo; Inagaki, Takeshi; Yoshida, Ayano; Kano, Yuka; Kimura, Hiroshi; Tanaka, Toshiya; Tsutsumi, Shuichi; Nakao, Mitsuyoshi; Doi, Takefumi; Fukami, Kiyoko; Osborne, Timothy F; Kodama, Tatsuhiko; Aburatani, Hiroyuki; Sakai, Juro

    2015-11-19

    Bivalent H3K4me3 and H3K27me3 chromatin domains in embryonic stem cells keep active developmental regulatory genes expressed at very low levels and poised for activation. Here, we show an alternative and previously unknown bivalent modified histone signature in lineage-committed mesenchymal stem cells and preadipocytes that pairs H3K4me3 with H3K9me3 to maintain adipogenic master regulatory genes (Cebpa and Pparg) expressed at low levels yet poised for activation when differentiation is required. We show lineage-specific gene-body DNA methylation recruits H3K9 methyltransferase SETDB1, which methylates H3K9 immediately downstream of transcription start sites marked with H3K4me3 to establish the bivalent domain. At the Cebpa locus, this prevents transcription factor C/EBPβ binding, histone acetylation, and further H3K4me3 deposition and is associated with pausing of RNA polymerase II, which limits Cebpa gene expression and adipogenesis. PMID:26590716

  18. Whole genome sequencing of a banana wild relative Musa itinerans provides insights into lineage-specific diversification of the Musa genus

    PubMed Central

    Wu, Wei; Yang, Yu-Lan; He, Wei-Ming; Rouard, Mathieu; Li, Wei-Ming; Xu, Meng; Roux, Nicolas; Ge, Xue-Jun

    2016-01-01

    Crop wild relatives are valuable resources for future genetic improvement. Here, we report the de novo genome assembly of Musa itinerans, a disease-resistant wild banana relative in subtropical China. The assembled genome size was 462.1 Mb, covering 75.2% of the genome (615.2Mb) and containing 32, 456 predicted protein-coding genes. Since the approximate divergence around 5.8 million years ago, the genomes of Musa itinerans and Musa acuminata have shown conserved collinearity. Gene family expansions and contractions enrichment analysis revealed that some pathways were associated with phenotypic or physiological innovations. These include a transition from wood to herbaceous in the ancestral Musaceae, intensification of cold and drought tolerances, and reduced diseases resistance genes for subtropical marginally distributed Musa species. Prevalent purifying selection and transposed duplications were found to facilitate the diversification of NBS-encoding gene families for two Musa species. The population genome history analysis of M. itinerans revealed that the fluctuated population sizes were caused by the Pleistocene climate oscillations, and that the formation of Qiongzhou Strait might facilitate the population downsizing on the isolated Hainan Island about 10.3 Kya. The qualified assembly of the M. itinerans genome provides deep insights into the lineage-specific diversification and also valuable resources for future banana breeding. PMID:27531320

  19. Whole genome sequencing of a banana wild relative Musa itinerans provides insights into lineage-specific diversification of the Musa genus.

    PubMed

    Wu, Wei; Yang, Yu-Lan; He, Wei-Ming; Rouard, Mathieu; Li, Wei-Ming; Xu, Meng; Roux, Nicolas; Ge, Xue-Jun

    2016-01-01

    Crop wild relatives are valuable resources for future genetic improvement. Here, we report the de novo genome assembly of Musa itinerans, a disease-resistant wild banana relative in subtropical China. The assembled genome size was 462.1 Mb, covering 75.2% of the genome (615.2Mb) and containing 32, 456 predicted protein-coding genes. Since the approximate divergence around 5.8 million years ago, the genomes of Musa itinerans and Musa acuminata have shown conserved collinearity. Gene family expansions and contractions enrichment analysis revealed that some pathways were associated with phenotypic or physiological innovations. These include a transition from wood to herbaceous in the ancestral Musaceae, intensification of cold and drought tolerances, and reduced diseases resistance genes for subtropical marginally distributed Musa species. Prevalent purifying selection and transposed duplications were found to facilitate the diversification of NBS-encoding gene families for two Musa species. The population genome history analysis of M. itinerans revealed that the fluctuated population sizes were caused by the Pleistocene climate oscillations, and that the formation of Qiongzhou Strait might facilitate the population downsizing on the isolated Hainan Island about 10.3 Kya. The qualified assembly of the M. itinerans genome provides deep insights into the lineage-specific diversification and also valuable resources for future banana breeding. PMID:27531320

  20. Copy number variation of E3 ubiquitin ligase genes in peripheral blood leukocyte and colorectal cancer.

    PubMed

    Bi, Haoran; Tian, Tian; Zhu, Lin; Zhou, Haibo; Hu, Hanqing; Liu, Yanhong; Li, Xia; Hu, Fulan; Zhao, Yashuang; Wang, Guiyu

    2016-01-01

    Given that E3 ubiquitin ligases (E3) regulate specific protein degradation in many cancer-related biological processes. E3 copy number variation (CNV) may affect the development and prognosis of colorectal cancer (CRC). Therefore, we detected CNVs of five E3 genes in 518 CRC patients and 518 age, gender and residence matched controls in China, and estimated the association between E3 gene CNVs and CRC risk and prognosis. We also estimated their interactions with environmental factors and CRC risk. We find a significant association between the CNVs of MDM2 and CRC risk (amp v.s. wt: odds ratio = 14.37, 95% confidence interval: 1.27, 163.74, P = 0.032), while SKP2 CNVs may significantly decrease CRC risk (del v.s. wt: odds ratio = 0.32, 95% confidence interval: 0.10, 1.00, P = 0.050). However, we find no significant association between the CNVs of other genes and CRC risk. The only significant gene-environment interaction effects are between SKP2 CNVs and consumption of fish and/or fruit (P = 0.014 and P = 0.035) and between FBXW7 CNVs and pork intake (P = 0.040). Finally, we find marginally significant association between β-TRCP CNVs and CRC prognosis (amp v.s. wt, hazard ratio = 0.42, 95% confidence interval: 0.19, 0.97, P = 0.050). PMID:27417709

  1. Copy number variation of E3 ubiquitin ligase genes in peripheral blood leukocyte and colorectal cancer

    PubMed Central

    Bi, Haoran; Tian, Tian; Zhu, Lin; Zhou, Haibo; Hu, Hanqing; Liu, Yanhong; Li, Xia; Hu, Fulan; Zhao, Yashuang; Wang, Guiyu

    2016-01-01

    Given that E3 ubiquitin ligases (E3) regulate specific protein degradation in many cancer-related biological processes. E3 copy number variation (CNV) may affect the development and prognosis of colorectal cancer (CRC). Therefore, we detected CNVs of five E3 genes in 518 CRC patients and 518 age, gender and residence matched controls in China, and estimated the association between E3 gene CNVs and CRC risk and prognosis. We also estimated their interactions with environmental factors and CRC risk. We find a significant association between the CNVs of MDM2 and CRC risk (amp v.s. wt: odds ratio = 14.37, 95% confidence interval: 1.27, 163.74, P = 0.032), while SKP2 CNVs may significantly decrease CRC risk (del v.s. wt: odds ratio = 0.32, 95% confidence interval: 0.10, 1.00, P = 0.050). However, we find no significant association between the CNVs of other genes and CRC risk. The only significant gene-environment interaction effects are between SKP2 CNVs and consumption of fish and/or fruit (P = 0.014 and P = 0.035) and between FBXW7 CNVs and pork intake (P = 0.040). Finally, we find marginally significant association between β-TRCP CNVs and CRC prognosis (amp v.s. wt, hazard ratio = 0.42, 95% confidence interval: 0.19, 0.97, P = 0.050). PMID:27417709

  2. Integrative View of α2,3-Sialyltransferases (ST3Gal) Molecular and Functional Evolution in Deuterostomes: Significance of Lineage-Specific Losses

    PubMed Central

    Petit, Daniel; Teppa, Elin; Mir, Anne-Marie; Vicogne, Dorothée; Thisse, Christine; Thisse, Bernard; Filloux, Cyril; Harduin-Lepers, Anne

    2015-01-01

    Sialyltransferases are responsible for the synthesis of a diverse range of sialoglycoconjugates predicted to be pivotal to deuterostomes’ evolution. In this work, we reconstructed the evolutionary history of the metazoan α2,3-sialyltransferases family (ST3Gal), a subset of sialyltransferases encompassing six subfamilies (ST3Gal I–ST3Gal VI) functionally characterized in mammals. Exploration of genomic and expressed sequence tag databases and search of conserved sialylmotifs led to the identification of a large data set of st3gal-related gene sequences. Molecular phylogeny and large scale sequence similarity network analysis identified four new vertebrate subfamilies called ST3Gal III-r, ST3Gal VII, ST3Gal VIII, and ST3Gal IX. To address the issue of the origin and evolutionary relationships of the st3gal-related genes, we performed comparative syntenic mapping of st3gal gene loci combined to ancestral genome reconstruction. The ten vertebrate ST3Gal subfamilies originated from genome duplication events at the base of vertebrates and are organized in three distinct and ancient groups of genes predating the early deuterostomes. Inferring st3gal gene family history identified also several lineage-specific gene losses, the significance of which was explored in a functional context. Toward this aim, spatiotemporal distribution of st3gal genes was analyzed in zebrafish and bovine tissues. In addition, molecular evolutionary analyses using specificity determining position and coevolved amino acid predictions led to the identification of amino acid residues with potential implication in functional divergence of vertebrate ST3Gal. We propose a detailed scenario of the evolutionary relationships of st3gal genes coupled to a conceptual framework of the evolution of ST3Gal functions. PMID:25534026

  3. Directing lineage specification of human mesenchymal stem cells by decoupling electrical stimulation and physical patterning on unmodified graphene

    NASA Astrophysics Data System (ADS)

    Balikov, Daniel A.; Fang, Brian; Chun, Young Wook; Crowder, Spencer W.; Prasai, Dhiraj; Lee, Jung Bok; Bolotin, Kiril I.; Sung, Hak-Joon

    2016-07-01

    The organization and composition of the extracellular matrix (ECM) have been shown to impact the propagation of electrical signals in multiple tissue types. To date, many studies with electroactive biomaterial substrates have relied upon passive electrical stimulation of the ionic media to affect cell behavior. However, development of cell culture systems in which stimulation can be directly applied to the material - thereby isolating the signal to the cell-material interface and cell-cell contracts - would provide a more physiologically-relevant paradigm for investigating how electrical cues modulate lineage-specific stem cell differentiation. In the present study, we have employed unmodified, directly-stimulated, (un)patterned graphene as a cell culture substrate to investigate how extrinsic electrical cycling influences the differentiation of naïve human mesenchymal stem cells (hMSCs) without the bias of exogenous biochemicals. We first demonstrated that cyclic stimulation does not deteriorate the cell culture media or result in cytotoxic pH, which are critical experiments for correct interpretation of changes in cell behavior. We then measured how the expression of osteogenic and neurogenic lineage-specific markers were altered simply by exposure to electrical stimulation and/or physical patterns. Expression of the early osteogenic transcription factor RUNX2 was increased by electrical stimulation on all graphene substrates, but the mature marker osteopontin was only modulated when stimulation was combined with physical patterns. In contrast, the expression of the neurogenic markers MAP2 and β3-tubulin were enhanced in all electrical stimulation conditions, and were less responsive to the presence of patterns. These data indicate that specific combinations of non-biological inputs - material type, electrical stimulation, physical patterns - can regulate hMSC lineage specification. This study represents a substantial step in understanding how the interplay of

  4. Directing lineage specification of human mesenchymal stem cells by decoupling electrical stimulation and physical patterning on unmodified graphene.

    PubMed

    Balikov, Daniel A; Fang, Brian; Chun, Young Wook; Crowder, Spencer W; Prasai, Dhiraj; Lee, Jung Bok; Bolotin, Kiril I; Sung, Hak-Joon

    2016-07-14

    The organization and composition of the extracellular matrix (ECM) have been shown to impact the propagation of electrical signals in multiple tissue types. To date, many studies with electroactive biomaterial substrates have relied upon passive electrical stimulation of the ionic media to affect cell behavior. However, development of cell culture systems in which stimulation can be directly applied to the material - thereby isolating the signal to the cell-material interface and cell-cell contracts - would provide a more physiologically-relevant paradigm for investigating how electrical cues modulate lineage-specific stem cell differentiation. In the present study, we have employed unmodified, directly-stimulated, (un)patterned graphene as a cell culture substrate to investigate how extrinsic electrical cycling influences the differentiation of naïve human mesenchymal stem cells (hMSCs) without the bias of exogenous biochemicals. We first demonstrated that cyclic stimulation does not deteriorate the cell culture media or result in cytotoxic pH, which are critical experiments for correct interpretation of changes in cell behavior. We then measured how the expression of osteogenic and neurogenic lineage-specific markers were altered simply by exposure to electrical stimulation and/or physical patterns. Expression of the early osteogenic transcription factor RUNX2 was increased by electrical stimulation on all graphene substrates, but the mature marker osteopontin was only modulated when stimulation was combined with physical patterns. In contrast, the expression of the neurogenic markers MAP2 and β3-tubulin were enhanced in all electrical stimulation conditions, and were less responsive to the presence of patterns. These data indicate that specific combinations of non-biological inputs - material type, electrical stimulation, physical patterns - can regulate hMSC lineage specification. This study represents a substantial step in understanding how the interplay of

  5. Understanding stress-induced immunosuppression: exploration of cytokine and chemokine gene profiles in chicken peripheral leukocytes.

    PubMed

    Shini, S; Huff, G R; Shini, A; Kaiser, P

    2010-04-01

    At present, the poultry meat and egg industry has gained a lot of ground, being viewed as a provider of a healthy alternative to red meat and other protein sources. If this trend is to be maintained, solutions must be found to improve resistance of chickens to disease, which often is weakened by stressful conditions. In poultry, stress-induced immunosuppression is manifested by failures in vaccination and increased morbidity and mortality of flocks. Currently, several modern cellular and molecular approaches are being used to explore the status of the immune system during stress and disease. It is likely that these new techniques will lead to the development of new strategies for preventing and controlling immunosuppression in poultry. Using quantitative reverse transcription-PCR assays, a broad spectrum of cytokine, chemokine, and their receptor genes can be quantified in birds and then be used as markers to assess the effects of stress on the immune system. Currently, we are investigating immune and endocrine interactions in the chicken, in particular the cells and molecules that are known to be involved in such interactions in mammals. We have evaluated the effects of corticosterone administration in drinking water on peripheral lymphocyte and heterophil cytokine and chemokine gene profiles. In particular, there seems to be effects on cytokine and chemokine mRNA expression levels in both lymphocytes and heterophils, especially expression of the proinflammatory cytokines interleukin (IL)-1beta, IL-6, and IL-18 and chemokines C-C motif, ligand 1 inflammatory (CCLi1); C-C motif, ligand 2 inflammatory (CCLi2); C-C motif, ligand 5 (CCL5); C-C motif, ligand 16 (CCL16); C-X-C motif ligand 1 inflammatory (CXCLi1); and C-X-C motif ligand 2 inflammatory (CXCLi2), which are initially upregulated and are potentially involved in modulating the adaptive immune response. A chronic treatment with corticosterone downregulates proinflammatory cytokines and chemokines, suggesting

  6. Functional microRNAs and target sites are created by lineage-specific transposition

    PubMed Central

    Spengler, Ryan M.; Oakley, Clayton K.; Davidson, Beverly L.

    2014-01-01

    Transposable elements (TEs) account for nearly one-half of the sequence content in the human genome, and de novo germline transposition into regulatory or coding sequences of protein-coding genes can cause heritable disorders. TEs are prevalent in and around protein-coding genes, providing an opportunity to impart regulation. Computational studies reveal that microRNA (miRNA) genes and miRNA target sites reside within TE sequences, but there is little experimental evidence supporting a role for TEs in the birth of miRNAs, or as platform for gene regulation by miRNAs. In this work, we validate miRNAs and target sites derived from TE families prevalent in the human genome, including the ancient long interspersed nuclear element 2 (LINE2/L2), mammalian-wide interspersed repeat (MIR) retrotransposons and the primate-specific Alu family. We show that genes with 3′ untranslated region (3′ UTR) MIR elements are enriched for let-7 targets and that these sites are conserved and responsive to let-7 expression. We also demonstrate that 3′ UTR-embedded Alus are a source of miR-24 and miR-122 target sites and that a subset of active genomic Alus provide for de novo target site creation. Finally, we report that although the creation of miRNA genes by Alu elements is relatively uncommon relative to their overall genomic abundance, Alu-derived miR-1285-1 is efficiently processed from its genomic locus and regulates genes with target sites contained within homologous elements. Taken together, our data provide additional evidence for TEs as a source for miRNAs and miRNA target sites, with instances of conservation through the course of mammalian evolution. PMID:24234653

  7. Secretory leukocyte protease inhibitor gene deletion alters bleomycin-induced lung injury, but not development of pulmonary fibrosis.

    PubMed

    Habgood, Anthony N; Tatler, Amanda L; Porte, Joanne; Wahl, Sharon M; Laurent, Geoffrey J; John, Alison E; Johnson, Simon R; Jenkins, Gisli

    2016-06-01

    Idiopathic pulmonary fibrosis is a progressive, fatal disease with limited treatment options. Protease-mediated transforming growth factor-β (TGF-β) activation has been proposed as a pathogenic mechanism of lung fibrosis. Protease activity in the lung is tightly regulated by protease inhibitors, particularly secretory leukocyte protease inhibitor (SLPI). The bleomycin model of lung fibrosis was used to determine the effect of increased protease activity in the lungs of Slpi(-/-) mice following injury. Slpi(-/-), and wild-type, mice received oropharyngeal administration of bleomycin (30 IU) and the development of pulmonary fibrosis was assessed. Pro and active forms of matrix metalloproteinase (MMP)-2 and MMP-9 were measured. Lung fibrosis was determined by collagen subtype-specific gene expression, hydroxyproline concentration, and histological assessment. Alveolar TGF-β activation was measured using bronchoalveolar lavage cell pSmad2 levels and global TGF-β activity was assessed by pSmad2 immunohistochemistry. The active-MMP-9 to pro-MMP-9 ratio was significantly increased in Slpi(-/-) animals compared with wild-type animals, demonstrating enhanced metalloproteinase activity. Wild-type animals showed an increase in TGF-β activation following bleomycin, with a progressive and sustained increase in collagen type I, alpha 1 (Col1α1), III, alpha 1(Col3α1), IV, alpha 1(Col4α1) mRNA expression, and a significant increase in total lung collagen 28 days post bleomycin. In contrast Slpi(-/-) mice showed no significant increase of alveolar TGF-β activity following bleomycin, above their already elevated levels, although global TGF-β activity did increase. Slpi(-/-) mice had impaired collagen gene expression but animals demonstrated minimal reduction in lung fibrosis compared with wild-type animals. These data suggest that enhanced proteolysis does not further enhance TGF-β activation, and inhibits sustained Col1α1, Col3α1, and Col4α1 gene expression

  8. Identification of novel allelic variants of integrin beta 2 (ITGB2) gene and screening for Bubaline leukocyte adhesion deficiency syndrome in Indian water buffaloes (Bubalus bubalis).

    PubMed

    Sharma, Deepak; Kumar, Subodh; Deb, Sitangsu M; Mitra, Abhijit; Niranjan, Saket K; Naskar, Soumen; Sharma, Arjava

    2009-01-01

    A fragment of 570 bp corresponding to exon 5 and 6 of integrin beta 2 (ITGB2) gene was amplified for screening D128G mutation in one hundred and fifty two buffaloes (Bubalus bubalis) which causes bovine leukocyte adhesion deficiency syndrome (BLAD) in cattle, as well as to ascertain polymorphism. TaqI PCR-RFLP revealed no such mutation thus indicating the absence of bubaline leukocyte adhesion deficiency (BuLAD) allele in animals under study. However, the polymorphism studies using MspI restriction enzyme revealed two genotypic patterns viz. AA pattern (bands of 293, 141, 105, and 31 bp) and BB pattern (bands of 293, 105, 77, 64, and 31 bp). The sequences of A and B alleles were submitted to the GenBank (EU853307 and AY821799). PMID:19544212

  9. Lineage-specific interface proteins match up the cell cycle and differentiation in embryo stem cells.

    PubMed

    Re, Angela; Workman, Christopher T; Waldron, Levi; Quattrone, Alessandro; Brunak, Søren

    2014-09-01

    The shortage of molecular information on cell cycle changes along embryonic stem cell (ESC) differentiation prompts an in silico approach, which may provide a novel way to identify candidate genes or mechanisms acting in coordinating the two programs. We analyzed germ layer specific gene expression changes during the cell cycle and ESC differentiation by combining four human cell cycle transcriptome profiles with thirteen in vitro human ESC differentiation studies. To detect cross-talk mechanisms we then integrated the transcriptome data that displayed differential regulation with protein interaction data. A new class of non-transcriptionally regulated genes was identified, encoding proteins which interact systematically with proteins corresponding to genes regulated during the cell cycle or cell differentiation, and which therefore can be seen as interface proteins coordinating the two programs. Functional analysis gathered insights in fate-specific candidates of interface functionalities. The non-transcriptionally regulated interface proteins were found to be highly regulated by post-translational ubiquitylation modification, which may synchronize the transition between cell proliferation and differentiation in ESCs. PMID:25173649

  10. Impact of Tissue-Specific Stem Cells on Lineage-Specific Differentiation: A Focus on the Musculoskeletal System

    PubMed Central

    Pizzute, Tyler; Lynch, Kevin; Pei, Ming

    2014-01-01

    Tissue-specific stem cells are found throughout the body and, with proper intervention and environmental cues, these stem cells exercise their capabilities for differentiation into several lineages to form cartilage, bone, muscle, and adipose tissue in vitro and in vivo. Interestingly, it has been widely demonstrated that they do not differentiate with the same efficacy during lineage-specific differentiation studies, as the tissue-specific stem cells are generally more effective when differentiating toward the tissues from which they were derived. This review focuses on four mesodermal lineages for tissue-specific stem cell differentiation: adipogenesis, chondrogenesis, myogenesis, and osteogenesis. It is intended to give insight into current multilineage differentiation and comparative research, highlight and contrast known trends regarding differentiation, and introduce supporting evidence which demonstrates particular tissue-specific stem cells’ superiority in lineage-specific differentiation, along with their resident tissue origins and natural roles. In addition, some epigenetic and transcriptomic differences between stem cells which may explain the observed trends are discussed. PMID:25113801

  11. Functional Genetic Diversity among Mycobacterium tuberculosis Complex Clinical Isolates: Delineation of Conserved Core and Lineage-Specific Transcriptomes during Intracellular Survival

    PubMed Central

    Homolka, Susanne; Niemann, Stefan; Russell, David G.; Rohde, Kyle H.

    2010-01-01

    Tuberculosis exerts a tremendous burden on global health, with ∼9 million new infections and ∼2 million deaths annually. The Mycobacterium tuberculosis complex (MTC) was initially regarded as a highly homogeneous population; however, recent data suggest the causative agents of tuberculosis are more genetically and functionally diverse than appreciated previously. The impact of this natural variation on the virulence and clinical manifestations of the pathogen remains largely unknown. This report examines the effect of genetic diversity among MTC clinical isolates on global gene expression and survival within macrophages. We discovered lineage-specific transcription patterns in vitro and distinct intracellular growth profiles associated with specific responses to host-derived environmental cues. Strain comparisons also facilitated delineation of a core intracellular transcriptome, including genes with highly conserved regulation across the global panel of clinical isolates. This study affords new insights into the genetic information that M. tuberculosis has conserved under selective pressure during its long-term interactions with its human host. PMID:20628579

  12. Human Lineage-Specific Transcriptional Regulation through GA-Binding Protein Transcription Factor Alpha (GABPa).

    PubMed

    Perdomo-Sabogal, Alvaro; Nowick, Katja; Piccini, Ilaria; Sudbrak, Ralf; Lehrach, Hans; Yaspo, Marie-Laure; Warnatz, Hans-Jörg; Querfurth, Robert

    2016-05-01

    A substantial fraction of phenotypic differences between closely related species are likely caused by differences in gene regulation. While this has already been postulated over 30 years ago, only few examples of evolutionary changes in gene regulation have been verified. Here, we identified and investigated binding sites of the transcription factor GA-binding protein alpha (GABPa) aiming to discover cis-regulatory adaptations on the human lineage. By performing chromatin immunoprecipitation-sequencing experiments in a human cell line, we found 11,619 putative GABPa binding sites. Through sequence comparisons of the human GABPa binding regions with orthologous sequences from 34 mammals, we identified substitutions that have resulted in 224 putative human-specific GABPa binding sites. To experimentally assess the transcriptional impact of those substitutions, we selected four promoters for promoter-reporter gene assays using human and African green monkey cells. We compared the activities of wild-type promoters to mutated forms, where we have introduced one or more substitutions to mimic the ancestral state devoid of the GABPa consensus binding sequence. Similarly, we introduced the human-specific substitutions into chimpanzee and macaque promoter backgrounds. Our results demonstrate that the identified substitutions are functional, both in human and nonhuman promoters. In addition, we performed GABPa knock-down experiments and found 1,215 genes as strong candidates for primary targets. Further analyses of our data sets link GABPa to cognitive disorders, diabetes, KRAB zinc finger (KRAB-ZNF), and human-specific genes. Thus, we propose that differences in GABPa binding sites played important roles in the evolution of human-specific phenotypes. PMID:26814189

  13. Human Lineage-Specific Transcriptional Regulation through GA-Binding Protein Transcription Factor Alpha (GABPa)

    PubMed Central

    Perdomo-Sabogal, Alvaro; Nowick, Katja; Piccini, Ilaria; Sudbrak, Ralf; Lehrach, Hans; Yaspo, Marie-Laure; Warnatz, Hans-Jörg; Querfurth, Robert

    2016-01-01

    A substantial fraction of phenotypic differences between closely related species are likely caused by differences in gene regulation. While this has already been postulated over 30 years ago, only few examples of evolutionary changes in gene regulation have been verified. Here, we identified and investigated binding sites of the transcription factor GA-binding protein alpha (GABPa) aiming to discover cis-regulatory adaptations on the human lineage. By performing chromatin immunoprecipitation-sequencing experiments in a human cell line, we found 11,619 putative GABPa binding sites. Through sequence comparisons of the human GABPa binding regions with orthologous sequences from 34 mammals, we identified substitutions that have resulted in 224 putative human-specific GABPa binding sites. To experimentally assess the transcriptional impact of those substitutions, we selected four promoters for promoter-reporter gene assays using human and African green monkey cells. We compared the activities of wild-type promoters to mutated forms, where we have introduced one or more substitutions to mimic the ancestral state devoid of the GABPa consensus binding sequence. Similarly, we introduced the human-specific substitutions into chimpanzee and macaque promoter backgrounds. Our results demonstrate that the identified substitutions are functional, both in human and nonhuman promoters. In addition, we performed GABPa knock-down experiments and found 1,215 genes as strong candidates for primary targets. Further analyses of our data sets link GABPa to cognitive disorders, diabetes, KRAB zinc finger (KRAB-ZNF), and human-specific genes. Thus, we propose that differences in GABPa binding sites played important roles in the evolution of human-specific phenotypes. PMID:26814189

  14. Identification of focally amplified lineage-specific super-enhancers in human epithelial cancers.

    PubMed

    Zhang, Xiaoyang; Choi, Peter S; Francis, Joshua M; Imielinski, Marcin; Watanabe, Hideo; Cherniack, Andrew D; Meyerson, Matthew

    2016-02-01

    Whole-genome analysis approaches are identifying recurrent cancer-associated somatic alterations in noncoding DNA regions. We combined somatic copy number analysis of 12 tumor types with tissue-specific epigenetic profiling to identify significant regions of focal amplification harboring super-enhancers. Copy number gains of noncoding regions harboring super-enhancers near KLF5, USP12, PARD6B and MYC are associated with overexpression of these cancer-related genes. We show that two distinct focal amplifications of super-enhancers 3' to MYC in lung adenocarcinoma (MYC-LASE) and endometrial carcinoma (MYC-ECSE) are physically associated with the MYC promoter and correlate with MYC overexpression. CRISPR/Cas9-mediated repression or deletion of a constituent enhancer within the MYC-LASE region led to significant reductions in the expression of MYC and its target genes and to the impairment of anchorage-independent and clonogenic growth, consistent with an oncogenic function. Our results suggest that genomic amplification of super-enhancers represents a common mechanism to activate cancer driver genes in multiple cancer types. PMID:26656844

  15. 5-Hydroxymethylcytosine Remodeling Precedes Lineage Specification during Differentiation of Human CD4(+) T Cells.

    PubMed

    Nestor, Colm E; Lentini, Antonio; Hägg Nilsson, Cathrine; Gawel, Danuta R; Gustafsson, Mika; Mattson, Lina; Wang, Hui; Rundquist, Olof; Meehan, Richard R; Klocke, Bernward; Seifert, Martin; Hauck, Stefanie M; Laumen, Helmut; Zhang, Huan; Benson, Mikael

    2016-07-12

    5-methylcytosine (5mC) is converted to 5-hydroxymethylcytosine (5hmC) by the TET family of enzymes as part of a recently discovered active DNA de-methylation pathway. 5hmC plays important roles in regulation of gene expression and differentiation and has been implicated in T cell malignancies and autoimmunity. Here, we report early and widespread 5mC/5hmC remodeling during human CD4(+) T cell differentiation ex vivo at genes and cell-specific enhancers with known T cell function. We observe similar DNA de-methylation in CD4(+) memory T cells in vivo, indicating that early remodeling events persist long term in differentiated cells. Underscoring their important function, 5hmC loci were highly enriched for genetic variants associated with T cell diseases and T-cell-specific chromosomal interactions. Extensive functional validation of 22 risk variants revealed potentially pathogenic mechanisms in diabetes and multiple sclerosis. Our results support 5hmC-mediated DNA de-methylation as a key component of CD4(+) T cell biology in humans, with important implications for gene regulation and lineage commitment. PMID:27346350

  16. Association of human leukocyte antigen DP/DQ gene polymorphisms with chronic hepatitis B in Chinese Han and Uygur populations.

    PubMed

    Xiang, Xin; Guo, Yuxuan; Yang, Li; Ge, Qinghui; Mijit, Sadatgul; Xu, Feili

    2016-09-01

    Several genome-wide association studies (GWAS) have shown that human leukocyte antigen (HLA) DP/DQ gene polymorphisms are associated with susceptibility to chronic hepatitis B virus (HBV) infection. We clarified the roles of the HLA-DP/DQ gene in HBV infection in different nationalities. Three single nucleotide polymorphisms (SNPs) in HLA-DP (rs9277471, rs9277535 and rs9277542) and the SNP rs9272346 in HLA-DQ were studied. In total, 779 patients were recruited to this study, including 400 Chinese Han and 399 Uygurs. The rs9277535 variant genotypes were directly associated with HBV persistence compared to healthy controls in an additive model of the Chinese Han population (odds ratio [OR]=1.88, 95% confidence interval [CI]=1.03-3.41, P=0.040), and in a recessive model of the Chinese female population (OR=2.02, 95% CI=1.26-3.24, P=0.003). In addition, rs9277471 and rs9277542 variant genotypes significantly decreased the risk of HBV infection compared to healthy controls in an additive model of the Chinese Han population (OR=0.53, 95% CI=0.29-0.98, P=0.042; OR=0.53, 95% CI=0.29-0.97, P=0.039) and in a dominant model of the Chinese female population (OR=0.50, 95% CI=0.31-0.80, P=0.004; OR=0.49, 95% CI=0.31-0.79, P=0.003). The GG genotype of rs9277346 was associated with HBV infection in the Chinese Han population (additive model: OR=0.38, 95%CI=017-0.82, P=0.014; recessive model: OR=0.41, 95% CI=0.19-0.86, P=0.019) and in males (additive model: OR=0.31, 95% CI=0.14-0.65, P=0.002; dominant model: OR=0.65, 95% CI=0.43-0.97, P=0.034; recessive model: OR=0.36, 95% CI=0.18-0.73, P=0.005). In addition, allele G of rs9277346 was marginally related to a reduction in risk for HBV infection in the Uygur population. Our study suggests that HLA-DP/DQ polymorphisms can affect susceptibility and resistance to HBV infection in Chinese populations, and are possibly linked to race and sex. PMID:27291710

  17. ZINC-INDUCED FACILITATOR-LIKE family in plants: lineage-specific expansion in monocotyledons and conserved genomic and expression features among rice (Oryza sativa) paralogs

    PubMed Central

    2011-01-01

    Background Duplications are very common in the evolution of plant genomes, explaining the high number of members in plant gene families. New genes born after duplication can undergo pseudogenization, neofunctionalization or subfunctionalization. Rice is a model for functional genomics research, an important crop for human nutrition and a target for biofortification. Increased zinc and iron content in the rice grain could be achieved by manipulation of metal transporters. Here, we describe the ZINC-INDUCED FACILITATOR-LIKE (ZIFL) gene family in plants, and characterize the genomic structure and expression of rice paralogs, which are highly affected by segmental duplication. Results Sequences of sixty-eight ZIFL genes, from nine plant species, were comparatively analyzed. Although related to MSF_1 proteins, ZIFL protein sequences consistently grouped separately. Specific ZIFL sequence signatures were identified. Monocots harbor a larger number of ZIFL genes in their genomes than dicots, probably a result of a lineage-specific expansion. The rice ZIFL paralogs were named OsZIFL1 to OsZIFL13 and characterized. The genomic organization of the rice ZIFL genes seems to be highly influenced by segmental and tandem duplications and concerted evolution, as rice genome contains five highly similar ZIFL gene pairs. Most rice ZIFL promoters are enriched for the core sequence of the Fe-deficiency-related box IDE1. Gene expression analyses of different plant organs, growth stages and treatments, both from our qPCR data and from microarray databases, revealed that the duplicated ZIFL gene pairs are mostly co-expressed. Transcripts of OsZIFL4, OsZIFL5, OsZIFL7, and OsZIFL12 accumulate in response to Zn-excess and Fe-deficiency in roots, two stresses with partially overlapping responses. Conclusions We suggest that ZIFL genes have different evolutionary histories in monocot and dicot lineages. In rice, concerted evolution affected ZIFL duplicated genes, possibly maintaining similar

  18. Lineage-Specific Methyltransferases Define the Methylome of the Globally Disseminated Escherichia coli ST131 Clone

    PubMed Central

    Forde, Brian M.; Phan, Minh-Duy; Gawthorne, Jayde A.; Ashcroft, Melinda M.; Stanton-Cook, Mitchell; Sarkar, Sohinee; Peters, Kate M.; Chan, Kok-Gan; Chong, Teik Min; Yin, Wai-Fong; Upton, Mathew

    2015-01-01

    ABSTRACT Escherichia coli sequence type 131 (ST131) is a clone of uropathogenic E. coli that has emerged rapidly and disseminated globally in both clinical and community settings. Members of the ST131 lineage from across the globe have been comprehensively characterized in terms of antibiotic resistance, virulence potential, and pathogenicity, but to date nothing is known about the methylome of these important human pathogens. Here we used single-molecule real-time (SMRT) PacBio sequencing to determine the methylome of E. coli EC958, the most-well-characterized completely sequenced ST131 strain. Our analysis of 52,081 methylated adenines in the genome of EC958 discovered three m6A methylation motifs that have not been described previously. Subsequent SMRT sequencing of isogenic knockout mutants identified the two type I methyltransferases (MTases) and one type IIG MTase responsible for m6A methylation of novel recognition sites. Although both type I sites were rare, the type IIG sites accounted for more than 12% of all methylated adenines in EC958. Analysis of the distribution of MTase genes across 95 ST131 genomes revealed their prevalence is highly conserved within the ST131 lineage, with most variation due to the presence or absence of mobile genetic elements on which individual MTase genes are located. PMID:26578678

  19. New Functional Signatures for Understanding Melanoma Biology from Tumor Cell Lineage-Specific Analysis.

    PubMed

    Rambow, Florian; Job, Bastien; Petit, Valérie; Gesbert, Franck; Delmas, Véronique; Seberg, Hannah; Meurice, Guillaume; Van Otterloo, Eric; Dessen, Philippe; Robert, Caroline; Gautheret, Daniel; Cornell, Robert A; Sarasin, Alain; Larue, Lionel

    2015-10-27

    Molecular signatures specific to particular tumor types are required to design treatments for resistant tumors. However, it remains unclear whether tumors and corresponding cell lines used for drug development share such signatures. We developed similarity core analysis (SCA), a universal and unsupervised computational framework for extracting core molecular features common to tumors and cell lines. We applied SCA to mRNA/miRNA expression data from various sources, comparing melanoma cell lines and metastases. The signature obtained was associated with phenotypic characteristics in vitro, and the core genes CAPN3 and TRIM63 were implicated in melanoma cell migration/invasion. About 90% of the melanoma signature genes belong to an intrinsic network of transcription factors governing neural development (TFAP2A, DLX2, ALX1, MITF, PAX3, SOX10, LEF1, and GAS7) and miRNAs (211-5p, 221-3p, and 10a-5p). The SCA signature effectively discriminated between two subpopulations of melanoma patients differing in overall survival, and classified MEKi/BRAFi-resistant and -sensitive melanoma cell lines. PMID:26489459

  20. Development of Peptide-Based Lineage-Specific Serology for Chronic Chagas Disease: Geographical and Clinical Distribution of Epitope Recognition

    PubMed Central

    Bhattacharyya, Tapan; Falconar, Andrew K.; Luquetti, Alejandro O.; Costales, Jaime A.; Grijalva, Mario J.; Lewis, Michael D.; Messenger, Louisa A.; Tran, Trang T.; Ramirez, Juan-David; Guhl, Felipe; Carrasco, Hernan J.; Diosque, Patricio; Garcia, Lineth; Litvinov, Sergey V.; Miles, Michael A.

    2014-01-01

    Background Chagas disease, caused by infection with the protozoan Trypanosoma cruzi, remains a serious public health issue in Latin America. Genetically diverse, the species is sub-divided into six lineages, known as TcI–TcVI, which have disparate geographical and ecological distributions. TcII, TcV, and TcVI are associated with severe human disease in the Southern Cone countries, whereas TcI is associated with cardiomyopathy north of the Amazon. T. cruzi persists as a chronic infection, with cardiac and/or gastrointestinal symptoms developing years or decades after initial infection. Identifying an individual's history of T. cruzi lineage infection directly by genotyping of the parasite is complicated by the low parasitaemia and sequestration in the host tissues. Methodology/Principal Findings We have applied here serology against lineage-specific epitopes of the T. cruzi surface antigen TSSA, as an indirect approach to allow identification of infecting lineage. Chagasic sera from chronic patients from a range of endemic countries were tested by ELISA against synthetic peptides representing lineage-specific TSSA epitopes bound to avidin-coated ELISA plates via a biotin labelled polyethylene glycol-glycine spacer to increase rotation and ensure each amino acid side chain could freely interact with their antibodies. 79/113 (70%) of samples from Brazil, Bolivia, and Argentina recognised the TSSA epitope common to lineages TcII/TcV/TcVI. Comparison with clinical information showed that a higher proportion of Brazilian TSSApep-II/V/VI responders had ECG abnormalities than non-responders (38% vs 17%; p<0.0001). Among northern chagasic sera 4/20 (20%) from Ecuador reacted with this peptide; 1/12 Venezuelan and 1/34 Colombian samples reacted with TSSApep-IV. In addition, a proposed TcI-specific epitope, described elsewhere, was demonstrated here to be highly conserved across lineages and therefore not applicable to lineage-specific serology. Conclusions

  1. A genome survey sequencing of the Java mouse deer (Tragulus javanicus) adds new aspects to the evolution of lineage specific retrotransposons in Ruminantia (Cetartiodactyla).

    PubMed

    Gallus, S; Kumar, V; Bertelsen, M F; Janke, A; Nilsson, M A

    2015-10-25

    Ruminantia, the ruminating, hoofed mammals (cow, deer, giraffe and allies) are an unranked artiodactylan clade. Around 50-60 million years ago the BovB retrotransposon entered the ancestral ruminantian genome through horizontal gene transfer. A survey genome screen using 454-pyrosequencing of the Java mouse deer (Tragulus javanicus) and the lesser kudu (Tragelaphus imberbis) was done to investigate and to compare the landscape of transposable elements within Ruminantia. The family Tragulidae (mouse deer) is the only representative of Tragulina and phylogenetically important, because it represents the earliest divergence in Ruminantia. The data analyses show that, relative to other ruminantian species, the lesser kudu genome has seen an expansion of BovB Long INterspersed Elements (LINEs) and BovB related Short INterspersed Elements (SINEs) like BOVA2. In comparison the genome of Java mouse deer has fewer BovB elements than other ruminants, especially Bovinae, and has in addition a novel CHR-3 SINE most likely propagated by LINE-1. By contrast the other ruminants have low amounts of CHR SINEs but high numbers of actively propagating BovB-derived and BovB-propagated SINEs. The survey sequencing data suggest that the transposable element landscape in mouse deer (Tragulina) is unique among Ruminantia, suggesting a lineage specific evolutionary trajectory that does not involve BovB mediated retrotransposition. This shows that the genomic landscape of mobile genetic elements can rapidly change in any lineage. PMID:26123917

  2. Autoimmune type 1 diabetes genetic susceptibility encoded by human leukocyte antigen DRB1 and DQB1 genes in Tunisia.

    PubMed

    Stayoussef, Mouna; Benmansour, Jihen; Al-Irhayim, Abdul-Qader; Said, Hichem B; Rayana, Chiheb B; Mahjoub, Touhami; Almawi, Wassim Y

    2009-08-01

    Human leukocyte antigen (HLA) class II genes contribute to the genetic susceptibility to type 1 diabetes (T1D), and susceptible alleles and haplotypes were implicated in the pathogenesis of T1D. This study investigated the heterogeneity in HLA class II haplotype distribution among Tunisian patients with T1D. This was a retrospective case control study done in Monastir in central Tunisia. The subjects comprised 88 T1D patients and 112 healthy controls. HLA-DRB1 and -DQB1 genotyping was done by PCR-sequence-specific priming. Significant DRB1 and DQB1 allelic differences were seen between T1D patients and controls; these differences comprised DRB1*030101 and DQB1*0302, which were higher in T1D patients than in control subjects, and DRB1*070101, DRB1*110101, DQB1*030101, and DQB1*060101, which were lower in T1D patients than in control subjects. In addition, the frequencies of DRB1*030101-DQB1*0201 and DRB1*040101-DQB1*0302 were higher in T1D patients than in control subjects, and the frequencies of DRB1*070101-DQB1*0201 and DRB1*110101-DQB1*030101 haplotypes were lower in T1D patients than in control subjects. Multiple logistic regression analysis revealed the positive association of DRB1*030101-DQB1*0201 and DRB1*040101-DQB1*0302 and the negative association of only DRB1*070101-DQB1*0201 haplotypes with T1D. Furthermore, a significantly increased prevalence of DRB1*030101-DQB1*0201 homozygotes was seen for T1D subjects than for control subjects. Our results confirm the association of specific HLA-DR and -DQ alleles and haplotypes with T1D in Tunisians. The identification of similar and unique haplotypes in Tunisians compared to other Caucasians highlights the need for evaluating the contribution of HLA class II to the genetic susceptibility to T1D with regard to haplotype usage and also to ethnic origin and racial background. PMID:19553558

  3. Autoimmune Type 1 Diabetes Genetic Susceptibility Encoded by Human Leukocyte Antigen DRB1 and DQB1 Genes in Tunisia▿

    PubMed Central

    Stayoussef, Mouna; Benmansour, Jihen; Al-Irhayim, Abdul-Qader; Said, Hichem B.; Rayana, Chiheb B.; Mahjoub, Touhami; Almawi, Wassim Y.

    2009-01-01

    Human leukocyte antigen (HLA) class II genes contribute to the genetic susceptibility to type 1 diabetes (T1D), and susceptible alleles and haplotypes were implicated in the pathogenesis of T1D. This study investigated the heterogeneity in HLA class II haplotype distribution among Tunisian patients with T1D. This was a retrospective case control study done in Monastir in central Tunisia. The subjects comprised 88 T1D patients and 112 healthy controls. HLA-DRB1 and -DQB1 genotyping was done by PCR-sequence-specific priming. Significant DRB1 and DQB1 allelic differences were seen between T1D patients and controls; these differences comprised DRB1*030101 and DQB1*0302, which were higher in T1D patients than in control subjects, and DRB1*070101, DRB1*110101, DQB1*030101, and DQB1*060101, which were lower in T1D patients than in control subjects. In addition, the frequencies of DRB1*030101-DQB1*0201 and DRB1*040101-DQB1*0302 were higher in T1D patients than in control subjects, and the frequencies of DRB1*070101-DQB1*0201 and DRB1*110101-DQB1*030101 haplotypes were lower in T1D patients than in control subjects. Multiple logistic regression analysis revealed the positive association of DRB1*030101-DQB1*0201 and DRB1*040101-DQB1*0302 and the negative association of only DRB1*070101-DQB1*0201 haplotypes with T1D. Furthermore, a significantly increased prevalence of DRB1*030101-DQB1*0201 homozygotes was seen for T1D subjects than for control subjects. Our results confirm the association of specific HLA-DR and -DQ alleles and haplotypes with T1D in Tunisians. The identification of similar and unique haplotypes in Tunisians compared to other Caucasians highlights the need for evaluating the contribution of HLA class II to the genetic susceptibility to T1D with regard to haplotype usage and also to ethnic origin and racial background. PMID:19553558

  4. Atypical Regulation of a Green Lineage-Specific B-Type Cyclin-Dependent Kinase1

    PubMed Central

    Corellou, Florence; Camasses, Alain; Ligat, Laetitia; Peaucellier, Gérard; Bouget, François-Yves

    2005-01-01

    Cyclin-dependent kinases (CDKs) are the main regulators of cell cycle progression in eukaryotes. The role and regulation of canonical CDKs, such as the yeast (Saccharomyces cerevisiae) Cdc2 or plant CDKA, have been extensively characterized. However, the function of the plant-specific CDKB is not as well understood. Besides being involved in cell cycle control, Arabidopsis (Arabidopsis thaliana) CDKB would integrate developmental processes to cell cycle progression. We investigated the role of CDKB in Ostreococcus (Ostreococcus tauri), a unicellular green algae with a minimal set of cell cycle genes. In this primitive alga, at the basis of the green lineage, CDKB has integrated two levels of regulations: It is regulated by Tyr phosphorylation like cdc2/CDKA and at the level of synthesis-like B-type CDKs. Furthermore, Ostreococcus CDKB/cyclin B accounts for the main peak of mitotic activity, and CDKB is able to rescue a yeast cdc28ts mutant. By contrast, Ostreococcus CDKA is not regulated by Tyr phosphorylation, and it exhibits a low and steady-state activity from DNA replication to exit of mitosis. This suggests that from a major role in the control of mitosis in green algae, CDKB has evolved in higher plants to assume other functions outside the cell cycle. PMID:15965018

  5. The evolution of lineage-specific clusters of single nucleotide substitutions in the human genome.

    PubMed

    Xu, Ke; Wang, Jianrong; Elango, Navin; Yi, Soojin V

    2013-10-01

    Genomic regions harboring large numbers of human-specific single nucleotide substitutions are of significant interest since they are potential genomic foci underlying the evolution of human-specific traits as well as human adaptive evolution. Previous studies aimed to identify such regions either used pre-defined genomic locations such as coding sequences and conserved genomic elements or employed sliding window methods. Such approaches may miss clusters of substitutions occurring in regions other than those pre-defined locations, or not be able to distinguish human-specific clusters of substitutions from regions of generally high substitution rates. Here, we conduct a 'maximal segment' analysis to scan the whole human genome to identify clusters of human-specific substitutions that occurred since the divergence of the human and the chimpanzee genomes. This method can identify species-specific clusters of substitutions while not relying on pre-defined regions. We thus identify thousands of clusters of human-specific single nucleotide substitutions. The evolution of such clusters is driven by a combination of several different evolutionary processes including increased regional mutation rate, recombination-associated processes, and positive selection. These newly identified regions of human-specific substitution clusters include large numbers of previously identified human accelerated regions, and exhibit significant enrichments of genes involved in several developmental processes. Our study provides a useful tool to study the evolution of the human genome. PMID:23770436

  6. Lineage-specific evolution of echinoderm mitochondrial ATP synthase subunit 8.

    PubMed

    De Giorgi, C; Martiradonna, A; Pesole, G; Saccone, C

    1997-06-01

    Peculiar evolutionary properties of the subunit 8 of mitochondrial ATP synthase (ATPase8) are revealed by comparative analyses carried out between both closely and distantly related species of echinoderms. The analysis of nucleotide substitution in the three echinoids demonstrated a relaxation of amino acid functional constraints. The deduced protein sequences display a well conserved domain at the N-terminus, while the central part is very variable. At the C-terminus, the broad distribution of positively charged amino acids, which is typical of other organisms, is not conserved in the two different echinoderm classes of the sea urchins and of the sea stars. Instead, a motif of three amino acids, so far not described elsewhere, is conserved in sea urchins and is found to be very similar to the motif present in the sea stars. Our results indicate that the N-terminal region seems to follow the same evolutionary pattern in different organisms, while the maintenance of the C-terminal part in a phylum-specific manner may reflect the co-evolution of mitochondrial and nuclear genes. PMID:9298708

  7. DNA methylation profiles in the human genes for tumor necrosis factors alpha and beta in subpopulations of leukocytes and in leukemias.

    PubMed Central

    Kochanek, S; Radbruch, A; Tesch, H; Renz, D; Doerfler, W

    1991-01-01

    The genomic sequencing technique has been applied to assess the state of methylation in the DNA from human leukocyte subpopulations from healthy individuals and in the DNA from several individuals with myeloid or lymphatic leukemias or non-Hodgkin lymphomas. Leukocyte populations were purified by the high-gradient magnetic cell sorting technique. In the human tumor necrosis factor alpha (TNF-alpha) gene segment between nucleotides 300 and 1150, the specific methylation profile in the DNA from human granulocytes and monocytes is maintained in three cases of myeloid leukemia. In one such case, all 5-methyl-2'-deoxycytidine residues have been replaced by cytidine. In a chronic lymphatic T-cell leukemia, all 5-methyl-2'-deoxycytidine residues have been substituted by cytidine. In normal B lymphocytes, in two cases of chronic lymphatic B-cell leukemias and two cases of non-Hodgkin lymphomas, all 5'-CG-3' sequences in this gene segment are devoid of methylation. In the TNF-beta gene, DNA methylation is decreased in several examples of acute or chronic myeloid leukemias in comparison to normal human granulocytes or monocytes, whose DNA is almost completely methylated between nucleotides 700 and 900. In human T and B lymphocytes, the main producers of TNF-beta, in three instances of chronic lymphatic leukemias and two cases of non-Hodgkin lymphomas, all 5'-CG-3' sequences are unmethylated in this region. The DNA from the human HeLa cell line is highly methylated at all 5'-CG-3' sequences in the TNF-alpha and -beta genes. The TNF-alpha gene is transcribed in the cells of one case of acute myeloid leukemia in which the analyzed region of the TNF-alpha gene is completely unmethylated. The TNF-beta gene is not transcribed in any of the malignant cells tested. Images PMID:2062856

  8. Parallel Evolution and Lineage-Specific Expansion of RNA Editing in Ctenophores.

    PubMed

    Kohn, Andrea B; Sanford, Rachel S; Yoshida, Masa-aki; Moroz, Leonid L

    2015-12-01

    RNA editing is a process of targeted alterations of nucleotides in all types of RNA molecules (e.g., rRNA, tRNA, mRNA, and miRNA). As a result, the transcriptional output differs from its genomic DNA template. RNA editing can be defined both by biochemical mechanisms and by enzymes that perform these reactions. There are high levels of RNA editing detected in the mammalian nervous system, suggesting that nervous systems use this mechanism to increase protein diversity, because the post-transcription modifications lead to new gene products with novel functions. By re-annotating the ctenophore genomes, we found that the number of predicted RNA-editing enzymes is comparable to the numbers in mammals, but much greater than in other non-bilaterian basal metazoans. However, the overall molecular diversity of RNA-editing enzymes in ctenophores is lower, suggesting a possible "compensation" by an expansion of the ADAT1-like subfamily in this lineage. In two genera of ctenophores, Pleurobrachia and Mnemiopsis, there are high levels of expression for RNA-editing enzymes in their aboral organs, the integrative center involved in control of locomotion and geotaxis. This finding supports the hypothesis that RNA editing is correlated with the complexity of tissues and behaviors. Smaller numbers of RNA-editing enzymes in Porifera and Placozoa also correlates with the primary absence of neural and muscular systems in these lineages. In ctenophores, the expansion of the RNA-editing machinery can also provide mechanisms that support the remarkable capacity for regeneration in these animals. In summary, despite their compact genomes, a wide variety of epigenomic mechanisms employed by ctenophores and other non-bilaterian basal metazoans can provide novel insights into the evolutionary origins of biological novelties. PMID:26089435

  9. Electrospun scaffolds for multiple tissues regeneration in vivo through topography dependent induction of lineage specific differentiation.

    PubMed

    Yin, Zi; Chen, Xiao; Song, Hai-Xin; Hu, Jia-Jie; Tang, Qiao-Mei; Zhu, Ting; Shen, Wei-Liang; Chen, Jia-Lin; Liu, Huanhuan; Heng, Boon Chin; Ouyang, Hong-Wei

    2015-03-01

    Physical topographic cues from various substrata have been shown to exert profound effects on the growth and differentiation of stem cells due to their niche-mimicking features. However, the biological function of different topographic materials utilized as bio-scaffolds in vivo have not been rigorously characterized. This study investigated the divergent differentiation pathways of mesenchymal stem cells (MSCs) and neo-tissue formation trigged by aligned and randomly-oriented fibrous scaffolds, both in vitro and in vivo. The aligned group was observed to form more mature tendon-like tissue in the Achilles tendon injury model, as evidenced by histological scoring and collagen I immunohistochemical staining data. In contrast, the randomly-oriented group exhibited much chondrogenesis and subsequent bone tissue formation through ossification. Additionally, X-ray imaging and osteocalcin immunohistochemical staining also demonstrated that osteogenesis in vivo is driven by randomly oriented topography. Furthermore, MSCs on the aligned substrate exhibited tenocyte-like morphology and enhanced tenogenic differentiation compared to cells grown on randomly-oriented scaffold. qRT-PCR analysis of osteogenic marker genes and alkaline phosphatase (ALP) staining demonstrated that MSCs cultured on randomly-oriented fiber scaffolds displayed enhanced osteogenic differentiation compared with cells cultured on aligned fiber scaffolds. Finally, it was demonstrated that cytoskeletal tension release abrogated the divergent differentiation pathways on different substrate topography. Collectively, these findings illustrate the relationship between topographic cues of the scaffold and their inductive role in tissue regeneration; thus providing an insight into future development of smart functionalized bio-scaffold design and its application in tissue engineering. PMID:25617136

  10. Late regulation of immune genes and microRNAs in circulating leukocytes in a pig model of influenza A (H1N2) infection

    PubMed Central

    Brogaard, Louise; Heegaard, Peter M. H.; Larsen, Lars E.; Mortensen, Shila; Schlegel, Michael; Dürrwald, Ralf; Skovgaard, Kerstin

    2016-01-01

    MicroRNAs (miRNAs) are a class of short regulatory RNA molecules which are implicated in modulating gene expression. Levels of circulating, cell-associated miRNAs in response to influenza A virus (IAV) infection has received limited attention so far. To further understand the temporal dynamics and biological implications of miRNA regulation in circulating leukocytes, we collected blood samples before and after (1, 3, and 14 days) IAV challenge of pigs. Differential expression of miRNAs and innate immune factor mRNA transcripts was analysed using RT-qPCR. A total of 20 miRNAs were regulated after IAV challenge, with the highest number of regulated miRNAs seen on day 14 after infection at which time the infection was cleared. Targets of the regulated miRNAs included genes involved in apoptosis and cell cycle regulation. Significant regulation of both miRNAs and mRNA transcripts at 14 days after challenge points to a protracted effect of IAV infection, potentially affecting the host’s ability to respond to secondary infections. In conclusion, experimental IAV infection of pigs demonstrated the dynamic nature of miRNA and mRNA regulation in circulating leukocytes during and after infection, and revealed the need for further investigation of the potential immunosuppressing effect of miRNA and innate immune signaling after IAV infection. PMID:26893019

  11. Whole blood transcriptional profiling reveals significant down-regulation of human leukocyte antigen class I and II genes in essential thrombocythemia, polycythemia vera and myelofibrosis.

    PubMed

    Skov, Vibe; Riley, Caroline Hasselbalch; Thomassen, Mads; Larsen, Thomas Stauffer; Jensen, Morten K; Bjerrum, Ole Weis; Kruse, Torben A; Hasselbalch, Hans Carl

    2013-10-01

    Gene expression profiling studies in the Philadelphia-negative chronic myeloproliferative neoplasms have revealed significant deregulation of several immune and inflammation genes that might be of importance for clonal evolution due to defective tumor immune surveillance. Other mechanisms might be down-regulation of major histocompatibility (MHC) class I and II genes, which are used by tumor cells to escape antitumor T-cell-mediated immune responses. We have performed whole blood transcriptional profiling of genes encoding human leukocyte antigen (HLA) class I and II molecules, β2-microglobulin and members of the antigen processing machinery of HLA class I molecules (LMP2, LMP7, TAP1, TAP2 and tapasin). The findings of significant down-regulation of several of these genes may possibly be of major importance for defective tumor immune surveillance. Since up-regulation of HLA genes is recorded during treatment with epigenome modulating agents (DNA-hypomethylators and DNA-hyperacetylators [histone deacetylase inhibitors]) and interferon-α2, our findings call for prospective transcriptional studies of HLA genes during treatment with these agents. PMID:23302045

  12. Systems-level studies of glycosyltransferase gene expression and enzyme activity that are associated with the selectin binding function of human leukocytes

    PubMed Central

    Marathe, Dhananjay D.; Chandrasekaran, E. V.; Lau, Joseph T. Y.; Matta, Khushi L.; Neelamegham, Sriram

    2008-01-01

    The application of systems biology methods in the emerging field of glycomics requires the collection and integration of glycosyltransferase data at the gene and enzyme level for the purpose of hypothesis generation. We systematically examined the relationship between gene expression, glycosyltransferase activity, glycan expression, and selectin-binding function in different systems, including human neutrophils, undifferentiated HL-60 (human promyelocytic cells), differentiated HL-60, and HL-60 synchronized in specific growth phases. Results demonstrate that 1) the sLeX (sialyl-Lewis-X) epitope is expressed in P-selectin glycoprotein ligand-1 (PSGL-1) from neutrophils at higher levels compared with HL-60. This variation may be due to differences in the relative activities of α1,3-fucosyltransferases and α2,3-sialyltransferases in these two cell types. 2) HL-60 cell differentiation along granulocyte lineage increased the activity of β1,4GalT and β1,3GlcNAcT by 1.6- to 3.2-fold. This may contribute to LacNAc chain extension as evidenced by the 1.7-fold increase in DSA-lectin (lectin recognizing LacNAc) binding to cells after differentiation. 3) The activity of enzymes contributing to sLeX formation in leukocytes likely varies as ST3[Galβ1,4GlcNAc] ≤ α1,3FT[sialyl-LacNAc] < β1,3GlcNAcT. 4) O-glycan specific glycosyltransferase activity does not undergo periodic variation with cell cycle phases. Overall, gene expression and enzyme activity data combined with knowledge of biochemistry can predict the resulting glycan structures and yield viable experimentally testable hypothesis.—Marathe, D. D., Chandrasekaran, E. V., Lau, J. T. Y., Matta, K. L., Neelamegham, S. Systems-level studies of glycosyltransferase gene expression and enzyme activity that are associated with the selectin binding function of human leukocytes. PMID:18716032

  13. Survey of Transcript Expression in Rainbow Trout Leukocytes Reveals a Major Contribution of Interferon-Responsive Genes in the Early Response to a Rhabdovirus Infection

    PubMed Central

    O'Farrell, Caroline; Vaghefi, Nikta; Cantonnet, Monique; Buteau, Bénédicte; Boudinot, Pierre; Benmansour, Abdenour

    2002-01-01

    Virus infections induce changes in the expression of host cell genes. A global knowledge of these modifications should help to better understand the virus/host cell interactions. To obtain a more comprehensive view of the rainbow trout response to a viral infection, we used the subtractive suppressive hybridization methodology in the viral hemorrhagic septicemia model of infection. We infected rainbow trout leukocytes with viral hemorrhagic septicemia virus (VHSV), and total RNA from infected and mock-infected cells was compared at 40 h postinfection. Twenty-four virus-induced genes were ultimately retrieved from the subtracted cDNA library, and their differential expression was further confirmed by semiquantitative reverse transcription-PCR and Northern blot analysis. Among these sequences, three were already described as VHSV-induced genes. Eight sequences with known homologs were extended to full-length cDNA using 5′ and 3′ rapid amplification of cDNA ends, and they were subsequently divided into three functional subsets. Four genes were homologous to mammalian interferon responsive genes, three were similar to chemo-attractant molecules (CXC chemokine, galectin), and two had nucleic acid binding domains. All of the virus-induced genes were also induced by rainbow trout interferon, indicating that the interferon pathway is the predominant component of the anti-VHSV response. They were also expressed in vivo in experimentally infected fish, indicating their biological relevance in natural infection. PMID:12134009

  14. Lineage-specific effects of Notch/Numb signaling in post-embryonic development of the Drosophila brain.

    PubMed

    Lin, Suewei; Lai, Sen-Lin; Yu, Huang-Hsiang; Chihara, Takahiro; Luo, Liqun; Lee, Tzumin

    2010-01-01

    Numb can antagonize Notch signaling to diversify the fates of sister cells. We report here that paired sister cells acquire different fates in all three Drosophila neuronal lineages that make diverse types of antennal lobe projection neurons (PNs). Only one in each pair of postmitotic neurons survives into the adult stage in both anterodorsal (ad) and ventral (v) PN lineages. Notably, Notch signaling specifies the PN fate in the vPN lineage but promotes programmed cell death in the missing siblings in the adPN lineage. In addition, Notch/Numb-mediated binary sibling fates underlie the production of PNs and local interneurons from common precursors in the lAL lineage. Furthermore, Numb is needed in the lateral but not adPN or vPN lineages to prevent the appearance of ectopic neuroblasts and to ensure proper self-renewal of neural progenitors. These lineage-specific outputs of Notch/Numb signaling show that a universal mechanism of binary fate decision can be utilized to govern diverse neural sibling differentiations. PMID:20023159

  15. Lineage-specific function of the noncoding Tsix RNA for Xist repression and Xi reactivation in mice

    PubMed Central

    Ohhata, Tatsuya; Senner, Claire E.; Hemberger, Myriam; Wutz, Anton

    2011-01-01

    The noncoding Tsix RNA is an antisense repressor of Xist and regulates X inactivation in mice. Tsix is essential for preventing the inactivation of the maternally inherited X chromosome in extraembryonic lineages where imprinted X-chromosome inactivation (XCI) occurs. Here we establish an inducible Tsix expression system for investigating Tsix function in development. We show that Tsix has a clear functional window in extraembryonic development. Within this window, Tsix can repress Xist, which is accompanied by DNA methylation of the Xist promoter. As a consequence of Xist repression, reactivation of the inactive X chromosome (Xi) is widely observed. In the parietal endoderm, Tsix represses Xist and causes reactivation of an Xi-linked GFP transgene throughout development, whereas Tsix progressively loses its Xist-repressing function from embryonic day 9.5 (E9.5) onward in trophoblast giant cells and spongiotrophoblast, suggesting that Tsix function depends on a lineage-specific environment. Our data also demonstrate that the maintenance of imprinted XCI requires Xist expression in specific extraembryonic tissues throughout development. This finding shows that reversible XCI is not exclusive to pluripotent cells, and that in some lineages cell differentiation is not accompanied by a stabilization of the Xi. PMID:21852535

  16. LATS-YAP/TAZ controls lineage specification by regulating TGFβ signaling and Hnf4α expression during liver development

    PubMed Central

    Lee, Da-Hye; Park, Jae Oh; Kim, Tae-Shin; Kim, Sang-Kyum; Kim, Tack-hoon; Kim, Min-chul; Park, Gun Soo; Kim, Jeong-Hwan; Kuninaka, Shinji; Olson, Eric N.; Saya, Hideyuki; Kim, Seon-Young; Lee, Ho; Lim, Dae-Sik

    2016-01-01

    The Hippo pathway regulates the self-renewal and differentiation of various adult stem cells, but its role in cell fate determination and differentiation during liver development remains unclear. Here we report that the Hippo pathway controls liver cell lineage specification and proliferation separately from Notch signalling, using mice and primary hepatoblasts with liver-specific knockout of Lats1 and Lats2 kinase, the direct upstream regulators of YAP and TAZ. During and after liver development, the activation of YAP/TAZ induced by loss of Lats1/2 forces hepatoblasts or hepatocytes to commit to the biliary epithelial cell (BEC) lineage. It increases BEC and fibroblast proliferation by up-regulating TGFβ signalling, but suppresses hepatoblast to hepatocyte differentiation by repressing Hnf4α expression. Notably, oncogenic YAP/TAZ activation in hepatocytes induces massive p53-dependent cell senescence/death. Together, our results reveal that YAP/TAZ activity levels govern liver cell differentiation and proliferation in a context-dependent manner. PMID:27358050

  17. Cyanobacterial symbionts diverged in the late Cretaceous towards lineage-specific nitrogen fixation factories in single-celled phytoplankton

    PubMed Central

    Cornejo-Castillo, Francisco M.; Cabello, Ana M.; Salazar, Guillem; Sánchez-Baracaldo, Patricia; Lima-Mendez, Gipsi; Hingamp, Pascal; Alberti, Adriana; Sunagawa, Shinichi; Bork, Peer; de Vargas, Colomban; Raes, Jeroen; Bowler, Chris; Wincker, Patrick; Zehr, Jonathan P.; Gasol, Josep M.; Massana, Ramon; Acinas, Silvia G.

    2016-01-01

    The unicellular cyanobacterium UCYN-A, one of the major contributors to nitrogen fixation in the open ocean, lives in symbiosis with single-celled phytoplankton. UCYN-A includes several closely related lineages whose partner fidelity, genome-wide expression and time of evolutionary divergence remain to be resolved. Here we detect and distinguish UCYN-A1 and UCYN-A2 lineages in symbiosis with two distinct prymnesiophyte partners in the South Atlantic Ocean. Both symbiotic systems are lineage specific and differ in the number of UCYN-A cells involved. Our analyses infer a streamlined genome expression towards nitrogen fixation in both UCYN-A lineages. Comparative genomics reveal a strong purifying selection in UCYN-A1 and UCYN-A2 with a diversification process ∼91 Myr ago, in the late Cretaceous, after the low-nutrient regime period occurred during the Jurassic. These findings suggest that UCYN-A diversified in a co-evolutionary process, wherein their prymnesiophyte partners acted as a barrier driving an allopatric speciation of extant UCYN-A lineages. PMID:27002549

  18. Cyanobacterial symbionts diverged in the late Cretaceous towards lineage-specific nitrogen fixation factories in single-celled phytoplankton.

    PubMed

    Cornejo-Castillo, Francisco M; Cabello, Ana M; Salazar, Guillem; Sánchez-Baracaldo, Patricia; Lima-Mendez, Gipsi; Hingamp, Pascal; Alberti, Adriana; Sunagawa, Shinichi; Bork, Peer; de Vargas, Colomban; Raes, Jeroen; Bowler, Chris; Wincker, Patrick; Zehr, Jonathan P; Gasol, Josep M; Massana, Ramon; Acinas, Silvia G

    2016-01-01

    The unicellular cyanobacterium UCYN-A, one of the major contributors to nitrogen fixation in the open ocean, lives in symbiosis with single-celled phytoplankton. UCYN-A includes several closely related lineages whose partner fidelity, genome-wide expression and time of evolutionary divergence remain to be resolved. Here we detect and distinguish UCYN-A1 and UCYN-A2 lineages in symbiosis with two distinct prymnesiophyte partners in the South Atlantic Ocean. Both symbiotic systems are lineage specific and differ in the number of UCYN-A cells involved. Our analyses infer a streamlined genome expression towards nitrogen fixation in both UCYN-A lineages. Comparative genomics reveal a strong purifying selection in UCYN-A1 and UCYN-A2 with a diversification process ∼91 Myr ago, in the late Cretaceous, after the low-nutrient regime period occurred during the Jurassic. These findings suggest that UCYN-A diversified in a co-evolutionary process, wherein their prymnesiophyte partners acted as a barrier driving an allopatric speciation of extant UCYN-A lineages. PMID:27002549

  19. The shell-forming proteome of Lottia gigantea reveals both deep conservations and lineage-specific novelties.

    PubMed

    Marie, Benjamin; Jackson, Daniel J; Ramos-Silva, Paula; Zanella-Cléon, Isabelle; Guichard, Nathalie; Marin, Frédéric

    2013-01-01

    Proteins that are occluded within the molluscan shell, the so-called shell matrix proteins (SMPs), are an assemblage of biomolecules attractive to study for several reasons. They increase the fracture resistance of the shell by several orders of magnitude, determine the polymorph of CaCO(3) deposited, and regulate crystal nucleation, growth initiation and termination. In addition, they are thought to control the shell microstructures. Understanding how these proteins have evolved is also likely to provide deep insight into events that supported the diversification and expansion of metazoan life during the Cambrian radiation 543 million years ago. Here, we present an analysis of SMPs isolated form the CaCO(3) shell of the limpet Lottia gigantea, a gastropod that constructs an aragonitic cross-lamellar shell. We identified 39 SMPs by combining proteomic analysis with genomic and transcriptomic database interrogations. Among these proteins are various low-complexity domain-containing proteins, enzymes such as peroxidases, carbonic anhydrases and chitinases, acidic calcium-binding proteins and protease inhibitors. This list is likely to contain the most abundant SMPs of the shell matrix. It reveals the presence of both highly conserved and lineage-specific biomineralizing proteins. This mosaic evolutionary pattern suggests that there may be an ancestral molluscan SMP set upon which different conchiferan lineages have elaborated to produce the diversity of shell microstructures we observe nowadays. PMID:23145877

  20. LATS-YAP/TAZ controls lineage specification by regulating TGFβ signaling and Hnf4α expression during liver development.

    PubMed

    Lee, Da-Hye; Park, Jae Oh; Kim, Tae-Shin; Kim, Sang-Kyum; Kim, Tack-Hoon; Kim, Min-Chul; Park, Gun Soo; Kim, Jeong-Hwan; Kuninaka, Shinji; Olson, Eric N; Saya, Hideyuki; Kim, Seon-Young; Lee, Ho; Lim, Dae-Sik

    2016-01-01

    The Hippo pathway regulates the self-renewal and differentiation of various adult stem cells, but its role in cell fate determination and differentiation during liver development remains unclear. Here we report that the Hippo pathway controls liver cell lineage specification and proliferation separately from Notch signalling, using mice and primary hepatoblasts with liver-specific knockout of Lats1 and Lats2 kinase, the direct upstream regulators of YAP and TAZ. During and after liver development, the activation of YAP/TAZ induced by loss of Lats1/2 forces hepatoblasts or hepatocytes to commit to the biliary epithelial cell (BEC) lineage. It increases BEC and fibroblast proliferation by up-regulating TGFβ signalling, but suppresses hepatoblast to hepatocyte differentiation by repressing Hnf4α expression. Notably, oncogenic YAP/TAZ activation in hepatocytes induces massive p53-dependent cell senescence/death. Together, our results reveal that YAP/TAZ activity levels govern liver cell differentiation and proliferation in a context-dependent manner. PMID:27358050

  1. Quantitative reconstruction of leukocyte subsets using DNA methylation

    PubMed Central

    2014-01-01

    Background Cell lineage-specific DNA methylation patterns distinguish normal human leukocyte subsets and can be used to detect and quantify these subsets in peripheral blood. We have developed an approach that uses DNA methylation to simultaneously quantify multiple leukocyte subsets, enabling investigation of immune modulations in virtually any blood sample including archived samples previously precluded from such analysis. Here we assess the performance characteristics and validity of this approach. Results Using Illumina Infinium HumanMethylation27 and VeraCode GoldenGate Methylation Assay microarrays, we measure DNA methylation in leukocyte subsets purified from human whole blood and identify cell lineage-specific DNA methylation signatures that distinguish human T cells, B cells, NK cells, monocytes, eosinophils, basophils and neutrophils. We employ a bioinformatics-based approach to quantify these cell types in complex mixtures, including whole blood, using DNA methylation at as few as 20 CpG loci. A reconstruction experiment confirms that the approach could accurately measure the composition of mixtures of human blood leukocyte subsets. Applying the DNA methylation-based approach to quantify the cellular components of human whole blood, we verify its accuracy by direct comparison to gold standard immune quantification methods that utilize physical, optical and proteomic characteristics of the cells. We also demonstrate that the approach is not affected by storage of blood samples, even under conditions prohibiting the use of gold standard methods. Conclusions Cell mixture distributions within peripheral blood can be assessed accurately and reliably using DNA methylation. Thus, precise immune cell differential estimates can be reconstructed using only DNA rather than whole cells. PMID:24598480

  2. A pursuit of lineage-specific and niche-specific proteome features in the world of archaea

    PubMed Central

    2012-01-01

    Background Archaea evoke interest among researchers for two enigmatic characteristics –a combination of bacterial and eukaryotic components in their molecular architectures and an enormous diversity in their life-style and metabolic capabilities. Despite considerable research efforts, lineage- specific/niche-specific molecular features of the whole archaeal world are yet to be fully unveiled. The study offers the first large-scale in silico proteome analysis of all archaeal species of known genome sequences with a special emphasis on methanogenic and sulphur-metabolising archaea. Results Overall amino acid usage in archaea is dominated by GC-bias. But the environmental factors like oxygen requirement or thermal adaptation seem to play important roles in selection of residues with no GC-bias at the codon level. All methanogens, irrespective of their thermal/salt adaptation, show higher usage of Cys and have relatively acidic proteomes, while the proteomes of sulphur-metabolisers have higher aromaticity and more positive charges. Despite of exhibiting thermophilic life-style, korarchaeota possesses an acidic proteome. Among the distinct trends prevailing in COGs (Cluster of Orthologous Groups of proteins) distribution profiles, crenarchaeal organisms display higher intra-order variations in COGs repertoire, especially in the metabolic ones, as compared to euryarchaea. All methanogens are characterised by a presence of 22 exclusive COGs. Conclusions Divergences in amino acid usage, aromaticity/charge profiles and COG repertoire among methanogens and sulphur-metabolisers, aerobic and anaerobic archaea or korarchaeota and nanoarchaeota, as elucidated in the present study, point towards the presence of distinct molecular strategies for niche specialization in the archaeal world. PMID:22691113

  3. Leukocyte accumulation promoting fibrin deposition is mediated in vivo by P-selectin on adherent platelets

    NASA Astrophysics Data System (ADS)

    Palabrica, Theresa; Lobb, Roy; Furie, Barbara C.; Aronovitz, Mark; Benjamin, Christopher; Hsu, Yen-Ming; Sajer, Susan A.; Furie, Bruce

    1992-10-01

    THE glycoprotein P-selectin is a cell adhesion molecule of stimulated platelets and endothelial cells, which mediates the interaction of these cells with neutrophils and monocytes1,2. It is a membrane component of cell storage granules3-6, and is a member of the selectin family which includes E-selectin and L-selectin7,8. P-selectin recognizes both lineage-specific carbohydrate ligands on monocytes and neutrophils, including the Lewis x antigen, sialic acid, and a protein component9-12. In inflammation and thrombosis, P-selectin may mediate the interaction of leukocytes with platelets bound in the region of tissue injury and with stimulated endothelium1,2. To evaluate the role of P-selectin in platelet-leukocyte adhesion in vivo, the accumulation of leukocytes within an experimental thrombus was explored in an arteriovenous shunt model in baboons13. A Dacron graft implanted within an arteriovenous shunt is thrombogenic, accumulating platelets and fibrin within its lumen. These bound platelets express P-selectin14. Here we show that antibody inhibition of leukocyte binding to P-selectin expressed on platelets immobilized on the graft blocks leukocyte accumulation and inhibits the deposition of fibrin within the thrombus. These results indicate that P-selectin is an important adhesion molecule on platelets, mediating platelet-leukocyte binding in vivo, that the presence of leukocytes in thrombi is mediated by P-selectin, and that these leukocytes promote fibrin deposition.

  4. Insights into leukocyte adhesion deficiency type 2 from a novel mutation in the GDP-fucose transporter gene.

    PubMed

    Hidalgo, Andrés; Ma, Songhui; Peired, Anna J; Weiss, Linnea A; Cunningham-Rundles, Charlotte; Frenette, Paul S

    2003-03-01

    Leukocyte adhesion deficiency type 2 (LADII) is characterized by defective selectin ligand formation, recurrent infection, and mental retardation. This rare syndrome has only been described in 2 kindreds of Middle Eastern descent who have differentially responded to exogenous fucose treatment. The molecular defect was recently ascribed to single and distinct missense mutations in a putative Golgi guanosine diphosphate (GDP)-fucose transporter. Here, we describe a patient of Brazilian origin with features of LADII. Sequencing of the GDP-fucose transporter revealed a novel single nucleotide deletion producing a shift in the open-reading frame and severe truncation of the polypeptide. Overexpression of the mutant protein in the patient's fibroblasts did not rescue fucosylation, suggesting that the deletion ablated the activity of the transporter. Administration of oral L-fucose to the patient produced molecular and clinical responses, as measured by the appearance of selectin ligands, normalization of neutrophil counts, and prevention of infectious recurrence. The lower neutrophil counts paralleled improved neutrophil interactions with activated endothelium in cremasteric venules of nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice. However, fucose supplementation induced autoimmune neutropenia and the appearance of H antigen on erythrocytes, albeit without evidence of intravascular hemolysis. The robust response to fucose despite a severely truncated transporter suggests alternative means to transport GDP-fucose into the Golgi complex. PMID:12406889

  5. Methylation of the leukocyte glucocorticoid receptor gene promoter in adults: associations with early adversity and depressive, anxiety and substance-use disorders.

    PubMed

    Tyrka, A R; Parade, S H; Welch, E S; Ridout, K K; Price, L H; Marsit, C; Philip, N S; Carpenter, L L

    2016-01-01

    Early adversity increases risk for developing psychopathology. Epigenetic modification of stress reactivity genes is a likely mechanism contributing to this risk. The glucocorticoid receptor (GR) gene is of particular interest because of the regulatory role of the GR in hypothalamic-pituitary-adrenal (HPA) axis function. Mounting evidence suggests that early adversity is associated with GR promoter methylation and gene expression. Few studies have examined links between GR promoter methylation and psychopathology, and findings to date have been mixed. Healthy adult participants (N=340) who were free of psychotropic medications reported on their childhood experiences of maltreatment and parental death and desertion. Lifetime depressive and anxiety disorders and past substance-use disorders were assessed using the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition. Methylation of exon 1F of the GR gene (NR3C1) was examined in leukocyte DNA via pyrosequencing. On a separate day, a subset of the participants (n=231) completed the dexamethasone/corticotropin-releasing hormone (Dex/CRH) test. Childhood adversity and a history of past substance-use disorder and current or past depressive or anxiety disorders were associated with lower levels of NR3C1 promoter methylation across the region as a whole and at individual CpG sites (P<0.05). The number of adversities was negatively associated with NR3C1 methylation in participants with no lifetime disorder (P=0.018), but not in those with a lifetime disorder. GR promoter methylation was linked to altered cortisol responses to the Dex/CRH test (P<0.05). This study presents evidence of reduced methylation of NR3C1 in association with childhood maltreatment and depressive, anxiety and substance-use disorders in adults. This finding stands in contrast to our prior work, but is consistent with emerging findings, suggesting complexity in the regulation of this gene. PMID

  6. Methylation of the leukocyte glucocorticoid receptor gene promoter in adults: associations with early adversity and depressive, anxiety and substance-use disorders

    PubMed Central

    Tyrka, A R; Parade, S H; Welch, E S; Ridout, K K; Price, L H; Marsit, C; Philip, N S; Carpenter, L L

    2016-01-01

    Early adversity increases risk for developing psychopathology. Epigenetic modification of stress reactivity genes is a likely mechanism contributing to this risk. The glucocorticoid receptor (GR) gene is of particular interest because of the regulatory role of the GR in hypothalamic–pituitary–adrenal (HPA) axis function. Mounting evidence suggests that early adversity is associated with GR promoter methylation and gene expression. Few studies have examined links between GR promoter methylation and psychopathology, and findings to date have been mixed. Healthy adult participants (N=340) who were free of psychotropic medications reported on their childhood experiences of maltreatment and parental death and desertion. Lifetime depressive and anxiety disorders and past substance-use disorders were assessed using the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition. Methylation of exon 1F of the GR gene (NR3C1) was examined in leukocyte DNA via pyrosequencing. On a separate day, a subset of the participants (n=231) completed the dexamethasone/corticotropin-releasing hormone (Dex/CRH) test. Childhood adversity and a history of past substance-use disorder and current or past depressive or anxiety disorders were associated with lower levels of NR3C1 promoter methylation across the region as a whole and at individual CpG sites (P<0.05). The number of adversities was negatively associated with NR3C1 methylation in participants with no lifetime disorder (P=0.018), but not in those with a lifetime disorder. GR promoter methylation was linked to altered cortisol responses to the Dex/CRH test (P<0.05). This study presents evidence of reduced methylation of NR3C1 in association with childhood maltreatment and depressive, anxiety and substance-use disorders in adults. This finding stands in contrast to our prior work, but is consistent with emerging findings, suggesting complexity in the regulation of this gene. PMID

  7. The Staphylococcus aureus lineage-specific markers collagen adhesin and toxic shock syndrome toxin 1 distinguish multilocus sequence typing clonal complexes within spa clonal complexes.

    PubMed

    Deurenberg, Ruud H; Rijnders, Michelle I A; Sebastian, Silvie; Welling, Maaike A; Beisser, Patrick S; Stobberingh, Ellen E

    2009-10-01

    Spa typing/based upon repeat pattern (BURP) sometimes cannot differentiate multilocus sequence typing (MLST) clonal complexes (CCs) within spa-CCs. It has been observed previously that virulence factors, such as collagen adhesin (CNA) and toxic shock syndrome toxin 1 (TSST-1), are associated with certain Staphylococcus aureus lineages. Analysis of methicillin-sensitive and methicillin-resistant S. aureus by spa typing/BURP and detection of CNA and TSST-1 observed an association between CNA and MLST CC1, 12, 22, 30, 45, 51, and 239 and between TSST-1 and MLST CC30. In spa-CC 012, associated with MLST CC7, CC15, and CC30, MLST CC30 could be distinguished from MLST CC7 and CC15 with CNA and TSST-1 as lineage-specific markers. Lineage-specific markers can overcome clustering of nonrelated MLST CCs into 1 spa-CC. PMID:19748421

  8. Profiling of the peripheral leukocyte transcriptome of dairy cattle reveals gene expression patterns associated with genetic potential for milk production

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mRNA transcript abundance in a cell or tissue is a heritable and quantitative trait that corresponds to the status of gene expression. This principle has been widely applied, using microarrays to identify core regulatory genes for physiological and pathological traits in plants and animals. G...

  9. [Investigation on bovine leukocyte adhesion deficiency].

    PubMed

    Ma, Jin-Zhu; Cui, Yu-Dong; Zhu, Zhan-Bo; Cao, Hong-Wei; Piao, Fan-Ze

    2006-10-01

    Bovine leukocyte adhesion deficiency (BLAD) is autosomal recessive disease. The pathogeny of BLAD is genic mutation of CD18-integrins on the leukocyte. In order to know the carrier and occurrence of bovine leukocyte adhesion deficiency (BLAD) among cows age from one to six years old in China, 1,000 cows were investigated by means of amplifying a CD18 gene fragment via reverse transcriptase-PCR followed by restriction digestion with Taq I. Results showed that 19 cows were BLAD carriers, indicating that the BLAD carrier rate was 1.9 percent. In addition, one cow was found to have BLAD. PMID:17035180

  10. A cleavage clock regulates features of lineage-specific differentiation in the development of a basal branching metazoan, the ctenophore Mnemiopsis leidyi

    PubMed Central

    2014-01-01

    DNA to cytoplasmic ratio, are critical for the appearance of lineage-specific differentiation. Conclusion Our work corroborates previous studies demonstrating that the cleavage program is causally involved in the spatial segregation and/or activation of factors that give rise to distinct cell types in ctenophore development. These factors are segregated independently to the appropriate lineage at the 8- and the 16-cell stages and have features of a clock, such that comb-plate-like cilia and light-emitting photoproteins appear at roughly the same developmental time in cleavage-arrested embryos as they do in untreated embryos. Nuclear division, which possibly affects DNA-cytoplasmic ratios, appears to be important in the timing of differentiation markers. Evidence suggests that the 60-cell stage, just prior to gastrulation, is the time of zygotic gene activation. Such cleavage-clock-regulated phenomena appear to be widespread amongst the Metazoa and these cellular and molecular developmental mechanisms probably evolved early in metazoan evolution. PMID:24485336

  11. Adenoviral gene delivery of elafin and secretory leukocyte protease inhibitor attenuates NF-kappa B-dependent inflammatory responses of human endothelial cells and macrophages to atherogenic stimuli.

    PubMed

    Henriksen, Peter A; Hitt, Mary; Xing, Zhou; Wang, Jun; Haslett, Chris; Riemersma, Rudolph A; Webb, David J; Kotelevtsev, Yuri V; Sallenave, Jean-Michel

    2004-04-01

    Atherosclerosis is a chronic inflammatory disease affecting arterial vessels. Strategies to reduce the inflammatory responses of endothelial cells and macrophages may slow lesion development and prevent complications such as plaque rupture. The human protease human neutrophil elastase (HNE), oxidized low density lipoprotein, LPS, and TNF-alpha were chosen as model stimuli of arterial wall inflammation and led to production of the chemokine IL-8 in endothelial cells. To counteract the activity of HNE, we have examined the effects of adenoviral gene delivery of the anti-elastases elafin, previously demonstrated within human atheroma, and murine secretory leukocyte protease inhibitor (SLPI), a related molecule, on the inflammatory responses of human endothelial cells and macrophages to atherogenic stimuli. We developed a technique of precomplexing adenovirus with cationic lipid to augment adenoviral infection efficiency in endothelial cells and to facilitate infection in macrophages. Elafin overexpression protected endothelial cells from HNE-induced IL-8 production and cytotoxicity. Elafin and murine SLPI also reduced endothelial IL-8 release in response to oxidized low density lipoprotein, LPS, and TNF-alpha and macrophage TNF-alpha production in response to LPS. This effect was associated with reduced activation of the inflammatory transcription factor NF-kappaB, through up-regulation of IkappaBalpha, in both cell types. Our work suggests a novel and extended anti-inflammatory role for these HNE inhibitors working as effectors of innate immunity to protect tissues against maladaptive inflammatory responses. Our findings indicate that elafin and SLPI may be gene therapy targets for the treatment of atheroma. PMID:15034071

  12. The African buffalo parasite Theileria. sp. (buffalo) can infect and immortalize cattle leukocytes and encodes divergent orthologues of Theileria parva antigen genes.

    PubMed

    Bishop, R P; Hemmink, J D; Morrison, W I; Weir, W; Toye, P G; Sitt, T; Spooner, P R; Musoke, A J; Skilton, R A; Odongo, D O

    2015-12-01

    African Cape buffalo (Syncerus caffer) is the wildlife reservoir of multiple species within the apicomplexan protozoan genus Theileria, including Theileria parva which causes East coast fever in cattle. A parasite, which has not yet been formally named, known as Theileria sp. (buffalo) has been recognized as a potentially distinct species based on rDNA sequence, since 1993. We demonstrate using reverse line blot (RLB) and sequencing of 18S rDNA genes, that in an area where buffalo and cattle co-graze and there is a heavy tick challenge, T. sp. (buffalo) can frequently be isolated in culture from cattle leukocytes. We also show that T. sp. (buffalo), which is genetically very closely related to T. parva, according to 18s rDNA sequence, has a conserved orthologue of the polymorphic immunodominant molecule (PIM) that forms the basis of the diagnostic ELISA used for T. parva serological detection. Closely related orthologues of several CD8 T cell target antigen genes are also shared with T. parva. By contrast, orthologues of the T. parva p104 and the p67 sporozoite surface antigens could not be amplified by PCR from T. sp. (buffalo), using conserved primers designed from the corresponding T. parva sequences. Collectively the data re-emphasise doubts regarding the value of rDNA sequence data alone for defining apicomplexan species in the absence of additional data. 'Deep 454 pyrosequencing' of DNA from two Theileria sporozoite stabilates prepared from Rhipicephalus appendiculatus ticks fed on buffalo failed to detect T. sp. (buffalo). This strongly suggests that R. appendiculatus may not be a vector for T. sp. (buffalo). Collectively, the data provides further evidence that T. sp. (buffalo). is a distinct species from T. parva. PMID:26543804

  13. The African buffalo parasite Theileria. sp. (buffalo) can infect and immortalize cattle leukocytes and encodes divergent orthologues of Theileria parva antigen genes

    PubMed Central

    Bishop, R.P.; Hemmink, J.D.; Morrison, W.I.; Weir, W.; Toye, P.G.; Sitt, T.; Spooner, P.R.; Musoke, A.J.; Skilton, R.A.; Odongo, D.O.

    2015-01-01

    African Cape buffalo (Syncerus caffer) is the wildlife reservoir of multiple species within the apicomplexan protozoan genus Theileria, including Theileria parva which causes East coast fever in cattle. A parasite, which has not yet been formally named, known as Theileria sp. (buffalo) has been recognized as a potentially distinct species based on rDNA sequence, since 1993. We demonstrate using reverse line blot (RLB) and sequencing of 18S rDNA genes, that in an area where buffalo and cattle co-graze and there is a heavy tick challenge, T. sp. (buffalo) can frequently be isolated in culture from cattle leukocytes. We also show that T. sp. (buffalo), which is genetically very closely related to T. parva, according to 18s rDNA sequence, has a conserved orthologue of the polymorphic immunodominant molecule (PIM) that forms the basis of the diagnostic ELISA used for T. parva serological detection. Closely related orthologues of several CD8 T cell target antigen genes are also shared with T. parva. By contrast, orthologues of the T. parva p104 and the p67 sporozoite surface antigens could not be amplified by PCR from T. sp. (buffalo), using conserved primers designed from the corresponding T. parva sequences. Collectively the data re-emphasise doubts regarding the value of rDNA sequence data alone for defining apicomplexan species in the absence of additional data. ‘Deep 454 pyrosequencing’ of DNA from two Theileria sporozoite stabilates prepared from Rhipicephalus appendiculatus ticks fed on buffalo failed to detect T. sp. (buffalo). This strongly suggests that R. appendiculatus may not be a vector for T. sp. (buffalo). Collectively, the data provides further evidence that T. sp. (buffalo). is a distinct species from T. parva. PMID:26543804

  14. Inhibition of bacterial cell wall-induced leukocyte recruitment and hepatic granuloma formation by TGF-beta gene transfer.

    PubMed

    Song, X; Zeng, L; Pilo, C M; Zagorski, J; Wahl, S M

    1999-10-01

    Intraperitoneal injection of streptococcal cell walls (SCW) into Lewis rats results in dissemination of SCW to the liver, spleen, bone marrow, and peripheral joints. The uptake of SCW by Kupffer cells in the liver initiates a chain of events largely mediated by T lymphocytes and macrophages. Local synthesis and secretion of cytokines and growth factors in response to the persistent SCW lead to the evolution and maintenance of a chronic T cell-dependent granulomatous response and result in granuloma formation and irreversible hepatic fibrosis. In an attempt to impede the development of the chronic granulomatous lesions in the liver, we injected a plasmid DNA encoding TGF-beta 1 i.m. to the SCW animals to determine the effect of TGF-beta 1 gene transfer on the course of liver inflammation and fibrosis. A single injection of plasmid DNA encoding TGF-beta 1 resulted in virtual abolition of the development of the SCW-induced hepatic granuloma formation and matrix expansion. TGF-beta 1 DNA not only reduced key proinflammatory cytokines including TNF-alpha, IL-1 beta, IFN-gamma, and IL-18, but also inhibited both CXC and CC chemokine production, thereby blocking inflammatory cell recruitment and accumulation in the liver. Moreover, TGF-beta 1 gene delivery inhibited its own expression in the liver tissue, which is otherwise up-regulated in SCW-injected animals. Our study suggests that TGF-beta 1 gene transfer suppresses hepatic granuloma formation by blocking the recruitment of inflammatory cells to the liver, and thus may provide a new approach to the control of hepatic granulomatous and fibrotic diseases. PMID:10491005

  15. Human leukocyte antigen-G allele polymorphisms have evolved following three different evolutionary lineages based on intron sequences.

    PubMed

    Cervera, Isabel; Herraiz, Miguel Angel; Peñaloza, Jorge; Barbolla, Maria Luz; Jurado, Maria Luisa; Macedo, Jacqueline; Vidart, José Antonio; Martinez-Laso, Jorge

    2010-11-01

    Human leukocyte antigen (HLA)-G alleles follow a different pattern of polymorphism generation from those of the HLA classical I alleles. These polymorphisms have been defined as a result of random permitted point mutations in exons. However, this polymorphism maintenance could have an evolutionary specific pathways based on noncoding regions as introns, 14-bp deletion/insertion (exon 8), or promoter regions. Therefore a systematic sequencing study of HLA-G alleles was done obtaining the complete genomic sequence of 16 different HLA-G alleles: nine alleles were intron and exon confirmatory sequences, four were exon confirmatory and new intron described sequences, and three were new alleles. A 14-bp deletion/insertion polymorphism was also sequenced in these alleles. These sequences, together with those previously published, were compared, and phylogenetic and molecular evolutionary analyses were performed. Results showed the presence of three major specific evolutionary patterns, tentatively named lineages, and the other four as minor lineages (only one allele). The relative age of the major lineages could also be established based on the number of lineage-specific positions and the number of alleles of each lineage. Two main mechanisms are clearly defined in the generation of the lineages (introns), gene conversion, and/or convergent evolution following specific patterns. PMID:20650296

  16. Reconstruction of cyclooxygenase evolution in animals suggests variable, lineage-specific duplications, and homologs with low sequence identity.

    PubMed

    Havird, Justin C; Kocot, Kevin M; Brannock, Pamela M; Cannon, Johanna T; Waits, Damien S; Weese, David A; Santos, Scott R; Halanych, Kenneth M

    2015-04-01

    Cyclooxygenase (COX) enzymatically converts arachidonic acid into prostaglandin G/H in animals and has importance during pregnancy, digestion, and other physiological functions in mammals. COX genes have mainly been described from vertebrates, where gene duplications are common, but few studies have examined COX in invertebrates. Given the increasing ease in generating genomic data, as well as recent, although incomplete descriptions of potential COX sequences in Mollusca, Crustacea, and Insecta, assessing COX evolution across Metazoa is now possible. Here, we recover 40 putative COX orthologs by searching publicly available genomic resources as well as ~250 novel invertebrate transcriptomic datasets. Results suggest the common ancestor of Cnidaria and Bilateria possessed a COX homolog similar to those of vertebrates, although such homologs were not found in poriferan and ctenophore genomes. COX was found in most crustaceans and the majority of molluscs examined, but only specific taxa/lineages within Cnidaria and Annelida. For example, all octocorallians appear to have COX, while no COX homologs were found in hexacorallian datasets. Most species examined had a single homolog, although species-specific COX duplications were found in members of Annelida, Mollusca, and Cnidaria. Additionally, COX genes were not found in Hemichordata, Echinodermata, or Platyhelminthes, and the few previously described COX genes in Insecta lacked appreciable sequence homology (although structural analyses suggest these may still be functional COX enzymes). This analysis provides a benchmark for identifying COX homologs in future genomic and transcriptomic datasets, and identifies lineages for future studies of COX. PMID:25758350

  17. Leukocyte relaxation properties.

    PubMed Central

    Sung, K L; Dong, C; Schmid-Schönbein, G W; Chien, S; Skalak, R

    1988-01-01

    Study of the mechanical properties of leukocytes is useful to understand their passage through narrow capillaries and interaction with other cells. Leukocytes are known to be viscoelastic and their properties have been established by micropipette aspiration techniques. Here, the recovery of leukocytes to their normal spherical form is studied after prolonged deformation in a pipette which is large enough to permit complete entry of the leukocyte. The recovery history is characterized by the time history of the major diameter (d1) and minor diameter (d2). When the cell is removed from the pipette, it shows initially a small rapid recoil followed by a slower asymptotic recovery to the spherical shape. In the presence of cell activation and formation of pseudopods, the time history for recovery is prolonged compared with passive cell recovery. If a protopod pre-existed during the holding period, the recovery only begins when the protopod starts to retract. Images FIGURE 1 PMID:3207829

  18. The expression of two novel orange-spotted grouper (Epinephelus coioides) TNF genes in peripheral blood leukocytes, various organs, and fish larvae.

    PubMed

    Lam, Freda Wai-San; Wu, Szu-Yin; Lin, Shih-Jie; Lin, Chin-Chou; Chen, Yi-Ming; Wang, Han-Ching; Chen, Tzong-Yueh; Lin, Han-Tso; Lin, John Han-You

    2011-02-01

    The tumour necrosis factor (TNF) super-family is a group of important cytokines involved in inflammation, apoptosis, cell proliferation, and the general stimulation of the immune system. The TNF gene has been cloned in some bony fish; however, its counterparts are still unidentified in the majority of fish species. In this study, we cloned gTNF-1 and gTNF-2 from the orange-spotted grouper (Epinephelus coioides), an economically important farmed fish. Both genes include 4 exons and 3 introns and encoded 253 and 241 amino acid proteins with a molecular weight of approximately 27 and 26 kDa, respectively. The identity of the putative amino acid sequences between gTNF-1 and gTNF-2 was only 38%. The positions of cysteine residues, a protease cleavage site, and a transmembrane domain sequence derived from gTNF-1 and gTNF-2 were similar to those in other fish and mammalian TNF-α. The mRNA expression levels of the 2 gTNF molecules were evaluated in unstimulated/stimulated peripheral blood leukocytes, various organs, and fish larvae. Following lipopolysaccharide (LPS) treatment, gTNF-2 was expressed at higher levels, was up-regulated more quickly, and was more sensitive to the immune response than gTNF-1. gTNF-1 was constitutively expressed in the thymus, brain, and spleen, but it was also expressed in the heart, head kidney, and trunk kidney after LPS stimulation. gTNF-2 was constitutively expressed in the thymus, head kidney, trunk kidney, spleen, and intestine; further, gTNF-2 was highly expressed in all organs post-LPS stimulation. Finally, the gTNF expression levels were evaluated at various developmental stages in grouper larvae. A higher variation of gTNF expression levels was observed in fish larvae from a contaminated hatchery. This study revealed the different expression patterns of gTNF-1 and gTNF-2. In addition, gTNF-2 was more sensitive to pathogens than gTNF-1; therefore, it may be an appropriate marker for pathogen invasion and the evaluation of the larval

  19. Abnormalities of the TITF-1 lineage-specific oncogene in NSCLC: Implications in lung cancer pathogenesis and prognosis

    PubMed Central

    Tang, Ximing; Kadara, Humam; Behrens, Carmen; Liu, Diane D.; Xiao, Yun; Rice, David; Gazdar, Adi F.; Fujimoto, Junya; Moran, Cesar; Varella-Garcia, Marileila; Lee, J. Jack; Hong, Waun Ki; Wistuba, Ignacio I.

    2011-01-01

    PURPOSE Emerging evidence suggests that aberrant expression of oncogenes contributes to development of lung malignancy. The thyroid transcription factor 1 (TITF-1) gene functions as a lineage survival gene abnormally expressed in a significant fraction of NSCLCs, in particular lung adenocarcinomas. EXPERIMENTAL DESIGN To better characterize TITF-1 abnormality: patterns in NSCLC, we studied TITF-1’s gene copy number using fluorescent in situ hybridization (FISH) and quantitative PCR, as well as its protein expression by immunohistochemistry analysis in a tissue microarray comprised of surgically resected NSCLC (N=321) including 204 adenocarcinomas and 117 squamous cell carcinomas (SCCs). TITF-1 copy number and protein expression were correlated with patients’ clinicopathologic characteristics, and in a subset of adenocarcinomas with EGFR and KRAS mutation status. RESULTS We found that increased TITF-1 protein expression was prevalent in lung adenocarcinomas only and was significantly associated with female gender (p<0.001), never smokers (p=0.004), presence of EGFR mutations (p=0.05) and better overall survival (all stages, p=0.0478. stages I and II, p=0.002). TITF-1 copy number gain (CBG) was detected by FISH analysis in both adenocarcinomas (18.9%; high CNG, 8.3%) and SCCs (20.1%; high CNG, 3.0%), and correlated significantly with the protein product (p=0.004) and presence of KRAS mutations (p=0.008) in lung adenocarcinomas. Moreover, multivariate analysis revealed that TITF-1 copy number gain was an independent predictor of poor survival of NSCLC (p=0.039). CONCLUSIONS Our integrative study demonstrates that the protein versus genomic expression patterns of TITF-1 have opposing roles in lung cancer prognosis and may occur preferentially in different subsets of NSCLC patients with distinct oncogene mutations. PMID:21257719

  20. The ubiquitous transcription factor CTCF promotes lineage-specific epigenomic remodeling and establishment of transcriptional networks driving cell differentiation

    PubMed Central

    Dubois-Chevalier, Julie; Staels, Bart; Lefebvre, Philippe; Eeckhoute, Jérôme

    2015-01-01

    Cell differentiation relies on tissue-specific transcription factors (TFs) that cooperate to establish unique transcriptomes and phenotypes. However, the role of ubiquitous TFs in these processes remains poorly defined. Recently, we have shown that the CCCTC-binding factor (CTCF) is required for adipocyte differentiation through epigenomic remodelling of adipose tissue-specific enhancers and transcriptional activation of Peroxisome proliferator-activated receptor gamma (PPARG), the main driver of the adipogenic program (PPARG), and its target genes. Here, we discuss how these findings, together with the recent literature, illuminate a functional role for ubiquitous TFs in lineage-determining transcriptional networks. PMID:25565413

  1. Lineage-affiliated transcription factors bind the Gata3 Tce1 enhancer to mediate lineage-specific programs

    PubMed Central

    Ohmura, Sakie; Mizuno, Seiya; Oishi, Hisashi; Ku, Chia-Jui; Hermann, Mary; Hosoya, Tomonori; Takahashi, Satoru; Engel, James Douglas

    2016-01-01

    The transcription factor GATA3 is essential for the genesis and maturation of the T cell lineage, and GATA3 dysregulation has pathological consequences. Previous studies have shown that GATA3 function in T cell development is regulated by multiple signaling pathways and that the Notch nuclear effector, RBP-J, binds specifically to the Gata3 promoter. We previously identified a T cell–specific Gata3 enhancer (Tce1) lying 280 kb downstream from the structural gene and demonstrated in transgenic mice that Tce1 promoted T lymphocyte–specific transcription of reporter genes throughout T cell development; however, it was not clear if Tce1 is required for Gata3 transcription in vivo. Here, we determined that the canonical Gata3 promoter is insufficient for Gata3 transcriptional activation in T cells in vivo, precluding the possibility that promoter binding by a host of previously implicated transcription factors alone is responsible for Gata3 expression in T cells. Instead, we demonstrated that multiple lineage-affiliated transcription factors bind to Tce1 and that this enhancer confers T lymphocyte–specific Gata3 activation in vivo, as targeted deletion of Tce1 in a mouse model abrogated critical functions of this T cell–regulatory element. Together, our data show that Tce1 is both necessary and sufficient for critical aspects of Gata3 T cell–specific transcriptional activity. PMID:26808502

  2. Lineage-affiliated transcription factors bind the Gata3 Tce1 enhancer to mediate lineage-specific programs.

    PubMed

    Ohmura, Sakie; Mizuno, Seiya; Oishi, Hisashi; Ku, Chia-Jui; Hermann, Mary; Hosoya, Tomonori; Takahashi, Satoru; Engel, James Douglas

    2016-03-01

    The transcription factor GATA3 is essential for the genesis and maturation of the T cell lineage, and GATA3 dysregulation has pathological consequences. Previous studies have shown that GATA3 function in T cell development is regulated by multiple signaling pathways and that the Notch nuclear effector, RBP-J, binds specifically to the Gata3 promoter. We previously identified a T cell-specific Gata3 enhancer (Tce1) lying 280 kb downstream from the structural gene and demonstrated in transgenic mice that Tce1 promoted T lymphocyte-specific transcription of reporter genes throughout T cell development; however, it was not clear if Tce1 is required for Gata3 transcription in vivo. Here, we determined that the canonical Gata3 promoter is insufficient for Gata3 transcriptional activation in T cells in vivo, precluding the possibility that promoter binding by a host of previously implicated transcription factors alone is responsible for Gata3 expression in T cells. Instead, we demonstrated that multiple lineage-affiliated transcription factors bind to Tce1 and that this enhancer confers T lymphocyte-specific Gata3 activation in vivo, as targeted deletion of Tce1 in a mouse model abrogated critical functions of this T cell-regulatory element. Together, our data show that Tce1 is both necessary and sufficient for critical aspects of Gata3 T cell-specific transcriptional activity. PMID:26808502

  3. Single-cell sequencing analysis characterizes common and cell-lineage-specific mutations in a muscle-invasive bladder cancer

    PubMed Central

    2012-01-01

    Background Cancers arise through an evolutionary process in which cell populations are subjected to selection; however, to date, the process of bladder cancer, which is one of the most common cancers in the world, remains unknown at a single-cell level. Results We carried out single-cell exome sequencing of 66 individual tumor cells from a muscle-invasive bladder transitional cell carcinoma (TCC). Analyses of the somatic mutant allele frequency spectrum and clonal structure revealed that the tumor cells were derived from a single ancestral cell, but that subsequent evolution occurred, leading to two distinct tumor cell subpopulations. By analyzing recurrently mutant genes in an additional cohort of 99 TCC tumors, we identified genes that might play roles in the maintenance of the ancestral clone and in the muscle-invasive capability of subclones of this bladder cancer, respectively. Conclusions This work provides a new approach of investigating the genetic details of bladder tumoral changes at the single-cell level and a new method for assessing bladder cancer evolution at a cell-population level. PMID:23587365

  4. Smad2 and Smad3 have differential sensitivity in relaying TGFβ signaling and inversely regulate early lineage specification

    PubMed Central

    Liu, Ling; Liu, Xu; Ren, Xudong; Tian, Yue; Chen, Zhenyu; Xu, Xiangjie; Du, Yanhua; Jiang, Cizhong; Fang, Yujiang; Liu, Zhongliang; Fan, Beibei; Zhang, Quanbin; Jin, Guohua; Yang, Xiao; Zhang, Xiaoqing

    2016-01-01

    The transforming growth factor beta (TGFβ) related signaling is one of the most important signaling pathways regulating early developmental events. Smad2 and Smad3 are structurally similar and it is mostly considered that they are equally important in mediating TGFβ signals. Here, we show that Smad3 is an insensitive TGFβ transducer as compared with Smad2. Smad3 preferentially localizes within the nucleus and is thus sequestered from membrane signaling. The ability of Smad3 in oligomerization with Smad4 upon agonist stimulation is also impaired given its unique linker region. Smad2 mediated TGFβ signaling plays a crucial role in epiblast development and patterning of three germ layers. However, signaling unrelated nuclear localized Smad3 is dispensable for TGFβ signaling-mediated epiblast specification, but important for early neural development, an event blocked by TGFβ/Smad2 signaling. Both Smad2 and Smad3 bind to the conserved Smads binding element (SBE), but they show nonoverlapped target gene binding specificity and differential transcriptional activity. We conclude that Smad2 and Smad3 possess differential sensitivities in relaying TGFβ signaling and have distinct roles in regulating early developmental events. PMID:26905010

  5. Smad2 and Smad3 have differential sensitivity in relaying TGFβ signaling and inversely regulate early lineage specification.

    PubMed

    Liu, Ling; Liu, Xu; Ren, Xudong; Tian, Yue; Chen, Zhenyu; Xu, Xiangjie; Du, Yanhua; Jiang, Cizhong; Fang, Yujiang; Liu, Zhongliang; Fan, Beibei; Zhang, Quanbin; Jin, Guohua; Yang, Xiao; Zhang, Xiaoqing

    2016-01-01

    The transforming growth factor beta (TGFβ) related signaling is one of the most important signaling pathways regulating early developmental events. Smad2 and Smad3 are structurally similar and it is mostly considered that they are equally important in mediating TGFβ signals. Here, we show that Smad3 is an insensitive TGFβ transducer as compared with Smad2. Smad3 preferentially localizes within the nucleus and is thus sequestered from membrane signaling. The ability of Smad3 in oligomerization with Smad4 upon agonist stimulation is also impaired given its unique linker region. Smad2 mediated TGFβ signaling plays a crucial role in epiblast development and patterning of three germ layers. However, signaling unrelated nuclear localized Smad3 is dispensable for TGFβ signaling-mediated epiblast specification, but important for early neural development, an event blocked by TGFβ/Smad2 signaling. Both Smad2 and Smad3 bind to the conserved Smads binding element (SBE), but they show nonoverlapped target gene binding specificity and differential transcriptional activity. We conclude that Smad2 and Smad3 possess differential sensitivities in relaying TGFβ signaling and have distinct roles in regulating early developmental events. PMID:26905010

  6. Lineage-Specific Responses of Tooth Shape in Murine Rodents (Murinae, Rodentia) to Late Miocene Dietary Change in the Siwaliks of Pakistan

    PubMed Central

    Kimura, Yuri; Jacobs, Louis L.; Flynn, Lawrence J.

    2013-01-01

    Past ecological responses of mammals to climate change are recognized in the fossil record by adaptive significance of morphological variations. To understand the role of dietary behavior on functional adaptations of dental morphology in rodent evolution, we examine evolutionary change of tooth shape in late Miocene Siwalik murine rodents, which experienced a dietary shift toward C4 diets during late Miocene ecological change indicated by carbon isotopic evidence. Geometric morphometric analysis in the outline of upper first molars captures dichotomous lineages of Siwalik murines, in agreement with phylogenetic hypotheses of previous studies (two distinct clades: the Karnimata and Progonomys clades), and indicates lineage-specific functional responses to mechanical properties of their diets. Tooth shapes of the two clades are similar at their sympatric origin but deviate from each other with decreasing overlap through time. Shape change in the Karnimata clade is associated with greater efficiency of propalinal chewing for tough diets than in the Progonomys clade. Larger body mass in Karnimata may be related to exploitation of lower-quality food items, such as grasses, than in smaller-bodied Progonomys. The functional and ecophysiological aspects of Karnimata exploiting C4 grasses are concordant with their isotopic dietary preference relative to Progonomys. Lineage-specific selection was differentially greater in Karnimata, and a faster rate of shape change toward derived Karnimata facilitated inclusion of C4 grasses in the diet. Sympatric speciation in these clades is most plausibly explained by interspecific competition on resource utilization between the two, based on comparisons of our results with the carbon isotope data. Interspecific competition with Karnimata may have suppressed morphological innovation of the Progonomys clade. Pairwise analyses of morphological and carbon isotope data can uncover ecological causes of sympatric speciation and define

  7. Lineage-Specific Evolution of the Complex Nup160 Hybrid Incompatibility Between Drosophila melanogaster and Its Sister Species

    PubMed Central

    Tang, Shanwu; Presgraves, Daven C.

    2015-01-01

    Two genes encoding protein components of the nuclear pore complex Nup160 and Nup96 cause lethality in F2-like hybrid genotypes between Drosophila simulans and Drosophila melanogaster. In particular, D. simulans Nup160 and Nup96 each cause inviability when hemizygous or homozygous in species hybrids that are also hemizygous (or homozygous) for the D. melanogaster X chromosome. The hybrid lethality of Nup160, however, is genetically complex, depending on one or more unknown additional factors in the autosomal background. Here we study the genetics and evolution of Nup160-mediated hybrid lethality in three ways. First, we test for variability in Nup160-mediated hybrid lethality within and among the three species of the D. simulans clade— D. simulans, D. sechellia, and D. mauritiana. We show that the hybrid lethality of Nup160 is fixed in D. simulans and D. sechellia but absent in D. mauritiana. Second, we explore how the hybrid lethality of Nup160 depends on other loci in the autosomal background. We find that D. simulans Nup160-mediated hybrid lethality does not depend on the presence of D. melanogaster Nup96, and we find that D. simulans and D. mauritiana are functionally differentiated at Nup160 as well as at other autosomal factor(s). Finally, we use population genetics data to show that Nup160 has experienced histories of recurrent positive selection both before and after the split of the three D. simulans clade species ∼240,000 years ago. Our genetic results suggest that a hybrid lethal Nup160 allele evolved before the split of the three D. simulans clade species, whereas the other autosomal factor(s) evolved more recently. PMID:26022241

  8. Downhill running and exercise in hot environments increase leukocyte Hsp72 (HSPA1A) and Hsp90α (HSPC1) gene transcripts.

    PubMed

    Tuttle, James A; Castle, Paul C; Metcalfe, Alan J; Midgley, Adrian W; Taylor, Lee; Lewis, Mark P

    2015-04-15

    Stressors within humans and other species activate Hsp72 and Hsp90α mRNA transcription, although it is unclear which environmental temperature or treadmill gradient induces the largest increase. To determine the optimal stressor for priming the Hsp system, physically active but not heat-acclimated participants (19.8 ± 1.9 and 20.9 ± 3.6 yr) exercised at lactate threshold in either temperate (20°C, 50% relative humidity; RH) or hot (30°C, 50% RH) environmental conditions. Within each condition, participants completed a flat running (temperate flat or hot flat) and a downhill running (temperate downhill or hot downhill) experimental trial in a randomized counterbalanced order separated by at least 7 days. Venous blood samples were taken immediately before (basal), immediately after exercise, and 3 and 24 h postexercise. RNA was extracted from leukocytes and RT-quantitative PCR conducted to determine Hsp72 and Hsp90α mRNA relative expression. Leukocyte Hsp72 mRNA was increased immediately after exercise following downhill running (1.9 ± 0.9-fold) compared with flat running (1.3 ± 0.4-fold; P = 0.001) and in hot (1.9 ± 0.6-fold) compared with temperate conditions (1.1 ± 0.5-fold; P = 0.003). Leukocyte Hsp90α mRNA increased immediately after exercise following downhill running (1.4 ± 0.8-fold) compared with flat running (0.9 ± 0.6-fold; P = 0.002) and in hot (1.6 ± 1.0-fold) compared with temperate conditions (0.9 ± 0.6-fold; P = 0.003). Downhill running and exercise in hot conditions induced the largest stimuli for leukocyte Hsp72 and Hsp90α mRNA increases. PMID:25722377

  9. Biomechanics of leukocyte rolling.

    PubMed

    Sundd, Prithu; Pospieszalska, Maria K; Cheung, Luthur Siu-Lun; Konstantopoulos, Konstantinos; Ley, Klaus

    2011-01-01

    Leukocyte rolling on endothelial cells and other P-selectin substrates is mediated by P-selectin binding to P-selectin glycoprotein ligand-1 expressed on the tips of leukocyte microvilli. Leukocyte rolling is a result of rapid, yet balanced formation and dissociation of selectin-ligand bonds in the presence of hydrodynamic shear forces. The hydrodynamic forces acting on the bonds may either increase (catch bonds) or decrease (slip bonds) their lifetimes. The force-dependent 'catch-slip' bond kinetics are explained using the 'two pathway model' for bond dissociation. Both the 'sliding-rebinding' and the 'allosteric' mechanisms attribute 'catch-slip' bond behavior to the force-induced conformational changes in the lectin-EGF domain hinge of selectins. Below a threshold shear stress, selectins cannot mediate rolling. This 'shear-threshold' phenomenon is a consequence of shear-enhanced tethering and catch bond-enhanced rolling. Quantitative dynamic footprinting microscopy has revealed that leukocytes rolling at venular shear stresses (>0.6 Pa) undergo cellular deformation (large footprint) and form long tethers. The hydrodynamic shear force and torque acting on the rolling cell are thought to be synergistically balanced by the forces acting on tethers and stressed microvilli, however, their relative contribution remains to be determined. Thus, improvement beyond the current understanding requires in silico models that can predict both cellular and microvillus deformation and experiments that allow measurement of forces acting on individual microvilli and tethers. PMID:21515934

  10. Substitution of Aspartate for glycine 1018 in the Type III procollagen (COL3AI) gene causes type IV Ehlers-Danlos Syndrome: The mutated allele is present in most blood leukocytes of the asymptomatic and mosaic mother

    SciTech Connect

    Kontusaari, S.; Tromp, G.; Kuivaniemi, H.; Prockop, D.J. ); Stolle, C. ); Pope, F.M.

    1992-09-01

    A proband with arterial ruptures and skin changes characteristic of the type IV variant of Ehlers-Danlos syndrome was found to have a single-base mutation in the type III procollagen gene, which converted the codon for glycine at amino position 1018 to a codon for aspartate. (Amino acid positions are numbered by the standard convention in which the first glycine of the triple-helical domain of an [alpha] chain is number 1. The numbers of positions in the [alpha]1(III) chains can be converted to positions in the human pro[alpha](III) chain by adding 167.). Nucleotide sequencing of overlapping PCR products in which the two alleles were distinguished demonstrated that the mutation of glycine 1018 was the only mutation that changed the primary structure of type III procollagen. The glycine substitution markedly decreased the amount of type III procollagen secreted into the medium by cultured skin fibroblasts from the proband. It is surprising that the same mutation was found in about 94% of the peripheral blood leukocytes from the proband's asymptomatic 72-year-old mother. Other tissues from the mother contained the mutated allele; it was present in 0%-100% of different samples of hair cells and in about 40% of cells from the oral epithelium. Therefore, the mother was a mosaic for the mutation. Since the mutated allele was present in cells derived from all three germ layers, the results indicated that the mutation arose by the late blastocyst stage of development. The results also indicate that assays of blood leukocytes do not always reveal mosaicism or predict phenotypic involvement of tissues, such as blood vessels, that are derived from the same embryonic cells as are leukocytes. 66 refs., 6 figs., 1 tab.

  11. Postnatal stem cell survival: does the niche, a rare harbor where to resist the ebb tide of differentiation, also provide lineage-specific instructions?

    PubMed

    Kindler, Vincent

    2005-10-01

    Postnatal stem cells regulate the homeostasis of the majority of our tissues. They continuously generate new progenitors and mature, functional cells to replace old cells, which cannot assume the tissue function anymore and are eliminated. Blood, skin, gut mucosa, muscle, cartilage, nerves, cornea, retina, liver, and many other structures are regulated by stem cells. As a result of their ability to produce large numbers of functionally mature cells, postnatal stem cells represent a promising tool for regenerative therapy. Indeed, unmanipulated stem cells or their progeny amplified in vitro are already used in some clinical applications to restore the function of injured or genetically deficient tissues. However, despite our cumulating understanding concerning postnatal stem cells, many aspects of their functionality remain unclear. For instance, in most tissues, we cannot reliably define the phenotype of the postnatal stem cells sustaining its survival. We do not know to which extent the environment surrounding the stem cell-the niche-which is a key actor insuring stem cell self-maintenance, is also implicated in the maintenance of stem cell lineage specificity. Moreover, we have to clarify whether postnatal stem cells are capable of undertaking "transdifferentiation", that is, the conversion of one cell type into another under physiological conditions. Answering these questions should help us to draw a more accurate picture of postnatal stem cell biology and should lead to the design of safe, effective therapies. PMID:16199730

  12. Towards a computational model of leukocyte adhesion cascade: Leukocyte rolling

    NASA Astrophysics Data System (ADS)

    Khismatullin, Damir

    2005-11-01

    Recruitment of leukocytes into sites of acute and chronic inflammation is a vital component of the innate immune response in humans and plays an important role in cardiovascular diseases, such as ischemia-reperfusion injury and atherosclerosis. Leukocytes extravasate into the inflamed tissue through a multi-step process called "leukocyte adhesion cascade", which involves initial contact of a leukocyte with activated endothelium (tethering), leukocyte rolling, firm adhesion, and transendothelial migration. Recently we developed a fully three-dimensional CFD model of receptor-mediated leukocyte adhesion to endothelium in a parallel-plate flow chamber. The model treats the leukocyte as a viscoelastic cell with the nucleus located in the intracellular space and cylindrical microvilli distributed over the cell membrane. Leukocyte-endothelial adhesion is assumed to be mediated by adhesion molecules expressed on the tips of cell microvilli and on endothelium. We show that the model can predict both shape changes and velocities of rolling leukocytes under physiological flow conditions. Results of this study also indicate that viscosity of the cytoplasm is a critical parameter of leukocyte adhesion, affecting the cell's ability to roll on endothelium. This work is supported by NIH Grant HL- 57446 and NCSA Grant BCS040006 and utilized the NCSA IBM p690.

  13. Gene therapy for canine leukocyte adhesion deficiency with lentiviral vectors using the murine stem cell virus and human phosphoglycerate kinase promoters.

    PubMed

    Hunter, Michael J; Zhao, Huifen; Tuschong, Laura M; Bauer, Thomas R; Burkholder, Tanya H; Persons, Derek A; Hickstein, Dennis D

    2011-06-01

    Children with leukocyte adhesion deficiency type 1 (LAD-1) and dogs with canine LAD (CLAD) develop life-threatening bacterial infections due to mutations in the leukocyte integrin CD18. Here, we compared the human phosphoglycerate kinase (hPGK) promoter to the murine stem cell virus (MSCV) promoter/enhancer in a self-inactivating HIV-1-derived lentiviral vector to treat animals with CLAD. Four CLAD dogs were infused with CD34(+) cells transduced with the hPGK vector, and two CLAD dogs received MSCV vector-transduced CD34(+) cells. Infusions were preceded by a nonmyeloablative dose of 200 cGy total body irradiation. Comparable numbers of transduced cells were infused in each group of animals. Only one of four CLAD animals treated with the hPGK-cCD18 vector had reversal of CLAD, whereas both MSCV-cCD18 vector-treated dogs had reversal of the phenotype. Correction of CLAD depends both upon the percentage of CD18(+) myeloid cells and the level of expression of CD18 on individual myeloid cells. In this regard, the hPGK promoter directed low levels of expression of CD18 on neutrophils compared to the MSCV promoter, likely contributing to the suboptimal clinical outcome with the hPGK vector. PMID:21275758

  14. Gene Therapy for Canine Leukocyte Adhesion Deficiency with Lentiviral Vectors Using the Murine Stem Cell Virus and Human Phosphoglycerate Kinase Promoters

    PubMed Central

    Zhao, Huifen; Tuschong, Laura M.; Bauer, Thomas R.; Burkholder, Tanya H.; Persons, Derek A.; Hickstein, Dennis D.

    2011-01-01

    Abstract Children with leukocyte adhesion deficiency type 1 (LAD-1) and dogs with canine LAD (CLAD) develop life-threatening bacterial infections due to mutations in the leukocyte integrin CD18. Here, we compared the human phosphoglycerate kinase (hPGK) promoter to the murine stem cell virus (MSCV) promoter/enhancer in a self-inactivating HIV-1–derived lentiviral vector to treat animals with CLAD. Four CLAD dogs were infused with CD34+ cells transduced with the hPGK vector, and two CLAD dogs received MSCV vector–transduced CD34+ cells. Infusions were preceded by a nonmyeloablative dose of 200 cGy total body irradiation. Comparable numbers of transduced cells were infused in each group of animals. Only one of four CLAD animals treated with the hPGK-cCD18 vector had reversal of CLAD, whereas both MSCV-cCD18 vector–treated dogs had reversal of the phenotype. Correction of CLAD depends both upon the percentage of CD18+ myeloid cells and the level of expression of CD18 on individual myeloid cells. In this regard, the hPGK promoter directed low levels of expression of CD18 on neutrophils compared to the MSCV promoter, likely contributing to the suboptimal clinical outcome with the hPGK vector. PMID:21275758

  15. Bovine leukocyte adhesion deficiency (BLAD): a review.

    PubMed

    Nagahata, Hajime

    2004-12-01

    Bovine leukocyte adhesion deficiency (BLAD) in Holstein cattle is an autosomal recessive congenital disease characterized by recurrent bacterial infections, delayed wound healing and stunted growth, and is also associated with persistent marked neutrophilia. The molecular basis of BLAD is a single point mutation (adenine to guanine) at position 383 of the CD18 gene, which caused an aspartic acid to glycine substitution at amino acid 128 (D128G) in the adhesion molecule CD18. Neutrophils from BLAD cattle have impaired expression of the beta2 integrin (CD11a,b,c/CD18) of the leukocyte adhesion molecule. Abnormalities in a wide spectrum of adherence dependent functions of leukocytes have been fully characterized. Cattle affected with BLAD have severe ulcers on oral mucous membranes, severe periodontitis, loss of teeth, chronic pneumonia and recurrent or chronic diarrhea. Affected cattle die at an early age due to the infectious complications. Holstein bulls, including carrier sires that had a mutant BLAD gene in heterozygote were controlled from dairy cattle for a decade. The control of BLAD in Holstein cattle by publishing the genotypes and avoiding the mating between BLAD carriers was found to be successful. This paper provides an overview of the genetic disease BLAD with reference to the disease in Holstein cattle. PMID:15644595

  16. Cryopreservation of Human Mucosal Leukocytes

    PubMed Central

    Shu, Zhiquan; Levy, Claire N.; Ferre, April L.; Hartig, Heather; Fang, Cifeng; Lentz, Gretchen; Fialkow, Michael; Kirby, Anna C.; Adams Waldorf, Kristina M.; Veazey, Ronald S.; Germann, Anja; von Briesen, Hagen; McElrath, M. Juliana; Dezzutti, Charlene S.; Sinclair, Elizabeth; Baker, Chris A. R.; Shacklett, Barbara L.; Gao, Dayong; Hladik, Florian

    2016-01-01

    Background Understanding how leukocytes in the cervicovaginal and colorectal mucosae respond to pathogens, and how medical interventions affect these responses, is important for developing better tools to prevent HIV and other sexually transmitted infections. An effective cryopreservation protocol for these cells following their isolation will make studying them more feasible. Methods and Findings To find an optimal cryopreservation protocol for mucosal mononuclear leukocytes, we compared cryopreservation media and procedures using human vaginal leukocytes and confirmed our results with endocervical and colorectal leukocytes. Specifically, we measured the recovery of viable vaginal T cells and macrophages after cryopreservation with different cryopreservation media and handling procedures. We found several cryopreservation media that led to recoveries above 75%. Limiting the number and volume of washes increased the fraction of cells recovered by 10–15%, possibly due to the small cell numbers in mucosal samples. We confirmed that our cryopreservation protocol also works well for both endocervical and colorectal leukocytes. Cryopreserved leukocytes had slightly increased cytokine responses to antigenic stimulation relative to the same cells tested fresh. Additionally, we tested whether it is better to cryopreserve endocervical cells on the cytobrush or in suspension. Conclusions Leukocytes from cervicovaginal and colorectal tissues can be cryopreserved with good recovery of functional, viable cells using several different cryopreservation media. The number and volume of washes has an experimentally meaningful effect on the percentage of cells recovered. We provide a detailed, step-by-step protocol with best practices for cryopreservation of mucosal leukocytes. PMID:27232996

  17. Superoxide production by phagocytic leukocytes.

    PubMed

    Drath, D B; Karnovsky, M L

    1975-01-01

    Mononuclear phagocytic leukocytes, as well as polymorphonuclear leukocytes, produce and release superoxide at rest, and this is stimulated by phagocytosis. Of the mouse monocytic cells studied, alveolar macrophages released the largest amounts of superoxide during phagocytosis, followed by normal peritoneal macrophages. Casein-elicited and "activated" macrophages released smaller quantities. In the guinea pig, polymorphonuclear leukocytes and casein-elicited macrophages were shown to release superoxide during phagocytosis whereas alveolar macrophages did not. Superoxide release accounted for only a small fraction of the respiratory burst of phagocytosis in all but the normal mouse peritoneal macrophage, the guinea pig polymorphonuclear leukocyte, and probably the mouse alveolar macrophage. There are obviously considerable species differences in O2-release by various leukocytes that might reflect both the production and/or destruction (e.g. by dismutase) of that substance. PMID:804030

  18. Leukocyte biophysics. An invited review.

    PubMed

    Schmid-Schönbein, G W

    1990-10-01

    The biophysical properties of leukocytes in the passive and active state are discussed. In the passive unstressed state, leukocytes are spherical with numerous membrane folds. Passive leukocytes exhibit viscoelastic properties, and the stress is carried largely by the cell cytoplasm and the nucleus. The membrane is highly deformable in shearing and bending, but resists area expansion. Membrane tension can usually be neglected but plays a role in cases of large deformation when the membrane becomes unfolded. The constant membrane area constraint is a determinant of phagocytic capacity, spreading of cells, and passage through narrow pores. In the active state, leukocytes undergo large internal cytoplasmic deformation, pseudopod projection, and granule redistribution. Several different measurements for assessment of biophysical properties and the internal cytoplasmic deformation in form of strain and strain rate tensors are presented. The current theoretical models for active cytoplasmic motion in leukocytes are discussed in terms of specific macromolecular reactions. PMID:1705479

  19. Human uterus myoma and gene expression profiling: A novel in vitro model for studying secretory leukocyte protease inhibitor-mediated tumor invasion.

    PubMed

    Mikami, Yoshikazu; Fukushima, Atsushi; Komiyama, Yusuke; Iwase, Takashi; Tsuda, Hiromasa; Higuchi, Yasuhiko; Hayakawa, Satoshi; Kuyama, Kayo; Komiyama, Kazuo

    2016-08-28

    Secretory leukocyte protease inhibitor (SLPI) is a serine protease inhibitor that diminishes tissue destruction during inflammation. A recent report revealed high levels of SLPI expression in the oral carcinoma cell. In addition, overexpression of SLPI up-regulates metastasis in lung carcinoma cells. On the other hand, matrix metalloproteinases (MMPs) are proteinases that participate in extracellular matrix degradation. SLPI and MMPs are involved as accelerators of the tumor invasion process; however, their exact roles are not fully understood. Understanding the mechanism of tumor invasion requires models that take the effect of microenvironmental factors into account. In one such in vitro model, different carcinoma cells have been shown to invade myoma tissue in highly distinct patterns. We have used this myoma model, as it provides a more natural stroma-like environment, to investigate the role of SLPI in tumor invasion. Our results indicate that the model provides a relevant matrix for tumor invasion studies, and that SLPI is important for the invasion of oral carcinoma Ca9-22 cells in conjunction with MMPs. Furthermore, using bioinformatics analysis, we have identified candidates as key molecules involved in SLPI-mediated tumor invasion. PMID:27238568

  20. Serum Leukocyte Immunoglobulin-Like Receptor A3 (LILRA3) Is Increased in Patients with Multiple Sclerosis and Is a Strong Independent Indicator of Disease Severity; 6.7kbp LILRA3 Gene Deletion Is Not Associated with Diseases Susceptibility

    PubMed Central

    An, Hongyan; Lim, Chai; Guillemin, Gilles J.; Vollmer-Conna, Ute; Rawlinson, William; Bryant, Katherine; Tedla, Nicodemus

    2016-01-01

    Leukocyte immunoglobulin-like receptor A3 (LILRA3) is a soluble immune regulatory molecule primarily expressed by monocytes and macrophages. A homozygous 6.7kbp LILRA3 gene deletion that removes the first seven of its eight exons is predicted to lead to lack of LILRA3 protein, although this has not been experimentally confirmed. Moreover, there are conflicting results with regards to the link between the LILRA3 homozygous genetic deletion and susceptibility to multiple sclerosis (MS) in different European populations. The aim of this study was to investigate whether LILRA3 gene deletion is associated with MS susceptibility in a North American cohort of European ancestry and assess if serum LILRA3 protein level is a marker of clinical subtype and/or disease severity in MS. A total of 456 patients with MS and 99 unrelated healthy controls were genotyped for the 6.7kbp LILRA3 gene deletion and levels of LILRA3 protein in sera determined by in-house sandwich ELISA. We showed that LILRA3 gene deletion was not associated with MS susceptibility and did not affect the age of disease onset, clinical subtype or disease severity. However, we discovered for the first time that homozygous LILRA3 gene deletion results in lack of production of LILRA3 protein. Importantly, LILRA3 protein level was significantly increased in sera of patients with MS when compared with control subjects, particularly in more severe type primary progressive MS. Multiple regression analysis showed that LILRA3 level in serum was one of the strongest independent markers of disease severity in MS, which potentially can be used as a diagnostic marker. PMID:26871720

  1. Isolation of Leukocytes from the Human Maternal-fetal Interface.

    PubMed

    Xu, Yi; Plazyo, Olesya; Romero, Roberto; Hassan, Sonia S; Gomez-Lopez, Nardhy

    2015-01-01

    Pregnancy is characterized by the infiltration of leukocytes in the reproductive tissues and at the maternal-fetal interface (decidua basalis and decidua parietalis). This interface is the anatomical site of contact between maternal and fetal tissues; therefore, it is an immunological site of action during pregnancy. Infiltrating leukocytes at the maternal-fetal interface play a central role in implantation, pregnancy maintenance, and timing of delivery. Therefore, phenotypic and functional characterizations of these leukocytes will provide insight into the mechanisms that lead to pregnancy disorders. Several protocols have been described in order to isolate infiltrating leukocytes from the decidua basalis and decidua parietalis; however, the lack of consistency in the reagents, enzymes, and times of incubation makes it difficult to compare these results. Described herein is a novel approach that combines the use of gentle mechanical and enzymatic dissociation techniques to preserve the viability and integrity of extracellular and intracellular markers in leukocytes isolated from the human tissues at the maternal-fetal interface. Aside from immunophenotyping, cell culture, and cell sorting, the future applications of this protocol are numerous and varied. Following this protocol, the isolated leukocytes can be used to determine DNA methylation, expression of target genes, in vitro leukocyte functionality (i.e., phagocytosis, cytotoxicity, T-cell proliferation, and plasticity, etc.), and the production of reactive oxygen species at the maternal-fetal interface. Additionally, using the described protocol, this laboratory has been able to describe new and rare leukocytes at the maternal-fetal interface. PMID:26067211

  2. Effect of the level of maternal energy intake prepartum on immunometabolic markers, polymorphonuclear leukocyte function, and neutrophil gene network expression in neonatal Holstein heifer calves.

    PubMed

    Osorio, J S; Trevisi, E; Ballou, M A; Bertoni, G; Drackley, J K; Loor, J J

    2013-06-01

    A conventional approach in dairy cow nutrition programs during late gestation is to feed moderate-energy diets. The effects of the maternal plane of nutrition on immune function and metabolism in newborn calves are largely unknown. Holstein cows (n=20) were fed a controlled-energy (CON) diet (1.24 Mcal/kg) for the entire dry period (~50 d) or the CON diet during the first 29 d of the dry period followed by a moderate-energy (OVE) diet (1.47 Mcal/kg) during the last 21 d prepartum. All calves were weighed at birth before first colostrum intake. Calves chosen for this study (n=6 per maternal diet) had blood samples harvested before colostrum feeding (d 0) and at 2 and 7 d of age. Blood samples were used to determine metabolites, acute-phase proteins, oxidative stress markers, hormones, phagocytic capacity of polymorphonuclear leukocytes (PMN) and monocytes, and total RNA was isolated from PMN. Calves from OVE dams weighed, on average, 5kg less at birth (44.0 vs. 48.6kg) than calves from CON dams. Blood glucose concentration in OVE calves had a more pronounced increase between 0 and 2 d than CON, at which point phagocytosis by PMN averaged 85% in OVE and 62% in CON. Compared with CON, calves from OVE had greater expression of TLR4, but lower expression of PPARA and PPARD at birth. Expression of PPARG and RXRA decreased between 0 and 2 d in both groups. Concentrations of leptin, cholesterol, ceruloplasmin, reactive oxygen metabolites, myeloperoxidase, retinol, tocopherol, IgG, and total protein, as well as expression of SOD2 and SELL increased markedly by 2 d in both groups; whereas, cortisol, albumin, acid-soluble protein, NEFA, insulin, as well as expression of IL6, TLR4, IL1R2, LTC4S, and ALOX5 decreased by 2 d. By 7 d of age, the concentration of haptoglobin was greater than precolostrum and was lower for OVE than CON calves. Our data provide evidence for a carry-over effect of maternal energy overfeeding during the last 3 wk before calving on some measurements of

  3. Novel Mutants Define Genes Required for the Expression of Human Histocompatibility Leukocyte Antigen DM: Evidence for Loci on Human Chromosome 6p

    PubMed Central

    Fling, Steven P.; Rak, Jennifer; Muczynski, Kimberly A.; Arp, Benjamin; Pious, Donald

    1997-01-01

    We and others have shown that the products of the HLA-DM locus are required for the intracellular assembly of major histocompatibility complex class II molecules with cognate peptides for antigen presentation. HLA-DM heterodimers mediate the dissociation of invariant chain (Ii)-derived class II–associated Ii peptides (CLIP) from class II molecules and facilitate the loading of class II molecules with antigenic peptides. Here we describe novel APC mutants with defects in the formation of class II–peptide complexes. These mutants express class II molecules which are conformationally altered, and an aberrantly high percentage of these class II molecules are associated with Ii-derived CLIP. This phenotype resembles that of DM null mutants. However, we show that the defects in two of these new mutants do not map to the DM locus. Nevertheless, our evidence suggests that the antigen processing defective phenotype in these mutants results from deficient DM expression. These mutants thus appear to define genes in which mutations have differential effects on the expression of conventional class II molecules and DM molecules. Our data are most consistent with these factors mapping to human chromosome 6p. Previous data have suggested that the expression of DM and class II genes are coordinately regulated. The results reported here suggest that DM and class II can also be differentially regulated, and that this differential regulation has significant effects on class II–restricted antigen processing. PMID:9348304

  4. Human Leukocyte Antigen (HLA) A*1101-restricted Epstein-Barr Virus-specific T-cell Receptor Gene Transfer to Target Nasopharyngeal Carcinoma

    PubMed Central

    Zheng, Yong; Parsonage, Greg; Zhuang, Xiaodong; Machado, Lee R; James, Christine H.; Salman, Asmaa; Searle, Peter F.; Hui, Edwin P.; Chan, Anthony T.C.; Lee, Steven P.

    2015-01-01

    Infusing virus-specific T cells is effective treatment for rare Epstein-Barr virus (EBV)-associated post-transplant lymphomas and more limited success has been reported using this approach to treat a far more common EBV-associated malignancy, nasopharyngeal carcinoma (NPC). However, current approaches using EBV-transformed lymphoblastoid cell lines to reactivate EBV-specific T cells for infusion take 2 to 3 months of in vitro culture and favour outgrowth of T cells targeting viral antigens expressed within EBV+ lymphomas but not in NPC. Here we explore T-cell receptor (TCR) gene transfer to rapidly and reliably generate T cells specific for the NPC-associated viral protein LMP2. We cloned a HLA A*1101-restricted TCR, which would be widely applicable since 40% of NPC patients carry this HLA allele. Studying both the wild-type and modified forms we have optimised expression of the TCR and demonstrated high avidity antigen-specific function (proliferation, cytotoxicity, cytokine release) in both CD8+ and CD4+ T cells. The engineered T cells also inhibited LMP2+ epithelial tumour growth in a mouse model. Furthermore, transduced T cells from patients with advanced NPC lysed LMP2-expressing NPC cell lines. Using this approach, within a few days large numbers of high avidity LMP2-specific T cells can be generated reliably to treat NPC, thus providing an ideal clinical setting to test TCR gene transfer without the risk of autoimmunity through targeting self-antigens. PMID:25711537

  5. Elemental composition of leukocyte subfractions

    NASA Astrophysics Data System (ADS)

    Admans, L. L.; Spyrou, N. M.

    2003-01-01

    The aim of this preliminary investigation was to determine the elemental concentration of various subfractions of leukocytes in a normal subject. Little work has been published on the elemental composition of these subfractions. First, a reliable technique for separation of these subfractions had to be established so that it could be applied to the determination of elemental concentrations in leukocyte subfractions from patients undergoing heart bypass surgery. Instrumental neutron activation analysis (INAA) utilising short irradiation and counting was the technique employed. Various washing media were examined during the separation of the leukocyte subfractions, for contamination of these small samples of peripheral blood mononuclear cells (PBMC) and polymorphonuclearcytes (PMN). Early results showed Mg and Se were present in these subfractions. Possibilities for further work are also discussed.

  6. A genome-scale in vivo loss-of-function screen identifies Phf6 as a lineage-specific regulator of leukemia cell growth

    PubMed Central

    Meacham, Corbin E.; Lawton, Lee N.; Soto-Feliciano, Yadira M.; Pritchard, Justin R.; Joughin, Brian A.; Ehrenberger, Tobias; Fenouille, Nina; Zuber, Johannes; Williams, Richard T.; Young, Richard A.

    2015-01-01

    We performed a genome-scale shRNA screen for modulators of B-cell leukemia progression in vivo. Results from this work revealed dramatic distinctions between the relative effects of shRNAs on the growth of tumor cells in culture versus in their native microenvironment. Specifically, we identified many “context-specific” regulators of leukemia development. These included the gene encoding the zinc finger protein Phf6. While inactivating mutations in PHF6 are commonly observed in human myeloid and T-cell malignancies, we found that Phf6 suppression in B-cell malignancies impairs tumor progression. Thus, Phf6 is a “lineage-specific” cancer gene that plays opposing roles in developmentally distinct hematopoietic malignancies. PMID:25737277

  7. Defects in the synthetic pathway prevent DIF-1 mediated stalk lineage specification cascade in the non-differentiating social amoeba, Acytostelium subglobosum.

    PubMed

    Mohri, Kurato; Hata, Takashi; Kikuchi, Haruhisa; Oshima, Yoshiteru; Urushihara, Hideko

    2014-01-01

    Separation of somatic cells from germ-line cells is a crucial event for multicellular organisms, but how this step was achieved during evolution remains elusive. In Dictyostelium discoideum and many other dictyostelid species, solitary amoebae gather and form a multicellular fruiting body in which germ-line spores and somatic stalk cells differentiate, whereas in Acytostelium subglobosum, acellular stalks form and all aggregated amoebae become spores. In this study, because most D. discoideum genes known to be required for stalk cell differentiation have homologs in A. subglobosum, we inferred functional variations in these genes and examined conservation of the stalk cell specification cascade of D. discoideum mediated by the polyketide differentiation-inducing factor-1 (DIF-1) in A. subglobosum. Through heterologous expression of A. subglobosum orthologs of DIF-1 biosynthesis genes in D. discoideum, we confirmed that two of the three genes were functional equivalents, while DIF-methyltransferase (As-dmtA) involved at the final step of DIF-1 synthesis was not. In fact, DIF-1 activity was undetectable in A. subglobosum lysates and amoebae of this species were not responsive to DIF-1, suggesting a lack of DIF-1 production in this species. On the other hand, the molecular function of an A. subglobosum ortholog of DIF-1 responsive transcription factor was equivalent with that of D. discoideum and inhibition of polyketide synthesis caused developmental arrest in A. subglobosum, which could not be rescued by DIF-1 addition. These results suggest that non-DIF-1 polyketide cascades involving downstream transcription factors are required for fruiting body development of A. subglobosum. PMID:24876391

  8. Selective Harvesting of Marginating-pulmonary Leukocytes.

    PubMed

    Shaashua, Lee; Sorski, Liat; Melamed, Rivka; Ben-Eliyahu, Shamgar

    2016-01-01

    Marginating-pulmonary (MP) leukocytes are leukocytes that adhere to the inner endothelium of the lung capillaries. MP-leukocytes were shown to exhibit unique composition and characteristics compared to leukocytes of other immune compartments. Evidence suggests higher cytotoxicity of natural killer cells, and a distinct pro- and anti-inflammatory profile of the MP-leukocyte population compared to circulating or splenic immunocytes. The method presented herein enables selective harvesting of MP-leukocytes by forced perfusion of the lungs in either mice or rats. In contrast to other methods used to extract lung-leukocytes, such as tissue grinding and biological degradation, this method exclusively yields leukocytes from the lung capillaries, uncontaminated with parenchymal, interstitial, and broncho-alveolar cells. In addition, the perfusion technique better preserves the integrity and the physiological milieu of MP-leukocytes, without inducing physiological responses due to tissue processing. This unique MP leukocyte population is strategically located to identify and react towards abnormal circulating cells, as all circulating malignant cells and infected cells are detained while passing through the lung capillaries, physically interacting with endothelial cells and resident leukocytes,. Thus, selective harvesting of MP-leukocytes and their study under various conditions may advance our understanding of their biological and clinical significance, specifically with respect to controlling circulating aberrant cells and lung-related diseases. PMID:27023665

  9. Lineage-Specific Effector Signatures of Invariant NKT Cells Are Shared amongst γδ T, Innate Lymphoid, and Th Cells.

    PubMed

    Lee, You Jeong; Starrett, Gabriel J; Lee, Seungeun Thera; Yang, Rendong; Henzler, Christine M; Jameson, Stephen C; Hogquist, Kristin A

    2016-08-15

    Invariant NKT cells differentiate into three predominant effector lineages in the steady state. To understand these lineages, we sorted undifferentiated invariant NK T progenitor cells and each effector population and analyzed their transcriptional profiles by RNAseq. Bioinformatic comparisons were made to effector subsets among other lymphocytes, specifically Th cells, innate lymphoid cells (ILC), and γδ T cells. Myc-associated signature genes were enriched in NKT progenitors, like in other hematopoietic progenitors. Only NKT1 cells, but not NKT2 and NKT17 cells, had transcriptome similarity to NK cells and were also similar to other IFN-γ-producing lineages such as Th1, ILC1, and intraepithelial γδ T cells. NKT2 and NKT17 cells were similar to their analogous subsets of γδ T cells and ILCs, but surprisingly, not to Th2 and Th17 cells. We identified a set of genes common to each effector lineage regardless of Ag receptor specificity, suggesting the use of conserved regulatory cores for effector function. PMID:27385777

  10. Effect of human polymorphonuclear and mononuclear leukocytes on chromosomal and plasmid DNA of Escherichia coli. Role of acid DNase

    SciTech Connect

    Rozenberg-Arska, M.; van Strijp, J.A.; Hoekstra, W.P.; Verhoef, J.

    1984-05-01

    Phagocytosis and killing by polymorphonuclear and mononuclear leukocytes are important host resistance factors against invading microorganisms. Evidence showing that killing is rapidly followed by degradation of bacterial components is limited. Therefore, we studied the fate of Escherichia coli DNA following phagocytosis of E. coli by polymorphonuclear and mononuclear leukocytes. (/sup 3/H)Thymidine-labeled, unencapsulated E. coli PC2166 and E. coli 048K1 were incubated in serum, washed, and added to leukocytes. Uptake and killing of the bacteria and degradation of DNA were measured. Although phagocytosis and killing by mononuclear leukocytes was less efficient than that by polymorphonuclear leukocytes, only mononuclear leukocytes were able to degrade E. coli PC2166 DNA. Within 2 h, 60% of the radioactivity added to mononuclear leukocytes was released into the supernate, of which 40% was acid soluble. DNA of E. coli 048K1 was not degraded. To further analyze the capacity of mononuclear leukocytes to degrade E. coli DNA, chromosomal and plasmid DNA was isolated from ingested bacteria and subjected to agarose gel-electrophoresis. Only chromosomal DNA was degraded after phagocytosis. Plasmid DNA of E. coli carrying a gene coding for ampicillin resistance remained intact for a 2-h period after ingestion, and was still able to transform recipient E. coli cells after this period. Although we observed no DNA degradation during phagocytosis by polymorphonuclear leukocytes, lysates of both polymorphonuclear and mononuclear leukocytes contained acid-DNase activity with a pH optimum of 4.9. However, the DNase activity of mononuclear leukocytes was 20 times higher than that of polymorphonuclear leukocytes. No difference was observed between DNase activity from polymorphonuclear and mononuclear leukocytes from a chronic granulomatous disease patient with DNase activity from control polymorphonuclear and mononuclear leukocytes.

  11. Leukocyte margination at arteriole shear rate

    PubMed Central

    Takeishi, Naoki; Imai, Yohsuke; Nakaaki, Keita; Yamaguchi, Takami; Ishikawa, Takuji

    2014-01-01

    Abstract We numerically investigated margination of leukocytes at arteriole shear rate in straight circular channels with diameters ranging from 10 to 22 μm. Our results demonstrated that passing motion of RBCs effectively induces leukocyte margination not only in small channels but also in large channels. A longer time is needed for margination to occur in a larger channel, but once a leukocyte has marginated, passing motion of RBCs occurs continuously independent of the channel diameter, and leukocyte margination is sustained for a long duration. We also show that leukocytes rarely approach the wall surface to within a microvillus length at arteriole shear rate. PMID:24907300

  12. A Model of Canine Leukocyte Telomere Dynamics

    PubMed Central

    Benetos, Athanase; Kimura, Masayuki; Labat, Carlos; Buchoff, Gerald M.; Huber, Shell; Labat, Laura; Lu, Xiaobin; Aviv, Abraham

    2011-01-01

    Summary Recent studies have found associations of leukocyte telomere length (TL) with diseases of aging and with longevity. However, it is unknown whether birth leukocyte TL or its age-dependent attrition— the two factors that determine leukocyte TL dynamics— explains these associations, since acquiring this information entails monitoring individuals over their entire life course. We tested in dogs a model of leukocyte TL dynamics, based on the following premises: (i) TL is synchronized among somatic tissues; (ii) TL in skeletal muscle, which is largely post-mitotic, is a measure of TL in early development; (iii) the difference between TL in leukocytes and muscle (ΔLMTL) is the extent of leukocyte TL shortening since early development. Using this model, we observed in 83 dogs (ages 4–42 months) no significant change with age in TLs of skeletal muscle and a shorter TL in leukocytes than in skeletal muscle (P<0.0001). Age explained 43% of the variation in ΔLMTL (P<0.00001) but only 6% of the variation in leukocyte TL (P=0.035) among dogs. Accordingly, muscle TL and ΔLMTL provide the two essential factors of leukocyte TL dynamics in the individual dog. When applied to humans, the partition of the contribution of leukocyte TL during early development versus telomere shortening afterward might provide information about whether the individual’s longevity is calibrated to either one or both factors that define leukocyte TL dynamics. PMID:21917112

  13. [Detection and typing for swine leukocyte antigen].

    PubMed

    Li, Hua; Luo, Huai-Rong; Zhang, Ya-Ping; Qiu, Xiang-Pin; Ye, Chun

    2004-03-01

    Traditionally the cluster of swine leukocyte antigen (SLA) was typed by serological, cytological and biochemical methods. Many special molecular typing methods have been developed with the progress of molecular biological technology, such as PCR-RFLP, PCR-SSCP , MS and DNA sequencing. Here we discussed the advantages and disadvantages of each method based on the polymorphic and conservative (from the functional aspect, such as supertype and supermotif) characteristics of SLA, and illustrated the development of typing for SLA in the future. In addition, we pointed out the editorial mistakes about the serological haplotype of SLA in reference book and emphasized that the accurate polymorphism of SLA-DQB gene must be based on the cloning sequencing. PMID:15639990

  14. Comparative Genomics of Carriage and Disease Isolates of Streptococcus pneumoniae Serotype 22F Reveals Lineage-Specific Divergence and Niche Adaptation.

    PubMed

    Cleary, David W; Devine, Vanessa T; Jefferies, Johanna M C; Webb, Jeremy S; Bentley, Stephen D; Gladstone, Rebecca A; Faust, Saul N; Clarke, Stuart C

    2016-01-01

    Streptococcus pneumoniae is a major cause of meningitis, sepsis, and pneumonia worldwide. Pneumococcal conjugate vaccines have been part of the United Kingdom's childhood immunization program since 2006 and have significantly reduced the incidence of disease due to vaccine efficacy in reducing carriage in the population. Here we isolated two clones of 22F (an emerging serotype of clinical concern, multilocus sequence types 433 and 698) and conducted comparative genomic analysis on four isolates, paired by Sequence Type (ST) with one of each pair being derived from carriage and the other disease (sepsis). The most compelling observation was of nonsynonymous mutations in pgdA, encoding peptidoglycan N-acetylglucosamine deacetylase A, which was found in the carriage isolates of both ST433 and 698. Deacetylation of pneumococcal peptidoglycan is known to enable resistance to lysozyme upon invasion. Althought no other clear genotypic signatures related to disease or carriage could be determined, additional intriguing comparisons between the two STs were possible. These include the presence of an intact prophage, in addition to numerous additional phage insertions, within the carriage isolate of ST433. Contrasting gene repertoires related to virulence and colonization, including bacteriocins, lantibiotics, and toxin--antitoxin systems, were also observed. PMID:27016484

  15. Comparative Genomics of Carriage and Disease Isolates of Streptococcus pneumoniae Serotype 22F Reveals Lineage-Specific Divergence and Niche Adaptation

    PubMed Central

    Cleary, David W.; Devine, Vanessa T.; Jefferies, Johanna M.C.; Webb, Jeremy S.; Bentley, Stephen D.; Gladstone, Rebecca A.; Faust, Saul N.; Clarke, Stuart C.

    2016-01-01

    Streptococcus pneumoniae is a major cause of meningitis, sepsis, and pneumonia worldwide. Pneumococcal conjugate vaccines have been part of the United Kingdom’s childhood immunization program since 2006 and have significantly reduced the incidence of disease due to vaccine efficacy in reducing carriage in the population. Here we isolated two clones of 22F (an emerging serotype of clinical concern, multilocus sequence types 433 and 698) and conducted comparative genomic analysis on four isolates, paired by Sequence Type (ST) with one of each pair being derived from carriage and the other disease (sepsis). The most compelling observation was of nonsynonymous mutations in pgdA, encoding peptidoglycan N-acetylglucosamine deacetylase A, which was found in the carriage isolates of both ST433 and 698. Deacetylation of pneumococcal peptidoglycan is known to enable resistance to lysozyme upon invasion. Althought no other clear genotypic signatures related to disease or carriage could be determined, additional intriguing comparisons between the two STs were possible. These include the presence of an intact prophage, in addition to numerous additional phage insertions, within the carriage isolate of ST433. Contrasting gene repertoires related to virulence and colonization, including bacteriocins, lantibiotics, and toxin-–antitoxin systems, were also observed. PMID:27016484

  16. A morphogenetic survey on ciliate plankton from a mountain lake pinpoints the necessity of lineage-specific barcode markers in microbial ecology

    PubMed Central

    Stoeck, Thorsten; Breiner, Hans-Werner; Filker, Sabine; Ostermaier, Veronika; Kammerlander, Barbara; Sonntag, Bettina

    2014-01-01

    Analyses of high-throughput environmental sequencing data have become the ‘gold-standard’ to address fundamental questions of microbial diversity, ecology and biogeography. Findings that emerged from sequencing are, e.g. the discovery of the extensive ‘rare microbial biosphere’ and its potential function as a seed-bank. Even though applied since several years, results from high-throughput environmental sequencing have hardly been validated. We assessed how well pyrosequenced amplicons [the hypervariable eukaryotic V4 region of the small subunit ribosomal RNA (SSU rRNA) gene] reflected morphotype ciliate plankton. Moreover, we assessed if amplicon sequencing had the potential to detect the annual ciliate plankton stock. In both cases, we identified significant quantitative and qualitative differences. Our study makes evident that taxon abundance distributions inferred from amplicon data are highly biased and do not mirror actual morphotype abundances at all. Potential reasons included cell losses after fixation, cryptic morphotypes, resting stages, insufficient sequence data availability of morphologically described species and the unsatisfying resolution of the V4 SSU rRNA fragment for accurate taxonomic assignments. The latter two underline the necessity of barcoding initiatives for eukaryotic microbes to better and fully exploit environmental amplicon data sets, which then will also allow studying the potential of seed-bank taxa as a buffer for environmental changes. PMID:23848238

  17. The Genome of Mycobacterium Africanum West African 2 Reveals a Lineage-Specific Locus and Genome Erosion Common to the M. tuberculosis Complex

    PubMed Central

    Bentley, Stephen D.; Comas, Iñaki; Bryant, Josephine M.; Walker, Danielle; Smith, Noel H.; Harris, Simon R.; Thurston, Scott; Gagneux, Sebastien; Wood, Jonathan; Antonio, Martin; Quail, Michael A.; Gehre, Florian; Adegbola, Richard A.; Parkhill, Julian; de Jong, Bouke C.

    2012-01-01

    Background M. africanum West African 2 constitutes an ancient lineage of the M. tuberculosis complex that commonly causes human tuberculosis in West Africa and has an attenuated phenotype relative to M. tuberculosis. Methodology/Principal Findings In search of candidate genes underlying these differences, the genome of M. africanum West African 2 was sequenced using classical capillary sequencing techniques. Our findings reveal a unique sequence, RD900, that was independently lost during the evolution of two important lineages within the complex: the “modern” M. tuberculosis group and the lineage leading to M. bovis. Closely related to M. bovis and other animal strains within the M. tuberculosis complex, M. africanum West African 2 shares an abundance of pseudogenes with M. bovis but also with M. africanum West African clade 1. Comparison with other strains of the M. tuberculosis complex revealed pseudogenes events in all the known lineages pointing toward ongoing genome erosion likely due to increased genetic drift and relaxed selection linked to serial transmission-bottlenecks and an intracellular lifestyle. Conclusions/Significance The genomic differences identified between M. africanum West African 2 and the other strains of the Mycobacterium tuberculosis complex may explain its attenuated phenotype, and pave the way for targeted experiments to elucidate the phenotypic characteristic of M. africanum. Moreover, availability of the whole genome data allows for verification of conservation of targets used for the next generation of diagnostics and vaccines, in order to ensure similar efficacy in West Africa. PMID:22389744

  18. The analysis of novel distal Cebpa enhancers and silencers using a transcriptional model reveals the complex regulatory logic of hematopoietic lineage specification.

    PubMed

    Bertolino, Eric; Reinitz, John; Manu

    2016-05-01

    C/EBPα plays an instructive role in the macrophage-neutrophil cell-fate decision and its expression is necessary for neutrophil development. How Cebpa itself is regulated in the myeloid lineage is not known. We decoded the cis-regulatory logic of Cebpa, and two other myeloid transcription factors, Egr1 and Egr2, using a combined experimental-computational approach. With a reporter design capable of detecting both distal enhancers and silencers, we analyzed 46 putative cis-regulatory modules (CRMs) in cells representing myeloid progenitors, and derived early macrophages or neutrophils. In addition to novel enhancers, this analysis revealed a surprisingly large number of silencers. We determined the regulatory roles of 15 potential transcriptional regulators by testing 32,768 alternative sequence-based transcriptional models against CRM activity data. This comprehensive analysis allowed us to infer the cis-regulatory logic for most of the CRMs. Silencer-mediated repression of Cebpa was found to be effected mainly by TFs expressed in non-myeloid lineages, highlighting a previously unappreciated contribution of long-distance silencing to hematopoietic lineage resolution. The repression of Cebpa by multiple factors expressed in alternative lineages suggests that hematopoietic genes are organized into densely interconnected repressive networks instead of hierarchies of mutually repressive pairs of pivotal TFs. More generally, our results demonstrate that de novo cis-regulatory dissection is feasible on a large scale with the aid of transcriptional modeling. Current address: Department of Biology, University of North Dakota, 10 Cornell Street, Stop 9019, Grand Forks, ND 58202-9019, USA. PMID:26945717

  19. Long-term outcome and lineage-specific chimerism in 194 patients with Wiskott-Aldrich syndrome treated by hematopoietic cell transplantation in the period 1980-2009: an international collaborative study.

    PubMed

    Moratto, Daniele; Giliani, Silvia; Bonfim, Carmem; Mazzolari, Evelina; Fischer, Alain; Ochs, Hans D; Cant, Andrew J; Thrasher, Adrian J; Cowan, Morton J; Albert, Michael H; Small, Trudy; Pai, Sung-Yun; Haddad, Elie; Lisa, Antonella; Hambleton, Sophie; Slatter, Mary; Cavazzana-Calvo, Marina; Mahlaoui, Nizar; Picard, Capucine; Torgerson, Troy R; Burroughs, Lauri; Koliski, Adriana; Neto, Jose Zanis; Porta, Fulvio; Qasim, Waseem; Veys, Paul; Kavanau, Kristina; Hönig, Manfred; Schulz, Ansgar; Friedrich, Wilhelm; Notarangelo, Luigi D

    2011-08-11

    In this retrospective collaborative study, we have analyzed long-term outcome and donor cell engraftment in 194 patients with Wiskott-Aldrich syndrome (WAS) who have been treated by hematopoietic cell transplantation (HCT) in the period 1980- 2009. Overall survival was 84.0% and was even higher (89.1% 5-year survival) for those who received HCT since the year 2000, reflecting recent improvement of outcome after transplantation from mismatched family donors and for patients who received HCT from an unrelated donor at older than 5 years. Patients who went to transplantation in better clinical conditions had a lower rate of post-HCT complications. Retrospective analysis of lineage-specific donor cell engraftment showed that stable full donor chimerism was attained by 72.3% of the patients who survived for at least 1 year after HCT. Mixed chimerism was associated with an increased risk of incomplete reconstitution of lymphocyte count and post-HCT autoimmunity, and myeloid donor cell chimerism < 50% was associated with persistent thrombocytopenia. These observations indicate continuous improvement of outcome after HCT for WAS and may have important implications for the development of novel protocols aiming to obtain full correction of the disease and reduce post-HCT complications. PMID:21659547

  20. Glycobiology of leukocyte trafficking in inflammation.

    PubMed

    Wright, Rachael D; Cooper, Dianne

    2014-12-01

    To fulfill their potential, leukocytes must be able to exit the vasculature and reach the site of inflammation within the tissue. This process of leukocyte extravasation is a tightly regulated sequence of events that is governed by a host of cell adhesion molecules, cytokines, chemokines and lipid mediators. Of major importance to this process and the function of many of the proteins and lipids involved is the posttranslational modification of these moieties by glycosylation. The glycosylation process is coordinated by multiple enzymes that add and remove saccharides to/from glycan structures on proteins and lipids, resulting in a unique molecular signature that affords specificity to the molecules involved in leukocyte recruitment. This review will discuss how glycosylation impacts the function of these key molecules involved in the recruitment of leukocytes during inflammation and the function of specific lectins (carbohydrate-binding proteins) that have a role in leukocyte trafficking. PMID:25258391

  1. Disseminated Intravascular Coagulation Induced with Leukocyte Procoagulant

    PubMed Central

    Kociba, Gary J.; Griesemer, Richard A.

    1972-01-01

    The procoagulant activity of rabbit peritoneal leukocytes significantly increased when the leukocytes were incubated in suspension cultures at 37 C for 24 hours. Intravenous infusions of Iysates of 232 × 106 rabbit leukocytes which had been incubated in cultures at 37 C for 24 hours produced disseminated intravascular coagulation and vasculitis involving the pulmonary arteries in normal rabbits. Intraaortic infusions of lysates of 230 × 106 similarly incubated leukocytes produced renal thrombosis and renal cortical necrosis in normal rabbits. These observations suggest that the procoagulant of granulocytic leukocytes could play a role in the generalized Shwartzman reaction and other syndromes of disseminated intravascular coagulation. ImagesFig 3Fig 4Fig 5Fig 6Fig 7Fig 1Fig 2 PMID:5086898

  2. Leukocyte rheology in recent stroke.

    PubMed

    Ernst, E; Matrai, A; Paulsen, F

    1987-01-01

    Eighteen patients with recent ischemic stroke were compared with an equal number of matched controls. Standardized suspensions of red cells as well as of red and white cells were filtered in a new filtration apparatus capable of discriminating between cell deformability and filter occlusion. Results show that red cell deformability, although slightly lower than in controls, is not significantly altered in stroke patients. Filter occlusion, however, was significantly higher in patients when red and white cell suspensions were filtered, but not when red cell suspensions were used, suggesting that white cell filterability is impaired after stroke, which could be due to decreased deformability and/or increased adhesiveness of leukocytes. Slowed white cell passage may also occur in the living microcirculation and may present an obstacle to nutritive flow in exchange vessels, possibly contributing to local ischemia and tissue necrosis after stroke. PMID:3810770

  3. Longitudinal evaluation of leukocyte transcripts in killer whales (Orcinus Orca)

    USGS Publications Warehouse

    Sitt, Tatjana; Bowen, Lizabeth; Lee, Chia-Shan; Blanchard, Myra; McBain, James; Dold, Christopher; Stott, Jeffrey L.

    2016-01-01

    Early identification of illness and/or presence of environmental and/or social stressors in free-ranging and domestic cetaceans is a priority for marine mammal health care professionals. Incorporation of leukocyte gene transcript analysis into the diagnostic tool kit has the potential to augment classical diagnostics based upon ease of sample storage and shipment, inducible nature and well-defined roles of transcription and associated downstream actions. Development of biomarkers that could serve to identify “insults” and potentially differentiate disease etiology would be of great diagnostic value. To this end, a modest number of peripheral blood leukocyte gene transcripts were selected for application to a domestic killer whale population with a focus on broad representation of inducible immunologically relevant genes. Normalized leukocyte transcript values, longitudinally acquired from 232 blood samples derived from 26 clinically healthy whales, were not visibly influenced temporally nor by sex or the specific Park in which they resided. Stability in leukocyte transcript number during periods of health enhances their potential use in diagnostics through identification of outliers. Transcript levels of two cytokine genes, IL-4 and IL-17, were highly variable within the group as compared to the other transcripts. IL-4 transcripts were typically absent. Analysis of transcript levels on the other genes of interest, on an individual animal basis, identified more outliers than were visible when analyzed in the context of the entire population. The majority of outliers (9 samples) were low, though elevated transcripts were identified for IL-17 from 2 animals and one each for Cox-2 and IL-10. The low number of outliers was not unexpected as sample selection was intentionally directed towards animals that were clinically healthy at the time of collection. Outliers may reflect animals experiencing subclinical disease that is transient and self-limiting. The

  4. Longitudinal evaluation of leukocyte transcripts in killer whales (Orcinus Orca).

    PubMed

    Sitt, Tatjana; Bowen, Lizabeth; Lee, Chia-Shan; Blanchard, Myra T; McBain, James; Dold, Christopher; Stott, Jeffrey L

    2016-07-01

    Early identification of illness and/or presence of environmental and/or social stressors in free-ranging and domestic cetaceans is a priority for marine mammal health care professionals. Incorporation of leukocyte gene transcript analysis into the diagnostic tool kit has the potential to augment classical diagnostics based upon ease of sample storage and shipment, inducible nature and well-defined roles of transcription and associated downstream actions. Development of biomarkers that could serve to identify "insults" and potentially differentiate disease etiology would be of great diagnostic value. To this end, a modest number of peripheral blood leukocyte gene transcripts were selected for application to a domestic killer whale population with a focus on broad representation of inducible immunologically relevant genes. Normalized leukocyte transcript values, longitudinally acquired from 232 blood samples derived from 26 clinically healthy whales, were not visibly influenced temporally nor by sex or the specific Park in which they resided. Stability in leukocyte transcript number during periods of health enhances their potential use in diagnostics through identification of outliers. Transcript levels of two cytokine genes, IL-4 and IL-17, were highly variable within the group as compared to the other transcripts. IL-4 transcripts were typically absent. Analysis of transcript levels on the other genes of interest, on an individual animal basis, identified more outliers than were visible when analyzed in the context of the entire population. The majority of outliers (9 samples) were low, though elevated transcripts were identified for IL-17 from 2 animals and one each for Cox-2 and IL-10. The low number of outliers was not unexpected as sample selection was intentionally directed towards animals that were clinically healthy at the time of collection. Outliers may reflect animals experiencing subclinical disease that is transient and self-limiting. The immunologic

  5. Leukocyte Activation in Obese Patients

    PubMed Central

    Minervino, Daniele; Gumiero, Daniela; Nicolazzi, Maria Anna; Carnicelli, Annamaria; Fuorlo, Mariella; Guidone, Caterina; Di Gennaro, Leonardo; Fattorossi, Andrea; Mingrone, Geltrude; Landolfi, Raffaele

    2015-01-01

    Abstract The rising prevalence of obesity is a major global health problem. In severe obesity, bariatric surgery (BS) allows to obtain a significant weight loss and comorbidities improvement, among them one of the factors is the thrombotic risk. In this observational study, we measured indices of leukocyte activation in severely obese patients as markers of increased thrombotic risk in relation with serum markers of inflammation before and after BS. Frequency of polymorphonuclear neutrophil-platelet (PLT) and monocyte (MONO)-PLT aggregates as well as of tissue factor (TF) expressing MONOs was measured in the peripheral blood of 58 consecutive obese patients and 30 healthy controls. In 31 of the 58 obese patients, data obtained at the enrollment were compared with those obtained at 3, 6, and 12 months after BS. Compared with healthy controls, obese patients showed a higher frequency of polymorphonuclear leukocyte (PMNL)-PLT aggregates (7.47 ± 2.45 [6.82–8.11]% vs 5.85 ± 1.89 [5.14–6.55]%, P = 0.001), MONO-PLT aggregates (12.31 ± 7.33 [10.38–14.24]% vs 8.14 ± 2.22 [7.31–8.97]%, P < 0.001), and TF expressing MONOs (4.01 ± 2.11 [3.45–4.56]% vs 2.64 ± 1.65 [2.02–3.25]%, P = 0.002). PMNL-PLT and MONO-PLT aggregate frequency was positively correlated with TF expressing MONOs (R2 = 0.260, P = 0.049 and R2 = 0.318, P = 0.015, respectively). BS was performed in 31 patients and induced a significant reduction of the body mass index, and waist and hip circumferences. These effects were associated with a significant decrease of PMNL-PLT aggregates at 12 months (7.58 ± 2.27 [6.75–8.42]% vs 4.47 ± 1.11 [3.93–5.01]%, P < 0.001), and a reduction of TF expressing MONOs at 6 (3.82 ± 2.04 [3.07–4.57]% vs 1.60 ± 1.69 [0.30–2.90]%, P = 0.008) and 12 months (3.82 ± 2.04 [3.07–4.57]% vs 1.71 ± 0.54 [1.45–1.97]%, P = 0.001) after BS. These data suggest that leukocyte

  6. Highly specific blockade of CCR5 inhibits leukocyte trafficking and reduces mucosal inflammation in murine colitis

    PubMed Central

    Mencarelli, Andrea; Cipriani, Sabrina; Francisci, Daniela; Santucci, Luca; Baldelli, Franco; Distrutti, Eleonora; Fiorucci, Stefano

    2016-01-01

    Targeted disruption of leukocyte trafficking to the gut represents a promising approach for the treatment of inflammatory bowel diseases (IBDs). CCR5, the shared receptor for MIP1α and β and RANTES, is expressed by multiple leukocytes. Here, we aimed to determine the role of CCR5 in mediating leukocyte trafficking in models of colitis, and evaluate the therapeutic potential of maraviroc, an orally active CCR5 antagonist used in the treatment of CCR5-tropic HIV. Acute and chronic colitis were induced by administration of DSS or TNBS to wild-type and CCR5−/− mice or adoptive transfer of splenic naïve CD4+ T-cells from wild type or CCR5−/− mice into RAG-1−/−. CCR5 gene ablation reduced the mucosal recruitment and activation of CCR5-bearing CD4+ and CD11b+ leukocytes, resulting in profound attenuation of signs and symptoms of inflammation in the TNBS and transfer models of colitis. In the DSS/TNBS colitis and in the transfer model, maraviroc attenuated development of intestinal inflammation by selectively reducing the recruitment of CCR5 bearing leukocytes. In summary, CCR5 regulates recruitment of blood leukocytes into the colon indicating that targeting CCR5 may offer therapeutic options in IBDs. PMID:27492684

  7. Highly specific blockade of CCR5 inhibits leukocyte trafficking and reduces mucosal inflammation in murine colitis.

    PubMed

    Mencarelli, Andrea; Cipriani, Sabrina; Francisci, Daniela; Santucci, Luca; Baldelli, Franco; Distrutti, Eleonora; Fiorucci, Stefano

    2016-01-01

    Targeted disruption of leukocyte trafficking to the gut represents a promising approach for the treatment of inflammatory bowel diseases (IBDs). CCR5, the shared receptor for MIP1α and β and RANTES, is expressed by multiple leukocytes. Here, we aimed to determine the role of CCR5 in mediating leukocyte trafficking in models of colitis, and evaluate the therapeutic potential of maraviroc, an orally active CCR5 antagonist used in the treatment of CCR5-tropic HIV. Acute and chronic colitis were induced by administration of DSS or TNBS to wild-type and CCR5(-/-) mice or adoptive transfer of splenic naïve CD4(+) T-cells from wild type or CCR5(-/-) mice into RAG-1(-/-). CCR5 gene ablation reduced the mucosal recruitment and activation of CCR5-bearing CD4(+) and CD11b(+) leukocytes, resulting in profound attenuation of signs and symptoms of inflammation in the TNBS and transfer models of colitis. In the DSS/TNBS colitis and in the transfer model, maraviroc attenuated development of intestinal inflammation by selectively reducing the recruitment of CCR5 bearing leukocytes. In summary, CCR5 regulates recruitment of blood leukocytes into the colon indicating that targeting CCR5 may offer therapeutic options in IBDs. PMID:27492684

  8. 21 CFR 864.7660 - Leukocyte alkaline phosphatase test.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Leukocyte alkaline phosphatase test. 864.7660... Leukocyte alkaline phosphatase test. (a) Identification. A leukocyte alkaline phosphatase test is a device used to identify the enzyme leukocyte alkaline phosphatase in neutrophilic granulocytes...

  9. 21 CFR 864.7660 - Leukocyte alkaline phosphatase test.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Leukocyte alkaline phosphatase test. 864.7660... Leukocyte alkaline phosphatase test. (a) Identification. A leukocyte alkaline phosphatase test is a device used to identify the enzyme leukocyte alkaline phosphatase in neutrophilic granulocytes...

  10. Indium-111 autologous leukocyte imaging in pancreatitis

    SciTech Connect

    Anderson, J.R.; Spence, R.A.; Laird, J.D.; Ferguson, W.R.; Kennedy, T.L.

    1986-03-01

    Thirty-nine patients with acute pancreatitis have been assessed using a prognostic factor grading system, abdominal ultrasound, and autologous leukocyte imaging. Both prognostic factor grading and leukocyte imaging can accurately assess the severity of the disease early in its course. All patients with a negative indium-labeled leukocyte image recovered without sequelae, whereas five of the 12 patients with a positive image developed complications, including two deaths. Abdominal ultrasound is of no value in assessing severity, but is a useful method of detecting those patients with gallstone-associated disease. In patients with suspected abscess formation following acute pancreatitis, indium leukocyte imaging does not differentiate between fat necrosis and abscess formation. In this situation, computerized tomography should be carried out before laparotomy is undertaken.

  11. Coagulant Activity of Leukocytes. TISSUE FACTOR ACTIVITY

    PubMed Central

    Niemetz, J.

    1972-01-01

    Peritoneal leukocytes harvested from rabbits which have received two spaced doses of endotoxin have significantly greater (10-fold) coagulant activity than leukocytes from control rabbits. The coagulant activity accelerates the clotting of normal plasma and activates factor X in the presence of factor VII and calcium and is therefore regarded as tissue factor. A total of 40-80 mg tissue factor activity was obtained from the peritoneal cavity of single endotoxin-treated rabbits. In leukocyte subcellular fractions, separated by centrifugation, the specific tissue factor activity sedimented mainly at 14,500 g and above. The procoagulant activity was destroyed after heating for 10 min at 65°C but was preserved at lower temperatures. Polymyxin B, when given with the first dose of endotoxin, reduced both the number of peritoneal leukocytes and their tissue factor activity by two-thirds. When given immediately before the second dose of endotoxin, polymyxin B had no inhibitory effect. PMID:4333021

  12. Leukocyte filtration in lung transplantation.

    PubMed

    Kurusz, Mark; Roach, John D; Vertrees, Roger A; Girouard, Mark K; Lick, Scott D

    2002-05-01

    Controlled reperfusion of the transplanted lung has been used in nine consecutive patients to decrease manifestations of lung reperfusion injury. An extracorporeal circuit containing a roller pump, heat exchanger and leukodepleting filter is primed with substrate-enhanced reperfusion solution mixed with approximately 2000 ml of the patient's blood. This solution is slowly recirculated to remove leukocytes prior to reperfusion. When the pulmonary anastomoses are completed, the pulmonary artery is cannulated through the untied anastomosis using a catheter containing a pressure lumen for measurement of infusion pressure. An atrial clamp is left in place on the patient's native atrial cuff to decrease the risk of systemic air embolism during the brief period of reperfusion from the extracorporeal reservoir. During reperfusion, the water bath to the heat exchanger is kept at 35 degrees C and the flow rate for reperfusion solution is between 150 and 200 m/min, keeping the pulmonary artery pressure <14 mmHg. Eight of nine patients were ventilated on 40% inspired oxygen within a few hours of operation and 7/9 were extubated on or before postoperative day 1. Six of nine patients are long-term survivors. PMID:12009087

  13. Myeloid differentiation architecture of leukocyte transcriptome dynamics in perceived social isolation

    PubMed Central

    Cole, Steven W.; Capitanio, John P.; Chun, Katie; Arevalo, Jesusa M. G.; Ma, Jeffrey; Cacioppo, John T.

    2015-01-01

    To define the cellular mechanisms of up-regulated inflammatory gene expression and down-regulated antiviral response in people experiencing perceived social isolation (loneliness), we conducted integrative analyses of leukocyte gene regulation in humans and rhesus macaques. Five longitudinal leukocyte transcriptome surveys in 141 older adults showed up-regulation of the sympathetic nervous system (SNS), monocyte population expansion, and up-regulation of the leukocyte conserved transcriptional response to adversity (CTRA). Mechanistic analyses in a macaque model of perceived social isolation confirmed CTRA activation and identified selective up-regulation of the CD14++/CD16− classical monocyte transcriptome, functional glucocorticoid desensitization, down-regulation of Type I and II interferons, and impaired response to infection by simian immunodeficiency virus (SIV). These analyses identify neuroendocrine-related alterations in myeloid cell population dynamics as a key mediator of CTRA transcriptome skewing, which may both propagate perceived social isolation and contribute to its associated health risks. PMID:26598672

  14. Myeloid differentiation architecture of leukocyte transcriptome dynamics in perceived social isolation.

    PubMed

    Cole, Steven W; Capitanio, John P; Chun, Katie; Arevalo, Jesusa M G; Ma, Jeffrey; Cacioppo, John T

    2015-12-01

    To define the cellular mechanisms of up-regulated inflammatory gene expression and down-regulated antiviral response in people experiencing perceived social isolation (loneliness), we conducted integrative analyses of leukocyte gene regulation in humans and rhesus macaques. Five longitudinal leukocyte transcriptome surveys in 141 older adults showed up-regulation of the sympathetic nervous system (SNS), monocyte population expansion, and up-regulation of the leukocyte conserved transcriptional response to adversity (CTRA). Mechanistic analyses in a macaque model of perceived social isolation confirmed CTRA activation and identified selective up-regulation of the CD14(++)/CD16(-) classical monocyte transcriptome, functional glucocorticoid desensitization, down-regulation of Type I and II interferons, and impaired response to infection by simian immunodeficiency virus (SIV). These analyses identify neuroendocrine-related alterations in myeloid cell population dynamics as a key mediator of CTRA transcriptome skewing, which may both propagate perceived social isolation and contribute to its associated health risks. PMID:26598672

  15. The multiple faces of leukocyte interstitial migration

    PubMed Central

    Lämmermann, Tim; Germain, Ronald N.

    2014-01-01

    Spatiotemporal control of leukocyte dynamics within tissues is critical for successful innate and adaptive immune responses. Homeostatic trafficking and coordinated infiltration into and within sites of inflammation and infection rely on signaling in response to extracellular cues that in turn controls a variety of intracellular protein networks regulating leukocyte motility, migration, chemotaxis, positioning, and cell–cell interaction. In contrast to mesenchymal cells, leukocytes migrate in an amoeboid fashion by rapid cycles of actin polymerization and actomyosin contraction, and their migration in tissues is generally referred to as low adhesive and nonproteolytic. The interplay of actin network expansion, contraction, and adhesion shapes the exact mode of amoeboid migration, and in this review, we explore how leukocyte subsets potentially harness the same basic biomechanical mechanisms in a cell-type-specific manner. Most of our detailed understanding of these processes derives from in vitro migration studies in three-dimensional gels and confined spaces that mimic geometrical aspects of physiological tissues. We summarize these in vitro results and then critically compare them to data from intravital imaging of leukocyte interstitial migration in mouse tissues. We outline the technical challenges of obtaining conclusive mechanistic results from intravital studies, discuss leukocyte migration strategies in vivo, and present examples of mode switching during physiological interstitial migration. These findings are also placed in the context of leukocyte migration defects in primary immunodeficiencies. This overview of both in vitro and in vivo studies highlights recent progress in understanding the molecular and biophysical mechanisms that shape robust leukocyte migration responses in physiologically complex and heterogeneous environments. PMID:24573488

  16. Leukocyte chemoattractant activity of diacylglycerol

    SciTech Connect

    Wright, T.M.; Hoffman, R.D.; Nishijima, J.; Shin, H.S.

    1986-03-05

    Phosphatidylinositol breakdown with the generation of 1,2-diacylglycerol (1,2-DG) and inositol phosphates occurs in response to receptor mediated stimulation of lymphocytes and polymorphonuclear neutrophils (PMN). In the authors attempt to demonstrate the direct role of 1,2-DG in cell migration, they have found 1,2 dioctanoyl glycerol (1,2-C8DG) to be a chemoattractant for 6C3HED, a mouse thymic lymphoma, and human peripheral blood PMN's. The chemoattractant activity for both cell types was observed at concentrations from 0.5 to 10mM in an under agarose assay. The maximum effect of 1,2-C8DG on 6C3HED cells was similar to that of 1mM lysophosphatidylcholine and the maximum effect of 1,2-C8DG on PMN's was similar to that of 10/sup -7/M f-met-leu-phe. Other 1,2-DG's with acyl chains ranging from 6 to 18 carbons in length and 1-oleoyl-2-acetyl-glycerol were also chemoattractants for 6C3HED, although their activities were less than 1,2-C8DG. In addition, phorbol myristate acetate (PMA), another activator of protein kinase C, was a chemoattractant for 6C3HED and human PMN's. PMA was more potent than 1,2-C8DG for both 6C3HED and PMN's with chemoattractant activity in the range of 30nM to 1..mu..M. These studies support the direct role of 1,2-DG in the transduction of chemotactic stimuli in leukocytes and further suggest that the formation of diacylglycerol represents a common step in the migratory responses of lymphoid and myeloid cells.

  17. Leukocyte Recruitment and Ischemic Brain Injury

    PubMed Central

    Yilmaz, Gokhan

    2010-01-01

    Leukocytes are recruited into the cerebral microcirculation following an ischemic insult. The leukocyte–endothelial cell adhesion manifested within a few hours after ischemia (followed by reperfusion, I/R) largely reflects an infiltration of neutrophils, while other leukocyte populations appear to dominate the adhesive interactions with the vessel wall at 24 h of reperfusion. The influx of rolling and adherent leukocytes is accompanied by the recruitment of adherent platelets, which likely enhances the cytotoxic potential of the leukocytes to which they are attached. The recruitment of leukocytes and platelets in the postischemic brain is mediated by specific adhesion glycoproteins expressed by the activated blood cells and on cerebral microvascular endothelial cells. This process is also modulated by different signaling pathways (e.g., CD40/CD40L, Notch) and cytokines (e.g., RANTES) that are activated/released following I/R. Some of the known risk factors for cardiovascular disease, including hypercholesterolemia and obesity appear to exacerbate the leukocyte and platelet recruitment elicited by brain I/R. Although lymphocyte–endothelial cell and –platelet interactions in the postischemic cerebral microcirculation have not been evaluated to date, recent evidence in experimental animals implicate both CD4+ and CD8+ T-lymphocytes in the cerebral microvascular dysfunction, inflammation, and tissue injury associated with brain I/R. Evidence implicating regulatory T-cells as cerebroprotective modulators of the inflammatory and tissue injury responses to brain I/R support a continued focus on leukocytes as a target for therapeutic intervention in ischemic stroke. PMID:19579016

  18. Halloysite Nanotube Coatings Suppress Leukocyte Spreading.

    PubMed

    Hughes, Andrew D; Marsh, Graham; Waugh, Richard E; Foster, David G; King, Michael R

    2015-12-22

    The nanoscale topography of adhesive surfaces is known to be an important factor governing cellular behavior. Previous work has shown that surface coatings composed of halloysite nanotubes enhance the adhesion, and therefore capture of, rare target cells such as circulating tumor cells. Here we demonstrate a unique feature of these coatings in their ability to reduce the adhesion of leukocytes and prevent leukocyte spreading. Surfaces were prepared with coatings of halloysite nanotubes and functionalized for leukocyte adhesion with E-selectin, and the dilution of nanotube concentration revealed a threshold concentration below which cell spreading became comparable to smooth surfaces. Evaluation of surface roughness characteristics determined that the average distance between discrete surface features correlated with adhesion metrics, with a separation distance of ∼2 μm identified as the critical threshold. Computational modeling of the interaction of leukocytes with halloysite nanotube-coated surfaces of varying concentrations demonstrates that the geometry of the cell surface and adhesive counter-surface produces a significantly diminished effective contact area compared to a leukocyte interacting with a smooth surface. PMID:26605493

  19. Osteomyelitis: diagnosis with In-111-labeled leukocytes

    SciTech Connect

    Schauwecker, D.S.

    1989-04-01

    In a retrospective review, 485 patients with suspected osteomyelitis were studied. Of these, 453 patients were studied with both bone and indium-111 leukocyte scanning (173 sequentially and 280 simultaneously). The ability to determine that the infection was in bone rather than in adjacent soft tissue was greater with simultaneous bone scan and In-111 leukocyte studies than with sequential studies. The locations of suspected osteomyelitis were divided into central (containing active bone marrow), peripheral (hands and feet), and middle (between central and peripheral). Specificity remained high (about 90%) regardless of the location. Overall sensitivity was significantly lower in the central location than in the peripheral or middle location. Determination of whether the In-111 leukocyte activity was in bone or adjacent soft tissue was also more difficult when the infection was in the central location. For acute osteomyelitis, sensitivity was high regardless of the location. For chronic osteomyelitis, sensitivity was lower in the central location.

  20. Leukocyte migration inhibition in nickel dermatitis.

    PubMed

    Mirza, A M; Perera, M G; Maccia, C A; Dziubynskyj, O G; Bernstein, I L

    1975-01-01

    Leukocyte migration inhibitory factor assay was employed as an in vitro diagnostic aid in nickel dermatitis, the second most common contact dermatitis in North America. 15 patch test-positive and 5 patch test-negative patients, all giving a past history suggestive of nickel dermatitis, were investigated. Significant inhibition of leukocyte migration in both groups of patients was obtained only with nickel sulfate-albumin conjugate and not with unconjugated nickel sulfate. Specificity of this system was tested by utilizing an unrelated metallic albumin complex, and no inhibition was found. When patch testing is equivocal or contraindicated, this in vitro technique may be a practical alternative. PMID:1102460

  1. The melanocortin system in leukocyte biology.

    PubMed

    Catania, Anna

    2007-02-01

    The melanocortin system is composed of the melanocortin peptides, adrenocorticotropic hormone and alpha-, beta-, and gamma-melanocyte-stimulating hormone, the melanocortin receptors (MCRs), and the endogenous antagonists agouti- and agouti-related protein. Melanocortin peptides exert multiple effects upon the host, including anti-inflammatory and immunomodulatory effects. Leukocytes are a source of melanocortins and a major target for these peptides. Because of reduced translocation of the nuclear factor NF-kappaB to the nucleus, MCR activation by their ligands causes a collective reduction of the most important molecules involved in the inflammatory process. This review examines how melanocortin peptides and their receptors participate in leukocyte biology. PMID:17041004

  2. Genetics Home Reference: leukocyte adhesion deficiency type 1

    MedlinePlus

    ... adhesion deficiency type 1 leukocyte adhesion deficiency type 1 Enable Javascript to view the expand/collapse boxes. ... All Close All Description Leukocyte adhesion deficiency type 1 is a disorder that causes the immune system ...

  3. Halo sign on indium-111 leukocyte scan in gangrenous cholecystitis

    SciTech Connect

    Bauman, J.M.; Boykin, M.; Hartshorne, M.F.; Cawthon, M.A.; Landry, A.J.

    1986-02-01

    A 56-year-old man with a long history of Crohn's disease was evaluated by In-111 labeled leukocyte scanning. A halo of leukocyte activity was seen around the gallbladder fossa. A gangrenous gallbladder was removed at surgery.

  4. Position 156 influences the peptide repertoire and tapasin dependency of human leukocyte antigen B*44 allotypes

    PubMed Central

    Badrinath, Soumya; Saunders, Philippa; Huyton, Trevor; Aufderbeck, Susanne; Hiller, Oliver; Blasczyk, Rainer; Bade-Doeding, Christina

    2012-01-01

    Background Polymorphic differences between donor and recipient human leukocyte antigen class I molecules can result in graft-versus-host disease due to distinct peptide presentation. As part of the peptide-loading complex, tapasin plays an important role in selecting peptides from the pool of potential ligands. Class I polymorphisms can significantly alter the tapasin-mediated interaction with the peptide-loading complex and although most class I allotypes are highly dependent upon tapasin, some are able to load peptides independently of tapasin. Several human leukocyte antigen B*44 allotypes differ exclusively at position 156 (B*44:02156Asp, 44:03156Leu, 44:28156Arg, 44:35156Glu). From these alleles, only the high tapasin-dependency of human leukocyte antigen B*44:02 has been reported. Design and Methods We investigated the influence of position 156 polymorphisms on both the requirement of tapasin for efficient surface expression of each allotype and their peptide features. Genes encoding human leukocyte antigen B*44 variants bearing all possible substitutions at position 156 were lentivirally transduced into human leukocyte antigen class I-negative LCL 721.221 cells and the tapasin-deficient cell line LCL 721.220. Results Exclusively human leukocyte antigen B*44:28156Arg was expressed on the surface of tapasin-deficient cells, suggesting that the remaining B*44/156 variants are highly tapasin-dependent. Our computational analysis suggests that the tapasin-independence of human leukocyte antigen B*44:28156Arg is a result of stabilization of the peptide binding region and generation of a more peptide receptive state. Sequencing of peptides eluted from human leukocyte antigen B*44 molecules by liquid chromatography-electrospray ionization-mass spectrometry (LTQ-Orbitrap) demonstrated that both B*44:02 and B*44:28 share the same overall peptide motif and a certain percentage of their individual peptide repertoires in the presence and/or absence of tapasin

  5. Oxygen radical production by avian leukocytes.

    PubMed

    Conlon, P; Smith, D; Gowlett, T

    1991-04-01

    Oxygen radical production by heterophils of red-tailed hawks and chickens, and by neutrophils of calves, was evaluated in a chemiluminescence microassay. Leukocytes were isolated by centrifugation of blood in capillary tubes and then challenged with opsonized zymosan in the presence of luminol. Avian heterophils produced significantly fewer oxygen radicals than did bovine neutrophils. PMID:1884301

  6. MICROWAVES, HYPERTHERMIA, AND HUMAN LEUKOCYTE FUNCTION

    EPA Science Inventory

    The objective of this study is to determine whether exposure to microwaves (2450 MHz) affects the function of human leukocytes in the resting state and during antigenic or mitogenic challenge. This publication is a summary report of the construction and calibration of a waveguide...

  7. Bovine leukocyte adhesion deficiency: in vitro assessment of neutrophil function and leukocyte integrin expression.

    PubMed Central

    Olchowy, T W; Bochsler, P N; Neilsen, N R; Welborn, M G; Slauson, D O

    1994-01-01

    Bovine leukocyte adhesion deficiency (BLAD) was identified in a two-month-old Holstein heifer calf using DNA-polymerase chain reaction analysis of the affected calf and other clinical parameters. Neutrophil integrin expression (CD18, CD11a, CD11c), aggregation, and transendothelial migration were studied in vitro. Neutrophils were isolated from the affected calf and from normal, healthy, age-matched control Holstein calves. Neutrophils isolated from the affected BLAD calf had decreased expression of leukocyte integrins on their cell surface, decreased ability to aggregate in response to chemotactic stimuli, and decreased ability to migrate across bovine endothelial cell monolayers in vitro. Transendothelial migration of neutrophils from normal calves was reduced to levels comparable to the BLAD neutrophils by treatment with an anti-CD18 monoclonal antibody (MAb 60.3). Peripheral-blood lymphocytes from the BLAD calf also expressed negligible levels of leukocyte integrins, similar to their neutrophil counterparts. Our experimental findings in vitro correlate well with the clinical observations of decreased leukocyte trafficking and diminished host defense in leukocyte adhesion-deficient animals. The syndrome of BLAD may be a suitable model for one of the human leukocyte adhesion deficiency disorders. Images Fig. 4. PMID:7911733

  8. Bovine leukocyte adhesion deficiency: in vitro assessment of neutrophil function and leukocyte integrin expression.

    PubMed

    Olchowy, T W; Bochsler, P N; Neilsen, N R; Welborn, M G; Slauson, D O

    1994-04-01

    Bovine leukocyte adhesion deficiency (BLAD) was identified in a two-month-old Holstein heifer calf using DNA-polymerase chain reaction analysis of the affected calf and other clinical parameters. Neutrophil integrin expression (CD18, CD11a, CD11c), aggregation, and transendothelial migration were studied in vitro. Neutrophils were isolated from the affected calf and from normal, healthy, age-matched control Holstein calves. Neutrophils isolated from the affected BLAD calf had decreased expression of leukocyte integrins on their cell surface, decreased ability to aggregate in response to chemotactic stimuli, and decreased ability to migrate across bovine endothelial cell monolayers in vitro. Transendothelial migration of neutrophils from normal calves was reduced to levels comparable to the BLAD neutrophils by treatment with an anti-CD18 monoclonal antibody (MAb 60.3). Peripheral-blood lymphocytes from the BLAD calf also expressed negligible levels of leukocyte integrins, similar to their neutrophil counterparts. Our experimental findings in vitro correlate well with the clinical observations of decreased leukocyte trafficking and diminished host defense in leukocyte adhesion-deficient animals. The syndrome of BLAD may be a suitable model for one of the human leukocyte adhesion deficiency disorders. PMID:7911733

  9. Nicotinamide phosphoribosyltransferase leukocyte overexpression in Graves' opthalmopathy.

    PubMed

    Sawicka-Gutaj, Nadia; Budny, Bartłomiej; Zybek-Kocik, Ariadna; Sowiński, Jerzy; Ziemnicka, Katarzyna; Waligórska-Stachura, Joanna; Ruchała, Marek

    2016-08-01

    To investigate the role of NAMPT/visfatin in euthyroid patients with Graves' disease without (GD) and with Graves' ophthalmopathy (GO), we analyzed NAMPT leukocyte expression and its serum concentration. This was a single-center, cross-sectional study with consecutive enrollment. In total, 149 patients diagnosed with Graves' disease were enrolled in the study. We excluded subjects with hyper- or hypothyroidism, diabetes mellitus, other autoimmune disorders, active neoplastic disease, and infection. The control group was recruited among healthy volunteers adjusted for age, sex, and BMI with normal thyroid function and negative thyroid antibodies. Serum levels of visfatin, TSH, FT4, FT3, antibodies against TSH receptor (TRAb), antithyroperoxidase antibodies, antithyroglobulin antibodies, fasting glucose, and insulin were measured. NAMPT mRNA leukocyte expression was assessed using RT-qPCR. NAMPT/visfatin serum concentration was higher in GD (n = 44) and GO (n = 49) patients than in the control group (n = 40) (p = 0.0275). NAMPT leukocyte expression was higher in patients with GO (n = 30) than in GD patients (n = 27) and the control group (n = 29) (p < 0.0001). Simple linear regression analysis revealed that NAMPT/visfatin serum concentration was significantly associated with GD (β = 1.5723; p = 0.021). When NAMPT leukocyte expression was used as a dependent variable, simple regression analysis found association with TRAb, fasting insulin level, HOMA-IR, GD, and GO. In the stepwise multiple regression analysis, we confirmed the association between higher serum NAMPT/visfatin level and GD (coefficient = 1.5723; p = 0.0212), and between NAMPT leukocyte expression and GO (coefficient = 2.4619; p = 0.0001) and TRAb (coefficient = 0.08742; p = 0.006). Increased NAMPT leukocyte expression in patients with GO might suggest a presently undefined role in the pathogenesis of GO. PMID:26767650

  10. Evaluation of leptin receptor expression on buffalo leukocytes.

    PubMed

    De Matteis, Giovanna; Grandoni, Francesco; Scatà, Maria Carmela; Catizone, Angela; Reale, Anna; Crisà, Alessandra; Moioli, Bianca

    2016-09-01

    Experimental evidences support a direct role for leptin in immunity. Besides controlling food intake and energy expenditure, leptin was reported to be involved in the regulation of the immune system in ruminants. The aim of this work was to highlight the expression of leptin receptor (LEPR) on Bubalus bubalis immune cells using a multi-approach assessment: flow cytometry, confocal microscopy and gene expression analysis. Flow cytometric analysis of LEPR expression showed that peripheral blood monocytes were the predominant cells expressing LEPR. This result was corroborated by confocal microscopy and RT-PCR analysis. Moreover, among lymphocytes, LEPR was mainly expressed by B lymphocytes and Natural Killer cells. Evidence of LEPR expression on buffalo blood leukocytes showed to be a good indicator of the responsivity of these cells to leptin, so confirming the involvement of leptin in buffalo immune response. PMID:27436440

  11. Delay discounting, genetic sensitivity, and leukocyte telomere length.

    PubMed

    Yim, Onn-Siong; Zhang, Xing; Shalev, Idan; Monakhov, Mikhail; Zhong, Songfa; Hsu, Ming; Chew, Soo Hong; Lai, Poh San; Ebstein, Richard P

    2016-03-01

    In a graying world, there is an increasing interest in correlates of aging, especially those found in early life. Leukocyte telomere length (LTL) is an emerging marker of aging at the cellular level, but little is known regarding its link with poor decision making that often entails being overly impatient. Here we investigate the relationship between LTL and the degree of impatience, which is measured in the laboratory using an incentivized delay discounting task. In a sample of 1,158 Han Chinese undergraduates, we observe that steeper delay discounting, indexing higher degree of impatience, is negatively associated with LTL. The relationship is robust after controlling for health-related variables, as well as risk attitude-another important determinant of decision making. LTL in females is more sensitive to impatience than in males. We then asked if genes possibly modulate the effect of impatient behavior on LTL. The oxytocin receptor gene (OXTR) polymorphism rs53576, which has figured prominently in investigations of social cognition and psychological resources, and the estrogen receptor β gene (ESR2) polymorphism rs2978381, one of two gonadal sex hormone genes, significantly mitigate the negative effect of impatience on cellular aging in females. The current results contribute to understanding the relationship between preferences in decision making, particularly impatience, and cellular aging, for the first time to our knowledge. Notably, oxytocin and estrogen receptor polymorphisms temper accelerated cellular aging in young females who tend to make impatient choices. PMID:26903639

  12. Indium-111 leukocyte scanning and fracture healing

    SciTech Connect

    Mead, L.P.; Scott, A.C.; Bondurant, F.J.; Browner, B.D. )

    1990-01-01

    This study was undertaken to determine the specificity of indium-111 leukocyte scans for osteomyelitis when fractures are present. Midshaft tibial osteotomies were performed in 14 New Zealand white rabbits, seven of which were infected postoperatively with Staphylococcus aureus per Norden's protocol. All 14 rabbits were scanned following injection with 75 microCi of indium 111 at 72 h after osteotomy and at weekly intervals for 4 weeks. Before the rabbits were killed, the fracture sites were cultured to document the presence or absence of infection. The results of all infected osteotomy sites were positive, whereas no positive scans were found in the noninfected osteotomies. We concluded from this study that uncomplicated fracture healing does not result in a positive indium-111 leukocyte scan.

  13. Getting Leukocytes to the Site of Inflammation

    PubMed Central

    Muller, W. A.

    2013-01-01

    There is no “response” in either the innate or adaptive immune response unless leukocytes cross blood vessels. They do this through the process of diapedesis, in which the leukocyte moves in ameboid fashion through tightly apposed endothelial borders (paracellular transmigration) and in some cases through the endothelial cell itself (transcellular migration). This review summarizes the steps leading up to diapedesis, then focuses on the molecules and mechanisms responsible for transendothelial migration. Surprisingly, many of the same molecules and mechanisms that regulate paracellular migration also control transcellular migration, including a major role for membrane from the recently described lateral border recycling compartment. A hypothesis that integrates the various known mechanisms of transmigration is proposed. PMID:23345459

  14. Comparison of technetium-99m-HM-PAO leukocytes with indium-111-oxine leukocytes for localizing intraabdominal sepsis

    SciTech Connect

    Mountford, P.J.; Kettle, A.G.; O'Doherty, M.J.; Coakley, A.J. )

    1990-03-01

    Technetium-99m-HM-PAO (({sup 99m}Tc)HM-PAO) leukocyte and indium-111-oxine (111In-oxine) leukocyte scanning were carried out simultaneously in 41 patients at 4 hr and 24 hr after reinjection to determine whether the 4-hr {sup 99m}Tc scan could replace the 24-hr {sup 111}In scan for detecting intraabdominal sepsis. Abdominal infection was confirmed in 12 cases. The 4-hr {sup 99}Tc-leukocyte scan, the 4-hr {sup 111}In-leukocyte scan, and the 24-hr {sup 111}In-leukocyte scan yielded a sensitivity of 100%, 67%, and 100%, respectively, and a specificity of 62%, 90%, and 86%, respectively. The 24-hr {sup 99m}Tc-leukocyte scan also produced a sensitivity of 100%, but it was falsely positive in all 29 cases without infection due to physiologic bowel uptake. False-positive 4-hr {sup 99m}Tc-leukocyte scans were also produced by physiologic bowel uptake in seven cases all of whom had true-negative 4-hr and 24-hr {sup 111}In-leukocyte scans. Because of the high incidence of false-positive 4-hr ({sup 99m}Tc)HM-PAO leukocyte scans, it was concluded that they could not replace 24-hr {sup 111}In-leukocyte scans for detecting intraabdominal sepsis, and that serial {sup 99m}Tc leukocyte scans starting earlier than 4 hr after reinjection must be evaluated.

  15. Computational modeling of leukocyte adhesion cascade (LAC)

    NASA Astrophysics Data System (ADS)

    Sarkar, Kausik

    2005-11-01

    In response to an inflammation in the body, leukocytes (white blood cell) interact with the endothelium (interior wall of blood vessel) through a series of steps--capture, rolling, adhesion and transmigration--critical for proper functioning of the immune system. We are numerically simulating this process using a Front-tracking finite-difference method. The viscoelastcity of the cell membrane, cytoplasm and nucleus are incorporated and allowed to change with time in response to the cell surface molecular chemistry. The molecular level forces due to specific ligand-receptor interactions are accounted for by stochastic spring-peeling model. Even though leukocyte rolling has been investigated through various models, the transitioning through subsequent steps, specifically firm adhesion and transmigration through endothelial layer, has not been modeled. The change of viscoelastic properties due to the leukocyte activation is observed to play a critical role in mediating the transition from rolling to transmigration. We will provide details of our approach and discuss preliminary results.

  16. Cell adhesion molecules mediate radiation-induced leukocyte adhesion to the vascular endothelium.

    PubMed

    Hallahan, D; Kuchibhotla, J; Wyble, C

    1996-11-15

    The predominant early histological changes in irradiated tissues are edema and leukocyte infiltration. Cell adhesion molecules (CAMs) are required for the extravasation of leukocytes from the circulation. To study the role of CAMs in the pathogenesis of radiation-mediated inflammation, we quantified the expression of P-selectin, E-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 glycoproteins on the surface of irradiated human endothelial cells. We found that E-selectin and ICAM-1 expression increased after irradiation, whereas there was no increased expression of other cytokine-inducible adhesion molecules (P-selectin or vascular cell adhesion molecule-1). We found a dose- and time-dependent increase in radiation-induced expression of both E-selectin and ICAM-1. Furthermore, the threshold dose for E-selectin expression was 1 Gy, whereas the threshold dose for ICAM-1 synthesis was 5 Gy of X-rays. Northern blot analysis of RNA from irradiated endothelial cells demonstrated that ICAM-1 is expressed at 3-6 h following irradiation. No de novo protein synthesis was required for increased ICAM-1 mRNA expression. The 1.1-kb segment of the 5' untranslated region of the ICAM-1 gene was sufficient for X-ray induction of chloramphenicol acetyltransferase reporter gene expression. We measured whether ICAM-1 mediates adhesion of leukocyte to the irradiated endothelium and found that leukocyte adhesion occurred concurrently with ICAM-1 induction. Radiation-mediated leukocyte adhesion was prevented by anti-ICAM-1 blocking antibodies. These data indicate that ICAM-1 participates in the inflammatory response to ionizing radiation. Moreover, radiation induction of these CAMs occurs in the absence of tumor necrosis factor and interleukin 1 production. PMID:8912850

  17. Intracytoplasmic inclusions in circulating leukocytes from an eastern box turtle (Terrapene carolina carolina) with iridoviral infection.

    PubMed

    Allender, Matthew C; Fry, Michael M; Irizarry, Armando R; Craig, Linden; Johnson, April J; Jones, Michael

    2006-07-01

    A free-ranging adult female eastern box turtle (Terrapene carolina carolina) was presented to the University of Tennessee in October 2003 because of suspected trauma and blindness. Physical examination revealed lethargy, clear ocular and nasal discharges, and white oral and laryngeal plaques. Intracytoplasmic inclusions within heterophils and large mononuclear leukocytes were observed on routine blood smear examination. Postmortem findings included necrosis of epithelial and parenchymal cells with intracytoplasmic inclusions. Ultrastructurally, the leukocyte inclusions consisted of variably electron-dense granular material and viral particles consistent with the Iridoviridae family of viruses. The virus shared 100% sequence identity to a 420-base pair sequence of frog virus 3 (family Iridoviridae, genus Ranavirus) as determined by polymerase chain reaction and gene sequencing targeting a portion of the Ranavirus major capsid protein gene. PMID:17092902

  18. Long Telomeres in Blood Leukocytes Are Associated with a High Risk of Ascending Aortic Aneurysm

    PubMed Central

    Huusko, Tuija J.; Santaniemi, Merja; Kakko, Sakari; Taskinen, Panu; Ukkola, Olavi; Kesäniemi, Y. Antero; Savolainen, Markku J.; Salonurmi, Tuire

    2012-01-01

    Ascending aortic aneurysm is a connective tissue disorder. Even though multiple novel gene mutations have been identified, risk profiling and diagnosis before rupture still represent a challenge. There are studies demonstrating shorter telomere lengths in the blood leukocytes of abdominal aortic aneurysm patients. The aim of this study was to measure whether relative telomere lengths are changed in the blood leukocytes of ascending aortic aneurysm patients. We also studied the expression of telomerase in aortic tissue samples of ascending aortic aneurysms. Relative lengths of leukocyte telomeres were determined from blood samples of patients with ascending aortic aneurysms and compared with healthy controls. Telomerase expression, both at the level of mRNA and protein, was quantified from the aortic tissue samples. Mean relative telomere length was significantly longer in ascending aortic aneurysm blood samples compared with controls (T/S ratio 0.87 vs. 0.61, p<0.001). Expressions of telomerase mRNA and protein were elevated in the aortic aneurysm samples (p<0.05 and p<0.01). Our study reveals a significant difference in the mean length of blood leukocyte telomeres in ascending aortic aneurysm and controls. Furthermore, expression of telomerase, the main compensating factor for telomere loss, is elevated at both the mRNA and protein level in the samples of aneurysmal aorta. Further studies will be needed to confirm if this change in telomere length can serve as a tool for assessing the risk of ascending aortic aneurysm. PMID:23209831

  19. Blood leukocyte and spleen lymphocyte immune response of spleen lymphocytes and whole blood leukocytes of hamsters

    SciTech Connect

    Peters, B.A.; Sothmann, M.; Wehrenberg, W.B. )

    1989-01-01

    This study was designed to evaluate the effects of chronic physical activity on the immune response of spleen lymphocytes and whole blood leukocytes of hamsters. Animals were kept sedentary or allowed to exercise spontaneously on running wheels for eight weeks. Physically active animals averaged 12 kilometers per day. The immune response of spleen lymphocytes whole blood leukocytes was evaluated by {sup 3}H-thymidine incorporation in response to Concanavalin A or lipopolysaccharide. There was no treatment effect between physically active and sedentary hamster in response of spleen lymphocytes. The immune response of whole blood leukocytes to these mitogens was significantly greater in physically active vs. sedentary hamsters. These results demonstrate that chronic physical activity has the capacity to modulate immunoresponses.

  20. Intravital Microscopy of Leukocyte-endothelial and Platelet-leukocyte Interactions in Mesenterial Veins in Mice.

    PubMed

    Herr, Nadine; Mauler, Maximilian; Bode, Christoph; Duerschmied, Daniel

    2015-01-01

    Intravital microscopy is a method that can be used to investigate different processes in different regions and vessels in living animals. In this protocol, we describe intravital microscopy of mesentery veins. This can be performed in a short period of time with reproducible results showing leukocyte-endothelial interactions in vivo. We describe an inflammatory setting after LPS challenge of the endothelium. But in this model one can apply many different types of inflammatory conditions, like bacterial, chemical or biological and investigate the administration of drugs and their direct effects on the living animal and its impact on leukocyte recruitment. This protocol has been applied successfully to a number of different treatments of mice and their effects on inflammatory response in vessels. Herein, we describe the visualization of leukocytes and platelets by fluorescently labeling these with rhodamine 6G. Additionally, any specific imaging can be performed using targeted fluorescently labeled molecules. PMID:26325284

  1. Leukocyte emigration in normal calves and calves with leukocyte adhesion deficiency.

    PubMed

    Nagahata, H; Masuyama, A; Masue, M; Yuki, M; Higuchi, H; Ohtsuka, H; Kurosawa, T; Sato, H; Noda, H

    1997-12-01

    The emigration of leukocytes from calves with beta 2 integrin deficiency (BLAD) into bronchoalveolar spaces and scraped tissues was compared to that of normal calves. Polymorphonuclear neutrophils were found in bronchoalveolar lavage fluid from BLAD-affected calves showing chronic pneumonia. The neutrophils were complement receptor type 3 (CR3)-negative when characterized by flow cytometric analysis using anti-CD18 monoclonal antibody. Chemiluminescent response mediated by CR3 in neutrophils isolated from bronchoalveolar lavage fluid from BLAD-calves showed similar findings obtained from CR3-deficient neutrophils. Neutrophils from normal calves migrated into scraped tissue which was prepared in an upper gluteal surface area, whereas few leukocytes from calves with BLAD migrated to the scraped tissue, evaluated by skin window (Rebuck) method. These findings confirmed the extravasation of CR3-deficient leukocytes into bronchoalveolar lumen in BLAD calves, and demonstrated in vivo characteristics of extravasating property of normal and CR3-deficient neutrophils into scraped tissues. PMID:9450245

  2. Correlation of MLH1 and MGMT methylation levels between peripheral blood leukocytes and colorectal tissue DNA samples in colorectal cancer patients

    PubMed Central

    LI, XIA; WANG, YIBAINA; ZHANG, ZUOMING; YAO, XIAOPING; GE, JIE; ZHAO, YASHUANG

    2013-01-01

    CpG island methylation in the promoter regions of the DNA mismatch repair gene mutator L homologue 1 (MLH1) and DNA repair gene O6-methylguanine-DNA methyltransferase (MGMT) genes has been shown to occur in the leukocytes of peripheral blood and colorectal tissue. However, it is unclear whether the methylation levels in the blood leukocytes and colorectal tissue are correlated. The present study analyzed and compared the levels of MGMT and MLH1 gene methylation in the leukocytes of peripheral blood and colorectal tissues obtained from patients with colorectal cancer (CRC). The methylation levels of MGMT and MLH1 were examined using methylation-sensitive high-resolution melting (MS-HRM) analysis. A total of 44 patients with CRC were selected based on the MLH1 and MGMT gene methylation levels in the leukocytes of the peripheral blood. Corresponding colorectal tumor and normal tissues were obtained from each patient and the DNA methylation levels were determined. The correlation coefficients were evaluated using Spearman’s rank test. Agreement was determined by generalized κ-statistics. Spearman’s rank correlation coefficients (r) for the methylation levels of the MGMT and MLH1 genes in the leukocytes of the peripheral blood and normal colorectal tissue were 0.475 and 0.362, respectively (P=0.001 and 0.016, respectively). The agreement of the MGMT and MLH1 gene methylation levels in the leukocytes of the peripheral blood and normal colorectal tissue were graded as fair and poor (κ=0.299 and 0.126, respectively). The methylation levels of MGMT and MLH1 were moderately and weakly correlated between the patient-matched leukocytes and the normal colorectal tissue, respectively. Blood-derived DNA methylation measurements may not always represent the levels of normal colorectal tissue methylation. PMID:24179526

  3. Sequence and expression analysis of rainbow trout CXCR2, CXCR3a and CXCR3b aids interpretation of lineage-specific conversion, loss and expansion of these receptors during vertebrate evolution.

    PubMed

    Xu, Qiaoqing; Li, Ronggai; Monte, Milena M; Jiang, Yousheng; Nie, Pin; Holland, Jason W; Secombes, Chris J; Wang, Tiehui

    2014-08-01

    The chemokine receptors CXCR1-3 bind to 11 chemokines (CXCL1-11) that are clustered on the same chromosome in mammals but are largely missing in ray-finned fish. A second CXCR1/2, and a CXCR3a and CXCR3b gene have been cloned in rainbow trout. Analysis of CXCR1-R3 genes in lobe-finned fish, ray-finned fish and tetrapod genomes revealed that the teleostomian ancestor likely possessed loci containing both CXCR1 and CXCR2, and CXCR3a and CXCR3b. Based on this synteny analysis the first trout CXCR1/2 gene was renamed CXCR1, and the new gene CXCR2. The CXCR1/R2 locus was shown to have further expanded in ray-finned fish. In relation to CXCR3, mammals appear to have lost CXCR3b and birds both CXCR3a and CXCR3b during evolution. Trout CXCR1-R3 have distinct tissue expression patterns and are differentially modulated by PAMPs, proinflammatory cytokines and infections. They are highly expressed in macrophages and neutrophils, with CXCR1 and CXCR2 also expressed in B-cells. PMID:24613851

  4. An epigenomic signature of postprandial hyperglycemia in peripheral blood leukocytes.

    PubMed

    Shim, Sung-Mi; Cho, Yoon-Kyung; Hong, Eun-Jung; Han, Bok-Ghee; Jeon, Jae-Pil

    2016-03-01

    Postprandial hyperglycemia is known to be one of the earliest signs of abnormal glucose homeostasis associated with type 2 diabetes. This study aimed to assess clinical significance of a 1-h postprandial glucose level for the development of diabetes, and identify epigenetic biomarkers of postprandial hyperglycemia. We analyzed clinical data from the oral glucose tolerance tests for healthy subjects (n=4502). The ratio (Glu60/Glu0) of 1-h glucose levels to fasting glucose levels was significantly associated with an insulin sensitive index (QUICKI, quantitative insulin sensitivity check index) (β=0.055, P=1.25E-04) as well as a risk of future pre-diabetic and diabetic conversion. Next, DNA methylation profile analyses of 24 matched pairs of the high and low Glu60/Glu0 ratio subjects showed that specific DNA methylation levels in the promoter region of an olfactory receptor gene (olfactory receptor gene family10 member A4, OR10A4) were associated with the Glu60/Glu0 ratios (β=0.337, P=0.03). Moreover, acute oral glucose challenges decreased the DNA methylation levels of OR10A4 but not the global DNA methylation in peripheral leukocytes of healthy subjects (n=7), indicating that OR10A4 is a specific epigenomic target of postprandial hyperglycemia. This work suggests possible relevance of olfactory receptor genes to an earlier molecular biomarker of peripheral hyperglycemia and diabetic conversion. PMID:26632885

  5. Indium-111 leukocyte imaging in patients with rheumatoid arthritis

    SciTech Connect

    Uno, K.; Matsui, N.; Nohira, K.; Suguro, T.; Kitakata, Y.; Uchiyama, G.; Miyoshi, T.; Uematsu, S.; Inoue, S.; Arimizu, N.

    1986-03-01

    This study evaluates the usefulness of labeled leukocyte imaging in patients with rheumatoid arthritis. In 33 patients, the incidence of pain and swelling in 66 wrist joints and 66 knee joints was compared with the accumulation of (/sup 111/In)leukocytes. No accumulation of (/sup 111/In)leukocytes was seen in any of the patients' wrists (0/12) or knee joints (0/14) when both pain and swelling were absent. In contrast, 93% (25/27) of wrist joints and 80% (24/30) of knee joints with both pain and swelling were positive by (/sup 111/In)leukocyte scintigraphy. There was little correlation between the stage of the disease, as determined by radiography, and (/sup 111/In)leukocyte accumulation. This study suggests that (/sup 111/In)leukocyte imaging may be a reliable procedure for monitoring the activity of rheumatoid arthritis, especially for confirming the lack of an ongoing inflammatory response.

  6. Cervical leukocytes and spontaneous preterm birth

    PubMed Central

    Hunter, Patricia J.; Sheikh, Sairah; David, Anna L.; Peebles, Donald M.; Klein, Nigel

    2016-01-01

    The objective was to characterise cervical leukocyte populations and inflammatory mediators associated with term and recurrent spontaneous preterm birth (SPTB) in pregnant women with a history of SPTB. A prospective observational study was undertaken on 120 women with a history of SPTB. A cytobrush was used to sample cells from the cervix at 12–25 weeks’ gestation. Cells were enumerated and characterised by flow cytometry. Cytokines and chemokines were also measured. Participants were then grouped according to delivery at term (>36 + 6 weeks), late SPTB (34–36 + 6 weeks) or early SPTB (<34 weeks). Differences in leukocyte sub-populations, cytokine and chemokine levels were compared with outcome. Cervical leukocytes comprised up to 60% of the host-derived cells. Most of these (90–100%) were polymorphonuclear cells (PMN). Most of the remaining cells were mucosal macrophages expressing CD68 and CD103 in addition to markers shared with blood-borne monocytes. Failure to detect cervical macrophages in at least 250,000 cervical epithelial cells was a feature of women who experienced early SPTB (6 out of 6 cases, 95% CI 61–100%) compared with 34% (30 out of 88 cases, 95% CI 25–43%, P < 0.001) of women delivering after 34 weeks. CCL2 (MCP-1) was also low in SPTB before 34 weeks and levels above 75 ng/g and/or the presence of macrophages increased the specificity for birth after 34 weeks from 66% to 82% (55 out of 67 cases, 95% CI 73–91%). Absence of cervical macrophages and low CCL2 may be features of pregnancies at risk of early SPTB. PMID:26637953

  7. Imaging of Leukocyte Trafficking in Alzheimer's Disease.

    PubMed

    Pietronigro, Enrica; Zenaro, Elena; Constantin, Gabriela

    2016-01-01

    Alzheimer's disease (AD) is the most common neurodegenerative disorder and is characterized by a progressive decline of cognitive functions. The neuropathological features of AD include amyloid beta (Aβ) deposition, intracellular neurofibrillary tangles derived from the cytoskeletal hyperphosphorylated tau protein, amyloid angiopathy, the loss of synapses, and neuronal degeneration. In the last decade, inflammation has emerged as a key feature of AD, but most studies have focused on the role of microglia-driven neuroinflammation mechanisms. A dysfunctional blood-brain barrier has also been implicated in the pathogenesis of AD, and several studies have demonstrated that the vascular deposition of Aβ induces the expression of adhesion molecules and alters the expression of tight junction proteins, potentially facilitating the transmigration of circulating leukocytes. Two-photon laser scanning microscopy (TPLSM) has become an indispensable tool to dissect the molecular mechanisms controlling leukocyte trafficking in the central nervous system (CNS). Recent TPLSM studies have shown that vascular deposition of Aβ in the CNS promotes intraluminal neutrophil adhesion and crawling on the brain endothelium and also that neutrophils extravasate in the parenchyma preferentially in areas with Aβ deposits. These studies have also highlighted a role for LFA-1 integrin in neutrophil accumulation in the CNS of AD-like disease models, revealing that LFA-1 inhibition reduces the corresponding cognitive deficit and AD neuropathology. In this article, we consider how current imaging techniques can help to unravel new inflammation mechanisms in the pathogenesis of AD and identify novel therapeutic strategies to treat the disease by interfering with leukocyte trafficking mechanisms. PMID:26913031

  8. A role for leukocyte-endothelial adhesion mechanisms in epilepsy

    PubMed Central

    Fabene, Paolo F.; Mora, Graciela Navarro; Martinello, Marianna; Rossi, Barbara; Merigo, Flavia; Ottoboni, Linda; Bach, Simona; Angiari, Stefano; Benati, Donatella; Chakir, Asmaa; Zanetti, Lara; Schio, Federica; Osculati, Antonio; Marzola, Pasquina; Nicolato, Elena; Homeister, Jonathon W.; Xia, Lijun; Lowe, John B.; McEver, Rodger P.; Osculati, Francesco; Sbarbati, Andrea; Butcher, Eugene C.; Constantin, Gabriela

    2009-01-01

    The mechanisms involved in the pathogenesis of epilepsy, a chronic neurological disorder that affects approximately 1 percent of the world population, are not well understood1–3. Using a mouse model of epilepsy, we show that seizures induce elevated expression of vascular cell adhesion molecules and enhanced leukocyte rolling and arrest in brain vessels mediated by the leukocyte mucin P-selectin glycoprotein ligand-1 (PSGL-1) and leukocyte integrins α4β1 and αLβ2. Inhibition of leukocyte-vascular interactions either with blocking antibodies, or in mice genetically deficient in functional PSGL-1, dramatically reduced seizures. Treatment with blocking antibodies following acute seizures prevented the development of epilepsy. Neutrophil depletion also inhibited acute seizure induction and chronic spontaneous recurrent seizures. Blood-brain barrier (BBB) leakage, which is known to enhance neuronal excitability, was induced by acute seizure activity but was prevented by blockade of leukocyte-vascular adhesion, suggesting a pathogenetic link between leukocyte-vascular interactions, BBB damage and seizure generation. Consistent with potential leukocyte involvement in the human, leukocytes were more abundant in brains of epileptics than of controls. Our results suggest leukocyte-endothelial interaction as a potential target for the prevention and treatment of epilepsy. PMID:19029985

  9. Imputing amino acid polymorphisms in human leukocyte antigens.

    PubMed

    Jia, Xiaoming; Han, Buhm; Onengut-Gumuscu, Suna; Chen, Wei-Min; Concannon, Patrick J; Rich, Stephen S; Raychaudhuri, Soumya; de Bakker, Paul I W

    2013-01-01

    DNA sequence variation within human leukocyte antigen (HLA) genes mediate susceptibility to a wide range of human diseases. The complex genetic structure of the major histocompatibility complex (MHC) makes it difficult, however, to collect genotyping data in large cohorts. Long-range linkage disequilibrium between HLA loci and SNP markers across the major histocompatibility complex (MHC) region offers an alternative approach through imputation to interrogate HLA variation in existing GWAS data sets. Here we describe a computational strategy, SNP2HLA, to impute classical alleles and amino acid polymorphisms at class I (HLA-A, -B, -C) and class II (-DPA1, -DPB1, -DQA1, -DQB1, and -DRB1) loci. To characterize performance of SNP2HLA, we constructed two European ancestry reference panels, one based on data collected in HapMap-CEPH pedigrees (90 individuals) and another based on data collected by the Type 1 Diabetes Genetics Consortium (T1DGC, 5,225 individuals). We imputed HLA alleles in an independent data set from the British 1958 Birth Cohort (N = 918) with gold standard four-digit HLA types and SNPs genotyped using the Affymetrix GeneChip 500 K and Illumina Immunochip microarrays. We demonstrate that the sample size of the reference panel, rather than SNP density of the genotyping platform, is critical to achieve high imputation accuracy. Using the larger T1DGC reference panel, the average accuracy at four-digit resolution is 94.7% using the low-density Affymetrix GeneChip 500 K, and 96.7% using the high-density Illumina Immunochip. For amino acid polymorphisms within HLA genes, we achieve 98.6% and 99.3% accuracy using the Affymetrix GeneChip 500 K and Illumina Immunochip, respectively. Finally, we demonstrate how imputation and association testing at amino acid resolution can facilitate fine-mapping of primary MHC association signals, giving a specific example from type 1 diabetes. PMID:23762245

  10. Roscovitine blocks leukocyte extravasation by inhibition of cyclin-dependent kinases 5 and 9

    PubMed Central

    Berberich, Nina; Uhl, Bernd; Joore, Jos; Schmerwitz, Ulrike K; Mayer, Bettina A; Reichel, Christoph A; Krombach, Fritz; Zahler, Stefan; Vollmar, Angelika M; Fürst, Robert

    2011-01-01

    BACKGROUND AND PURPOSE Roscovitine, a cyclin-dependent kinase (CDK) inhibitor that induces tumour cell death, is under evaluation as an anti-cancer drug. By triggering leukocyte apoptosis, roscovitine can also enhance the resolution of inflammation. Beyond death-inducing properties, we tested whether roscovitine affects leukocyte-endothelial cell interaction, a vital step in the onset of inflammation. EXPERIMENTAL APPROACH Leukocyte-endothelial cell interactions were evaluated in venules of mouse cremaster muscle, using intravital microscopy. In primary human endothelial cells, we studied the influence of roscovitine on adhesion molecules and on the nuclear factor-κB (NF-κB) pathway. A cellular kinome array, in vitro CDK profiling and RNAi methods were used to identify targets of roscovitine. KEY RESULTS In vivo, roscovitine attenuated the tumour necrosis factor-α (TNF-α)-induced leukocyte adherence to and transmigration through, the endothelium. In vitro, roscovitine strongly inhibited TNF-α-evoked expression of endothelial adhesion molecules (E-selectin, intercellular cell adhesion molecule, vascular cell adhesion molecule). Roscovitine blocked NF-κB-dependent gene transcription, but not the NF-κB activation cascade [inhibitor of κB (IκB) kinase activity, IκB-α degradation, p65 translocation]. Using a cellular kinome array and an in vitro CDK panel, we found that roscovitine inhibited protein kinase A, ribosomal S6 kinase and CDKs 2, 5, 7 and 9. Experiments using kinase inhibitors and siRNA showed that the decreased endothelial activation was due solely to blockade of CDK5 and CDK9 by roscovitine. CONCLUSIONS AND IMPLICATIONS Our study highlights a novel mode of action for roscovitine, preventing endothelial activation and leukocyte-endothelial cell interaction by inhibition of CDK5 and 9. This might expand its usage as a promising anti-inflammatory compound. PMID:21391976

  11. Leukocyte Mitochondrial DNA Copy Number Is Associated with Chronic Obstructive Pulmonary Disease

    PubMed Central

    Liu, Shih-Feng; Kuo, Ho-Chang; Tseng, Ching-Wan; Huang, Hung-Tu; Chen, Yung-Che; Tseng, Chia-Cheng; Lin, Meng-Chih

    2015-01-01

    Background Oxidative stress is known to be involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). Evidence suggests that leukocytes mitochondria DNA (mtDNA) is susceptible to undergo mutations, insertions, or depletion in response to reactive oxidative stress (ROS). We hypothesize that mtDNA copy number is associated with the development of COPD. Methodology/Principal Findings Relative mtDNA copy number was measured by a quantitative real-time PCR assay using DNA extracted from peripheral leukocytes. MtDNA copy number of peripheral leukocytes in the COPD group (n = 86) is significantly decreased compared with non-smoker group (n = 77) (250.3± 21.5 VS. 464.2± 49.9, P<0.001). MtDNA copy number in the COPD group was less than that in the healthy smoking group, but P value nearly achieved significance (250.3± 21.5 VS. 404.0± 76.7, P = 0.08) MtDNA copy number has no significance with age, gender, body mass index, current smoking, and pack-years in COPD group, healthy smoker group and no smoker group, respectively. Serum glutathione level in the COPD group is significantly decreased compared with healthy smoker and non-smoker groups (4.5± 1.3 VS. 6.2± 1.9 and 4.5± 1.3 VS. 7.1±1.1 mU/mL; P<0.001 respectively). Pearson correlation test shows a significant liner correlation between mtDNA copy number and serum glutathione level (R = 0.2, P = 0.009). Conclusions/Significance COPD is associated with decreased leukocyte mtDNA copy number and serum glutathione. COPD is a regulatory disorder of leukocytes mitochondria. However, further studies are needed to determine the real mechanisms about the gene and the function of mitochondria. PMID:26394041

  12. Factors affecting leukocyte count in healthy adults.

    PubMed

    Carel, R S; Eviatar, J

    1985-09-01

    The relationships between white blood cell (WBC) count, smoking, and other health variables were determined among 35,000 apparently healthy men and women. The effect of smoking on the WBC count was greater than that of all other variables. The leukocyte level and the variance in WBC count values increased with increased smoking intensity. The relationship between smoking intensity and leukocyte level is expressed quantitatively by the following regression equation: WBC (10(3)/mm3) = 7.1 + 0.05(SM), where SM has seven values according to the smoking level. Multiple regression analysis with additional variables other than smoking did not much improve the predictive value of the equation. The effect of smoking on WBC count could be only partially explained by an inflammatory process, e.g., chronic bronchitis. Relationships of statistical significance (but mostly with r values of less than 0.10) were found between WBC count and the following variables: hemoglobin, heart rate, weight (or Quetelet index), cholesterol, uric acid, creatinine, sex, ethnic origin, systolic blood pressure, height, blood sugar, and diastolic blood pressure. The normal WBC count range for smokers differs from that of nonsmokers and is shifted to the right according to the smoking level. This may have both a diagnostic and prognostic significance in different clinical settings. PMID:4070192

  13. [A diagnostic analysis of a genetic mutation associated with a deficiency of leukocyte adhesion in cattle].

    PubMed

    Kostetskiĭ, I E; Kirilenko, S M; Glazko, V I; Sozinov, A A

    1996-01-01

    Bovine leukocyte adhesion (BLAD) is a recessive autosomal disease in Holstein-Friesian cattle caused by point mutation in CD18 gene encoding neutrophil-surface glycoprotein. To determine BLAD carriers, the convenient primers were chosen to amplify the mutant region of gene with the following restriction analysis. A screening program for BLAD has been initiated. Among 190 animals from different Ukrainian farms 6 were heterozygous according to the tested trait, i.e., BLAD deficient. No homozygous BLAD carriers were detected. PMID:9281202

  14. Optimization of the PCR test for the mutation causing bovine leukocyte adhesion deficiency.

    PubMed

    Mirck, M H; Von Bannisseht-Wijsmuller, T; Timmermans-Besselink, W J; Van Luijk, J H; Buntjer, J B; Lenstra, J A

    1995-07-01

    The recent emergence of the bovine leukocyte adhesion deficiency (BLAD) demonstrated the risks of narrowing the genetic basis of a population. About 6% of the Holstein-Friesian cattle now descends from one bull who was a heterozygous BLAD carrier. Crossing his descendants resulted in the birth of homozygous BLAD calves with a life expectancy of < 1 year. The BLAD syndrome is caused by a point mutation in the gene coding for CD18, a subunit of the beta 2 integrins on the surface of leukocytes. By using a PCR-RFLP test, large numbers of cattle are now being screened in several countries to eradicate the mutant allele. We describe an optimization of the PCR primer set that has led to an improvement of the test. PMID:7580848

  15. The effects of stress on the enzymes of peripheral leukocytes

    NASA Technical Reports Server (NTRS)

    Leise, E. M.; Gray, I.

    1973-01-01

    Previous work showed an early response of rabbit and human leukocyte enzymes to the stress of bacterial infection. Since these represented a mixed population of leukocytes and since polymorphonuclear leukocytes (PMN) increased in these preparations, it was necessary to establish whether the observed increase in lactate dehydrenase (LDH) and protein was the result of an increase in any one particular cell type or in all cells. The need for the development of a simple reproducible method for the differential separation of peripheral leukocytes for the furtherance of our own studies was apparent. It was also becoming increasingly apparent that morphologically similar cells, such as small lymphocytes (L) and macrophages, were capable of different biological functions. A dextran gradient centrifugation method was developed which has provided an easily reproducible technique for separating L from PMN. During the course of this work, in which over 250 rabbits were examined, the pattern of daily leukocyte protein and enzyme variation became increasingly more apparent. This information could have some impact on future work with leukocyte enzymes, by our group and by other workers. The differences in normal protein and enzyme levels maintained by some individuals, and some inbred strains, were evaluated and reported separately. It has been shown that one type of leukocyte may react more to a given stress than other leukocytes.

  16. Alteration of rat polymorphonuclear leukocyte function after thermal injury.

    PubMed

    Gruber, D F; D'Alesandro, M M

    1989-01-01

    One portion of host defense to bacterial challenge(s) involves the activation and infiltration of endogenous polymorphonuclear leukocytes. Thermal injuries are frequently associated with immunologic abnormalities including alterations of polymorphonuclear leukocyte-associated nonspecific resistance. We examined isolated peripheral rat polymorphonuclear leukocytes for alterations in membrane potential, oxidative capability, and locomotor function after the experimental application of 20% full-thickness body surface area thermal injury. Thermal injury resulted in significant reductions of peripheral red blood cell concentration(s) and increases in leukocyte and platelet concentrations for 42 days after injury. In addition to the quantitative changes, polymorphonuclear leukocytes also demonstrated altered qualitative functions. Compared with phorbol myristate acetate-induced activation of normal cells, polymorphonuclear leukocyte membranes from thermal-injured animals were electrophysiologically less responsive for 3 weeks after injury. The ability of polymorphonuclear leukocytes to produce intracellular H2O2, a measure of oxidative function, was also significantly decreased for 7 days after injury. The paradox in this paradigm of thermal injury was the demonstration of peripheral polymorphonuclear leukocyte quantitative increases with concurrent significant qualitative impairment. Qualitative lesions included altered states of membrane depolarization and depressed oxidative capability that may individually, or collectively, reduce nonspecific immune capabilities of the host to levels that are inadequate to combat infection. PMID:2793916

  17. [Bovine leukocyte adhesion deficiency: clinical picture and differential diagnosis].

    PubMed

    Lienau, A; Stöber, M; Kehrli, M E; Tammen, I; Schwenger, B; Kuczka, A; Pohlenz, J

    1994-10-01

    The pathological clinical and laboratory findings obtained in 50 calves and young cattle affected with Bovine Leukocyte Adhesion Deficiency are compared with those found in 114 calves and young cattle showing marked neutrophil leukocytosis of other origin (age: < 2 years; leukocyte count: > 30,000 per microl; percentage of lymphocytes: < 55%). PMID:7851303

  18. 21 CFR 864.7675 - Leukocyte peroxidase test.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Leukocyte peroxidase test. 864.7675 Section 864... peroxidase test. (a) Identification. A leukocyte peroxidase test is a device used to distinguish certain... peroxidase activity as evidenced by staining. The results of this test are used in the differential...

  19. 21 CFR 864.7675 - Leukocyte peroxidase test.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Leukocyte peroxidase test. 864.7675 Section 864... peroxidase test. (a) Identification. A leukocyte peroxidase test is a device used to distinguish certain... peroxidase activity as evidenced by staining. The results of this test are used in the differential...

  20. 21 CFR 864.7675 - Leukocyte peroxidase test.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Leukocyte peroxidase test. 864.7675 Section 864... peroxidase test. (a) Identification. A leukocyte peroxidase test is a device used to distinguish certain... peroxidase activity as evidenced by staining. The results of this test are used in the differential...

  1. 21 CFR 864.7675 - Leukocyte peroxidase test.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Leukocyte peroxidase test. 864.7675 Section 864... peroxidase test. (a) Identification. A leukocyte peroxidase test is a device used to distinguish certain... peroxidase activity as evidenced by staining. The results of this test are used in the differential...

  2. 21 CFR 864.7675 - Leukocyte peroxidase test.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Leukocyte peroxidase test. 864.7675 Section 864... peroxidase test. (a) Identification. A leukocyte peroxidase test is a device used to distinguish certain... peroxidase activity as evidenced by staining. The results of this test are used in the differential...

  3. Degradation of thyroid hormones by phagocytosing human leukocytes.

    PubMed

    Klebanoff, S J; Green, W L

    1973-01-01

    Thyroxine (T(4)) and triiodothyronine (T(9)) are rapidly degraded by a purified preparation of myeloperoxidase (MPO) and H(2)O(2) with the formation of iodide and material which remains at the origin on paper chromatography. Deiodination by MPO and H(2)O(2) occurs more readily at pH 7.0 than at pH 5.0 in contrast to iodination by this system which is known to occur more readily at pH 5.0 than at pH 7.0. Degradation is inhibited by azide, cyanide, ascorbic acid, and propylthiouracil. Methimazole stimulates deiodination by MPO and H(2)O(2) but inhibits this reaction when MPO is replaced by lactoperoxidase or horseradish peroxidase.Intact human leukocytes, in the resting state, degrade T(4) and T(3) slowly: degradation, however, is increased markedly during phagocytosis of preopsonized particles. Serum inhibits this reaction. T(3) can be detected as a minor product of T(4) degradation. Proteolytic digestion of the reaction products increases the recovery of monoiodotyrosine. The fixation of iodine in the cytoplasm of leukocytes which contain ingested bacteria was detected radioautographically. Chronic granulomatous disease leukocytes, which are deficient in H(2)O(2) formation, degrade T(4) and T(3) poorly during phagocytosis. MPO-deficient leukocytes degrade the thyroid hormones at a slower rate than do normal leukocytes although considerable degradation is still observed. Azide, cyanide, ascorbic acid, and propylthiouracil which inhibit certain peroxidasecatalyzed reactions inhibit degradation by normal leukocytes; however, inhibition is incomplete. Formation of iodinated origin material is inhibited to a greater degree by azide, cyanide, and propylthiouracil than is deiodination. Methimazole inhibits the formation of iodinated origin material by both normal and MPO-deficient leukocytes. However, deiodination by normal leukocytes is stimulated and that of MPO-deficient leukocytes is unaffected by methimazole. Hypoxia inhibits the degradation of T(4) and T(3) by

  4. [Polar coordinates representation based leukocyte segmentation of microscopic cell images].

    PubMed

    Gu, Guanghua; Cui, Dong; Hao, Lianwang

    2010-12-01

    We propose an algorithm for segmentation of the overlapped leukocyte in the microscopic cell image. The histogram of the saturation channel in the cell image is smoothed to obtain the meaningful global valley point by the fingerprint smoothing method, and then the nucleus can be segmented. A circular region, containing the entire regions of the leukocyte, is marked off according to the equivalent sectional radius of the nucleus. Then, the edge of the overlapped leukocyte is represented by polar coordinates. The overlapped region by the change of the polar angle of the edge pixels is determined, and the closed edge of the leukocyte integrating the gradient information of the overlapped region is reconstructed. Finally, the leukocyte is exactly extracted. The experimental results show that our method has good performance in terms of recall ratio, precision ratio and pixel error ratio. PMID:21374971

  5. Cannabinoid-receptor expression in human leukocytes.

    PubMed

    Bouaboula, M; Rinaldi, M; Carayon, P; Carillon, C; Delpech, B; Shire, D; Le Fur, G; Casellas, P

    1993-05-15

    Marijuana and many of its constituent cannabinoids influence the central nervous system (CNS), probably through the cannabinoid receptor, which has recently been cloned in rat and human. While numerous reports have also described effects of cannabinoids on the immune system, the observation of both mRNA and cannabinoid receptor has hitherto been exclusively confined to the brain, a reported detection in the testis being the sole example of its presence at the periphery. Here we report the expression of the cannabinoid receptor on human immune tissues using a highly sensitive polymerase-chain-reaction-based method for mRNA quantification. We show that, although present in a much lower abundance than in brain, cannabinoid receptor transcripts are found in human spleen, tonsils and peripheral blood leukocytes. The distribution pattern displays important variations of the mRNA level for the cannabinoid receptor among the main human blood cell subpopulations. The rank order of mRNA levels in these cells is B cells > natural killer cells > or = polymorphonuclear neutrophils > or = T8 cells > monocytes > T4 cells. Cannabinoid-receptor mRNA, which is also found in monocytic, as well as T and B leukemia cell lines but not in Jurkat cells, presents a great diversity of expression on these cells as well, B-cell lines expressing a much higher level than T-cell lines. The cannabinoid receptor PCR products from leukocytes and brain are identical both in size and sequence suggesting a strong similarity between central and peripheral cannabinoid receptors. The expression of this receptor was demonstrated on membranes of the myelomonocytic U937 cells using the synthetic cannabinoid [3H]CP-55940 as ligand. The Kd determined from Scatchard analysis was 0.1 nM and the Bmax for membranes was 525 fmol/mg protein. The demonstration of cannabinoid-receptor expression at both mRNA and protein levels on human leukocytes provides a molecular basis for cannabinoid action on these cells. PMID

  6. Margination of leukocytes in blood flow through small tubes.

    PubMed

    Goldsmith, H L; Spain, S

    1984-03-01

    Leukocyte margination in the vessels of the microcirculation has been attributed to a flow-dependent interaction with red cells. To determine the extent of this effect, experiments with human blood were done in 100- to 180-micron tubes to detect changes in cell distribution as a function of hematocrit and flow rate. Using a flow visualization technique, the leukocyte concentration distribution was determined in 45% ghost cell suspensions. Migration of cells toward the wall was observed at centerline velocities greater than 1 mm sec-1 and increased with increasing flow rate. The effect was probably due to a more rapid inward migration of ghosts than leukocytes because of fluid inertia and cell density differences. Experiments were therefore carried out in whole blood at hematocrits from 20 to 60%, measuring the number concentration of leukocytes and erythrocytes within the tube, nt, and comparing it to that in the infusing reservoir, no, (Fahraeus effect). At mean tube shear rates G less than 100 sec-1, nt/no less than 1 for both leukocytes and erythrocytes showing net migration of cells away from the wall, although at nearly all hematocrits there was an enrichment of leukocytes relative to erythrocytes in the tubes. At G less than 50 sec-1, nt/no remained less than 1 for erythrocytes but increased to greater than 1 for leukocytes showing migration toward the wall, the increase being greatest at 20% hematocrit in the 100-micron tubes. The nature of the effect was revealed by cine films which showed that, as the flow rate decreased, erythrocytes formed rouleaux which migrated inward creating a core and displacing leukocytes to the periphery. In control experiments using washed blood cells in phosphate buffer-albumin, nt/no less than 1 for both leukocytes and erythrocytes at all G and hematocrits, and leukocytes were now distributed. Cine films of washed blood confirmed that, in the absence of rouleaux, no significant inward migration of erythrocytes occurred. PMID

  7. Age-Related Changes following In Vitro Stimulation with Rhodococcus equi of Peripheral Blood Leukocytes from Neonatal Foals

    PubMed Central

    Kachroo, Priyanka; Ivanov, Ivan; Seabury, Ashley G.; Liu, Mei; Chowdhary, Bhanu P.; Cohen, Noah D.

    2013-01-01

    Rhodococcus equi is an intracellular bacterium primarily known as an equine pathogen that infects young foals causing a pyogranulomatuous pneumonia. The molecular mechanisms mediating the immune response of foals to R. equi are not fully elucidated. Hence, global genomic high-throughput tools like gene expression microarrays might identify age-related gene expression signatures and molecular pathways that contribute to the immune mechanisms underlying the inherent susceptibility of foals to disease caused by R. equi. The objectives of this study were 2-fold: 1) to compare the expression profiles at specific ages of blood leukocytes from foals stimulated with virulent R. equi with those of unstimulated leukocytes; and, 2) to characterize the age-related changes in the gene expression profile associated with blood leukocytes in response to stimulation with virulent R. equi. Peripheral blood leukocytes were obtained from 6 foals within 24 hours (h) of birth (day 1) and 2, 4, and 8 weeks after birth. The samples were split, such that half were stimulated with live virulent R. equi, and the other half served as unstimulated control. RNA was extracted and the generated cDNA was labeled with fluorescent dyes for microarray hybridizations using an equine microarray. Our findings suggest that there is age-related differential expression of genes involved in host immune response and immunity. We found induction of genes critical for host immunity against pathogens (MHC class II) only at the later time-points (compared to birth). While it appears that foals up to 8-weeks of age are able to initiate a protective inflammatory response against the bacteria, relatively decreased expression of various other immune-related genes points toward inherent diminished immune responses closer to birth. These genes and pathways may contribute to disease susceptibility in foals if infected early in life, and might thus be targeted for developing preventative or therapeutic strategies. PMID

  8. Factors influencing human leukocyte adherence in vitro.

    PubMed

    Stepniewicz, W; Tchórzewski, H; Luciak, M

    1983-01-01

    Studies were performed on factors influencing leucocyte adherence in vitro. Blood condensation was found to increase leukocyte adherence. Addition of heparin, dextran or ethanol caused a significant reduction of white blood cell count in blood samples in comparison with blood mixed with sodium EDTA or ACD solution. This suggests the existence of two granulocyte subpopulations; viz, rapidly adhering and slowly adhering. Heparin enhanced granulocyte adherence, while dextran and ethanol decreased it. Five-day storage of ACD blood led to a decrease in granulocyte adherence, while addition of heparin or histamine to ACD blood prevented this change to occur. The glucose concentration of 1,000 mg/dl augmented granulocyte adherence, while higher glucose concentrations induced its progressive fall below the control values. There was no significant change of lymphocyte adherence during the experiments. PMID:6194070

  9. Leukocytic oxygen activation and microbicidal oxidative toxins.

    PubMed

    Hurst, J K; Barrette, W C

    1989-01-01

    Following a brief introduction of cellular response to stimulation comprising leukocyte activation, three major areas are discussed: (1) the neutrophil oxidase; (2) myeloperoxidase (MPO)-dependent oxidative microbicidal reactions; and (3) MPO-independent oxidative reactions. Topics included in section (A) are current views on the activation mechanism, redox composition, structural and topographic organization of the oxidase, and its respiratory products. In section (B), emphasis is placed on recent research on cidal mechanisms of HOCl, including the oxidative biochemistry of active chlorine compounds, identification of sites of lesions in bacteria, and attendant metabolic consequences. In section (C), we review the (bio)chemistry of H2O2 and .OH microbicidal reactions, with particular attention being given to addressing the controversial issue of probe methods to identify .OH radical and critical assessment of the recent proposal that MPO-independent killing arises from site-specific metal-catalyzed Fenton-type chemistry. PMID:2548810

  10. Leukocyte-associated immunoglobulin-like receptor-1 is expressed on human megakaryocytes and negatively regulates the maturation of primary megakaryocytic progenitors and cell line

    SciTech Connect

    Xue, Jiangnan; Zhang, Xiaoshu; Zhao, Haiya; Fu, Qiang; Cao, Yanning; Wang, Yuesi; Feng, Xiaoying; Fu, Aili

    2011-02-04

    Research highlights: {yields} LAIR-1 is expressed on human megakaryocytes from an early stage. {yields} Up-regulation of LAIR-1 negatively regulates megakaryocytic differentiation of cell line. {yields} LAIR-1 negatively regulates the differentiation of primary megakaryocytic progenitors. -- Abstract: Leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) is an inhibitory collagen receptor which belongs to the immunoglobulin (Ig) superfamily. Although the inhibitory function of LAIR-1 has been extensively described in multiple leukocytes, its role in megakaryocyte (MK) has not been explored so far. Here, we show that LAIR-1 is expressed on human bone marrow CD34{sup +}CD41a{sup +} and CD41a{sup +}CD42b{sup +} cells. LAIR-1 is also detectable in a fraction of human cord blood CD34{sup +} cell-derived MK that has morphological characteristics of immature MK. In megakaryoblastic cell line Dami, the membrane protein expression of LAIR-1 is up-regulated significantly when cells are treated with phorbol ester phorbol 12-myristate 13-acetate (PMA). Furthermore, cross-linking of LAIR-1 in Dami cells with its natural ligand or anti-LAIR-1 antibody leads to the inhibition of cell proliferation and PMA-promoted differentiation when examined by the MK lineage-specific markers (CD41a and CD42b) and polyploidization. In addition, we also observed that cross-linking of LAIR-1 results in decreased MK generation from primary human CD34{sup +} cells cultured in a cytokines cocktail that contains TPO. These results suggest that LAIR-1 is a likely candidate for an early marker of MK differentiation, and provide initial evidence indicating that LAIR-1 serves as a negative regulator of megakaryocytopoiesis.

  11. Endosymbiotic origin and differential loss of eukaryotic genes.

    PubMed

    Ku, Chuan; Nelson-Sathi, Shijulal; Roettger, Mayo; Sousa, Filipa L; Lockhart, Peter J; Bryant, David; Hazkani-Covo, Einat; McInerney, James O; Landan, Giddy; Martin, William F

    2015-08-27

    Chloroplasts arose from cyanobacteria, mitochondria arose from proteobacteria. Both organelles have conserved their prokaryotic biochemistry, but their genomes are reduced, and most organelle proteins are encoded in the nucleus. Endosymbiotic theory posits that bacterial genes in eukaryotic genomes entered the eukaryotic lineage via organelle ancestors. It predicts episodic influx of prokaryotic genes into the eukaryotic lineage, with acquisition corresponding to endosymbiotic events. Eukaryotic genome sequences, however, increasingly implicate lateral gene transfer, both from prokaryotes to eukaryotes and among eukaryotes, as a source of gene content variation in eukaryotic genomes, which predicts continuous, lineage-specific acquisition of prokaryotic genes in divergent eukaryotic groups. Here we discriminate between these two alternatives by clustering and phylogenetic analysis of eukaryotic gene families having prokaryotic homologues. Our results indicate (1) that gene transfer from bacteria to eukaryotes is episodic, as revealed by gene distributions, and coincides with major evolutionary transitions at the origin of chloroplasts and mitochondria; (2) that gene inheritance in eukaryotes is vertical, as revealed by extensive topological comparison, sparse gene distributions stemming from differential loss; and (3) that continuous, lineage-specific lateral gene transfer, although it sometimes occurs, does not contribute to long-term gene content evolution in eukaryotic genomes. PMID:26287458

  12. Association between Kinin B1 Receptor Expression and Leukocyte Trafficking across Mouse Mesenteric Postcapillary Venules

    PubMed Central

    McLean, Peter G.; Ahluwalia, Amrita; Perretti, Mauro

    2000-01-01

    Using intravital microscopy, we examined the role played by B1 receptors in leukocyte trafficking across mouse mesenteric postcapillary venules in vivo. B1 receptor blockade attenuated interleukin (IL)-1β–induced (5 ng intraperitoneally, 2 h) leukocyte–endothelial cell interactions and leukocyte emigration (∼50% reduction). The B1 receptor agonist des-Arg9bradykinin (DABK), although inactive in saline- or IL-8–treated mice, caused marked neutrophil rolling, adhesion, and emigration 24 h after challenge with IL-1β (when the cellular response to IL-1β had subsided). Reverse transcriptase polymerase chain reaction and Western blot revealed a temporal association between the DABK-induced response and upregulation of mesenteric B1 receptor mRNA and de novo protein expression after IL-1β treatment. DABK-induced leukocyte trafficking was antagonized by the B1 receptor antagonist des-arg10HOE 140 but not by the B2 receptor antagonist HOE 140. Similarly, DABK effects were maintained in B2 receptor knockout mice. The DABK-induced responses involved the release of neuropeptides from C fibers, as capsaicin treatment inhibited the responses. Treatment with the neurokinin (NK)1 and NK3 receptor antagonists attenuated the responses, whereas NK2, calcitonin gene-related peptide, or platelet-activating factor receptor antagonists had no effect. Substance P caused leukocyte recruitment that, similar to DABK, was inhibited by NK1 and NK3 receptor blockade. Mast cell depletion using compound 48/80 reduced DABK-induced leukocyte trafficking, and DABK treatment was shown histologically to induce mast cell degranulation. DABK-induced trafficking was inhibited by histamine H1 receptor blockade. Our findings provide clear evidence that B1 receptors play an important role in the mediation of leukocyte–endothelial cell interactions in postcapillary venules, leading to leukocyte recruitment during an inflammatory response. This involves activation of C fibers and mast cells

  13. Treatment of Obstructive Sleep Apnea Alters Cancer-associated Transcriptional Signatures in Circulating Leukocytes

    PubMed Central

    Gharib, Sina A.; Seiger, Ashley N.; Hayes, Amanda L.; Mehra, Reena; Patel, Sanjay R.

    2014-01-01

    Rationale: Obstructive sleep apnea (OSA) has been associated with a number of chronic disorders that may improve with effective therapy. However, the molecular pathways affected by continuous positive airway pressure (CPAP) treatment are largely unknown. We sought to assess the system-wide consequences of CPAP therapy by transcriptionally profiling peripheral blood leukocytes (PBLs). Methods: Subjects in whom severe OSA was diagnosed were treated with CPAP, and whole-genome expression measurement of PBLs was performed at baseline and following therapy. We used gene set enrichment analysis (GSEA) to identify pathways that were differentially enriched. Network analysis was then applied to highlight key drivers of processes influenced by CPAP. Results: Eighteen subjects with significant OSA underwent CPAP therapy and microarray analysis of their PBLs. Treatment with CPAP improved apnea-hypopnea index (AHI), daytime sleepiness, and blood pressure, but did not affect anthropometric measures. GSEA revealed a number of enriched gene sets, many of which were involved in neoplastic processes and displayed downregulated expression patterns in response to CPAP. Network analysis identified several densely connected genes that are important modulators of cancer and tumor growth. Conclusions: Effective therapy of OSA with CPAP is associated with alterations in circulating leukocyte gene expression. Functional enrichment and network analyses highlighted transcriptional suppression in cancer-related pathways, suggesting potentially novel mechanisms linking OSA with neoplastic signatures. Citation: Gharib SA; Seiger AN; Hayes AL; Mehra R; Patel SR. Treatment of obstructive sleep apnea alters cancer-associated transcriptional signatures in circulating leukocytes. SLEEP 2014;37(4):709-714. PMID:24688164

  14. Functional Mineralocorticoid Receptors in Human Vascular Endothelial Cells Regulate ICAM-1 Expression and Promote Leukocyte Adhesion

    PubMed Central

    Caprio, Massimiliano; Newfell, Brenna G.; la Sala, Andrea; Baur, Wendy; Fabbri, Andrea; Rosano, Giuseppe; Mendelsohn, Michael E.; Jaffe, Iris Z.

    2008-01-01

    In clinical trials, aldosterone antagonists decrease cardiovascular mortality and ischemia by unknown mechanisms. The steroid hormone aldosterone acts by binding to the mineralocorticoid receptor (MR), a ligand-activated transcription factor. In humans, aldosterone causes MR-dependent endothelial cell (EC) dysfunction and in animal models, aldosterone increases vascular macrophage infiltration and atherosclerosis. MR antagonists inhibit these effects without changing blood pressure, suggesting a direct role for vascular MR in EC function and atherosclerosis. Whether human vascular EC express functional MR is not known. Here we show that human coronary artery and aortic EC express MR mRNA and protein and that EC MR mediates aldosterone-dependent gene transcription. Human EC also express the enzyme 11-beta hydroxysteroid dehydrogenase-2(11βHSD2) and inhibition of 11βHSD2 in aortic EC enhances gene transactivation by cortisol, supporting that EC 11βHSD2 is functional. Furthermore, aldosterone stimulates transcription of the proatherogenic leukocyte-EC adhesion molecule Intercellular Adhesion Molecule-1(ICAM1) gene and protein expression on human coronary artery EC, an effect inhibited by the MR antagonist spironolactone and by MR knock-down with siRNA. Cell adhesion assays demonstrate that aldosterone promotes leukocyte-EC adhesion, an effect that is inhibited by spironolactone and ICAM1 blocking antibody, supporting that aldosterone induction of EC ICAM1 surface expression via MR mediates leukocyte-EC adhesion. These data show that aldosterone activates endogenous EC MR and proatherogenic gene expression in clinically important human EC. These studies describe a novel mechanism by which aldosterone may influence ischemic cardiovascular events and support a new explanation for the decrease in ischemic events in patients treated with aldosterone antagonists. PMID:18467630

  15. Impact of gene family evolutionary histories on phylogenetic species tree inference by gene tree parsimony.

    PubMed

    Shi, Tao

    2016-03-01

    Complicated history of gene duplication and loss brings challenge to molecular phylogenetic inference, especially in deep phylogenies. However, phylogenomic approaches, such as gene tree parsimony (GTP), show advantage over some other approaches in its ability to use gene families with duplications. GTP searches the 'optimal' species tree by minimizing the total cost of biological events such as duplications, but accuracy of GTP and phylogenetic signal in the context of different gene families with distinct histories of duplication and loss are unclear. To evaluate how different evolutionary properties of different gene families can impact on species tree inference, 3900 gene families from seven angiosperms encompassing a wide range of gene content, lineage-specific expansions and contractions were analyzed. It was found that the gene content and total duplication number in a gene family strongly influence species tree inference accuracy, with the highest accuracy achieved at either very low or very high gene content (or duplication number) and lowest accuracy centered in intermediate gene content (or duplication number), as the relationship can fit a binomial regression. Besides, for gene families of similar level of average gene content, those with relatively higher lineage-specific expansion or duplication rates tend to show lower accuracy. Additional correlation tests support that high accuracy for those gene families with large gene content may rely on abundant ancestral copies to provide many subtrees to resolve conflicts, whereas high accuracy for single or low copy gene families are just subject to sequence substitution per se. Very low accuracy reached by gene families of intermediate gene content or duplication number can be due to insufficient subtrees to resolve the conflicts from loss of alternative copies. As these evolutionary properties can significantly influence species tree accuracy, I discussed the potential weighting of the duplication cost by

  16. Metabolomics profiling reveals novel markers for leukocyte telomere length.

    PubMed

    Zierer, Jonas; Kastenmüller, Gabi; Suhre, Karsten; Gieger, Christian; Codd, Veryan; Tsai, Pei-Chien; Bell, Jordana; Peters, Annette; Strauch, Konstantin; Schulz, Holger; Weidinger, Stephan; Mohney, Robert P; Samani, Nilesh J; Spector, Tim; Mangino, Massimo; Menni, Cristina

    2016-01-01

    Leukocyte telomere length (LTL) is considered one of the most predictive markers of biological aging. The aim of this study was to identify novel pathways regulating LTL using a metabolomics approach. To this end, we tested associations between 280 blood metabolites and LTL in 3511 females from TwinsUK and replicated our results in the KORA cohort. We furthermore tested significant metabolites for associations with several aging-related phenotypes, gene expression markers and epigenetic markers to investigate potential underlying pathways. Five metabolites were associated with LTL: Two lysolipids, 1-stearoylglycerophosphoinositol (P=1.6×10(-5)) and 1-palmitoylglycerophosphoinositol (P=1.6×10(-5)), were found to be negatively associated with LTL and positively associated with phospholipase A2 expression levels suggesting an involvement of fatty acid metabolism and particularly membrane composition in biological aging. Moreover, two gamma-glutamyl amino acids, gamma-glutamyltyrosine (P=2.5×10(-6)) and gamma-glutamylphenylalanine (P=1.7×10(-5)), were negatively correlated with LTL. Both are products of the glutathione cycle and markers for increased oxidative stress. Metabolites were also correlated with functional measures of aging, i.e. higher blood pressure and HDL cholesterol levels and poorer lung, liver and kidney function. Our results suggest an involvement of altered fatty acid metabolism and increased oxidative stress in human biological aging, reflected by LTL and age-related phenotypes of vital organ systems. PMID:26797767

  17. Metabolomics profiling reveals novel markers for leukocyte telomere length

    PubMed Central

    Zierer, Jonas; Kastenmüller, Gabi; Suhre, Karsten; Gieger, Christian; Codd, Veryan; Tsai, Pei-Chien; Bell, Jordana; Peters, Annette; Strauch, Konstantin; Schulz, Holger; Weidinger, Stephan; Mohney, Robert P.; Samani, Nilesh J.; Spector, Tim; Mangino, Massimo; Menni, Cristina

    2016-01-01

    Leukocyte telomere length (LTL) is considered one of the most predictive markers of biological aging. The aim of this study was to identify novel pathways regulating LTL using a metabolomics approach. To this end, we tested associations between 280 blood metabolites and LTL in 3511 females from TwinsUK and replicated our results in the KORA cohort. We furthermore tested significant metabolites for associations with several aging-related phenotypes, gene expression markers and epigenetic markers to investigate potential underlying pathways. Five metabolites were associated with LTL: Two lysolipids, 1-stearoylglycerophosphoinositol (P=1.6×10−5) and 1-palmitoylglycerophosphoinositol (P=1.6×10−5), were found to be negatively associated with LTL and positively associated with phospholipase A2 expression levels suggesting an involvement of fatty acid metabolism and particularly membrane composition in biological aging. Moreover, two gamma-glutamyl amino acids, gamma-glutamyltyrosine (P=2.5×10−6) and gamma-glutamylphenylalanine (P=1.7×10−5), were negatively correlated with LTL. Both are products of the glutathione cycle and markers for increased oxidative stress. Metabolites were also correlated with functional measures of aging, i.e. higher blood pressure and HDL cholesterol levels and poorer lung, liver and kidney function. Our results suggest an involvement of altered fatty acid metabolism and increased oxidative stress in human biological aging, reflected by LTL and age-related phenotypes of vital organ systems. PMID:26797767

  18. Characterization of Leukocyte Formin FMNL1 Expression in Human Tissues.

    PubMed

    Gardberg, Maria; Heuser, Vanina D; Iljin, Kristiina; Kampf, Caroline; Uhlen, Mathias; Carpén, Olli

    2014-04-01

    Formins are cytoskeleton regulating proteins characterized by a common FH2 structural domain. As key players in the assembly of actin filaments, formins direct dynamic cytoskeletal processes that influence cell shape, movement and adhesion. The large number of formin genes, fifteen in the human, suggests distinct tasks and expression patterns for individual family members, in addition to overlapping functions. Several formins have been associated with invasive cell properties in experimental models, linking them to cancer biology. One example is FMNL1, which is considered to be a leukocyte formin and is known to be overexpressed in lymphomas. Studies on FMNL1 and many other formins have been hampered by a lack of research tools, especially antibodies suitable for staining paraffin-embedded formalin-fixed tissues. Here we characterize, using bioinformatics tools and a validated antibody, the expression pattern of FMNL1 in human tissues and study its subcellular distribution. Our results indicate that FMNL1 expression is not restricted to hematopoietic tissues and that neoexpression of FMNL1 can be seen in epithelial cancer. PMID:24700756

  19. Characterization of Leukocyte Formin FMNL1 Expression in Human Tissues

    PubMed Central

    Heuser, Vanina D.; Iljin, Kristiina; Kampf, Caroline; Uhlen, Mathias; Carpén, Olli

    2014-01-01

    Formins are cytoskeleton regulating proteins characterized by a common FH2 structural domain. As key players in the assembly of actin filaments, formins direct dynamic cytoskeletal processes that influence cell shape, movement and adhesion. The large number of formin genes, fifteen in the human, suggests distinct tasks and expression patterns for individual family members, in addition to overlapping functions. Several formins have been associated with invasive cell properties in experimental models, linking them to cancer biology. One example is FMNL1, which is considered to be a leukocyte formin and is known to be overexpressed in lymphomas. Studies on FMNL1 and many other formins have been hampered by a lack of research tools, especially antibodies suitable for staining paraffin-embedded formalin-fixed tissues. Here we characterize, using bioinformatics tools and a validated antibody, the expression pattern of FMNL1 in human tissues and study its subcellular distribution. Our results indicate that FMNL1 expression is not restricted to hematopoietic tissues and that neoexpression of FMNL1 can be seen in epithelial cancer. PMID:24700756

  20. Childhood Physical and Sexual Abuse History and Leukocyte Telomere Length among Women in Middle Adulthood

    PubMed Central

    Mason, Susan M.; Prescott, Jennifer; Tworoger, Shelley S.; DeVivo, Immaculata; Rich-Edwards, Janet W.

    2015-01-01

    Objective Abuse victimization in childhood is associated with a variety of age-related cardiometabolic diseases, but the mechanisms remain unknown. Telomeres, which form the protective caps at the ends of chromosomes, have been proposed as measures of biological age, and a growing body of research suggests that telomere attrition may help to explain relationships between stress and cardiometabolic degradation. We examined the association between childhood abuse victimization and leukocyte telomere length among 1,135 participants in the Nurses’ Health Study II (NHSII). Methods The NHSII ascertained physical and sexual child abuse histories in 2001. Telomere length was measured in genomic DNA extracted from peripheral blood leukocytes collected between 1996 and 1999. The ratio of telomere repeat copy number to a single gene copy number (T/S) was determined by a modified version of the quantitative real-time PCR telomere assay. Telomere length was log-transformed and corrected for assay variation across batch. We regressed telomere length on childhood abuse exposure variables and covariates using linear regression. Results We observed a reduction in telomere length associated with moderate physical abuse versus no physical abuse, but there was no evidence of a dose-response relationship for increased severity of physical abuse. No associations were noted for sexual abuse. Conclusions We found no evidence of an association between severity of childhood physical or sexual abuse and leukocyte telomere length in the NHSII. PMID:26053088

  1. Production of tumor necrosis factor alpha in human leukocytes stimulated by Cryptococcus neoformans.

    PubMed Central

    Levitz, S M; Tabuni, A; Kornfeld, H; Reardon, C C; Golenbock, D T

    1994-01-01

    Tumor necrosis factor alpha (TNF-alpha) is a key mediator of inflammation and may promote human immunodeficiency virus replication in latently infected cells. Since cryptococcosis often is associated with aberrations in the host inflammatory response and occurs preferentially in persons with AIDS, we defined the conditions under which human leukocytes produce TNF-alpha when stimulated by Cryptococcus neoformans. Peripheral blood mononuclear cells (PBMC) produced comparable amounts of TNF-alpha following stimulation with C. neoformans and lipopolysaccharide. Detectable TNF-alpha release in response to C. neoformans occurred only when fungi with small-sized capsules were used and complement-sufficient serum was added. Fractionation of PBMC established that monocytes were the predominant source of TNF-alpha. TNF-alpha gene expression and release occurred significantly later in PBMC stimulated with C. neoformans than in PBMC stimulated with LPS. C. neoformans was also a potent inducer of TNF-alpha from freshly isolated bronchoalveolar macrophages (BAM). Upon in vitro culture, BAM and monocytes bound greater numbers of fungal cells, yet their capacity to produce TNF-alpha following cryptococcal stimulation declined by 74 to 100%. However, this decline was reversed if the BAM and monocytes were cultured with gamma interferon. These data establish that C. neoformans can potently stimulate TNF-alpha release from human leukocytes. However, several variables profoundly affected the amount of TNF-alpha released, including the type of leukocyte and its state of activation, the size of the cryptococcal capsule, and the availability of opsonins. PMID:8168965

  2. Theileria induces oxidative stress and HIF1α activation that are essential for host leukocyte transformation.

    PubMed

    Medjkane, S; Perichon, M; Marsolier, J; Dairou, J; Weitzman, J B

    2014-04-01

    Complex links between infection and cancer suggest that we still can learn much about tumorigenesis by studying how infectious agents hijack the host cell machinery. We studied the effects of an intracellular parasite called Theileria that infects bovine leukocytes and turns them into invasive cancer-like cells. We investigated the host cells pathways that are deregulated in infected leukocytes and might link infection and lymphoproliferative disease. We show that intracellular Theileria parasites drive a Warburg-like phenotype in infected host leukocytes, characterized by increased expression of metabolic regulators, increased glucose uptake and elevated lactate production, which were lost when the parasite was eliminated. The cohabitation of the parasites within the host cells leads to disruption of the redox balance (as measured by reduced/oxidized glutathione ratio) and elevated ROS (reactive oxygen species) levels, associated with chronic stabilization of the hypoxia-inducible factor 1 alpha (HIF1α). Inhibition of HIF1α (pharmacologically or genetically), or treatment with antioxidants, led to a marked reduction in expression of aerobic glycolytic genes and inhibited the transformed phenotype. These data show that stabilization of HIF1α, following increased ROS production, modulates host glucose metabolism and is critical for parasite-induced transformation. Our study expands knowledge about the molecular strategy used by the parasite Theileria to induce the transformed phenotypes of infected cells via reprogramming of glucose metabolism and redox signaling. PMID:23665677

  3. Myeloperoxidase-mediated activation of xenobiotics by human leukocytes.

    PubMed

    Hofstra, A H; Uetrecht, J P

    1993-10-01

    Peripheral blood leukocytes contain a variety of enzymes that are capable of metabolising xenobiotics. The enzyme myeloperoxidase (MPO) appears to be the most important for drug metabolism. MPO is a peroxidase/oxidase and generates the powerful oxidant hypochlorous acid. MPO- or MPO-generated oxidants are capable of oxidizing a wide variety of compounds and a broad range of functional groups, especially those that contain nitrogen and sulfur. Leukocytes have a role in immune response; therefore, reactive intermediates generated by leukocyte metabolism of xenobiotics may have a role in idiosyncratic drug reactions, particularly those that are immune-mediated such as drug-induced lupus or agranulocytosis. PMID:8236277

  4. Uptake of indium-111-labeled leukocytes by brain metastasis

    SciTech Connect

    Balachandran, S.; Husain, M.M.; Adametz, J.R.; Pallin, J.S.; Angtuaco, T.L.; Boyd, C.M.

    1987-04-01

    Uptake of indium-labeled leukocytes was seen in two cases of histologically proven brain metastasis. In one, this led to misdiagnosis of the lesion as an abscess. On histological evaluation, a large number of white blood cells or macrophages was seen at the neoplastic sites. Reasons for leukocyte accumulation around metastatic brain neoplasms are discussed. In contrast to the current reports that indium-labeled leukocyte scans can differentiate intracranial infection from tumor, these cases demonstrate their lack of specificity in the detection of brain abscess.

  5. Role of CXCL5 in leukocyte recruitment to the lungs during secondhand smoke exposure.

    PubMed

    Balamayooran, Gayathriy; Batra, Sanjay; Cai, Shanshan; Mei, Junjie; Worthen, G Scott; Penn, Arthur L; Jeyaseelan, Samithamby

    2012-07-01

    Chronic obstructive pulmonary disease (COPD) is the third leading cause of mortality in the United States. The major cause of COPD is cigarette smoking. Extensive leukocyte influx into the lungs, mediated by chemokines, is a critical event leading to COPD. Although both resident and myeloid cells secrete chemokines in response to inflammatory stimuli, little is known about the role of epithelial-derived chemokines, such as CXC chemokine ligand (CXCL)5, in the pathogenesis of cigarette smoke-induced inflammation. To explore the role of CXCL5, we generated CXCL5 gene-deficient mice and exposed them to secondhand smoke (SHS) for 5 hours/day for 5 days/week up to 3 weeks (subacute exposure). We observed a reduced recruitment of leukocytes to the lungs of CXCL5(-/-) mice compared with their wild-type (WT) counterparts, and noted that macrophages comprised the predominant leukocytes recruited to the lungs. Irradiation experiments performed on CXCL5(-/-) or WT mice transplanted with WT or CXCL5(-/-) bone marrow revealed that resident but not hematopoietic cell-driven CXCL5 is important for mediating SHS-induced lung inflammation. Interestingly, we observed a significant reduction of monocyte chemotactic protein-1 (MCP-1/CC chemokine ligand 2) concentrations in the lungs of CXCL5(-/-) mice. The instillation of recombinant MCP-1 in CXCL5(-/-) mice reversed macrophage recruitment. Our results also show the reduced activation of NF-κB/p65 in the lungs, as well as the attenuated activation of C-Jun N-terminal kinase, p42/44, and p38 mitogen-activated protein kinases and the expression of intercellular adhesion molecule-1 in the lungs of SHS-exposed CXCL5(-/-) mice. Our findings suggest an important role for CXCL5 in augmenting leukocyte recruitment in SHS-induced lung inflammation, and provide novel insights into CXCL5-driven pathogenesis. PMID:22362385

  6. [Comparative transcriptome analysis of human aorta atherosclerotic lesions and peripheral blood leukocytes from essential hypertension patients].

    PubMed

    Timofeeva, A V; Goriunova, L E; Khaspekov, G L; Il'inskaia, O P; Sirotkin, V N; Andreeva, E R; Tararak, E M; Bulkina, O S; Buza, V V; Britareva, V V; Karpov, Iu A; Bibilashvili, R Sh

    2009-01-01

    One of the major cardiovascular risk factor which predisposes to and accelerates atherosclerosis is arterial hypertension (AH). To determine the molecular basis of the crosslink between AH and atherosclerosis for the development of new treatment strategies large-scale transcriptome analysis of the cells implicated in atherogenesis is needed. We used cDNA microarray technique for simultaneous analysis of gene expression in human abdominal aorta normal sites and atherosclerotic lesions of different histological types, as well as in peripheral blood leukocytes from patients with essential hypertension (EH) and donors. The microarray data were verified by quantitative RT-PCR (reverse transcription coupled with polymerase chain reaction) and immunohistochemical analysis. Differential expression of 40 genes has been found, among which twenty two genes demonstrated up-regulation and 18 genes demonstrated down-regulation in atherosclerotic aorta compared with normal vessel. New gene-candidates, implicated in atherogenesis, have been identified - FPRL2, CD37, CD53, RGS1, LCP1, SPI1, CTSA, EPAS1, FHL1, GEM, RHOB, SPARCL1, ITGA8, PLN, and COL14A1. These genes participate in cell migration and adhesion, phenotypic changes of smooth muscle cells, immune and inflammatory reactions, oxidative processes and extracellular matrix remodeling. We have found increased expression levels of CD53, SPI1, FPRL2, SPP1, CTSD, ACP5, LCP1, CTSA and LIPA genes in peripheral blood leukocytes from EH patients and in atherosclerotic lesions of human aorta. The majority of these genes significantly (p<0.005) positively (r>0.5) correlated with AH stage as well as with histological grading of atherosclerotic lesions. PMID:19772500

  7. Influence of Magnetite Nanoparticles on Human Leukocyte Activity

    NASA Astrophysics Data System (ADS)

    Džarová, Anežka; Dubničková, Martina; Závišová, Vlasta; Koneracká, Martina; Kopčanský, Peter; Gojzewski, Hubert; Timko, Milan

    2010-12-01

    Chemically synthesized magnetite particles coated by sodium oleate and PEG (MNP), and magnetosomes (MS) influence the process of phagocytosis and the metabolic activity (lysozyme and peroxidase activity) in leukocytes. Lysozyme activity is oxygen-independent liquidation mechanisms of engulfed microorganism, peroxidase activity is an oxygen-dependent mechanism. Both tested types of nanoparticles lysed leukocyte cells during incubation. MNP at concentrations of 10 and 20 μg/mL lysed almost all leukocytes and their cell viability was in the 14±0.05% range. On the other hand MS begin to influence leukocytes activity at the concentration of 1 μg/ml and this influence grows with increasing concentration up to 20 μg/ml. MS are more suitable for biological applications than MNP which are more aggressive material than MS. MS should not be used above 10 μg/mL.

  8. Osteomyelitis complicating fracture: pitfalls of /sup 111/In leukocyte scintigraphy

    SciTech Connect

    Kim, E.E.; Pjura, G.A.; Lowry, P.A.; Gobuty, A.H.; Traina, J.F.

    1987-05-01

    /sup 111/In-labeled leukocyte imaging has shown greater accuracy and specificity than alternative noninvasive methods in the detection of uncomplicated osteomyelitis. Forty patients with suspected osteomyelitis complicating fractures (with and without surgical intervention) were evaluated with /sup 111/In-labeled leukocytes. All five patients with intense focal uptake, but only one of 13 with no uptake, had active osteomyelitis. However, mild to moderate /sup 111/In leukocyte uptake, observed in 22 cases, indicated the presence of osteomyelitis in only four of these; the other false-positive results were observed in noninfected callus formation, heterotopic bone formation, myositis ossificans, and sickle-cell disease. These results suggest that /sup 111/In-labeled leukocyte imaging is useful for the evaluation of suspected osteomyelitis complicating fracture but must be used in conjunction with clinical and radiographic correlation to avoid false-positive results.

  9. Clinical imaging with indium-111 leukocytes: uptake in bowel infarction

    SciTech Connect

    Gray, H.W.; Cuthbert, I.; Richards, J.R.

    1981-08-01

    Leukocytes labeled with indium-111 accumulated in an area of small-bowel infarction, mimicking a paracolic abscess. Evidence of subacute bowel obstruction should alert the nuclear medicine physician to the former possibility.

  10. Neutrophil Leukocyte: Combustive Microbicidal Action and Chemiluminescence

    PubMed Central

    Allen, Robert C.

    2015-01-01

    Neutrophil leukocytes protect against a varied and complex array of microbes by providing microbicidal action that is simple, potent, and focused. Neutrophils provide such action via redox reactions that change the frontier orbitals of oxygen (O2) facilitating combustion. The spin conservation rules define the symmetry barrier that prevents direct reaction of diradical O2 with nonradical molecules, explaining why combustion is not spontaneous. In burning, the spin barrier is overcome when energy causes homolytic bond cleavage producing radicals capable of reacting with diradical O2 to yield oxygenated radical products that further participate in reactive propagation. Neutrophil mediated combustion is by a different pathway. Changing the spin quantum state of O2 removes the symmetry restriction to reaction. Electronically excited singlet molecular oxygen (1O2*) is a potent electrophilic reactant with a finite lifetime that restricts its radius of reactivity and focuses combustive action on the target microbe. The resulting exergonic dioxygenation reactions produce electronically excited carbonyls that relax by light emission, that is, chemiluminescence. This overview of neutrophil combustive microbicidal action takes the perspectives of spin conservation and bosonic-fermionic frontier orbital considerations. The necessary principles of particle physics and quantum mechanics are developed and integrated into a fundamental explanation of neutrophil microbicidal metabolism. PMID:26783542

  11. Production of monoclonal antibodies against canine leukocytes.

    PubMed

    Aguiar, Paulo Henrique Palis; Borges dos Santos, Roberto Robson; Lima, Carla Andrade; Rios de Sousa Gomes, Hilton; Larangeira, Daniela Farias; Santos, Patrícia Meira; Barrouin-Melo, Stella Maria; Conrado dos-Santos, Washington Luis; Pontes-de-Carvalho, Lain

    2004-04-01

    A panel of anti-canine leukocyte monoclonal antibodies (MAbs) was produced by immunizing BALB/c mice with canine peripheral blood mononuclear cells (PBMC), either resting or stimulated with concanavalin A (ConA). Three out of 28 clones-IH1, AB6, and HG6-screened by ELISA and producing antibody with the highest specificity for canine cell immunostaining, were subjected to three subsequent subcloning steps by limiting dilution, and selected for further characterization. These MAbs belonged to IgG1 (HG6 and IH1) and IgG2a (AB6) isotypes. The distribution of cell populations expressing the antigen recognized by the antibodies was identified by indirect immunoflorescence on canine PBMC and on tissue sections of lymph node, spleen, liver and skin. The possible crossreactivity with human PBMC was also examined in immunocytochemistry. One of the antibodies specifically recognized macrophages. The MAbs presented here can be foreseen as possible valuable diagnostic and research tools to study immune functions in dogs. PMID:15165486

  12. [A method for detecting bovine leukocyte adhesion deficiency in Holstein calves using polymerase chain reaction-single strand conformation polymorphism analysis].

    PubMed

    Li, Yan-Hua; Zhang, Sheng-Li; Han, Guang-Wen; Xue, Jian-Hua; Zhao, Peng

    2007-12-01

    Bovine leukocyte adhesion deficiency (BLAD) is an autosomal recessive disease caused by A-->G mutation in the CD18 gene. The disease is characterized by greatly reduced expression of the heterodimeric beta2 integrin adhesion molecules on leukocytes, resulting in multiple defects in leukocyte function and significant deficits on performance traits in Holstein cattle. In this study, primers were designed to amplify the CD18 gene fragment and BLAD was detected by PCR-SSCP in a simple, highly accurate and high throughput manner. Results were confirmed by DNA sequencing. This study provides a more reliable and useful method for extensive screening of BLAD and also offers a theoretical basis for molecular diagnosis in Holstein calves. PMID:18065382

  13. Lineage specific transcriptional regulation of DICER by MITF in melanocytes

    PubMed Central

    Levy, Carmit; Khaled, Mehdi; Robinson, Kathleen C.; Veguilla, Rosa A.; Chen, Po-Hao; Yokoyama, Satoru; Makino, Eiichi; Lu, Jun; Larue, Lionel; Beermann, Friedrich; Chin, Lynda; Bosenberg, Marcus; Song, Jun. S.; Fisher, David E.

    2010-01-01

    Summary DICER is a central regulator of microRNA maturation. However little is known about mechanisms regulating its expression in development or disease. While profiling miRNA expression in differentiating melanocytes, two populations were observed: some upregulated at the pre-miRNA stage, and others upregulated as “mature” miRNAs (with stable pre-miRNA levels). Conversion of pre-miRNAs to fully processed miRNAs appeared to be dependent upon stimulation of DICER expression—an event found to occur via direct transcriptional targeting of DICER by the melanocyte master transcriptional regulator MITF. MITF binds and activates a conserved regulatory element upstream of DICER’s transcriptional start site upon melanocyte differentiation. Targeted KO of DICER is lethal to melanocytes, at least partly via DICER-dependent processing of the pre-miRNA-17~92 cluster thus targeting BIM, a known pro-apoptotic regulator of melanocyte survival. These observations highlight a central mechanism underlying miRNA regulation which could exist for other cell types during development. PMID:20550935

  14. Nephric lineage specification by Pax2 and Pax8.

    PubMed

    Bouchard, Maxime; Souabni, Abdallah; Mandler, Markus; Neubüser, Annette; Busslinger, Meinrad

    2002-11-15

    The mammalian kidney develops in three successive steps from the initial pronephros via the mesonephros to the adult metanephros. Although the nephric lineage is specified during pronephros induction, no single regulator, including the transcription factor Pax2 or Pax8, has yet been identified to control this initial phase of kidney development. In this paper, we demonstrate that mouse embryos lacking both Pax2 and Pax8 are unable to form the pronephros or any later nephric structures. In these double-mutant embryos, the intermediate mesoderm does not undergo the mesenchymal-epithelial transitions required for nephric duct formation, fails to initiate the kidney-specific expression of Lim1 and c-Ret, and is lost by apoptosis 1 d after failed pronephric induction. Conversely, retroviral misexpression of Pax2 was sufficient to induce ectopic nephric structures in the intermediate mesoderm and genital ridge of chick embryos. Together, these data identify Pax2 and Pax8 as critical regulators that specify the nephric lineage. PMID:12435636

  15. Lineage-specific biomarkers predict response to FGFR inhibition.

    PubMed

    Loch, David C; Pollock, Pamela M

    2012-12-01

    In this issue of Cancer Discovery, Guagnano and colleagues use a large and diverse annotated collection of cancer cell lines, the Cancer Cell Line Encyclopedia, to correlate whole-genome expression and genomic alteration datasets with cell line sensitivity data to the novel pan-fibroblast growth factor receptor (FGFR) inhibitor NVP-BGJ398. Their findings underscore not only the preclinical use of such cell line panels in identifying predictive biomarkers, but also the emergence of the FGFRs as valid therapeutic targets, across an increasingly broad range of malignancies. PMID:23230185

  16. CD99 is essential for leukocyte diapedesis in vivo.

    PubMed

    Dufour, Eric M; Deroche, Alana; Bae, Youngmee; Muller, William A

    2008-11-01

    Recruitment of leukocytes into inflamed tissue requires migration of leukocytes from the blood stream across the endothelial lining and the basement membrane of the local blood vessels. CD99 in humans is a 32-kDa highly O-glycosylated cell surface protein expressed on most leukocytes. The authors recently found CD99 to be expressed in leukocytes and at human endothelial cell contacts. Human CD99 is involved in homophilic interaction between the two cell types and participates in the transendothelial migration of monocytes and polymorphonuclear neutrophils (PMNs) in vitro. To test the role of CD99 in vivo, the authors cloned murine CD99 (muCD99), expressed it in vitro, and generated a blocking monoclonal antibody against it. We first showed that muCD99 is expressed on mouse leukocytes as well as enriched at the endothelial cell borders. Transfection of cells with muCD99 imparts on them the ability to aggregate in a CD99-dependent homophilic manner. Cells expressing muCD99 did not bind to cells expressing murine or human platelet endothelial call adhesion molecule (PECAM) or human CD99. In the thioglycollate peritonitis model of inflammation, anti-CD99 monoclonal antibody blocked the recruitment of neutrophils and monocytes by over 40% and 80%, respectively, at 18 h. Microscopy showed that this blocking occurred at the luminal surface of venules. The authors conclude that CD99 plays a major role in the emigration of leukocytes in vivo. PMID:18923973

  17. Binding of Pasteurella haemolytica leukotoxin to bovine leukocytes.

    PubMed Central

    Brown, J F; Leite, F; Czuprynski, C J

    1997-01-01

    Pasteurella haemolytica is the principal bacterial pathogen in the bovine respiratory disease complex. This organism produces an exotoxin (referred to as leukotoxin) during logarithmic-phase growth that is a potent leukocyte-modulating agent. At low concentrations, it activates neutrophils and mononuclear phagocytes to release inflammatory mediators, while at the same time making these cells destined to undergo apoptotic cell death. At higher concentrations, the toxin causes rapid swelling and loss of cell viability. In this study, we demonstrated that toxin binding can be directly evaluated by flow cytometry with biologically active biotinylated leukotoxin. Leukotoxin binding was blocked by the addition of a neutralizing anti-leukotoxin monoclonal antibody and was not detected when bovine leukocytes were incubated with culture filtrates from a mutant strain of P. haemolytica that does not produce biologically active leukotoxin. In addition, treatment of bovine leukocytes with protease K eliminated subsequent binding of leukotoxin, suggesting that there is a protein on the leukocyte surface that is either a leukotoxin binding site or is required for stabilization of leukotoxin binding. We did not detect binding of biotinylated leukotoxin to porcine or human leukocytes, which have been reported previously to be resistant to the lytic effects of the leukotoxin. These findings suggest that there may be a specific binding site for P. haemolytica leukotoxin on bovine but not on porcine or human leukocytes and that it might be involved in the activation and lytic activities of the leukotoxin. PMID:9284143

  18. Airway distension promotes leukocyte recruitment in rat tracheal circulation.

    PubMed

    Lim, Lina H K; Wagner, Elizabeth M

    2003-11-01

    Mechanical distortion of blood vessels is known to activate endothelial cells. Whether airway distension likewise activates the vascular endothelium within the airway wall is unknown. Using intravital microscopy in the rat trachea, we investigated if airway distention with the application of positive end-expiratory pressure (PEEP) caused leukocyte recruitment to the airway. Tracheal postcapillary venules were visualized and leukocyte kinetics monitored in anesthetized, mechanically ventilated rats (80 breaths/minute, 6 ml/kg VT, 1 cm H(2)O PEEP). Leukocyte rolling velocity (Vwbc) and the number of adherent cells were not altered with normal ventilation over the course of 2 hours. Ventilation with sustained PEEP (8 cm H(2)O for 1 hour reduced Vwbc and increased adhesion, reaching a maximum at 1 hour of PEEP. Intermittent (2x and 5x) 8 cm H(2)O PEEP also induced a similar reduction in Vwbc, accompanied by an increase in adhesion. However, leukocyte recruitment after airway distension is localized to the airways because increased PEEP did not induce leukocyte recruitment in the mesenteric microcirculation or when PEEP was applied to the lung distal to the site of measurement. Pretreatment with endothelin receptor and selectin inhibitors blocked the effects of distension on leukocyte recruitment, suggesting their involvement in the proinflammatory response. PMID:12869357

  19. Physiological levels of testosterone kill salmonid leukocytes in vitro

    USGS Publications Warehouse

    Slater, C.H.; Schreck, C.B.

    1997-01-01

    Adult spring chinook salmon (Oncorhynchus tshawytscha) elaborate high plasma concentrations of testosterone during sexual maturation, and these levels of testosterone have been shown to reduce the salmonid immune response in vitro. Our search for the mechanism of testosterone's immunosuppressive action has led to the characterization of an androgen receptor in salmonid leukocytes. In the present study we examined the specific effects that testosterone had on salmonid leukocytes. Direct counts of viable leukocytes after incubation with and without physiological levels of testosterone demonstrate a significant loss of leukocytes in cultures exposed to testosterone. At least 5 days of contact with testosterone was required to produce significant immunosuppression and addition of a 'conditioned media' (supernatant from proliferating lymphocytes not exposed to testosterone) did not reverse the immunosuppressive effects of testosterone. These data lead us to conclude that testosterone may exert its immunosuppressive effects by direct action on salmonid leukocytes, through the androgen receptor described, and that this action leads to the death of a significant number of these leukocytes.

  20. Conserved globulin gene across eight grass genomes identify fundamental units of the loci encoding seed storage proteins.

    PubMed

    Gu, Yong Qiang; Wanjugi, Humphrey; Coleman-Derr, Devin; Kong, Xiuying; Anderson, Olin D

    2010-03-01

    The wheat high molecular weight (HMW) glutenins are important seed storage proteins that determine bread-making quality in hexaploid wheat (Triticum aestivum). In this study, detailed comparative sequence analyses of large orthologous HMW glutenin genomic regions from eight grass species, representing a wide evolutionary history of grass genomes, reveal a number of lineage-specific sequence changes. These lineage-specific changes, which resulted in duplications, insertions, and deletions of genes, are the major forces disrupting gene colinearity among grass genomes. Our results indicate that the presence of the HMW glutenin gene in Triticeae genomes was caused by lineage-specific duplication of a globulin gene. This tandem duplication event is shared by Brachypodium and Triticeae genomes, but is absent in rice, maize, and sorghum, suggesting the duplication occurred after Brachypodium and Triticeae genomes diverged from the other grasses ~35 Ma ago. Aside from their physical location in tandem, the sequence similarity, expression pattern, and conserved cis-acting elements responsible for endosperm-specific expression further support the paralogous relationship between the HMW glutenin and globulin genes. While the duplicated copy in Brachypodium has apparently become nonfunctional, the duplicated copy in wheat has evolved to become the HMW glutenin gene by gaining a central prolamin repetitive domain. PMID:19707805

  1. C-type natriuretic peptide inhibits leukocyte recruitment and platelet-leukocyte interactions via suppression of P-selectin expression

    PubMed Central

    Scotland, Ramona S.; Cohen, Marc; Foster, Paul; Lovell, Matthew; Mathur, Anthony; Ahluwalia, Amrita; Hobbs, Adrian J.

    2005-01-01

    The multifaceted process of immune cell recruitment to sites of tissue injury is key to the development of an inflammatory response and involved in the pathogenesis of numerous cardiovascular disorders. We recently identified C-type natriuretic peptide (CNP) as an important endothelium-derived mediator that regulates vascular tone and protects against myocardial ischemia/reperfusion injury. Herein, we investigated whether CNP inhibits leukocyte recruitment and platelet aggregation and thereby exerts a potential antiinflammatory influence on the blood vessel wall. We assessed the effects of CNP on leukocyte-endothelial cell interactions in mouse mesenteric postcapillary venules in vivo in animals with high basal leukocyte activation (endothelial nitric oxide synthase knockout mice, eNOS-/-) or under acute inflammatory conditions (induced by interleukin-1β or histamine). CNP suppressed basal leukocyte rolling in eNOS-/- mice in a rapid, reversible, and concentration-dependent manner. These effects of CNP were mimicked by the selective natriuretic peptide receptor-C agonist cANF4-23. CNP also suppressed leukocyte rolling induced by IL-1β or histamine, inhibited platelet-leukocyte interactions, and prevented thrombin-induced platelet aggregation of human blood. Furthermore, analysis of human umbilical vein endothelial cells, leukocytes, and platelets revealed that CNP selectively attenuates expression of P-selectin. Thus, CNP is a modulator of acute inflammation in the blood vessel wall characterized by leukocyte and platelet activation. These antiinflammatory effects appear to be mediated, at least in part, via suppression of P-selectin expression. These observations suggest that endothelial CNP might maintain an anti-atherogenic influence on the blood vessel wall and represent a target for therapeutic intervention in inflammatory cardiovascular disorders. PMID:16179391

  2. C-type natriuretic peptide inhibits leukocyte recruitment and platelet-leukocyte interactions via suppression of P-selectin expression

    NASA Astrophysics Data System (ADS)

    Scotland, Ramona S.; Cohen, Marc; Foster, Paul; Lovell, Matthew; Mathur, Anthony; Ahluwalia, Amrita; Hobbs, Adrian J.

    2005-10-01

    The multifaceted process of immune cell recruitment to sites of tissue injury is key to the development of an inflammatory response and involved in the pathogenesis of numerous cardiovascular disorders. We recently identified C-type natriuretic peptide (CNP) as an important endothelium-derived mediator that regulates vascular tone and protects against myocardial ischemia/reperfusion injury. Herein, we investigated whether CNP inhibits leukocyte recruitment and platelet aggregation and thereby exerts a potential antiinflammatory influence on the blood vessel wall. We assessed the effects of CNP on leukocyte-endothelial cell interactions in mouse mesenteric postcapillary venules in vivo in animals with high basal leukocyte activation (endothelial nitric oxide synthase knockout mice, eNOS-/-) or under acute inflammatory conditions (induced by interleukin-1 or histamine). CNP suppressed basal leukocyte rolling in eNOS-/- mice in a rapid, reversible, and concentration-dependent manner. These effects of CNP were mimicked by the selective natriuretic peptide receptor-C agonist cANF4-23. CNP also suppressed leukocyte rolling induced by IL-1 or histamine, inhibited platelet-leukocyte interactions, and prevented thrombin-induced platelet aggregation of human blood. Furthermore, analysis of human umbilical vein endothelial cells, leukocytes, and platelets revealed that CNP selectively attenuates expression of P-selectin. Thus, CNP is a modulator of acute inflammation in the blood vessel wall characterized by leukocyte and platelet activation. These antiinflammatory effects appear to be mediated, at least in part, via suppression of P-selectin expression. These observations suggest that endothelial CNP might maintain an anti-atherogenic influence on the blood vessel wall and represent a target for therapeutic intervention in inflammatory cardiovascular disorders. endothelium | natriuretic peptide receptor type C | atherosclerosis | thrombosis

  3. Rescue from acute neuroinflammation by pharmacological chemokine-mediated deviation of leukocytes

    PubMed Central

    2012-01-01

    Background Neutrophil influx is an important sign of hyperacute neuroinflammation, whereas the entry of activated lymphocytes into the brain parenchyma is a hallmark of chronic inflammatory processes, as observed in multiple sclerosis (MS) and its animal models of experimental autoimmune encephalomyelitis (EAE). Clinically approved or experimental therapies for neuroinflammation act by blocking leukocyte penetration of the blood brain barrier. However, in view of unsatisfactory results and severe side effects, complementary therapies are needed. We have examined the effect of chlorite-oxidized oxyamylose (COAM), a potent antiviral polycarboxylic acid on EAE. Methods EAE was induced in SJL/J mice by immunization with spinal cord homogenate (SCH) or in IFN-γ-deficient BALB/c (KO) mice with myelin oligodendrocyte glycoprotein peptide (MOG35-55). Mice were treated intraperitoneally (i.p.) with COAM or saline at different time points after immunization. Clinical disease and histopathology were compared between both groups. IFN expression was analyzed in COAM-treated MEF cell cultures and in sera and peritoneal fluids of COAM-treated animals by quantitative PCR, ELISA and a bioassay on L929 cells. Populations of immune cell subsets in the periphery and the central nervous system (CNS) were quantified at different stages of disease development by flow cytometry and differential cell count analysis. Expression levels of selected chemokine genes in the CNS were determined by quantitative PCR. Results We discovered that COAM (2 mg i.p. per mouse on days 0 and 7) protects significantly against hyperacute SCH-induced EAE in SJL/J mice and MOG35-55-induced EAE in IFN-γ KO mice. COAM deviated leukocyte trafficking from the CNS into the periphery. In the CNS, COAM reduced four-fold the expression levels of the neutrophil CXC chemokines KC/CXCL1 and MIP-2/CXCL2. Whereas the effects of COAM on circulating blood and splenic leukocytes were limited, significant alterations were

  4. Evolution in the Fast Lane: Rapidly Evolving Sex-Related Genes in Drosophila

    PubMed Central

    Haerty, Wilfried; Jagadeeshan, Santosh; Kulathinal, Rob J.; Wong, Alex; Ravi Ram, Kristipati; Sirot, Laura K.; Levesque, Lisa; Artieri, Carlo G.; Wolfner, Mariana F.; Civetta, Alberto; Singh, Rama S.

    2007-01-01

    A large portion of the annotated genes in Drosophila melanogaster show sex-biased expression, indicating that sex and reproduction-related genes (SRR genes) represent an appreciable component of the genome. Previous studies, in which subsets of genes were compared among few Drosophila species, have found that SRR genes exhibit unusual evolutionary patterns. Here, we have used the newly released genome sequences from 12 Drosophila species, coupled to a larger set of SRR genes, to comprehensively test the generality of these patterns. Among 2505 SRR genes examined, including ESTs with biased expression in reproductive tissues and genes characterized as involved in gametogenesis, we find that a relatively high proportion of SRR genes have experienced accelerated divergence throughout the genus Drosophila. Several testis-specific genes, male seminal fluid proteins (SFPs), and spermatogenesis genes show lineage-specific bursts of accelerated evolution and positive selection. SFP genes also show evidence of lineage-specific gene loss and/or gain. These results bring us closer to understanding the details of the evolutionary dynamics of SRR genes with respect to species divergence. PMID:18039869

  5. Insulin radioreceptor assay on murine splenic leukocytes and peripheral erythrocytes

    SciTech Connect

    Shimizu, F.; Kahn, R.

    1982-02-01

    Insulin radioreceptor assays were developed using splenic leukocytes and peripheral erythrocytes from individual mice. Splenic leukocytes were prepared using an NH/sub 4/Cl buffer which did not alter insulin binding, but gave much higher yields than density gradient methods. Mouse erythrocytes were isolated from heparinized blood by three passages over a Boyum gradient, and a similar buffer was used to separate cells from free (/sup 125/I)iodoinsulin at the end of the binding incubation. Insulin binding to both splenic leukocytes and peripheral erythrocytes had typical pH, temperature, and time dependencies, and increased linearly with an increased number of cells. Optimal conditions for the splenic leukocytes (6 x 10/sup 7//ml) consisted of incubation with (/sup 125/I)iodoinsulin at 15 C for 2 h in Hepes buffer, pH 8.0. In cells from 20 individual mice, the specific (/sup 125/I)iodoinsulin binding was 2.6 +/- 0.1% (SEM), and nonspecific binding was 0.3 +/- 0.04% (10.6% of total binding). Erythrocytes (2.8 x 10/sup 9//ml) were incubated with (/sup 125/)iodoinsulin at 15 C for 2 h in Hepes buffer, pH 8.2. In cells from 25 individual mice, the specific (/sup 125/I)iodoinsulin binding was 4.5 +/- 0.2%, and nonspecific binding was 0.7 +/- 0.03% (13.6% of total binding). In both splenic leukocytes and peripheral erythrocytes, analysis of equilibrium binding data produced curvilinear Scatchard plots with approximately 3500 binding sites/leukocyte and 20 binding sites/erythrocyte. These data demonstrate that adequate numbers of splenic leukocytes and peripheral erythrocytes can be obtained from individual mice to study insulin binding in a precise and reproducible manner.

  6. Thyrotropin Receptor Epitope and Human Leukocyte Antigen in Graves' Disease.

    PubMed

    Inaba, Hidefumi; De Groot, Leslie J; Akamizu, Takashi

    2016-01-01

    Graves' disease (GD) is an organ-specific autoimmune disease, and thyrotropin (TSH) receptor (TSHR) is a major autoantigen in this condition. Since the extracellular domain of human TSHR (TSHR-ECD) is shed into the circulation, TSHR-ECD is a preferentially immunogenic portion of TSHR. Both genetic factors and environmental factors contribute to development of GD. Inheritance of human leukocyte antigen (HLA) genes, especially HLA-DR3, is associated with GD. TSHR-ECD protein is endocytosed into antigen-presenting cells (APCs), and processed to TSHR-ECD peptides. These peptide epitopes bind to HLA-class II molecules, and subsequently the complex of HLA-class II and TSHR-ECD epitope is presented to CD4+ T cells. The activated CD4+ T cells secrete cytokines/chemokines that stimulate B-cells to produce TSAb, and in turn hyperthyroidism occurs. Numerous studies have been done to identify T- and B-cell epitopes in TSHR-ECD, including (1) in silico, (2) in vitro, (3) in vivo, and (4) clinical experiments. Murine models of GD and HLA-transgenic mice have played a pivotal role in elucidating the immunological mechanisms. To date, linear or conformational epitopes of TSHR-ECD, as well as the molecular structure of the epitope-binding groove in HLA-DR, were reported to be related to the pathogenesis in GD. Dysfunction of central tolerance in the thymus, or in peripheral tolerance, such as regulatory T cells, could allow development of GD. Novel treatments using TSHR antagonists or mutated TSHR peptides have been reported to be effective. We review and update the role of immunogenic TSHR epitopes and HLA in GD, and offer perspectives on TSHR epitope specific treatments. PMID:27602020

  7. Thyrotropin Receptor Epitope and Human Leukocyte Antigen in Graves’ Disease

    PubMed Central

    Inaba, Hidefumi; De Groot, Leslie J.; Akamizu, Takashi

    2016-01-01

    Graves’ disease (GD) is an organ-specific autoimmune disease, and thyrotropin (TSH) receptor (TSHR) is a major autoantigen in this condition. Since the extracellular domain of human TSHR (TSHR-ECD) is shed into the circulation, TSHR-ECD is a preferentially immunogenic portion of TSHR. Both genetic factors and environmental factors contribute to development of GD. Inheritance of human leukocyte antigen (HLA) genes, especially HLA-DR3, is associated with GD. TSHR-ECD protein is endocytosed into antigen-presenting cells (APCs), and processed to TSHR-ECD peptides. These peptide epitopes bind to HLA-class II molecules, and subsequently the complex of HLA-class II and TSHR-ECD epitope is presented to CD4+ T cells. The activated CD4+ T cells secrete cytokines/chemokines that stimulate B-cells to produce TSAb, and in turn hyperthyroidism occurs. Numerous studies have been done to identify T- and B-cell epitopes in TSHR-ECD, including (1) in silico, (2) in vitro, (3) in vivo, and (4) clinical experiments. Murine models of GD and HLA-transgenic mice have played a pivotal role in elucidating the immunological mechanisms. To date, linear or conformational epitopes of TSHR-ECD, as well as the molecular structure of the epitope-binding groove in HLA-DR, were reported to be related to the pathogenesis in GD. Dysfunction of central tolerance in the thymus, or in peripheral tolerance, such as regulatory T cells, could allow development of GD. Novel treatments using TSHR antagonists or mutated TSHR peptides have been reported to be effective. We review and update the role of immunogenic TSHR epitopes and HLA in GD, and offer perspectives on TSHR epitope specific treatments. PMID:27602020

  8. Association of the bovine leukocyte antigen major histocompatibility complex class II DRB3*4401 allele with host resistance to the Lone Star Tick, Amblyomma americanum

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The MHC of cattle, known as the bovine leukocyte antigen (BoLA) complex, plays an integral role in disease and parasite susceptibility, and immune responsiveness of the host. While susceptibility to tick infestation in cattle is believed to be heritable, genes that may be responsible for the manife...

  9. Crossing the Vascular Wall: Common and Unique Mechanisms Exploited by Different Leukocyte Subsets during Extravasation

    PubMed Central

    Schnoor, Michael; Alcaide, Pilar; Voisin, Mathieu-Benoit; van Buul, Jaap D.

    2015-01-01

    Leukocyte extravasation is one of the essential and first steps during the initiation of inflammation. Therefore, a better understanding of the key molecules that regulate this process may help to develop novel therapeutics for treatment of inflammation-based diseases such as atherosclerosis or rheumatoid arthritis. The endothelial adhesion molecules ICAM-1 and VCAM-1 are known as the central mediators of leukocyte adhesion to and transmigration across the endothelium. Engagement of these molecules by their leukocyte integrin receptors initiates the activation of several signaling pathways within both leukocytes and endothelium. Several of such events have been described to occur during transendothelial migration of all leukocyte subsets, whereas other mechanisms are known only for a single leukocyte subset. Here, we summarize current knowledge on regulatory mechanisms of leukocyte extravasation from a leukocyte and endothelial point of view, respectively. Specifically, we will focus on highlighting common and unique mechanisms that specific leukocyte subsets exploit to succeed in crossing endothelial monolayers. PMID:26568666

  10. DNA Methylation Analysis of HTR2A Regulatory Region in Leukocytes of Autistic Subjects.

    PubMed

    Hranilovic, Dubravka; Blazevic, Sofia; Stefulj, Jasminka; Zill, Peter

    2016-02-01

    Disturbed brain and peripheral serotonin homeostasis is often found in subjects with autism spectrum disorder (ASD). The role of the serotonin receptor 2A (HTR2A) in the regulation of central and peripheral serotonin homeostasis, as well as its altered expression in autistic subjects, have implicated the HTR2A gene as a major candidate for the serotonin disturbance seen in autism. Several studies, yielding so far inconclusive results, have attempted to associate autism with a functional SNP -1438 G/A (rs6311) in the HTR2A promoter region, while possible contribution of epigenetic mechanisms, such as DNA methylation, to HTR2A dysregulation in autism has not yet been investigated. In this study, we compared the mean DNA methylation within the regulatory region of the HTR2A gene between autistic and control subjects. DNA methylation was analysed in peripheral blood leukocytes using bisulfite conversion and sequencing of the HTR2A region containing rs6311 polymorphism. Autistic subjects of rs6311 AG genotype displayed higher mean methylation levels within the analysed region than the corresponding controls (P < 0.05), while there was no statistically significant difference for AA and GG carriers. Our study provides preliminary evidence for increased HTR2A promoter methylation in leukocytes of a portion of adult autistic subjects, indicating that epigenetic mechanisms might contribute to HTR2A dysregulation observed in individuals with ASD. PMID:26149086

  11. Age-related methylation profiles of equine blood leukocytes in the RNASEL locus.

    PubMed

    Ząbek, T; Semik, E; Szmatoła, T; Oklejewicz, B; Fornal, A; Bugno-Poniewierska, M

    2016-08-01

    Methylation profiles across three CpG islands of the RNASEL gene were determined in blood leukocyte samples of Anglo-Arabian and Hucul horses. Bisulfite sequencing revealed hypomethylated state of the RNASEL promoter coinciding with methylated CpG island placed inside the gene. Several CpG sites were identified for which the methylation state was influenced by DNA polymorphism. Two of them showed monoallelic methylation. One of the CpG sites revealed functional polymorphism. A number of partially methylated CpG sites have been observed in the promoter area of RNASEL, which were used for the comparison of breed- and age-related effects. Clone bisulfite sequencing of blood leukocyte samples collected at different ages from particular individuals of AA and HC breeds and, also, BSPCR sequencing of 50 samples of juvenile and old AA and HC horses revealed increased methylation in particular CpG sites during aging. The age-related heterogeneity of white blood cells was hypothesized as being one of the potential causes of observed variability of methylation profiles in the RNASEL promoter. PMID:26553552

  12. Extravasation of leukocytes in comparison to tumor cells

    PubMed Central

    Strell, Carina; Entschladen, Frank

    2008-01-01

    The multi-step process of the emigration of cells from the blood stream through the vascular endothelium into the tissue has been termed extravasation. The extravasation of leukocytes is fairly well characterized down to the molecular level, and has been reviewed in several aspects. Comparatively little is known about the extravasation of tumor cells, which is part of the hematogenic metastasis formation. Although the steps of the process are basically the same in leukocytes and tumor cells, i.e. rolling, adhesion, transmigration (diapedesis), the molecules that are involved are different. A further important difference is that leukocyte interaction with the endothelium changes the endothelial integrity only temporarily, whereas tumor cell interaction leads to an irreversible damage of the endothelial architecture. Moreover, tumor cells utilize leukocytes for their extravasation as linkers to the endothelium. Thus, metastasis formation is indirectly susceptible to localization signals that are literally specific for the immune system. We herein compare the extravasation of leukocytes and tumor cells with regard to the involved receptors and the localization signals that direct the cells to certain organs and sites of the body. PMID:19055814

  13. Platelets Guide Leukocytes to Their Sites of Extravasation

    PubMed Central

    Puhr-Westerheide, Daniel; Pörnbacher, Michaela; Lauber, Kirsten; Krombach, Fritz; Reichel, Christoph Andreas

    2016-01-01

    Effective immune responses require the directed migration of leukocytes from the vasculature to the site of injury or infection. How immune cells “find” their site of extravasation remains largely obscure. Here, we identified a previously unrecognized role of platelets as pathfinders guiding leukocytes to their exit points in the microvasculature: upon onset of inflammation, circulating platelets were found to immediately adhere at distinct sites in venular microvessels enabling these cellular blood components to capture neutrophils and, in turn, inflammatory monocytes via CD40-CD40L-dependent interactions. In this cellular crosstalk, ligation of PSGL-1 by P-selectin leads to ERK1/2 MAPK-dependent conformational changes of leukocyte integrins, which promote the successive extravasation of neutrophils and monocytes to the perivascular tissue. Conversely, blockade of this cellular partnership resulted in misguided, inefficient leukocyte responses. Our experimental data uncover a platelet-directed, spatiotemporally organized, multicellular crosstalk that is essential for effective trafficking of leukocytes to the site of inflammation. PMID:27152726

  14. Recent insights into endothelial control of leukocyte extravasation.

    PubMed

    Hordijk, Peter L

    2016-04-01

    In the process of leukocyte migration from the circulation across the vascular wall, the crosstalk with endothelial cells that line the blood vessels is essential. It is now firmly established that in endothelial cells important signaling events are initiated upon leukocyte adhesion that impinge on the regulation of cell-cell contact and control the efficiency of transendothelial migration. In addition, several external factors such as shear force and vascular stiffness were recently identified as important regulators of endothelial signaling and, consequently, leukocyte transmigration. Here, I review recent insights into endothelial signaling events that are linked to leukocyte migration across the vessel wall. In this field, protein phosphorylation and Rho-mediated cytoskeletal dynamics are still widely studied using increasingly sophisticated mouse models. In addition, activation of tyrosine phosphatases, changes in endothelial cell stiffness as well as different vascular beds have all been established as important factors in endothelial signaling and leukocyte transmigration. Finally, I address less-well-studied but interesting components in the endothelium that also control transendothelial migration, such as the ephrins and their Eph receptors, that provide novel insights in the complexity associated with this process. PMID:26794844

  15. Leukocyte transcriptome from chickens infected with avian pathogenic Escherichia coli identifies pathways associated with resistance

    PubMed Central

    Sandford, Erin E.; Orr, Megan; Shelby, Mandy; Li, Xianyao; Zhou, Huaijun; Johnson, Timothy J.; Kariyawasam, Subhashinie; Liu, Peng; Nolan, Lisa K.; Lamont, Susan J.

    2012-01-01

    Avian pathogenic Escherichia coli (APEC) causes colibacillosis, which is responsible for morbidity and mortality in chickens. Gene expression patterns have previously been demonstrated to differ between chicken populations that are resistant vs. susceptible to bacterial infection, but little is currently known about gene expression response to APEC. Increased understanding of gene expression patterns associated with resistance will facilitate genetic selection to increase resistance to APEC. Male broiler chicks were vaccinated at 2 weeks of age and challenged with APEC at 4 weeks of age. Peripheral blood leukocytes were collected at 1 and 5 day post-infection. Lesions on the liver, pericardium, and air sacs were used to assign a mild or severe pathology status to non-vaccinated, challenged chicks. Ten treatment groups were therefore generated with a priori factors of vaccination, challenge, day post-infection, and the a posteriori factor of pathology status. Global transcriptomic response was evaluated using the Agilent 44K chicken microarray. APEC infection resulted in more up-regulation than down-regulation of differentially expressed genes. Immune response and metabolic processes were enriched with differentially expressed genes. Although vaccination significantly reduced lesions in challenged bird, there was no detectable effect of vaccination on gene expression. This study investigated the transcriptomic differences in host responses associated with mild vs. severe pathology, in addition to the effects of vaccination and challenge, thus revealing genes and networks associated with response to APEC and providing a foundation for future studies on, and genetic selection for, genetic resistance to APEC. PMID:24371566

  16. Neutrophils Are Not Less Sensitive Than Other Blood Leukocytes to the Genomic Effects of Glucocorticoids

    PubMed Central

    Hirsch, Gaelle; Lavoie-Lamoureux, Anouk; Beauchamp, Guy; Lavoie, Jean-Pierre

    2012-01-01

    Background Neutrophils are generally considered less responsive to glucocorticoids compared to other inflammatory cells. The reported increase in human neutrophil survival mediated by these drugs partly supports this assertion. However, it was recently shown that dexamethasone exerts potent anti-inflammatory effects in equine peripheral blood neutrophils. Few comparative studies of glucocorticoid effects in neutrophils and other leukocytes have been reported and a relative insensitivity of neutrophils to these drugs could not be ruled out. Objective We assessed glucocorticoid-responsiveness in equine and human peripheral blood neutrophils and neutrophil-depleted leukocytes. Methods Blood neutrophils and neutrophil-depleted leukocytes were isolated from 6 healthy horses and 4 human healthy subjects. Cells were incubated for 5 h with or without LPS (100 ng/mL) alone or combined with hydrocortisone, prednisolone or dexamethasone (10−8 M and 10−6 M). IL-1β, TNF-α, IL-8, glutamine synthetase and GR-α mRNA expression was quantified by qPCR. Equine neutrophils were also incubated for 20 h with or without the three glucocorticoids and cell survival was assessed by flow cytometry and light microscopy on cytospin preparations. Results We found that glucocorticoids down-regulated LPS-induced pro-inflammatory mRNA expression in both cell populations and species. These drugs also significantly increased glutamine synthetase gene expression in both equine cell populations. The magnitude of glucocorticoid response between cell populations was generally similar in both species. We also showed that dexamethasone had a comparable inhibitory effect on pro-inflammatory gene expression in both human and equine neutrophils. As reported in other species, glucocorticoids significantly increase the survival in equine neutrophils. Conclusions Glucocorticoids exert genomic effects of similar magnitude on neutrophils and on other blood leukocytes. We speculate that the poor response to

  17. Higher circulating leukocytes in women with PCOS is reversed by aerobic exercise.

    PubMed

    Covington, Jeffrey D; Tam, Charmaine S; Pasarica, Magdalena; Redman, Leanne M

    2016-05-01

    Polycystic ovary syndrome (PCOS) is characterized by insulin resistance, elevated circulating leukocytes, and hypothesized to have higher adipose tissue inflammation. Aerobic exercise reduces circulating leukocytes and improves insulin sensitivity in obese individuals, but the effect of exercise on inflammation in PCOS is not known. We investigated circulating leukocytes, insulin sensitivity by euglycemic-hyperinsulinemic clamp, serum pro- and anti-inflammatory markers (hsCRP, TNF-α, total and high molecular weight adiponectin), and abdominal subcutaneous adipose tissue (SAT) gene expression of proinflammatory markers in 8 PCOS women and 8 obese control females matched for BMI. Additionally, in a prospective study, the 8 women with PCOS underwent a 16-week aerobic exercise regimen with the same measures performed post-intervention. Compared to controls, white blood cell counts (WBC) were 30% higher (p = 0.04) and circulating total adiponectin levels were 150% lower (p = 0.03) in women with PCOS at baseline/pre-exercise conditions. SAT gene expression of macrophage migration inhibitory factor (MIF, p < 0.01) and interleukin-6 (IL-6, p < 0.05) were also lower in women with PCOS. In response to 16 weeks of aerobic exercise, insulin sensitivity improved (p < 0.01) and WBC counts decreased (p = 0.02). The exercise-induced change in WBC and circulating neutrophils correlated inversely with changes in glucose disposal rate (r = -0.73, p = 0.03; and r = -0.82, p = 0.01, respectively). Aerobic exercise reduced serum leptin (p < 0.05) after 4 weeks, trended to reduce the ratio of leptin-to-high molecular weight adiponectin (p < 0.1) by the 8th week, and significantly increased serum dehydroepiandrosterone sulfate (DHEA-S, p < 0.001) after 16 weeks. In conclusion, women with PCOS have higher circulating leukocytes compared to controls, which can be reversed by aerobic exercise and is associated with improvements in insulin sensitivity. PMID:25446648

  18. Aberration of miRNAs Expression in Leukocytes from Sporadic Amyotrophic Lateral Sclerosis

    PubMed Central

    Chen, YongPing; Wei, QianQian; Chen, XuePing; Li, ChunYu; Cao, Bei; Ou, RuWei; Hadano, Shinji; Shang, Hui-Fang

    2016-01-01

    Background: Accumulating evidence indicates that miRNAs play an important role in the development of amyotrophic lateral sclerosis (ALS). Most of previous studies on miRNA dysregulation in ALS focused on the alterative expression in ALS animal model or in limited samples from European patients with ALS. In the present study, the miRNA expression profiles were investigated in Chinese ALS patients to explore leukocytes miRNAs as a potential biomarker for the diagnosis of ALS. Methods: We analyzed the expression profiles of 1733 human mature miRNAs using microarray technology in leukocytes obtained from 5 patients with sporadic ALS (SALS) and 5 healthy controls. An independent group of 83 SALS patients, 24 Parkinson's disease (PD) patients and 61 controls was used for validation by real-time polymerase chain reaction assay. Area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. In addition, target genes and signaling information of validated differential expression miRNAs were predicted using Bioinformatics. Results: Eleven miRNAs, including four over-expressed and seven under-expressed miRNAs detected in SALS patients compared to healthy controls were selected for validation. Four under-expressed microRNAs, including hsa-miR-183, hsa-miR-193b, hsa-miR-451, and hsa-miR-3935, were confirmed in validation stage by comparison of 83 SALS patients and 61 HCs. Moreover, we identified a miRNA panel (hsa-miR-183, hsa-miR-193b, hsa-miR-451, and hsa-miR-3935) having a high diagnostic accuracy of SALS (AUC 0.857 for the validation group). However, only hsa-miR-183 was significantly lower in SALS patients than that in PD patients and in HCs, while no differences were found between PD patients and HCs. By bioinformatics analysis, we obtained a large number of target genes and signaling information that are linked to neurodegeneration. Conclusion: This study provided evidence of abnormal miRNA expression patterns in the peripheral

  19. [Case presentation: bovine leukocyte adhesion deficiency (BLAD) in Switzerland].

    PubMed

    Meylan, M; Abegg, R; Sager, H; Jungi, T W; Martig, J

    1997-01-01

    Bovine Leukocyte Adhesion Deficiency (BLAD) Syndrome is a lethal congenital immunodeficiency caused by the strong reduction in the expression of leukocyte integrins (beta 2 integrins) on the surface of leukocytes. Therefore, neutrophils from BLAD animals lack the capacity to adhere to the endothelium, a necessary step in their emigration into foci of infection. Due to the virtual absence of neutrophil-mediated host defense, animals suffer from recurrent infection of the respiratory and gastrointestinal tracts and finally succumb to infections. A 14 days old Holstein-Friesian calf showing omphalophlebitis and leukocytosis, was referred to our clinic. It was found to suffer from several febrile episodes of infection. The tentative diagnosis BLAD could be confirmed for the first time in Switzerland by flow cytometry, pedigree analysis and by restriction fragment length polymorphism. PMID:9411734

  20. Manipulating leukocyte interactions in vivo through optogenetic chemokine release.

    PubMed

    Sarris, Milka; Olekhnovitch, Romain; Bousso, Philippe

    2016-06-01

    Light-mediated release of signaling ligands, such as chemoattractants, growth factors, and cytokines is an attractive strategy for investigation and therapeutic targeting of leukocyte communication and immune responses. We introduce a versatile optogenetic method to control ligand secretion, combining UV-conditioned endoplasmic reticulum-to-Golgi trafficking and a furin-processing step. As proof of principle, we achieved light-triggered chemokine secretion and demonstrated that a brief pulse of chemokine release can mediate a rapid flux of leukocyte contacts with target cells in vitro and in vivo. This approach opens new possibilities for dynamic investigation of leukocyte communication in vivo and may confer the potential to control the local release of soluble mediators in the context of immune cell therapies. PMID:27057000

  1. Improving diagnosis of appendicitis. Early autologous leukocyte scanning

    SciTech Connect

    DeLaney, A.R.; Raviola, C.A.; Weber, P.N.; McDonald, P.T.; Navarro, D.A.; Jasko, I. )

    1989-10-01

    A prospective nonrandomized study investigating the accuracy and utility of autologous leukocyte scanning in the diagnosis of appendicitis was performed. One hundred patients in whom the clinical diagnosis of appendicitis was uncertain underwent indium 111 oxyquinoline labelling of autologous leukocytes and underwent scanning 2 hours following reinjection. Of 32 patients with proved appendicitis, three scans revealed normal results (false-negative rate, 0.09). Of 68 patients without appendicitis, three scans had positive results (false-positive rate, 0.03; sensitivity, 0.91; specificity, 0.97; predictive value of positive scan, 0.94; predictive value of negative scan, 0.96; and overall accuracy, 0.95). Scan results altered clinical decisions in 19 patients. In 13 cases, the scan produced images consistent with diagnoses other than appendicitis, expediting appropriate management. Early-imaging In 111 oxyquinoline autologous leukocyte scanning is a practical and highly accurate adjunct for diagnosing appendicitis.

  2. Plastid-LCGbase: a collection of evolutionarily conserved plastid-associated gene pairs.

    PubMed

    Wang, Dapeng; Yu, Jun

    2015-01-01

    Plastids carry their own genetic material that encodes a variable set of genes that are limited in number but functionally important. Aside from orthology, the lineage-specific order and orientation of these genes are also relevant. Here, we develop a database, Plastid-LCGbase (http://lcgbase.big.ac.cn/plastid-LCGbase/), which focuses on organizational variability of plastid genes and genomes from diverse taxonomic groups. The current Plastid-LCGbase contains information from 470 plastid genomes and exhibits several unique features. First, through a genome-overview page generated from OrganellarGenomeDRAW, it displays general arrangement of all plastid genes (circular or linear). Second, it shows patterns and modes of all paired plastid genes and their physical distances across user-defined lineages, which are facilitated by a step-wise stratification of taxonomic groups. Third, it divides the paired genes into three categories (co-directionally-paired genes or CDPGs, convergently-paired genes or CPGs and divergently-paired genes or DPGs) and three patterns (separation, overlap and inclusion) and provides basic statistics for each species. Fourth, the gene pairing scheme is expandable, where neighboring genes can also be included in species-/lineage-specific comparisons. We hope that Plastid-LCGbase facilitates gene variation (insertion-deletion, translocation and rearrangement) and transcription-level studies of plastid genomes. PMID:25378306

  3. Evolution of new functions de novo and from preexisting genes.

    PubMed

    Andersson, Dan I; Jerlström-Hultqvist, Jon; Näsvall, Joakim

    2015-06-01

    How the enormous structural and functional diversity of new genes and proteins was generated (estimated to be 10(10)-10(12) different proteins in all organisms on earth [Choi I-G, Kim S-H. 2006. Evolution of protein structural classes and protein sequence families. Proc Natl Acad Sci 103: 14056-14061] is a central biological question that has a long and rich history. Extensive work during the last 80 years have shown that new genes that play important roles in lineage-specific phenotypes and adaptation can originate through a multitude of different mechanisms, including duplication, lateral gene transfer, gene fusion/fission, and de novo origination. In this review, we focus on two main processes as generators of new functions: evolution of new genes by duplication and divergence of pre-existing genes and de novo gene origination in which a whole protein-coding gene evolves from a noncoding sequence. PMID:26032716

  4. Upregulation of Leukocytic Syncytin-1 in Acute Myeloid Leukemia Patients.

    PubMed

    Sun, Yi; Zhu, Hongyan; Song, Jianxin; Jiang, Yaxian; Ouyang, Hongmei; Huang, Rongzhong; Zhang, Guiqian; Fan, Xin; Tao, Rui; Jiang, Jie; Niu, Hua

    2016-01-01

    BACKGROUND Syncytin-1, a cell membrane-localizing fusogen, is abnormally expressed in several cancers, including endometrial cancer, breast cancer, and leukemia. Although abnormal syncytin-1 expression has been detected in two-thirds of leukemia blood samples, its expression profile in acute leukemia patients has not yet been analyzed. MATERIAL AND METHODS Bone marrow samples from 50 acute myelogenous leukemia (AML) cases and 14 B-cell acute lymphocytic leukemia (B-cell ALL) patients were subjected to flow cytometry to assess leukocyte type distributions and leukocytic syncytin-1 surface expression. RT-PCR was applied to assess leukocytic syncytin-1 mRNA expression. Statistical analysis was applied to compare syncytin-1 expression between AML and B-cell ALL patients across blasts, granulocytes, lymphocytes, and monocytes as well as to determine clinical factors statistically associated with changes in syncytin-1 expression. RESULTS The leukocyte type distributions of the AML and B-cell ALL cohorts highly overlapped, with an observable difference in blast distribution between the 2 cohorts. The AML cohort displayed significantly greater syncytin-1 surface and mRNA expression (p<0.05). Syncytin-1 surface and mRNA expression was significantly increased across all 4 leukocyte types (p<0.05). The percentage of syncytin-1-expressing blasts was significantly greater in AML patients (p<0.05), with blasts showing the largest fold-change in syncytin-1 expression (p<0.05). M5, M5a, and M5b AML patients displayed significantly higher syncytin-1 surface expression relative to all other AML French-American-British (FAB) classifications (p<0.05). CONCLUSIONS These findings suggest leukocytic syncytin-1 expression may play a role in the development and/or maintenance of the AML phenotype and the acute monocytic leukemia phenotype in particular. PMID:27393911

  5. Upregulation of Leukocytic Syncytin-1 in Acute Myeloid Leukemia Patients

    PubMed Central

    Sun, Yi; Zhu, Hongyan; Song, Jianxin; Jiang, Yaxian; Ouyang, Hongmei; Huang, Rongzhong; Zhang, Guiqian; Fan, Xin; Tao, Rui; Jiang, Jie; Niu, Hua

    2016-01-01

    Background Syncytin-1, a cell membrane-localizing fusogen, is abnormally expressed in several cancers, including endometrial cancer, breast cancer, and leukemia. Although abnormal syncytin-1 expression has been detected in two-thirds of leukemia blood samples, its expression profile in acute leukemia patients has not yet been analyzed. Material/Methods Bone marrow samples from 50 acute myelogenous leukemia (AML) cases and 14 B-cell acute lymphocytic leukemia (B-cell ALL) patients were subjected to flow cytometry to assess leukocyte type distributions and leukocytic syncytin-1 surface expression. RT-PCR was applied to assess leukocytic syncytin-1 mRNA expression. Statistical analysis was applied to compare syncytin-1 expression between AML and B-cell ALL patients across blasts, granulocytes, lymphocytes, and monocytes as well as to determine clinical factors statistically associated with changes in syncytin-1 expression. Results The leukocyte type distributions of the AML and B-cell ALL cohorts highly overlapped, with an observable difference in blast distribution between the 2 cohorts. The AML cohort displayed significantly greater syncytin-1 surface and mRNA expression (p<0.05). Syncytin-1 surface and mRNA expression was significantly increased across all 4 leukocyte types (p<0.05). The percentage of syncytin-1-expressing blasts was significantly greater in AML patients (p<0.05), with blasts showing the largest fold-change in syncytin-1 expression (p<0.05). M5, M5a, and M5b AML patients displayed significantly higher syncytin-1 surface expression relative to all other AML French-American-British (FAB) classifications (p<0.05). Conclusions These findings suggest leukocytic syncytin-1 expression may play a role in the development and/or maintenance of the AML phenotype and the acute monocytic leukemia phenotype in particular. PMID:27393911

  6. Leukocyte segmentation and classification in blood-smear images.

    PubMed

    Ramoser, Herbert; Laurain, Vincent; Bischof, Horst; Ecker, Rupert

    2005-01-01

    The detection and classification of leukocytes in blood smear images is a routine task in medical diagnosis. In this paper we present a fully automated approach to leukocyte segmentation that is robust with respect to cell appearance and image quality. A set of features is used to describe cytoplasm and nucleus properties. Pairwise SVM classification is used to discriminate between different cell types. Evaluation on a set of 1166 images (13 classes) resulted in 95% correct segmentations and 75% to 99% correct classification (with reject option). PMID:17280945

  7. Detection of deep venous thrombosis by indium-111 leukocyte scintigraphy

    SciTech Connect

    D'Alonzo, W.A. Jr.; Alavi, A.

    1986-05-01

    Indium-111-labeled leukocyte ((/sup 111/In)WBC) scintigraphy has been used successfully for detection of inflammation. Occasionally, noninflammatory collections of white blood cells such as hematomas or hemorrhage have been localized. We report a case in which unsuspected femoral deep venous thrombosis was diagnosed on an (/sup 111/In)WBC leukocyte scan performed for detection of osteomyelitis. Readers are advised to avoid interpreting all vascular (/sup 111/In)WBC localization as necessarily infectious. This may be of particular significance in patients with vascular grafts.

  8. Therapeutic hypothermia impacts leukocyte kinetics after cardiac arrest

    PubMed Central

    Dufner, Matthias C.; Andre, Florian; Stiepak, Jan; Zelniker, Thomas; Chorianopoulos, Emmanuel; Preusch, Michael; Katus, Hugo A.

    2016-01-01

    Background Patients admitted to the hospital after primarily successful cardiopulmonary resuscitation (CPR) are at a very high risk for neurologic deficits and death. Targeted temperature management (TTM) for mild therapeutic hypothermia has been shown to improve survival compared to standard treatment. Acute cardiovascular events, such as myocardial infarction (MI), are a major cause for cardiac arrest (CA) in patients who undergo CPR. Recent findings have demonstrated the importance and impact of the leukocyte response following acute MI. Methods In this retrospective, single center study we enrolled 169 patients with CA due to non-traumatic causes and primarily successful CPR. A total of 111 subjects (66%) underwent TTM aiming for a target temperature of 32–34 °C. Results Analysis of 30 day follow up showed a significantly improved survival of all patients who received TTM compared to patients without hypothermia (P=0.0001). Furthermore TTM was an independent variable of good neurological outcome after 6 months (P=0.0030). Therapeutic hypothermia was found to be beneficial independent of differences in age and sex between both groups. While a higher rate of pneumonia was observed with TTM, this diagnosis had no additional impact on survival or neurological outcome. The beneficial effect on mortality remained significant in patients with the diagnosis of an acute cardiac event (P=0.0145). Next, we evaluated the kinetics of leukocytes in this group over the course of 7 days after CA. At presentation, patients showed a mean level of 16.5±6.7 of leukocytes per microliter. While this level stayed stable in the group of patients without hypothermia, patients who received TTM showed a significant decline of leukocyte levels resulting in significantly lower numbers of leukocytes on days 3 and 5 after CPR. Interestingly, these differences in leukocyte counts remained beyond the time period of TTM while C-reactive protein (CRP) levels were suppressed only during

  9. Increased microvascular density and enhanced leukocyte rolling and adhesion in the skin of VEGF transgenic mice.

    PubMed

    Detmar, M; Brown, L F; Schön, M P; Elicker, B M; Velasco, P; Richard, L; Fukumura, D; Monsky, W; Claffey, K P; Jain, R K

    1998-07-01

    Vascular endothelial growth factor (VEGF) has been implicated in the pathologic angiogenesis observed in psoriasis and other chronic inflammatory skin diseases that are characterized by enhanced expression of VEGF by epidermal keratinocytes and of VEGF receptors by tortuous microvessels in the upper dermis. To investigate the functional importance of chronic VEGF overexpression in vivo, we used a keratin 14 promoter expression cassette containing the gene for murine VEGF164 to selectively target VEGF expression to basal epidermal keratinocytes in transgenic mice. These mice demonstrated an increased density of tortuous cutaneous blood capillaries with elevated expression levels of the high affinity VEGF receptors, VEGFR-1 and VEGFR-2, most prominently during the neonatal period. In contrast, no abnormalities of lymphatic vessels were detected. In addition, the number of mast cells in the upper dermis was significantly increased in transgenic skin. Intravital fluorescence microscopy revealed highly increased leukocyte rolling and adhesion in postcapillary skin venules that were both inhibited after injection of blocking antibodies against E- and P-selectin. Combined blocking antibodies against intercellular adhesion molecule-1 and lymphocyte function-associated antigen-1 were without effect, whereas an anti-vascular cell adhesion molecule-1/VLA-4 antibody combination almost completely normalized the enhanced leukocyte adhesion in transgenic mice. This study reveals VEGF as a growth factor specific for blood vessels, but not lymphatic vessels, and demonstrates that chronic orthotopic overexpression of VEGF in the epidermis is sufficient to induce cardinal features of chronic skin inflammation, providing a molecular link between angiogenesis, mast cell accumulation, and leukocyte recruitment to sites of inflammation. PMID:9665379

  10. Effect of prenatal arsenic exposure on DNA methylation and leukocyte subpopulations in cord blood

    PubMed Central

    Kile, Molly L; Houseman, E Andres; Baccarelli, Andrea A; Quamruzzaman, Quazi; Rahman, Mahmuder; Mostofa, Golam; Cardenas, Andres; Wright, Robert O; Christiani, David C

    2014-01-01

    Prenatal arsenic exposure is associated with increased risk of disease in adulthood. This has led to considerable interest in arsenic’s ability to disrupt fetal programming. Many studies report that arsenic exposure alters DNA methylation in whole blood but these studies did not adjust for cell mixture. In this study, we examined the relationship between arsenic in maternal drinking water collected ≤ 16 weeks gestational age and DNA methylation in cord blood (n = 44) adjusting for leukocyte-tagged differentially methylated regions. DNA methylation was quantified using the Infinium HumanMethylation 450 BeadChip array. Recursively partitioned mixture modeling examined the relationship between arsenic and methylation at 473,844 CpG sites. Median arsenic concentration in water was 12 µg/L (range < 1- 510 µg/L). Log10 arsenic was associated with altered DNA methylation across the epigenome (P = 0.002); however, adjusting for leukocyte distributions attenuated this association (P = 0.013). We also observed that arsenic had a strong effect on the distribution of leukocytes in cord blood. In adjusted models, every log10 increase in maternal drinking water arsenic exposure was estimated to increase CD8+ T cells by 7.4% (P = 0.0004) and decrease in CD4+ T cells by 9.2% (P = 0.0002). These results show that prenatal exposure to arsenic had an exposure-dependent effect on specific T cell subpopulations in cord blood and altered DNA methylation in cord blood. Future research is needed to determine if these small changes in DNA methylation alter gene expression or are associated with adverse health effects. PMID:24525453

  11. Differential loss of embryonic globin genes during the radiation of placental mammals

    PubMed Central

    Opazo, Juan C.; Hoffmann, Federico G.; Storz, Jay F.

    2008-01-01

    The differential gain and loss of genes from homologous gene families represents an important source of functional variation among the genomes of different species. Differences in gene content between species are primarily attributable to lineage-specific gene gains via duplication and lineage-specific losses via deletion or inactivation. Here, we use a comparative genomic approach to investigate this process of gene turnover in the β-globin gene family of placental mammals. By analyzing genomic sequence data from representatives of each of the main superordinal clades of placental mammals, we were able to reconstruct pathways of gene family evolution during the basal radiation of this physiologically and morphologically diverse vertebrate group. Our analysis revealed that an initial expansion of the nonadult portion of the β-globin gene cluster in the ancestor of placental mammals was followed by the differential loss and retention of ancestral gene lineages, thereby generating variation in the complement of embryonic globin genes among contemporary species. The sorting of ε-, γ-, and η-globin gene lineages among the basal clades of placental mammals has produced species differences in the functional types of hemoglobin isoforms that can be synthesized during the course of embryonic development. PMID:18755893

  12. Dysregulated 14-3-3 Family in Peripheral Blood Leukocytes of Patients with Schizophrenia

    PubMed Central

    Qing, Ying; Sun, Liya; Yang, Chao; Jiang, Jie; Yang, Xuhan; Hu, Xiaowen; Cui, Donghong; Xu, Yifeng; He, Lin; Han, Dongmei; Wan, Chunling

    2016-01-01

    The 14-3-3 family, which is composed of seven distinct members in humans, plays important roles in the cell cycle, apoptosis, synaptic plasticity and neuronal differentiation and migration. Previous genetic and post-mortem gene expression studies have linked this family to schizophrenia. However, the direction of gene expression changes in these studies has been inconsistent, and reports of 14-3-3 gene expression in living schizophrenic patients are still lacking. Here, we assessed 14-3-3 gene and protein expression levels in peripheral blood leukocytes from drug-naïve first-episode schizophrenic patients and matched controls. mRNA and protein expression levels were quantified by qRT-PCR and UPLC-MRM/MS, respectively. Expression analysis revealed four downregulated and one upregulated mRNA transcripts as well as five downregulated protein levels of 14-3-3 isoforms in schizophrenia. Moreover, significant positive correlations between 14-3-3 mRNA and protein expression levels were found in schizophrenia, and we also identified negative correlations between ε, θ and ζ isoform expression levels and positive symptoms of schizophrenia. Our results suggest that gene and protein expression levels for the 14-3-3 family are dysregulated in schizophrenia, perhaps owing to specific regulatory mechanisms, and we also suggest that expression of the 14-3-3ε, θ and ζ isoform genes could be useful indicators of disease severity. PMID:27030512

  13. The Effect of Hemiscorpius lepturus (Scorpionida: Hemiscorpiidae) Venom on Leukocytes and the Leukocyte Subgroups in Peripheral Blood of Rat

    PubMed Central

    Ghafourian, Mehri; Ganjalikhanhakemi, Neda; Hemmati, Ali Asghar; Dehghani, Rouhullah; Kooti, Wesam

    2016-01-01

    Background: The aim of this study was to investigate the effect of Hemiscorpius lepturus venom on leukocytes and the leukocyte subgroups in peripheral blood of rat. Methods: In this experimental study, sixty N-Mari rats were divided into three groups of 20 rats. Then the rats in each group were divided into four subgroups based on the blood sampling time that was 2, 6, 24 and 48 hours after the venom injection, respectively. The control group did not receive anything, however, the first and the second experimental groups received 0.1 and 0.01mg/kg of venom, subcutaneously. In accordance with a designated four sampling times, the blood sampling was carried out in three groups. After RBC lysis, the leukocytes and leukocyte sub-populations were determined and counted using appropriate hematological standard methods. Results: The leukocyte and the neutrophil count at two (P<0.05), six (P<0.01) and 24 (P<0.05) hours after the venom injection showed a significant decline compared with the control group, this decrease was significant at the dose of 0.1 mg/kg until 48 hours after the venom injection (P<0.05). The lymphocyte count showed a significant decline throughout the all hours of the experiment, compared with the control group (P<0.05). Conclusion: Leukocytes are probably affected by the cytotoxicity effect of the H. lepturus venom in a dose-dependent manner. This could be a wakeup call for the medical staff to perform quick and accurate treatment in the least time possible. PMID:27308274

  14. Studies on gonococcus infection. XV. Identification of surface proteins of Neisseria gonorrhoeae correlated with leukocyte association.

    PubMed Central

    King, G J; Swanson, J

    1978-01-01

    Neisseria gonorrhoeae which exhibit high levels of leukocyte association have a surface protein which is considerably diminished in isogenic gonococci which exhibit low levels of leukocyte association (LA). The LA protein exhibits strain variation in molecular weight and immunogenicity. Membranes derived from LA+ and LA- organisms show quantitative differences in their adsorption to leukocytes; these differences are analogous to those found for the intact organisms regarding their association with leukocytes. Images PMID:211086

  15. Recent advances in microscopic techniques for visualizing leukocytes in vivo

    PubMed Central

    Jain, Rohit; Tikoo, Shweta; Weninger, Wolfgang

    2016-01-01

    Leukocytes are inherently motile and interactive cells. Recent advances in intravital microscopy approaches have enabled a new vista of their behavior within intact tissues in real time. This brief review summarizes the developments enabling the tracking of immune responses in vivo. PMID:27239292

  16. Leukocyte driven-decidual angiogenesis in early pregnancy.

    PubMed

    Lima, Patricia D A; Zhang, Jianhong; Dunk, Caroline; Lye, Stephen J; Croy, B Anne

    2014-11-01

    Successful pregnancy and long-term, post-natal maternal and offspring cardiac, vascular and metabolic health require key maternal cardiovascular adaptations over gestation. Within the pregnant decidualizing uterus, coordinated vascular, immunological and stromal cell changes occur. Considerable attention has been given to the roles of uterine natural killer (uNK) cells in initiating decidual spiral arterial remodeling, a process normally completed by mid-gestation in mice and in humans. However, leukocyte roles in much earlier, region specific, decidual vascular remodeling are now being defined. Interest in immune cell-promoted vascular remodeling is driven by vascular aberrations that are reported in human gestational complications such as infertility, recurrent spontaneous abortion, preeclampsia (PE) and fetal growth restriction. Appropriate maternal cardiovascular responses during pregnancy protect mothers and their children from later cardiovascular disease risk elevation. One of the earliest uterine responses to pregnancy in species with hemochorial placentation is stromal cell decidualization, which creates unique niches for angiogenesis and leukocyte recruitment. In early decidua basalis, the aspect of the implantation site that will cradle the developing placenta and provide the major blood vessels to support mature placental functions, leukocytes are greatly enriched and display specialized properties. UNK cells, the most abundant leukocyte subset in early decidua basalis, have angiogenic abilities and are essential for normal early decidual angiogenesis. The regulation of uNK cells and their roles in determining maternal and progeny cardiovascular health over pregnancy and postpartum are discussed. PMID:25066422

  17. Improved survival of newborns receiving leukocyte transfusions for sepsis

    SciTech Connect

    Cairo, M.S.; Rucker, R.; Bennetts, G.A.; Hicks, D.; Worcester, C.; Amlie, R.; Johnson, S.; Katz, J.

    1984-11-01

    To determine the role of polymorphonuclear (PMN) leukocyte transfusions in neonates with sepsis, 23 consecutive newborns were prospectively randomly selected during an 18-month period in a treatment plan to receive polymorphonuclear leukocyte transfusions with supportive care or supportive care alone. Thirteen neonates received transfusions every 12 hours for a total of five transfusions. Each transfusion consisting of 15 mL/kg of polymorphonuclear leukocytes was subjected to 1,500 rads of radiation. The polymorphonuclear leukocytes were obtained by continuous-flow centrifugation leukapheresis and contained 0.5 to 1.0 X 10(9) granulocytes per 15 mL with less than 10% lymphocytes. Positive findings on blood cultures were obtained in 14/23 patients and seven were randomly selected for each treatment group. Absolute granulocyte counts were less than 1,500/microL in 13 patients but tibial bone marrow examinations revealed that the neutrophil supply pool was depleted in only three patients. The survival was significantly greater in the treatment group compared with the group that did not receive transfusions.

  18. Recent advances in microscopic techniques for visualizing leukocytes in vivo.

    PubMed

    Jain, Rohit; Tikoo, Shweta; Weninger, Wolfgang

    2016-01-01

    Leukocytes are inherently motile and interactive cells. Recent advances in intravital microscopy approaches have enabled a new vista of their behavior within intact tissues in real time. This brief review summarizes the developments enabling the tracking of immune responses in vivo. PMID:27239292

  19. Intracellular Penetration and Activity of Gemifloxacin in Human Polymorphonuclear Leukocytes

    PubMed Central

    García, Isabel; Pascual, Alvaro; Ballesta, Sofía; Joyanes, Providencia; Perea, Evelio J.

    2000-01-01

    The intracellular penetration and activity of gemifloxacin in human polymorphonuclear leukocytes (PMN) were evaluated. Gemifloxacin reached intracellular concentrations eight times higher than extracellular concentrations. The uptake was rapid, reversible, and nonsaturable and was affected by environmental temperature, cell viability, and membrane stimuli. At therapeutic extracellular concentrations, gemifloxacin showed intracellular activity against Staphylococcus aureus. PMID:11036051

  20. Multiparticle adhesive dynamics: Hydrodynamic recruitment of rolling leukocytes

    PubMed Central

    King, Michael R.; Hammer, Daniel A.

    2001-01-01

    The slow rolling motion of leukocytes along the walls of blood vessels mediated by specific receptor-ligand adhesion is important in inflammation and occurs in postcapillary venules over a wide range of wall shear stresses and vessel diameters. The ability of hydrodynamic collisions between cells to induce capture of free-stream leukocytes to a selectin-bearing surface under shear flow was studied experimentally by using a cell-free assay. It was found that carbohydrate-coated spherical beads, representing model leukocytes, tend to attach to the adhesive wall 4–5 cell diameters up- or downstream of a slowly rolling or stationary adhesive bead. A key feature of such “hydrodynamic recruitment” is that only glancing, indirect collisions occurring close to the plane will result in downstream attachment. A direct numerical simulation of cell capture and rolling that includes multiparticle hydrodynamic interactions is shown to reproduce the observed behavior accurately. The theory predicts that hydrodynamic recruitment will occur in the absence of buoyancy effects and over a range of shear rates, suggesting that the mechanism may be important in vivo. This theory is supported by measurements of leukocyte capture in vivo using the hamster cheek pouch model. PMID:11752440

  1. Differential MSC activation leads to distinct mononuclear leukocyte binding mechanisms

    NASA Astrophysics Data System (ADS)

    Kota, Daniel J.; Dicarlo, Bryan; Hetz, Robert A.; Smith, Philippa; Cox, Charles S.; Olson, Scott D.

    2014-04-01

    Advances in the field of Multipotent Mesenchymal Stromal cell (MSC) biology have demonstrated that MSCs can improve disease outcome when `activated' to exert immunomodulatory effects. However, the precise mechanisms modulating MSC-immune cells interactions remain largely elusive. In here, we activated MSC based on a recent polarization paradigm, in which MSCs can be polarized towards a pro- or anti-inflammatory phenotype depending on the Toll-like receptor stimulated, to dissect the mechanisms through which MSCs physically interact with and modulate leukocytes in this context. Our data show that MSCs activated through the Toll-like receptor (TLR) 4 pathway increased VCAM-1 and ICAM-1 dependent binding of leukocytes. On the other hand, TLR3 stimulation strongly increases leukocytes affinity to MSC comparatively, through the formation of cable-like hyaluronic acid structures. In addition, TLR4 activation elicited secretion of pro-inflammatory mediators by MSCs, whereas TLR3-activated MSCs displayed a milder pro-inflammatory phenotype, similar to inactivated MSCs. However, the differently activated MSCs maintained their ability to suppress leukocyte activation at similar levels in our in vitro model, and this immunomodulatory property was shown here to be partially mediated by prostaglandin. These results reinforce the concept that alternate activation profiles control MSC responses and may impact the therapeutic use of MSCs.

  2. The spreading of leukocytes released from their liquid environment.

    PubMed

    Cuadra, M

    1978-08-15

    It is known that while leukocytes in conventional blood smears reveal their structures, in smears of erythrocyte-free suspension (EFS) they show a tendency to pyknosis. From the present study it appears that: 1. When any leukocyte suspension is subjected to drying on slides the cells undergo shrinkage (pyknosis in stained state) because their water content is extracted by the suspending fluid that is rendered increasingly hypertonic during drying. 2. When the leukocytes, which while floating within physiological fluids are ball-shaped, are entirely released from their suspending fluid, they suddenly spread into flat cells because the capillary force between the cell membranes and glass overcomes the surface tension of the cells. 3. In contrast to shrunken or even normal (spherical) cells, spread cells are able to display their morphology. Using the smear technique, the cell density of the suspension will determine whether spreading or shrinkage are to occur; with normal blood (high density) the release and consequently the spread are achieved; with EFS (usually of low density) an inadequate release and consequently shrinkage occur. 4. A device (cell spreader) for spreading leukocytes of EFS is presented. PMID:678682

  3. Leukocyte driven-decidual angiogenesis in early pregnancy

    PubMed Central

    Lima, Patricia DA; Zhang, Jianhong; Dunk, Caroline; Lye, Stephen J; Anne Croy, B

    2014-01-01

    Successful pregnancy and long-term, post-natal maternal and offspring cardiac, vascular and metabolic health require key maternal cardiovascular adaptations over gestation. Within the pregnant decidualizing uterus, coordinated vascular, immunological and stromal cell changes occur. Considerable attention has been given to the roles of uterine natural killer (uNK) cells in initiating decidual spiral arterial remodeling, a process normally completed by mid-gestation in mice and in humans. However, leukocyte roles in much earlier, region specific, decidual vascular remodeling are now being defined. Interest in immune cell-promoted vascular remodeling is driven by vascular aberrations that are reported in human gestational complications such as infertility, recurrent spontaneous abortion, preeclampsia (PE) and fetal growth restriction. Appropriate maternal cardiovascular responses during pregnancy protect mothers and their children from later cardiovascular disease risk elevation. One of the earliest uterine responses to pregnancy in species with hemochorial placentation is stromal cell decidualization, which creates unique niches for angiogenesis and leukocyte recruitment. In early decidua basalis, the aspect of the implantation site that will cradle the developing placenta and provide the major blood vessels to support mature placental functions, leukocytes are greatly enriched and display specialized properties. UNK cells, the most abundant leukocyte subset in early decidua basalis, have angiogenic abilities and are essential for normal early decidual angiogenesis. The regulation of uNK cells and their roles in determining maternal and progeny cardiovascular health over pregnancy and postpartum are discussed. PMID:25066422

  4. Uptake of radiolabeled leukocytes in prosthetic graft infection

    SciTech Connect

    Serota, A.I.; Williams, R.A.; Rose, J.G.; Wilson, S.E.

    1981-07-01

    The utility of radionuclide labeled leukocytes in the demonstration of infection within vascular prostheses was examined. The infrarenal aorta was replaced with a 3 cm Dacron graft in 12 dogs. On the third postoperative day, six of the animals received an intravenous injection of 10(8) Staphylococcus aureus. Labeled leukocyte scans were performed at postoperative days one and three, and then weekly for 8 weeks with indium-111 and technetium-99 labeled autologous leukocytes. When scans showed focal uptake of isotope in the area of prosthetic material, the grafts were aseptically excised and cultured on mannitol-salt agar. Both control and infected animals had retroperitoneal isotope activity in the immediate postoperative period that disappeared by the end of the first week. By the eighth postoperative week, all of the animals that received the bacteremic challenge had both radionuclide concentration in the region of the vascular prosthesis and S. aureus cultured subsequently from the perigraft tissues. None of the control animals had either radionuclide or bacteriologic evidence of infection at the eighth postoperative week. The radiolabeled leukocyte scan is a highly sensitive and specific technique, clinically applicable for the diagnosis of vascular prosthetic infections.

  5. Secretory leukocyte proteinase inhibitor is a major leukocyte elastase inhibitor in human neutrophils.

    PubMed

    Sallenave, J M; Si Tahar, M; Cox, G; Chignard, M; Gauldie, J

    1997-06-01

    Secretory leukocyte proteinase inhibitor (SLPI) is the main neutrophil elastase (HLE) inhibitor found in the upper airways during pulmonary inflammation. It has been shown to be synthesized and secreted in vitro by epithelial cells and has been localized in tracheal glands and bronchiolar epithelial cells by immunocytochemistry. In this study, using immunodetection and immunopurification techniques with specific anti-SLPI immunoglobulin G (IgG), we show that SLPI is present as a native 14-kDa molecule in neutrophil cytosol. In addition, we demonstrate that SLPI is the major inhibitor of HLE present in neutrophil cytosol because pre-incubation with specific anti-SLPI IgG was able to inhibit completely the anti-HLE activity of the cytosol. SLPI can be secreted (probably in an inactive form) by neutrophils and its secretion is enhanced when the cells are stimulated with phorbol myristate acetate (PMA). Elafin, an elastase-specific inhibitor, is also present in minute amounts in neutrophil cytosol and its secretion can be up-regulated. The presence of SLPI in the cytosol of neutrophils may serve as a protective screen against proteinases spilling from azurophilic granules. An alternative or supplementary role may be the maintenance of a differentiated phenotype. PMID:9201260

  6. Origins of major archaeal clades correspond to gene acquisitions from bacteria

    PubMed Central

    Nelson-Sathi, Shijulal; Sousa, Filipa L.; Röttger, Mayo; Lozada-Chávez, Nabor; Thiergart, Thorsten; Janssen, Arnold; Bryant, David; Landan, Giddy; Schönheit, Peter; Siebers, Bettina; McInerney, James O.; Martin, William F.

    2014-01-01

    The mechanisms that underlie the origin of major prokaryotic groups are poorly understood. In principle, the origin of both species and higher taxa among prokaryotes should entail similar mechanisms — ecological interactions with the environment paired with natural genetic variation involving lineage-specific gene innovations and lineage-specific gene acquisitions1,2,3,4. To investigate the origin of higher taxa in archaea, we have determined gene distributions and gene phylogenies for the 267,568 protein coding genes of 134 sequenced archaeal genomes in the context of their homologs from 1,847 reference bacterial genomes. Archaea-specific gene families define 13 traditionally recognized archaeal higher taxa in our sample. Here we report that the origins of these 13 groups unexpectedly correspond to 2,264 group-specific gene acquisitions from bacteria. Interdomain gene transfer is highly asymmetric, transfers from bacteria to archaea are more than 5-fold more frequent than vice versa. Gene transfers identified at major evolutionary transitions among prokaryotes specifically implicate gene acquisitions for metabolic functions from bacteria as key innovations in the origin of higher archaeal taxa. PMID:25317564

  7. In vivo compartmental analysis of leukocytes in mouse lungs.

    PubMed

    Patel, Brijesh V; Tatham, Kate C; Wilson, Michael R; O'Dea, Kieran P; Takata, Masao

    2015-10-01

    The lung has a unique structure consisting of three functionally different compartments (alveolar, interstitial, and vascular) situated in an extreme proximity. Current methods to localize lung leukocytes using bronchoalveolar lavage and/or lung perfusion have significant limitations for determination of location and phenotype of leukocytes. Here we present a novel method using in vivo antibody labeling to enable accurate compartmental localization/quantification and phenotyping of mouse lung leukocytes. Anesthetized C57BL/6 mice received combined in vivo intravenous and intratracheal labeling with fluorophore-conjugated anti-CD45 antibodies, and lung single-cell suspensions were analyzed by flow cytometry. The combined in vivo intravenous and intratracheal CD45 labeling enabled robust separation of the alveolar, interstitial, and vascular compartments of the lung. In naive mice, the alveolar compartment consisted predominantly of resident alveolar macrophages. The interstitial compartment, gated by events negative for both intratracheal and intravenous CD45 staining, showed two conventional dendritic cell populations, as well as a Ly6C(lo) monocyte population. Expression levels of MHCII on these interstitial monocytes were much higher than on the vascular Ly6C(lo) monocyte populations. In mice exposed to acid aspiration-induced lung injury, this protocol also clearly distinguished the three lung compartments showing the dynamic trafficking of neutrophils and exudative monocytes across the lung compartments during inflammation and resolution. This simple in vivo dual-labeling technique substantially increases the accuracy and depth of lung flow cytometric analysis, facilitates a more comprehensive examination of lung leukocyte pools, and enables the investigation of previously poorly defined "interstitial" leukocyte populations during models of inflammatory lung diseases. PMID:26254421

  8. Fecal Leukocytes in Children Infected with Diarrheagenic Escherichia coli▿

    PubMed Central

    Mercado, Erik H.; Ochoa, Theresa J.; Ecker, Lucie; Cabello, Martin; Durand, David; Barletta, Francesca; Molina, Margarita; Gil, Ana I.; Huicho, Luis; Lanata, Claudio F.; Cleary, Thomas G.

    2011-01-01

    The purpose of this study was to determine the presence and quantity of fecal leukocytes in children infected with diarrheagenic Escherichia coli and to compare these levels between diarrhea and control cases. We analyzed 1,474 stool samples from 935 diarrhea episodes and 539 from healthy controls of a cohort study of children younger than 2 years of age in Lima, Peru. Stools were analyzed for common enteric pathogens, and diarrheagenic E. coli isolates were studied by a multiplex real-time PCR. Stool smears were stained with methylene blue and read by a blinded observer to determine the number of polymorphonuclear leukocytes per high-power field (L/hpf). Fecal leukocytes at >10 L/hpf were present in 11.8% (110/935) of all diarrheal episodes versus 1.1% (6/539) in controls (P < 0.001). Among stool samples with diarrheagenic E. coli as the only pathogen isolated (excluding coinfection), fecal leukocytes at >10 L/hpf were present in 8.5% (18/212) of diarrhea versus 1.3% (2/157) of control samples (P < 0.01). Ninety-five percent of 99 diarrheagenic E. coli diarrhea samples were positive for fecal lactoferrin. Adjusting for the presence of blood in stools, age, sex, undernutrition, and breastfeeding, enterotoxigenic E. coli (ETEC) isolation as a single pathogen, excluding coinfections, was highly associated with the presence of fecal leukocytes (>10 L/hpf) with an odds ratio (OR) of 4.1 (95% confidence interval [CI], 1.08 to 15.51; P < 0.05). Although diarrheagenic E. coli was isolated with similar frequencies in diarrhea and control samples, clearly it was associated with a more inflammatory response during symptomatic infection; however, in general, these pathogens elicited a mild inflammatory response. PMID:21325554

  9. Fecal leukocytes in children infected with diarrheagenic Escherichia coli.

    PubMed

    Mercado, Erik H; Ochoa, Theresa J; Ecker, Lucie; Cabello, Martin; Durand, David; Barletta, Francesca; Molina, Margarita; Gil, Ana I; Huicho, Luis; Lanata, Claudio F; Cleary, Thomas G

    2011-04-01

    The purpose of this study was to determine the presence and quantity of fecal leukocytes in children infected with diarrheagenic Escherichia coli and to compare these levels between diarrhea and control cases. We analyzed 1,474 stool samples from 935 diarrhea episodes and 539 from healthy controls of a cohort study of children younger than 2 years of age in Lima, Peru. Stools were analyzed for common enteric pathogens, and diarrheagenic E. coli isolates were studied by a multiplex real-time PCR. Stool smears were stained with methylene blue and read by a blinded observer to determine the number of polymorphonuclear leukocytes per high-power field (L/hpf). Fecal leukocytes at >10 L/hpf were present in 11.8% (110/935) of all diarrheal episodes versus 1.1% (6/539) in controls (P < 0.001). Among stool samples with diarrheagenic E. coli as the only pathogen isolated (excluding coinfection), fecal leukocytes at >10 L/hpf were present in 8.5% (18/212) of diarrhea versus 1.3% (2/157) of control samples (P < 0.01). Ninety-five percent of 99 diarrheagenic E. coli diarrhea samples were positive for fecal lactoferrin. Adjusting for the presence of blood in stools, age, sex, undernutrition, and breastfeeding, enterotoxigenic E. coli (ETEC) isolation as a single pathogen, excluding coinfections, was highly associated with the presence of fecal leukocytes (>10 L/hpf) with an odds ratio (OR) of 4.1 (95% confidence interval [CI], 1.08 to 15.51; P < 0.05). Although diarrheagenic E. coli was isolated with similar frequencies in diarrhea and control samples, clearly it was associated with a more inflammatory response during symptomatic infection; however, in general, these pathogens elicited a mild inflammatory response. PMID:21325554

  10. Two-step model of leukocyte-endothelial cell interaction in inflammation: distinct roles for LECAM-1 and the leukocyte beta 2 integrins in vivo.

    PubMed Central

    von Andrian, U H; Chambers, J D; McEvoy, L M; Bargatze, R F; Arfors, K E; Butcher, E C

    1991-01-01

    The lectin homing receptor LECAM-1 (LAM-1, Leu8) and the beta 2 integrins, particularly Mac-1 (CD11b/CD18), participate in leukocyte-endothelial cell interactions in inflammation. LECAM-1 is rapidly shed while Mac-1 expression is dramatically increased upon neutrophil activation, suggesting functionally distinct roles for these molecules. Using intravital video microscopy, we have compared the effect of antibodies against LECAM-1 and CD18 on leukocyte interactions with rabbit mesenteric venules. Anti-LECAM-1 monoclonal antibody and its Fab fragments inhibited initial reversible leukocyte rolling along the vascular wall. Anti-CD18 monoclonal antibody had no effect on rolling but prevented subsequent firm attachment of leukocytes to venular endothelium. These results support a two-step model of leukocyte-endothelial cell interactions: reversible rolling mediated in part by LECAM-1 facilitates leukocyte recruitment by the local microenvironment and precedes activation-dependent firm attachment involving beta 2 integrins. Images PMID:1715568

  11. Childhood Adversity and Epigenetic Modulation of the Leukocyte Glucocorticoid Receptor: Preliminary Findings in Healthy Adults

    PubMed Central

    Tyrka, Audrey R.; Price, Lawrence H.; Marsit, Carmen; Walters, Oakland C.; Carpenter, Linda L.

    2012-01-01

    Background A history of early adverse experiences is an important risk factor for adult psychopathology. Changes in stress sensitivity and functioning of the hypothalamic-pituitary-adrenal (HPA) axis may underlie the association between stress and risk for psychiatric disorders. Preclinical work in rodents has linked low levels of maternal care to increased methylation of the promoter region of the glucocorticoid receptor (GR) gene, as well as to exaggerated hormonal and behavioral responses to stress. Recent studies have begun to examine whether early-life stress leads to epigenetic modifications of the GR gene in humans. Methods We examined the degree of methylation of a region of the promoter of the human GR gene (NR3C1) in leukocyte DNA from 99 healthy adults. Participants reported on their childhood experiences of parental behavior, parental death or desertion, and childhood maltreatment. On a separate day, participants completed the dexamethasone/corticotropin-releasing hormone (Dex/CRH) test, a standardized neuroendocrine challenge test. Results Disruption or lack of adequate nurturing, as measured by parental loss, childhood maltreatment, and parental care, was associated with increased NR3C1 promoter methylation (p<.05). In addition, NR3C1 promoter methylation was linked to attenuated cortisol responses to the Dex/CRH test (p<.05). Conclusions These findings suggest that childhood maltreatment or adversity may lead to epigenetic modifications of the human GR gene. Alterations in methylation of this gene could underlie the associations between childhood adversity, alterations in stress reactivity, and risk for psychopathology. PMID:22295073

  12. CD99 and CD99L2 act at the same site as, but independently of, PECAM-1 during leukocyte diapedesis.

    PubMed

    Bixel, M Gabriele; Li, Hang; Petri, Bjoern; Khandoga, Alexander G; Khandoga, Andrej; Zarbock, Alexander; Wolburg-Buchholz, Karen; Wolburg, Hartwig; Sorokin, Lydia; Zeuschner, Dagmar; Maerz, Sigrid; Butz, Stefan; Krombach, Fritz; Vestweber, Dietmar

    2010-08-19

    Leukocyte extravasation depends on various adhesion receptors at endothelial cell contacts. Here we have analyzed how mouse CD99 and CD99L2 cooperate with PECAM-1. We found that antibodies against mouse CD99 and PECAM-1 trap neutrophils between endothelial cells in in vitro transmigration assays. A sequential function, as has been suggested for human PECAM-1 and CD99, could not be demonstrated. In contrast to these in vitro results, blocking CD99 or CD99L2 or gene disruption of PECAM-1 trapped neutrophils in vivo between endothelial cells and the underlying basement membrane as revealed by electron microscopy and by 3-dimensional confocal fluorescence microscopy in the inflamed cremaster tissue. Leukocyte extravasation was inhibited in interleukin-1beta-inflamed peritoneum and in the cremaster by PECAM-1 gene disruption and was further attenuated by blocking antibodies against CD99 and CD99L2. In addition, CD99 and CD99L2 were required for leukocyte extravasation in the cremaster after stimulation with tumor necrosis factor-alpha, where the need for PECAM-1 is known to be bypassed. We conclude that CD99 and CD99L2 act independently of PECAM-1 in leukocyte extravasation and cooperate in an independent way to help neutrophils overcome the endothelial basement membrane. PMID:20479283

  13. Splenic Leukocytes Traffic to the Thyroid and Produce a Novel TSHβ Isoform during Acute Listeria monocytogenes Infection in Mice.

    PubMed

    Montufar-Solis, Dina; Klein, John R

    2016-01-01

    The thyroid stimulating hormone beta-subunit (TSHβ) with TSHα form a glycoprotein hormone that is produced by the anterior pituitary in the hypothalamus-pituitary-thyroid (HPT) axis. Although TSHβ has been known for many years to be made by cells of the immune system, the role of immune system TSH has remained unclear. Recent studies demonstrated that cells of the immune system produce a novel splice variant isoform of TSHβ (TSHβv), but little if any native TSHβ. Here, we show that within three days of systemic infection of mice with Listeria monocytogenes, splenic leukocytes synthesized elevated levels of TSHβv. This was accompanied by an influx of CD14+, Ly6C+, Ly6G+ cells into the thyroid of infected mice, and increased levels of intrathyroidal TSHβv gene expression. Adoptive transfer of carboxyfluorescein succinimidyl ester (CFSE)-labeled splenic leukocytes from infected mice into non-infected mice migrated into the thyroid as early as forty-eight hours post-cell transfer, whereas CFSE-labeled cells from non-infected mice failed to traffic to the thyroid. These findings demonstrate for the first time that during bacterial infection peripheral leukocytes produce elevated levels of TSHβv, and that spleen cells traffic to the thyroid where they produce TSHβv intrathyroidally. PMID:26771831

  14. Splenic Leukocytes Traffic to the Thyroid and Produce a Novel TSHβ Isoform during Acute Listeria monocytogenes Infection in Mice

    PubMed Central

    Montufar-Solis, Dina; Klein, John R.

    2016-01-01

    The thyroid stimulating hormone beta-subunit (TSHβ) with TSHα form a glycoprotein hormone that is produced by the anterior pituitary in the hypothalamus-pituitary-thyroid (HPT) axis. Although TSHβ has been known for many years to be made by cells of the immune system, the role of immune system TSH has remained unclear. Recent studies demonstrated that cells of the immune system produce a novel splice variant isoform of TSHβ (TSHβv), but little if any native TSHβ. Here, we show that within three days of systemic infection of mice with Listeria monocytogenes, splenic leukocytes synthesized elevated levels of TSHβv. This was accompanied by an influx of CD14+, Ly6C+, Ly6G+ cells into the thyroid of infected mice, and increased levels of intrathyroidal TSHβv gene expression. Adoptive transfer of carboxyfluorescein succinimidyl ester (CFSE)-labeled splenic leukocytes from infected mice into non-infected mice migrated into the thyroid as early as forty-eight hours post-cell transfer, whereas CFSE-labeled cells from non-infected mice failed to traffic to the thyroid. These findings demonstrate for the first time that during bacterial infection peripheral leukocytes produce elevated levels of TSHβv, and that spleen cells traffic to the thyroid where they produce TSHβv intrathyroidally. PMID:26771831

  15. Modulation of leukocyte adhesion in rat mesenteric venules by aspirin and salicylate.

    PubMed

    Asako, H; Kubes, P; Wallace, J; Wolf, R E; Granger, D N

    1992-07-01

    Erythrocyte velocity, vessel diameter, leukocyte rolling velocity, and number of adherent and emigrated leukocytes were measured in postcapillary venules both before and during superfusion of rat mesentery with either aspirin or sodium salicylate. In some experiments, animals were treated with either a leukotriene (LT)-synthesis inhibitor (L-663,536), an LTD4 antagonist (MK-571), an LTB4 antagonist (SC-41930), misoprostol, or prostaglandin (PG) I2, then the aspirin protocol was repeated. Superfusion of aspirin but not sodium salicylate resulted in increased leukocyte adherence and a reduced leukocyte rolling velocity but did not affect leukocyte emigration. Aspirin-induced leukocyte adhesion was effectively prevented by the LT-synthesis inhibitor and LTB4 antagonist but not by the LTD4 antagonist. Misoprostol and PGI2 also prevented the aspirin-induced adhesion responses. Superfusion of the mesentery with either platelet-activating factor (PAF) or LTB4 enhanced leukocyte adherence and emigration while reducing leukocyte rolling velocity. Sodium salicylate prevented all of the adhesion responses elicited by LTB4. Although salicylate did not affect the PAF-induced leukocyte adherence and rolling responses, it completely prevented the increased leukocyte emigration. These results indicate that aspirin promotes, whereas sodium salicylate inhibits, leukocyte-endothelial cell adhesive interactions at therapeutically relevant concentrations. PMID:1319367

  16. Combined bone scintigraphy and indium-111 leukocyte scans in neuropathic foot disease

    SciTech Connect

    Schauwecker, D.S.; Park, H.M.; Burt, R.W.; Mock, B.H.; Wellman, H.N.

    1988-10-01

    It is difficult to diagnose osteomyelitis in the presence of neurotrophic osteoarthropathy. We performed combined (99mTc)MDP bone scans and indium-111 (111In) leukocyte studies on 35 patients who had radiographic evidence of neuropathic foot disease and clinically suspected osteomyelitis. The (111In)leukocyte study determined if there was an infection and the bone scan provided the anatomic landmarks so that the infection could be localized to the bone or the adjacent soft tissue. Seventeen patients had osteomyelitis and all showed increased (111In)leukocyte activity localized to the bone, giving a sensitivity of 100%. Among the 18 patients without osteomyelitis, eight had no accumulation of (111In)leukocytes, seven had the (111In)leukocyte activity correctly localized to the soft tissue, two had (111In)leukocyte activity mistakenly attributed to the bone, and one had (111In)leukocyte accumulation in a proven neuroma which was mistakenly attributed to bone. These three false-positive results for osteomyelitis reduced the specificity to 83%. Considering only the 27 patients with a positive (111In)leukocyte study, the combined bone scan and (111In)leukocyte study correctly localized the infection to the soft tissues or bone in 89%. Uninfected neurotrophic osteoarthropathy does not accumulate (111In)leukocytes. We found the combined bone scan and (111In) leukocyte study useful for the detection and localization of infection to soft tissue or bone in patients with neuropathic foot disease.

  17. The effects of space flight on polymorphonuclear leukocyte response experiment MA-032

    NASA Technical Reports Server (NTRS)

    Martin, R. R.

    1976-01-01

    In a series of studies performed at intervals from 30 day before flight to 30 days after recovery, blood samples were obtained from the three astronauts of the Apollo Soyuz Test Project and from eight control subjects. To determine the effects of space flight on polymorphonuclear leukocytes, tests were performed on blood samples obtained as quickly as possible after splashdown and on the day following recovery. The astronauts' inhalation of propellant gases and the inception of corticosteroid therapy 1 day after recovery provided an additional opportunity to investigate the possible effects of these factors on leukocyte function. Data were obtained during each time period on the total leukocyte count, differential count, leukocyte adhesion, leukocyte migration and chemotaxis, phagocytosis, and histochemical staining for leukocyte acid and alkaline phosphatase. These observations present a variety of in vitro correlates to white blood cell function within the body. Taken together, they serve as a reasonable approximation of the effects of space flight on leukocyte function.

  18. Big insights from small volumes: deciphering complex leukocyte behaviors using microfluidics.

    PubMed

    Irimia, Daniel; Ellett, Felix

    2016-08-01

    Inflammation is an indispensable component of the immune response, and leukocytes provide the first line of defense against infection. Although the major stereotypic leukocyte behaviors in response to infection are well known, the complexities and idiosyncrasies of these phenotypes in conditions of disease are still emerging. Novel tools are indispensable for gaining insights into leukocyte behavior, and in the past decade, microfluidic technologies have emerged as an exciting development in the field. Microfluidic devices are readily customizable, provide tight control of experimental conditions, enable high precision of ex vivo measurements of individual as well as integrated leukocyte functions, and have facilitated the discovery of novel leukocyte phenotypes. Here, we review some of the most interesting insights resulting from the application of microfluidic approaches to the study of the inflammatory response. The aim is to encourage leukocyte biologists to integrate these new tools into increasingly more sophisticated experimental designs for probing complex leukocyte functions. PMID:27194799

  19. Endothelial Japanese encephalitis virus infection enhances migration and adhesion of leukocytes to brain microvascular endothelia via MEK-dependent expression of ICAM1 and the CINC and RANTES chemokines.

    PubMed

    Lai, Ching-Yi; Ou, Yen-Chuan; Chang, Cheng-Yi; Pan, Hung-Chuan; Chang, Chen-Jung; Liao, Su-Lan; Su, Hong-Lin; Chen, Chun-Jung

    2012-10-01

    Currently, the underlying mechanisms and the specific cell types associated with Japanese encephalitis-associated leukocyte trafficking are not understood. Brain microvascular endothelial cells represent a functional barrier and could play key roles in leukocyte central nervous system trafficking. We found that cultured brain microvascular endothelial cells were susceptible to Japanese encephalitis virus (JEV) infection with limited amplification. This type of JEV infection had negligible effects on cell viability and barrier integrity. Instead, JEV-infected endothelial cells attracted more leukocytes adhesion onto surfaces and the supernatants promoted chemotaxis of leukocytes. Infection with JEV was found to elicit the elevated production of intercellular adhesion molecule-1, cytokine-induced neutrophil chemoattractant-1, and regulated-upon-activation normal T-cell expressed and secreted, contributing to the aforementioned leukocyte adhesion and chemotaxis. We further demonstrated that extracellular signal-regulated kinase was a key upstream regulator which stimulated extensive endothelial gene induction by up-regulating cytosolic phospholipase A₂, NF-κB, and cAMP response element-binding protein via signals involving phosphorylation. These data suggest that JEV infection could activate brain microvascular endothelial cells and modify their characteristics without compromising the barrier integrity, making them favorable for the recruitment and adhesion of circulating leukocytes, thereby together with other unidentified barrier-disrupting mechanisms contributing to Japanese encephalitis and associated neuroinflammation. PMID:22845610

  20. Methodological aspects of anti-human leukocyte antigen antibody analysis in solid organ transplantation

    PubMed Central

    Lobashevsky, Andrew L

    2014-01-01

    Donor human leukocyte antigen (HLA)-specific antibodies (DSA) play an important role in solid organ transplantation. Preexisting IgG isotype DSA are considered a risk factor for antibody mediated rejection, graft failure or graft loss. The post-transplant development of DSA depends on multiple factors including immunogenicity of mismatched antigens, HLA class II typing of the recipient, cytokine gene polymorphisms, and cellular immunoregulatory mechanisms. De novo developed antibodies require special attention because not all DSA have equal clinical significance. Therefore, it is important for transplant clinicians and transplant immunologists to accurately characterize DSA. In this review, the contemporary immunological techniques for detection and characterization of anti-HLA antibodies and their pitfalls are described. PMID:25346888

  1. The colonization of the Pacific: the story according to human leukocyte antigens.

    PubMed Central

    Serjeantson, S W; Ryan, D P; Thompson, A R

    1982-01-01

    The human leukocyte antigen (HLA) distributions in 16 Pacific populations have been collated from published and unpublished reports. Gene frequency and linkage disequilibrium relationships among groups show that Australians and Papuans share a common ancestry, that coastal Melanesia has about 16% Austronesian admixture, and that Fiji is truly intermediate between Melanesia and Polynesia. In Polynesia, Cook Islanders show closer affinity with populations of Western Polynesia than with Maoris and Easter Islanders, in contrast to their linguistic affiliations, but otherwise HLA distributions show a clear division between the populations of Eastern and Western Polynesia. This study emphasizes the contribution the HLA system can make to anthropological studies and has provided a version of colonization of the Pacific compatible with theories based on prodigious efforts in many disciplines. PMID:7180847

  2. Genotyping of human leukocyte antigen (HLA) ancestral haplotypes as prognostic marker in cancer using PCR analysis.

    PubMed

    Villabona, Lisa; Andersson, Emilia; Marchesi, Maddalena; Masucci, Giuseppe V

    2014-01-01

    The major histocompatibility complex (MHC) comprises a set of genes that are essential to immunity and surveillance against neoplastic transformation. MHC antigens not only regulate antitumor immune responses in experimental animal models but also directly correlate with survival and prognosis of patients with various types of cancers. Effective recognition of tumor cells by effector T cells may be affected by the genotype and the extent of expression of human leukocyte antigen (HLA)-peptide complexes. Therefore, MHC antigens may serve as potential biomarkers for prognosis and allow selection of cancer patients for specific therapy. We describe PCR-based method to determine the HLA genotype in healthy individuals and patients using blood and tumor tissue as DNA source. PMID:24258987

  3. Appropriate clinical use of human leukocyte antigen typing for coeliac disease: an Australasian perspective

    PubMed Central

    Tye-Din, J A; Cameron, D J S; Daveson, A J; Day, A S; Dellsperger, P; Hogan, C; Newnham, E D; Shepherd, S J; Steele, R H; Wienholt, L; Varney, M D

    2015-01-01

    The past decade has seen human leukocyte antigen (HLA) typing emerge as a remarkably popular test for the diagnostic work-up of coeliac disease with high patient acceptance. Although limited in its positive predictive value for coeliac disease, the strong disease association with specific HLA genes imparts exceptional negative predictive value to HLA typing, enabling a negative result to exclude coeliac disease confidently. In response to mounting evidence that the clinical use and interpretation of HLA typing often deviates from best practice, this article outlines an evidence-based approach to guide clinically appropriate use of HLA typing, and establishes a reporting template for pathology providers to improve communication of results. PMID:25827511

  4. Derivation of Cinnamon Blocks Leukocyte Attachment by Interacting with Sialosides.

    PubMed

    Lin, Wei-Ling; Guu, Shih-Yun; Tsai, Chan-Chuan; Prakash, Ekambaranellore; Viswaraman, Mohan; Chen, Hsing-Bao; Chang, Chuan-Fa

    2015-01-01

    Molecules derived from cinnamon have demonstrated diverse pharmacological activities against infectious pathogens, diabetes and inflammatory diseases. This study aims to evaluate the effect of the cinnamon-derived molecule IND02 on the adhesion of leukocytes to host cells. The anti-inflammatory ability of IND02, a pentameric procyanidin type A polyphenol polymer isolated from cinnamon alcohol extract, was examined. Pretreatment with IND02 significantly reduced the attachment of THP-1 cells or neutrophils to TNF-α-activated HUVECs or E-selectin/ICAM-1, respectively. IND02 also reduced the binding of E-, L- and P-selectins with sialosides. Furthermore, IND02 could agglutinate human red blood cells (RBC), and the agglutination could be disrupted by sialylated glycoprotein. Our findings demonstrate that IND02, a cinnamon-derived compound, can interact with sialosides and block the binding of selectins and leukocytes with sialic acids. PMID:26076445

  5. Signaling networks regulating leukocyte podosome dynamics and function

    PubMed Central

    Dovas, Athanassios; Cox, Dianne

    2011-01-01

    Podosomes are ventral adhesion structures prominent in cells of the myeloid lineage. A common aspect of these cells is that they are highly motile and are required to traverse multiple tissue barriers in order to perform their functions. Recently podosomes have gathered attention from researchers as important cellular structures that can influence cell adhesion, motility and matrix remodeling. Adhesive and soluble ligands act via transmembrane receptors and propagate signals to the leukocyte cytoskeleton via small G proteins of the Rho family, tyrosine kinases and scaffold proteins and are able to induce podosome formation and rearrangements. Manipulation of the signals that regulate podosome formation and dynamics can therefore be a strategy to interfere with leukocyte functions in a multitude of pathological settings, such as infections, atherosclerosis and arthritis. Here, we review the major signaling molecules that act in the formation and regulation of podosomes. PMID:21342664

  6. Derivation of Cinnamon Blocks Leukocyte Attachment by Interacting with Sialosides

    PubMed Central

    Lin, Wei-Ling; Guu, Shih-Yun; Tsai, Chan-Chuan; Prakash, Ekambaranellore; Viswaraman, Mohan; Chen, Hsing-Bao; Chang, Chuan-Fa

    2015-01-01

    Molecules derived from cinnamon have demonstrated diverse pharmacological activities against infectious pathogens, diabetes and inflammatory diseases. This study aims to evaluate the effect of the cinnamon-derived molecule IND02 on the adhesion of leukocytes to host cells. The anti-inflammatory ability of IND02, a pentameric procyanidin type A polyphenol polymer isolated from cinnamon alcohol extract, was examined. Pretreatment with IND02 significantly reduced the attachment of THP-1 cells or neutrophils to TNF-α-activated HUVECs or E-selectin/ICAM-1, respectively. IND02 also reduced the binding of E-, L- and P-selectins with sialosides. Furthermore, IND02 could agglutinate human red blood cells (RBC), and the agglutination could be disrupted by sialylated glycoprotein. Our findings demonstrate that IND02, a cinnamon-derived compound, can interact with sialosides and block the binding of selectins and leukocytes with sialic acids. PMID:26076445

  7. Dimensions of religious involvement and leukocyte telomere length.

    PubMed

    Hill, Terrence D; Ellison, Christopher G; Burdette, Amy M; Taylor, John; Friedman, Katherine L

    2016-08-01

    Although numerous studies suggest that religious involvement is associated with a wide range of favorable health outcomes, it is unclear whether this general pattern extends to cellular aging. In this paper, we tested whether leukocyte telomere length varies according to several dimensions of religious involvement. We used cross-sectional data from the Nashville Stress and Health Study (2011-2014), a large probability sample of 1252 black and white adults aged 22 to 69 living in Davidson County, TN, USA. Leukocyte telomere length was measured using the monochrome multiplex quantitative polymerase chain reaction method with albumin as the single-copy reference sequence. Dimensions of religious involvement included religiosity, religious support, and religious coping. Our multivariate analyses showed that religiosity (an index of religious attendance, prayer frequency, and religious identity) was positively associated with leukocyte telomere length, even with adjustments for religious support, religious coping, age, gender, race, education, employment status, income, financial strain, stressful life events, marital status, family support, friend support, depressive symptoms, smoking, heavy drinking, and allostatic load. Unlike religiosity, religious support and religious coping were unrelated to leukocyte telomere length across models. Depressive symptoms, smoking, heavy drinking, and allostatic load failed to explain any of the association between religiosity and telomere length. To our knowledge, this is the first population-based study to link religious involvement and cellular aging. Although our data suggest that adults who frequently attend religious services, pray with regularity, and consider themselves to be religious tend to exhibit longer telomeres than those who attend and pray less frequently and do not consider themselves to be religious, additional research is needed to establish the mechanisms underlying this association. PMID:27174242

  8. Myxoma and Vaccinia Viruses Bind Differentially to Human Leukocytes

    PubMed Central

    Chan, Winnie M.; Bartee, Eric C.; Moreb, Jan S.; Dower, Ken; Connor, John H.

    2013-01-01

    Myxoma virus (MYXV) and vaccinia virus (VACV), two distinct members of the family Poxviridae, are both currently being developed as oncolytic virotherapeutic agents. Recent studies have demonstrated that ex vivo treatment with MYXV can selectively recognize and kill contaminating cancerous cells from autologous bone marrow transplants without perturbing the engraftment of normal CD34+ hematopoietic stem and progenitor cells. However, the mechanism(s) by which MYXV specifically recognizes and eliminates the cancer cells in the autografts is not understood. While little is known about the cellular attachment factor(s) exploited by MYXV for entry into any target cells, VACV has been shown to utilize cell surface glycosaminoglycans such as heparan sulfate (HS), the extracellular matrix protein laminin, and/or integrin β1. We have constructed MYXV and VACV virions tagged with the Venus fluorescent protein and compared their characteristics of binding to various human cancer cell lines as well as to primary human leukocytes. We report that the binding of MYXV or VACV to some adherent cell lines could be partially inhibited by heparin, but laminin blocked only VACV binding. In contrast to cultured fibroblasts, the binding of MYXV and VACV to a wide spectrum of primary human leukocytes could not be competed by either HS or laminin. Additionally, MYXV and VACV exhibited very different binding characteristics against certain select human leukocytes, suggesting that the two poxviruses utilize different cell surface determinants for the attachment to these cells. These results indicate that VACV and MYXV can exhibit very different oncolytic tropisms against some cancerous human leukocytes. PMID:23388707

  9. Indium-111 leukocyte scintigraphy in Wegener's granulomatosis involving the spleen

    SciTech Connect

    Morayati, S.J.; Fink-Bennett, D.

    1986-12-01

    Indium-111-labeled leukocyte scintigraphy was performed on a 44-yr-old man to exclude an occult abscess. Four- and twenty-four-hour images of the abdomen revealed splenic photopenia except for a rim of activity medially. A subsequent computed tomography (CT) study demonstrated necrosis or hemorrhage of the spleen except for a medial rim. Exploratory laparotomy demonstrated necrotizing vasculitis with granuloma formation consistent with Wegener's granulomatosis and a rim of viable splenic tissue corresponding to the radionuclide and CT studies.

  10. Indium-111 leukocyte localization in infected prosthetic graft

    SciTech Connect

    Purnell, G.L.; Walker, C.W.; Allison, J.W.; Dalrymple, G.V. )

    1990-08-01

    Infective endocarditis can be difficult to prove, even in the face of strong clinical suspicion. A case in which standard methods of diagnosis failed to demonstrate endocarditis in a patient with recurrent Staphylococcus aureus bacteremia and porcine aortic valve is reported. An In-111 labelled leukocyte SPECT study demonstrated uptake in the aortic root and leaflets, and autopsy demonstrated vegetations on the leaflets. In-111 may prove useful in demonstrating endocarditis in patients with prosthetic valve infection.

  11. Leukocyte Trafficking to the Small Intestine and Colon.

    PubMed

    Habtezion, Aida; Nguyen, Linh P; Hadeiba, Husein; Butcher, Eugene C

    2016-02-01

    Leukocyte trafficking to the small and large intestines is tightly controlled to maintain intestinal immune homeostasis, mediate immune responses, and regulate inflammation. A wide array of chemoattractants, chemoattractant receptors, and adhesion molecules expressed by leukocytes, mucosal endothelium, epithelium, and stromal cells controls leukocyte recruitment and microenvironmental localization in intestine and in the gut-associated lymphoid tissues (GALTs). Naive lymphocytes traffic to the gut-draining mesenteric lymph nodes where they undergo antigen-induced activation and priming; these processes determine their memory/effector phenotypes and imprint them with the capacity to migrate via the lymph and blood to the intestines. Mechanisms of T-cell recruitment to GALT and of T cells and plasmablasts to the small intestine are well described. Recent advances include the discovery of an unexpected role for lectin CD22 as a B-cell homing receptor GALT, and identification of the orphan G-protein-coupled receptor 15 (GPR15) as a T-cell chemoattractant/trafficking receptor for the colon. GPR15 decorates distinct subsets of T cells in mice and humans, a difference in species that could affect translation of the results of mouse colitis models to humans. Clinical studies with antibodies to integrin α4β7 and its vascular ligand mucosal vascular addressin cell adhesion molecule 1 are proving the value of lymphocyte trafficking mechanisms as therapeutic targets for inflammatory bowel diseases. In contrast to lymphocytes, cells of the innate immune system express adhesion and chemoattractant receptors that allow them to migrate directly to effector tissue sites during inflammation. We review the mechanisms for innate and adaptive leukocyte localization to the intestinal tract and GALT, and discuss their relevance to human intestinal homeostasis and inflammation. PMID:26551552

  12. A Genetic Mosaic Screen of Essential Zygotic Genes in Caenorhabditis Elegans

    PubMed Central

    Bucher, E. A.; Greenwald, I.

    1991-01-01

    We have devised a simple genetic mosaic screen, which circumvents the difficulties posed by phenotypic analysis of early lethal mutants, to analyze essential zygotic genes in Caenorhabditis elegans. The screen attempts to distinguish genes involved in cell type and/or lineage specific processes such as determination, differentiation or morphogenesis from genes involved in general processes such as intermediary metabolism by using the pattern of gene function to classify genes: genes required in one or a subset of early blastomeres may have specific functions, whereas genes required in all early blastomeres may have general functions. We found that 12 of 17 genes examined function in specific early blastomeres, suggesting that many zygotic genes contribute to specific early processes. We discuss the advantages and limitations of this screen, which is applicable to other regions of the C. elegans genome. PMID:2071016

  13. Leukocyte recovery from umbilical cord blood by poligeline.

    PubMed

    Perutelli, P; Catellani, S

    1999-02-01

    Umbilical cord blood (UCB) collected at delivery is a source of transplantable stem/progenitor cells; it represents an alternative to bone marrow to restore hematopoiesis in patients affected by malignant and non-malignant disease. Therefore, large-scale UCB banks would be a natural complement to bone marrow donor registries. Storage of unmanipulated whole UCB units requires a great number of liquid nitrogen containers. Separation of leukocytes allows UCB storage in smaller space, thus lowering banking costs; unfortunately, UCB processing may cause significant losses of stem cells. We report about the use of poligeline to remove erythrocytes from UCB units. After erythrocyte sedimentation at 1xg (30' or 40') or 50xg, leukocyte-rich supernatant was collected and centrifuged to recover the leukocyte pool in view of stem cell transplantation. Erythrocyte depletion was always satisfactory, ranging from 82.6% to 88.9%, but 1xg sedimentation for 40' enabled us to achieve the best CD34+ cell recovery (mean value 80.5%). The proposed UCB-processing method allowed us to lower the final sample volume down to 1/10 of the initial one, in this way making UCB banking feasible. Erythrocyte depletion took place directly in the collection bag, thus reducing microbial contamination risk. PMID:10193639

  14. Curcumin modulates leukocyte and platelet adhesion in murine sepsis

    PubMed Central

    Vachharajani, Vidula; Wang, Si-Wei; Mishra, Nilamadhab; El-Gazzar, Mohammad; Yoza, Barbara; McCall, Charles

    2010-01-01

    Objective Circulating cell-endothelial cell interaction in sepsis is a rate-determining factor in organ dysfunction, and interventions targeting this process have a potential therapeutic value. In this project, we examined whether curcumin, an active ingredient of turmeric and an anti-inflammatory agent, could disrupt interactions between circulating blood cells and endothelium and improve survival in a murine model of sepsis. Methods Mice were subjected to cecal ligation and puncture (CLP) to induce sepsis vs. sham surgery. We studied leukocyte and platelet adhesion in cerebral microcirculation using intravital fluorescent video microscopy technique, blood brain barrier dysfunction using Evans Blue leakage method, P-selectin expression using dual radiolabeling technique and survival in mice subjected to Sham, CLP and CLP with curcumin pre-treatment (CLP+Curcumin). Results Curcumin significantly attenuated leukocyte and platelet adhesion in cerebral microcirculation, Evans Blue leakage in the brain tissue and improved survival in mice with CLP. P-selectin expression in mice with CLP+Curcumin was significantly attenuated compared to CLP in various microcirculatory beds including brain. Reduction in platelet adhesion was predominantly via modulation of endothelium by curcumin. Conclusion Curcumin pre-treatment modulates leukocyte and platelet adhesion and blood brain barrier dysfunction in mice with CLP via P-selectin expression and improves survival in mice with CLP. PMID:20690979

  15. Exposure to mercury alters early activation events in fish leukocytes.

    PubMed Central

    MacDougal, K C; Johnson, M D; Burnett, K G

    1996-01-01

    Although fish in natural populations may carry high body burdens of both organic and inorganic mercury, the effects of this divalent metal on such lower vertebrates is poorly understood. In this report, inorganic mercury in the form of mercuric chloride (HgCl2) is shown to produce both high-dose inhibition and low-dose activation of leukocytes in a marine teleost fish, Sciaenops ocellatus. Concentrations of inorganic mercury > or = 10 microM suppressed DNA synthesis and induced rapid influx of radiolabeled calcium, as well as tyrosine phosphorylation of numerous cellular proteins. Lower concentrations (0.1-1 microM) of HgCl2 that activated cell growth also induced a slow sustained rise in intracellular calcium in cells loaded with the calcium indicator dye fura-2, but did not produce detectable tyrosine phosphorylation of leukocyte proteins. These studies support the possibility that subtoxic doses of HgCl2 may inappropriately activate teleost leukocytes, potentially altering the processes that regulate the magnitude and specificity of the fish immune response to environmental pathogens. Images Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. Figure 6. Figure 7. PMID:8930553

  16. Report: Nuclei segmentation of leukocytes in blood smear digital images.

    PubMed

    Abbas, Naveed; Mohamad, Dzulkifli; Abdullah, Abdul Hanan; Saba, Tanzila; Al-Rodhaan, Mznah; Al-Dhelaan, Abdullah

    2015-09-01

    The Leukocytes are differentiated from each other on the basis of their nuclei, demanded in many Medical studies, especially in all types of Leukemia by the Hematologists to note the disorder caused by specific type of Leukocyte. Leukemia is a life threatening disease. The work for diagnosing is manually carried out by the Hematologists involving much labor, time and human errors. The problems mentioned are easily addressed through computer vision techniques, but still accuracy and efficiency are demanded in terms of the basic and challenging step segmentation of Leukocyte's nuclei. The underlying study proposed better method in terms of accuracy and efficiency by designing a dynamic convolution filter for boosting low intensity values in the separated green channel of an RGB image and suppressing the high values in the same channel. The high values in the green channel become 255 (background) while the nuclei always have low values in the green channel and thus clearly appear as foreground. The proposed technique is tested on 365 images achieving an overall accuracy of 95.89%, while improving the efficiency by 10%. The proposed technique achieved its targets in a realistic way by improving the accuracy as well as the efficiency and both are highly required in the area. PMID:26408877

  17. Imaging leukocytes in vivo with third harmonic generation microscopy

    NASA Astrophysics Data System (ADS)

    Tsai, Cheng-Kun; Chen, Chien-Kuo; Chen, Yu-Shing; Wu, Pei-Chun; Hsieh, Tsung-Yuan; Liu, Han-Wen; Yeh, Chiou-Yueh; Lin, Win-Li; Chia, Jean-San; Liu, Tzu-Ming

    2013-02-01

    Without a labeling, we demonstrated that lipid granules in leukocytes have distinctive third harmonic generation (THG) contrast. Excited by a 1230nm femtosecond laser, THG signals were generated at a significantly higher level in neutrophils than other mononuclear cells, whereas signals in agranular lymphocytes were one order smaller. These characteristic THG features can also be observed in vivo to trace the newly recruited leukocytes following lipopolysaccharide (LPS) challenge. Furthermore, using video-rate THG microscopy, we also captured images of blood cells in human capillaries. Quite different from red-blood-cells, every now and then, round and granule rich blood cells with strong THG contrast appeared in circulation. The corresponding volume densities in blood, evaluated from their frequencies of appearance and the velocity of circulation, fall within the physiological range of human white blood cell counts. These results suggested that labeling-free THG imaging may provide timely tracing of leukocyte movement and hematology inspection without disturbing the normal cellular or physiological status.

  18. Influence of tetracyclines on human polymorphonuclear leukocyte function.

    PubMed Central

    Glette, J; Sandberg, S; Hopen, G; Solberg, C O

    1984-01-01

    Low concentrations of oxytetracycline, doxycycline, or minocycline (less than 10 micrograms/ml) did not influence in vitro polymorphonuclear leukocyte random migration, chemiluminescence, or glucose oxidation. At high concentrations of doxycycline or minocycline (greater than 10 micrograms/ml), chemiluminescence and glucose oxidation were impaired. High concentrations of doxycycline also reduced random migration. Oxytetracycline did not influence these functions in concentrations up to 100 micrograms/ml. The inhibiting effect of doxycycline and minocycline was abolished when 4 mM Mg2+ was added to the reaction mixture, and 4 mM Ca2+ partly restored minocycline-inhibited polymorphonuclear leukocyte functions. This indicates that the major effect of tetracyclines on in vitro polymorphonuclear leukocyte functions is mediated by their divalent cation chelating effect and that the results of in vitro experiments are highly dependent on the concentration of divalent cations in the reaction mixtures. The difference between the tetracyclines may be due to differences in lipid solubility, with solubility being highest for minocycline and lowest for oxytetracycline, or to different divalent cation chelating ability. PMID:6721468

  19. Tracking flow of leukocytes in blood for drug analysis

    NASA Astrophysics Data System (ADS)

    Basharat, Arslan; Turner, Wesley; Stephens, Gillian; Badillo, Benjamin; Lumpkin, Rick; Andre, Patrick; Perera, Amitha

    2011-03-01

    Modern microscopy techniques allow imaging of circulating blood components under vascular flow conditions. The resulting video sequences provide unique insights into the behavior of blood cells within the vasculature and can be used as a method to monitor and quantitate the recruitment of inflammatory cells at sites of vascular injury/ inflammation and potentially serve as a pharmacodynamic biomarker, helping screen new therapies and individualize dose and combinations of drugs. However, manual analysis of these video sequences is intractable, requiring hours per 400 second video clip. In this paper, we present an automated technique to analyze the behavior and recruitment of human leukocytes in whole blood under physiological conditions of shear through a simple multi-channel fluorescence microscope in real-time. This technique detects and tracks the recruitment of leukocytes to a bioactive surface coated on a flow chamber. Rolling cells (cells which partially bind to the bioactive matrix) are detected counted, and have their velocity measured and graphed. The challenges here include: high cell density, appearance similarity, and low (1Hz) frame rate. Our approach performs frame differencing based motion segmentation, track initialization and online tracking of individual leukocytes.

  20. Carbohydrate ligands for endothelial - Leukocyte adhesion molecule 1

    SciTech Connect

    Tiemeyer, M.; Swiedler, S.J.; Ishihara, Masayuki; Moreland, M.; Schweingruber, H.; Hirtzer, P.; Brandley, B.K. )

    1991-02-15

    The acute inflammatory response requires that circulating leukocytes bind to and penetrate the vascular wall to access the site of injury. Several receptors have been implicated in this interaction, including a family of putative carbohydrate-binding proteins. The authors report here the identification of an endogenous carbohydrate ligand for one of these receptors, endothelial-leukocyte adhesion molecule 1 (ELAM-1). Radiolabeled COS cells transfected with a plasmid containing the cDNA for ELAM-1 were used as probes to screen glycolipids extracted from human leukocytes. COS cells transfected with this plasmid adhered to a subset of sialylated glycolipids resolved on TLC plates or adsorbed on polyvinyl chloride microtiter wells. Adhesion to these glycolipids required calcium but was not inhibited by heparin, chondroitin sulfate, keratan sulfate, or yeast phosphomannan. Monosaccharide composition, linkage analysis, and fast atom bombardment mass spectrometry of the glycolipids indicate that the ligands for ELAM-1 are terminally sialylated lactosylceramides with a variable number of N-acetyllactosamine repeats and at least one fucosylated N-acetylglucosamine residue.

  1. Leukocyte trafficking: Can we bring the fight to the tumor?

    PubMed

    Pachynski, Russell; Nazha, Jonathon; Kohrt, Holbrook

    2016-03-01

    Control of leukocyte trafficking plays a critical role in the establishment of effective immune responses. It is now well established that the number or ratio of effector to suppressor immune cells within the tumor microenvironment can significantly impact tumor growth and clinical outcomes. Recently approved immunotherapies by the FDA, and those in development, aim to stimulate effector immune cell function. For example, many checkpoint inhibitors seek to stimulate an immune response to tumors by reversing T-cell exhaustion. However, activation of the immune response outside the tumor microenvironment can lead to sometimes fatal immune-mediated adverse events -- the result of "on-target, off-tumor" effects. Thus, control of localization of these activated effector cells remains a critical component of optimizing tumor response while minimizing immune-mediated adverse events. Chemokines and chemoattractants, along with their receptors on immune cells, govern leukocyte trafficking; thus, understanding their expression pattern in the context of the tumor microenvironment and developing approaches to favorably alter those should lead to improved efficacy of current immunotherapeutics. This review highlights the background of cancer immunotherapy, leukocyte trafficking, and some novel approaches being utilized to optimize recruitment of effector immune cells into the tumor microenvironment. Future combinatorial immunotherapy should incorporate therapeutics aimed at 1) favorably altering the tumor microenvironment, 2) activating effector immune cells, and 3) optimizing effector cell trafficking into tumors. PMID:27115171

  2. Influence of delivery on numbers of leukocytes, leukocyte subpopulations, and hematopoietic progenitor cells in human umbilical cord blood.

    PubMed

    Lim, F T; van Winsen, L; Willemze, R; Kanhai, H H; Falkenburg, J H

    1994-01-01

    Human umbilical cord blood (UCB) may be used as an alternative source of bone marrow repopulating cells in allogeneic bone marrow transplantation. The quality and quantity of UCB harvests for transplantation is affected by several factors. In this study we analyzed the influence of delivery, in particular stress during delivery, on the numbers of leukocytes and leukocyte subsets in UCB. Four groups of women with different types of deliveries were included in the study, and from each group samples of UCB were analyzed. Blood samples from healthy adults were used as control. In UCB there was a higher absolute number of leukocytes than in peripheral blood (PB). UCB leukocytes were highest after deliveries with a prolonged second stage of labor, which was mainly due to granulocytosis. The percentage of T cells in UCB was lower than in PB, in particular when stress during delivery was higher. In all groups, however, the absolute concentration of T cells per milliliter of UCB was higher than in adult PB. The differences in T cells in stressful deliveries were mainly due to a relative decrease in CD3+/CD4+ cells in UCB. The relative frequency and absolute concentration of the CD56+ cell population in UCB was higher than in PB, which was mostly due to an increase of CD2-/CD56+ cells, in particular in stressful deliveries. The absolute number of CD34+ cells as well as hematopoietic progenitor cells as determined in semisolid medium cultures was high in UCB and was increased in cases of prolonged secondary stage of labor. This study demonstrates that the quality of UCB transplants is influenced by the course of delivery, in particular by stress during delivery. PMID:7749120

  3. Building Developmental Gene Regulatory Networks

    PubMed Central

    Li, Enhu; Davidson, Eric H.

    2009-01-01

    Animal development is an elaborate process programmed by genomic regulatory instructions. Regulatory genes encode transcription factors and signal molecules, and their expression is under the control of cis-regulatory modules that define the logic of transcriptional responses to the inputs of other regulatory genes. The functional linkages amongst regulatory genes constitute the gene regulatory networks (GRNs) that govern cell specification and patterning in development. Constructing such networks requires identification of the regulatory genes involved and characterization of their temporal and spatial expression patterns. Interactions (activation/repression) among transcription factors or signals can be investigated by large-scale perturbation analysis, in which the function of each gene is specifically blocked. Resultant expression changes are then integrated to identify direct linkages, and to reveal the structure of the GRN. Predicted GRN linkages can be tested and verified by cis-regulatory analysis. The explanatory power of the GRN was shown in the lineage specification of sea urchin endomesoderm. Acquiring such networks is essential for a systematic and mechanistic understanding of the developmental process. PMID:19530131

  4. Role of nitric oxide in tumor microcirculation. Blood flow, vascular permeability, and leukocyte-endothelial interactions.

    PubMed Central

    Fukumura, D.; Yuan, F.; Endo, M.; Jain, R. K.

    1997-01-01

    The present study was designed to define the role of nitric oxide (NO) in tumor microcirculation, through the direct intravital microcirculatory observations after administration of NO synthase (NOS) inhibitor and NO donor both regionally and systemically. More specifically, we tested the following hypotheses: 1) endogenous NO derived from tumor vascular endothelium and/or tumor cells increases and/or maintains tumor blood flow, decreases leukocyte-endothelial interactions, and increases vascular permeability, 2) exogenous NO can increase tumor blood flow via vessel dilatation and decrease leukocyte-endothelial interactions, and 3) NO production and tissue responses to NO are tumor dependent. To this end, a murine mammary adenocarcinoma (MCaIV) and a human colon adenocarcinoma (LS174T) were implanted in the dorsal skinfold chamber in C3H and severe combined immunodeficient mice, respectively, and observed by means of intravital fluorescence microscopy. Both regional and systemic inhibition of endogenous NO by N omega-nitro-L-arginine methyl ester (L-NAME; 100 mumol/L superfusion or 10 mg/kg intravenously) significantly decreased vessel diameter and local blood flow rate. The diameter change was dominant on the arteriolar side. Superfusion of NO donor (spermine NO, 100 mumol/L) increased tumor vessel diameter and flow rate, whereas systemic injection of spermine NO (2.62 mg/kg) had no significant effect on these parameters. Rolling and stable adhesion of leukocytes were significantly increased by intravenous injection of L-NAME. In untreated animals, both MCaIV and LS174T tumor vessels were leaky to albumin. Systemic NO inhibition significantly attenuated tumor vascular permeability of MCaIV but not of LS174T tumor. Immunohistochemical studies, using polyclonal antibodies to endothelial NOS and inducible NOS, revealed a diffuse pattern of positive labeling in both MCaIV and LS174T tumors. Nitrite and nitrate levels in tumor interstitial fluid of MCaIV but not of LS

  5. Altered Immunogenicity of Donor Lungs via Removal of Passenger Leukocytes Using Ex Vivo Lung Perfusion.

    PubMed

    Stone, J P; Critchley, W R; Major, T; Rajan, G; Risnes, I; Scott, H; Liao, Q; Wohlfart, B; Sjöberg, T; Yonan, N; Steen, S; Fildes, J E

    2016-01-01

    Passenger leukocyte transfer from the donor lung to the recipient is intrinsically involved in acute rejection. Direct presentation of alloantigen expressed on donor leukocytes is recognized by recipient T cells, promoting acute cellular rejection. We utilized ex vivo lung perfusion (EVLP) to study passenger leukocyte migration from donor lungs into the recipient and to evaluate the effects of donor leukocyte depletion prior to transplantation. For this purpose, female pigs received male left lungs either following 3 h of EVLP or retrieved using standard protocols. Recipients were monitored for 24 h and sequential samples were collected. EVLP-reduced donor leukocyte transfer into the recipient and migration to recipient lymph nodes was markedly reduced. Recipient T cell infiltration of the donor lung was significantly diminished via EVLP. Donor leukocyte removal during EVLP reduces direct allorecognition and T cell priming, diminishing recipient T cell infiltration, the hallmark of acute rejection. PMID:26366523

  6. Endothelial reticulon-4B (Nogo-B) regulates ICAM-1-mediated leukocyte transmigration and acute inflammation.

    PubMed

    Di Lorenzo, Annarita; Manes, Thomas D; Davalos, Alberto; Wright, Paulette L; Sessa, William C

    2011-02-17

    The reticulon (Rtn) family of proteins are localized primarily to the endoplasmic reticulum (ER) of most cells. The Rtn-4 family, (aka Nogo) consists of 3 splice variants of a common gene called Rtn-4A, Rtn-4B, and Rtn-4C. Recently, we identified the Rtn-4B (Nogo-B) protein in endothelial and smooth muscle cells of the vessel wall, and showed that Nogo-B is a regulator of cell migration in vitro and vascular remodeling and angiogenesis in vivo. However, the role of Nogo-B in inflammation is still largely unknown. In the present study, we use 2 models of inflammation to show that endothelial Nogo-B regulates leukocyte transmigration and intercellular adhesion molecule-1 (ICAM-1)-dependent signaling. Mice lacking Nogo-A/B have a marked reduction in neutrophil and monocyte recruitment to sites of inflammation, while Nogo-A/B(-/-) mice engrafted with wild-type (WT) bone marrow still exhibit impaired inflammation compared with WT mice engrafted with Nogo-A/B(-/-) bone marrow, arguing for a critical role of host Nogo in this response. Using human leukocytes and endothelial cells, we show mechanistically that the silencing of Nogo-B with small interfering RNA (siRNA) impairs the transmigration of neutrophils and reduces ICAM-1-stimulated phosphorylation of vascular endothelial-cell cadherin (VE-cadherin). Our results reveal a novel role of endothelial Nogo-B in basic immune functions and provide a key link in the molecular network governing endothelial-cell regulation of diapedesis. PMID:21183689

  7. Bovine leukocyte adhesion deficiency: a brief overview of a modern disease and its implications.

    PubMed

    Gerardi, A S

    1996-01-01

    Bovine Leukocyte Adhesion Deficiency (BLAD) is a genetic disease of cattle affecting the hematopoietic system. In the last decade BLAD has become a disease of economic importance in the dairy industry. As such, this overview describes the chronological developments and thinking that led to the elucidation of BLAD as a distinct disease entity from previous models in canine and human populations. All species affected exhibit symptoms of chronic and recurrent infections. Necrotic and/or gangrenous infections of soft tissues are prevalent, as well as secondary infections with bacteria or fungi. Low birthweight and unthriftiness are key symptoms of neonates in all species affected by LAD. Dermatomycoses and impaired pus formation are also common findings. The physiological basis for BLAD is a deficiency in leukocyte (particularly neutrophil) chemotactic and phagocytic properties. The inhibition of diapedesis in the inflammatory response prevents normal immune reactions to invading pathogens. Chronic infections are a consequence of the faulty immune mechanisms. The biochemical etiology of BLAD involves cell surface glycoprotein molecules known as integrins. These are responsible for cell-cell interactions necessary for neutrophils to adhere to vascular endothelium in a normal individual. Experiments using monoclonal antibodies to block LFA-1, Mac-1, and p150,95 (three integrins vital for cell-cell interactions) mimic BLAD symptomatology and have led to the discovery of the reciprocal Intercellular Adhesion Molecule (ICAM). Through pedigree analysis and biochemical detection with restrictive endonucleases BLAD has been isolated genetically to a single gene locus. The economic significance and prophylaxis are briefly discussed. In addition, the beneficial aspects of the study of BLAD are addressed. There are advantages of producing a BLAD-like state in preventing transplant rejection, ischemia-reperfusion injury, and other scenarios arising from the deleterious effects of

  8. Leukocyte Populations in Human Preterm and Term Breast Milk Identified by Multicolour Flow Cytometry

    PubMed Central

    Trend, Stephanie; de Jong, Emma; Lloyd, Megan L.; Kok, Chooi Heen; Richmond, Peter; Doherty, Dorota A.; Simmer, Karen; Kakulas, Foteini; Strunk, Tobias; Currie, Andrew

    2015-01-01

    Background Extremely preterm infants are highly susceptible to bacterial infections but breast milk provides some protection. It is unknown if leukocyte numbers and subsets in milk differ between term and preterm breast milk. This study serially characterised leukocyte populations in breast milk of mothers of preterm and term infants using multicolour flow cytometry methods for extended differential leukocyte counts in blood. Methods Sixty mothers of extremely preterm (<28 weeks gestational age), very preterm (28–31 wk), and moderately preterm (32–36 wk), as well as term (37–41 wk) infants were recruited. Colostrum (d2–5), transitional (d8–12) and mature milk (d26–30) samples were collected, cells isolated, and leukocyte subsets analysed using flow cytometry. Results The major CD45+ leukocyte populations circulating in blood were also detectable in breast milk but at different frequencies. Progression of lactation was associated with decreasing CD45+ leukocyte concentration, as well as increases in the relative frequencies of neutrophils and immature granulocytes, and decreases in the relative frequencies of eosinophils, myeloid and B cell precursors, and CD16- monocytes. No differences were observed between preterm and term breast milk in leukocyte concentration, though minor differences between preterm groups in some leukocyte frequencies were observed. Conclusions Flow cytometry is a useful tool to identify and quantify leukocyte subsets in breast milk. The stage of lactation is associated with major changes in milk leukocyte composition in this population. Fresh preterm breast milk is not deficient in leukocytes, but shorter gestation may be associated with minor differences in leukocyte subset frequencies in preterm compared to term breast milk. PMID:26288195

  9. An Incidental Detection of Popliteal Vein Aneurysm during Labeled Leukocyte Scintigraphy

    PubMed Central

    Hassan, H. Abu.; Nazri, M.; Azman, R. R.

    2012-01-01

    Technetium (99mTc) exametazime (hexamethylpropyleneamine oxime, HMPAO) labeled leukocyte scintigraphy is mainly used to exclude occult infection in our institution. On review of previously published article, no case of popliteal venous aneurysm was ever diagnosed and detected on labeled leukocyte scintigraphy. We present a rare case of popliteal venous aneurysm which was detected on labeled leukocyte scintigraphy and was further confirmed with single-photon emission computed tomography and computed tomography fusion. PMID:23372443

  10. Evolution of the Vertebrate Resistin Gene Family

    PubMed Central

    Hu, Qingda; Tan, Huanran; Irwin, David M.

    2015-01-01

    Resistin (encoded by Retn) was previously identified in rodents as a hormone associated with diabetes; however human resistin is instead linked to inflammation. Resistin is a member of a small gene family that includes the resistin-like peptides (encoded by Retnl genes) in mammals. Genomic searches of available genome sequences of diverse vertebrates and phylogenetic analyses were conducted to determine the size and origin of the resistin-like gene family. Genes encoding peptides similar to resistin were found in Mammalia, Sauria, Amphibia, and Actinistia (coelacanth, a lobe-finned fish), but not in Aves or fish from Actinopterygii, Chondrichthyes, or Agnatha. Retnl originated by duplication and transposition from Retn on the early mammalian lineage after divergence of the platypus, but before the placental and marsupial mammal divergence. The resistin-like gene family illustrates an instance where the locus of origin of duplicated genes can be identified, with Retn continuing to reside at this location. Mammalian species typically have a single copy Retn gene, but are much more variable in their numbers of Retnl genes, ranging from 0 to 9. Since Retn is located at the locus of origin, thus likely retained the ancestral expression pattern, largely maintained its copy number, and did not display accelerated evolution, we suggest that it is more likely to have maintained an ancestral function, while Retnl, which transposed to a new location, displays accelerated evolution, and shows greater variability in gene number, including gene loss, likely evolved new, but potentially lineage-specific, functions. PMID:26076481

  11. Evolution of the Vertebrate Resistin Gene Family.

    PubMed

    Hu, Qingda; Tan, Huanran; Irwin, David M

    2015-01-01

    Resistin (encoded by Retn) was previously identified in rodents as a hormone associated with diabetes; however human resistin is instead linked to inflammation. Resistin is a member of a small gene family that includes the resistin-like peptides (encoded by Retnl genes) in mammals. Genomic searches of available genome sequences of diverse vertebrates and phylogenetic analyses were conducted to determine the size and origin of the resistin-like gene family. Genes encoding peptides similar to resistin were found in Mammalia, Sauria, Amphibia, and Actinistia (coelacanth, a lobe-finned fish), but not in Aves or fish from Actinopterygii, Chondrichthyes, or Agnatha. Retnl originated by duplication and transposition from Retn on the early mammalian lineage after divergence of the platypus, but before the placental and marsupial mammal divergence. The resistin-like gene family illustrates an instance where the locus of origin of duplicated genes can be identified, with Retn continuing to reside at this location. Mammalian species typically have a single copy Retn gene, but are much more variable in their numbers of Retnl genes, ranging from 0 to 9. Since Retn is located at the locus of origin, thus likely retained the ancestral expression pattern, largely maintained its copy number, and did not display accelerated evolution, we suggest that it is more likely to have maintained an ancestral function, while Retnl, which transposed to a new location, displays accelerated evolution, and shows greater variability in gene number, including gene loss, likely evolved new, but potentially lineage-specific, functions. PMID:26076481

  12. GATA1 and PU.1 Bind to Ribosomal Protein Genes in Erythroid Cells: Implications for Ribosomopathies

    PubMed Central

    Amanatiadou, Elsa P.; Papadopoulos, Giorgio L.; Strouboulis, John; Vizirianakis, Ioannis S.

    2015-01-01

    The clear connection between ribosome biogenesis dysfunction and specific hematopoiesis-related disorders prompted us to examine the role of critical lineage-specific transcription factors in the transcriptional regulation of ribosomal protein (RP) genes during terminal erythroid differentiation. By applying EMSA and ChIP methodologies in mouse erythroleukemia cells we show that GATA1 and PU.1 bind in vitro and in vivo the proximal promoter region of the RPS19 gene which is frequently mutated in Diamond-Blackfan Anemia. Moreover, ChIPseq data analysis also demonstrates that several RP genes are enriched as potential GATA1 and PU.1 gene targets in mouse and human erythroid cells, with GATA1 binding showing an association with higher ribosomal protein gene expression levels during terminal erythroid differentiation in human and mouse. Our results suggest that RP gene expression and hence balanced ribosome biosynthesis may be specifically and selectively regulated by lineage specific transcription factors during hematopoiesis, a finding which may be clinically relevant to ribosomopathies. PMID:26447946

  13. GATA1 and PU.1 Bind to Ribosomal Protein Genes in Erythroid Cells: Implications for Ribosomopathies.

    PubMed

    Amanatiadou, Elsa P; Papadopoulos, Giorgio L; Strouboulis, John; Vizirianakis, Ioannis S

    2015-01-01

    The clear connection between ribosome biogenesis dysfunction and specific hematopoiesis-related disorders prompted us to examine the role of critical lineage-specific transcription factors in the transcriptional regulation of ribosomal protein (RP) genes during terminal erythroid differentiation. By applying EMSA and ChIP methodologies in mouse erythroleukemia cells we show that GATA1 and PU.1 bind in vitro and in vivo the proximal promoter region of the RPS19 gene which is frequently mutated in Diamond-Blackfan Anemia. Moreover, ChIPseq data analysis also demonstrates that several RP genes are enriched as potential GATA1 and PU.1 gene targets in mouse and human erythroid cells, with GATA1 binding showing an association with higher ribosomal protein gene expression levels during terminal erythroid differentiation in human and mouse. Our results suggest that RP gene expression and hence balanced ribosome biosynthesis may be specifically and selectively regulated by lineage specific transcription factors during hematopoiesis, a finding which may be clinically relevant to ribosomopathies. PMID:26447946

  14. Imaging of Leukocyte Trafficking in Alzheimer’s Disease

    PubMed Central

    Pietronigro, Enrica; Zenaro, Elena; Constantin, Gabriela

    2016-01-01

    Alzheimer’s disease (AD) is the most common neurodegenerative disorder and is characterized by a progressive decline of cognitive functions. The neuropathological features of AD include amyloid beta (Aβ) deposition, intracellular neurofibrillary tangles derived from the cytoskeletal hyperphosphorylated tau protein, amyloid angiopathy, the loss of synapses, and neuronal degeneration. In the last decade, inflammation has emerged as a key feature of AD, but most studies have focused on the role of microglia-driven neuroinflammation mechanisms. A dysfunctional blood–brain barrier has also been implicated in the pathogenesis of AD, and several studies have demonstrated that the vascular deposition of Aβ induces the expression of adhesion molecules and alters the expression of tight junction proteins, potentially facilitating the transmigration of circulating leukocytes. Two-photon laser scanning microscopy (TPLSM) has become an indispensable tool to dissect the molecular mechanisms controlling leukocyte trafficking in the central nervous system (CNS). Recent TPLSM studies have shown that vascular deposition of Aβ in the CNS promotes intraluminal neutrophil adhesion and crawling on the brain endothelium and also that neutrophils extravasate in the parenchyma preferentially in areas with Aβ deposits. These studies have also highlighted a role for LFA-1 integrin in neutrophil accumulation in the CNS of AD-like disease models, revealing that LFA-1 inhibition reduces the corresponding cognitive deficit and AD neuropathology. In this article, we consider how current imaging techniques can help to unravel new inflammation mechanisms in the pathogenesis of AD and identify novel therapeutic strategies to treat the disease by interfering with leukocyte trafficking mechanisms. PMID:26913031

  15. Phosphorylation of leukocyte PECAM and its association with detergent-resistant membranes regulate transendothelial migration.

    PubMed

    Florey, Oliver; Durgan, Joanne; Muller, William

    2010-08-01

    Leukocyte migration across the endothelial lining is a critical step in the body's response to infection and inflammation. The homophilic interaction between endothelial PECAM and leukocyte PECAM is essential for this process. The molecular events that are triggered in the endothelial cell by PECAM engagement have been well characterized; however, the function of leukocyte PECAM remains to be elucidated. To study this, we first blocked leukocyte transmigration using anti-PECAM Ab and then specifically activated leukocyte PECAM. This was sufficient to overcome the block and promote transmigration, suggesting an active signaling role for leukocyte PECAM. Consistent with this, we found that ligation of leukocyte PECAM induces phosphorylation of two tyrosine residues on its cytoplasmic tail. By performing RNA interference-rescue experiments, we demonstrate that these phosphorylation events are indispensable for transendothelial migration. Finally, we show that leukocyte PECAM translocates to a detergent-resistant membrane (DRM) during transmigration. PECAM localized in DRMs displays reduced phosphorylation and does not support transmigration. Together, these data support a model whereby engagement of leukocyte PECAM induces its transient tyrosine phosphorylation and induction of downstream signals that drive transmigration. These signals are then downregulated following PECAM translocation to DRMs. PMID:20581150

  16. Kinetics of reversible-sequestration of leukocytes by the isolated perfused rat lung

    SciTech Connect

    Goliaei, B.

    1980-08-01

    The kinetics and morphology of sequestration and margination of rat leukocytes were studied using an isolated perfused and ventilated rat lung preparation. Whole rat blood, bone marrow suspension, or leukocyte suspensions, were used to perfuse the isolated rat lung. The lung was also perfused with latex particle suspensions and the passage of particles through the lung capillaries was studied. When a leukocyte suspension was perfused through the lung in the single-pass mode, the rate of sequestration decreased as more cells were perfused. In contrast, latex particles of a size comparable to that of leukocytes were totally stopped by the lung. When the leukocyte suspension was recirculated through the lung, cells were rapidly removed from circulation until a steady state was reached, after which no net removal of cells by the lung occurred. These results indicate that leukocytes are reversibly sequestered from circulation. The sequestered cells marginated and attached to the luminal surface of the endothelium of post-capillary venules and veins. A mathematical model was developed based on the assumption that the attachment and detachment of leukocytes to blood vessel walls follows first-order kinetics. The model correctly predicts the following characteristics of the system: (a) the kinetics of the sequestration of leukocytes by the lung; (b) the existence of a steady state when a suspension of leukocytes is recirculated through the lung; and (c) the independence of the fraction of cells remaining in circulation from the starting concentration for all values of starting concentration. (ERB)

  17. BK Nephritis and Venous Thrombosis in Renal Transplant Recipient Detected by 111In Leukocyte Imaging.

    PubMed

    Pucar, Darko; Klein, Kandace; Corley, James; Williams, Hadyn T

    2015-07-01

    Three months after deceased donor kidney transplant, a patient who presented with proteinuric renal dysfunction and fever of undetermined origin was found to have BK viruria by quantitative polymerase chain reaction analysis. An ¹¹¹In leukocyte scan showed increased renal transplant uptake consistent with nephritis and linear uptake in the knee. Venous duplex ultrasound revealed acute occlusive thrombosis in the superficial right lesser saphenous vein in the area of increased radiolabeled leukocyte uptake. This ¹¹¹In leukocyte scan performed for fever of undetermined origin demonstrated findings of BK nephritis in a renal transplant patient and associated acute venous thrombosis related to leukocyte colonization. PMID:26018698

  18. The Study of Leukocyte Functions in a Rotating Wall Vessel

    NASA Technical Reports Server (NTRS)

    Trial, JoAnn

    1998-01-01

    The objective of this study was to investigate the behavior of leukocytes under free-fall conditions in a rotating wall vessel. In such a vessel, the tendency of a cell to fall in response to gravity is opposed by the rotation of the vessel and the culture medium within, keeping the cells in suspension without fluid shear. Previous reports indicated that such functions as lymphocyte migration through collagen matrix or monocyte cytokine secretion are altered under these conditions, and these changes correlate with similar functional defects of cultured cells seen during spaceflight.

  19. Theileria-transformed bovine leukocytes have cancer hallmarks.

    PubMed

    Tretina, Kyle; Gotia, Hanzel T; Mann, David J; Silva, Joana C

    2015-07-01

    The genus Theileria includes tick-transmitted apicomplexan parasites of ruminants with substantial economic impact in endemic countries. Some species, including Theileria parva and Theileria annulata, infect leukocytes where they induce phenotypes that are shared with some cancers, most notably immortalization, hyperproliferation, and dissemination. Despite considerable research into the affected host signaling pathways, the parasite proteins directly responsible for these host phenotypes remain unknown. In this review we outline current knowledge on the manipulation of host cells by transformation-inducing Theileria, and we propose that comparisons between cancer biology and host-Theileria interactions can reveal chemotherapeutic targets against Theileria-induced pathogenesis based on cancer treatment approaches. PMID:25951781

  20. Association of interleukin-6 methylation in leukocyte DNA with serum level and the risk of ischemic heart disease.

    PubMed

    Yang, Qinghui; Zhao, Yushi; Zhang, Zhijie; Chen, Jianxin

    2016-07-01

    Background Interleukin-6 (IL-6), a multifunctional cytokine, plays an important role in the development of ischemic heart disease (IHD), and DNA hypomethylation of 2 CpGs, located downstream in the proximity of the IL-6 gene promoter, has been associated with risk factor for IHD. This study was to examine the association of blood leukocyte DNA methylation of the 2 CpGs in IL-6 with the risk of IHD and the serum IL-6 level. Methods IL-6 methylation levels of 582 cases and 673 controls were measured using the bisulfite pyrosequencing technology. Serum level of IL-6 was measured using enzyme-linked immunosorbent assay. Results The IL-6 methylation was significantly lower in IHD cases than in the controls, irrespective of CpG site. After multivariate adjustment, lower (< median) average IL-6 methylation was associated with an increased risk of IHD (OR 1.57, 95% CI 1.22-2.02, p < 0.001). Average IL-6 methylation level was inversely associated with serum IL-6 level (β = -1.02 pg/mL per increase in IL-6 methylation, p = 0.002) among IHD cases. This significant relationship was not observed among controls. Conclusions DNA hypomethylation of IL-6 gene measured in blood leukocytes was associated with increased risk of IHD. IL-6 demethylation may upregulate its expression, whereby exerting its risk effect on the development of IHD. PMID:26986049

  1. Out of the Water: Origin and Diversification of the LBD Gene Family

    PubMed Central

    Chanderbali, Andre S.; He, Fengmei; Soltis, Pamela S.; Soltis, Douglas E.

    2015-01-01

    LBD (LATERAL ORGAN BOUNDARIES DOMAIN) genes are essential to the developmental programs of many fundamental plant organs and function in some of the basic metabolic pathways of plants. However, our historical perspective on the roles of LBD genes during plant evolution has, heretofore, been fragmentary. Here, we show that the LBD gene family underwent an initial radiation that established five gene lineages in the ancestral genome of most charophyte algae and land plants. By inference, the LBD gene family originated after the emergence of the green plants (Viridiplantae), but prior to the diversification of most extant streptophytes. After this initial radiation, we find limited instances of gene family diversification in land plants until successive rounds of expansion in the ancestors of seed plants and flowering plants. The most dynamic phases of LBD gene evolution, therefore, trace to the aquatic ancestors of embryophytes followed by relatively recent lineage-specific expansions on land. PMID:25839188

  2. Out of the Water: Origin and Diversification of the LBD Gene Family.

    PubMed

    Chanderbali, Andre S; He, Fengmei; Soltis, Pamela S; Soltis, Douglas E

    2015-08-01

    LBD (lateral organ boundaries domain) genes are essential to the developmental programs of many fundamental plant organs and function in some of the basic metabolic pathways of plants. However, our historical perspective on the roles of LBD genes during plant evolution has, heretofore, been fragmentary. Here, we show that the LBD gene family underwent an initial radiation that established five gene lineages in the ancestral genome of most charophyte algae and land plants. By inference, the LBD gene family originated after the emergence of the green plants (Viridiplantae), but prior to the diversification of most extant streptophytes. After this initial radiation, we find limited instances of gene family diversification in land plants until successive rounds of expansion in the ancestors of seed plants and flowering plants. The most dynamic phases of LBD gene evolution, therefore, trace to the aquatic ancestors of embryophytes followed by relatively recent lineage-specific expansions on land. PMID:25839188

  3. Leukocyte Responsiveness to Exercise in Individuals Positive for Human Cytomegalovirus.

    PubMed

    Wilson, J N; Navalta, J W

    2016-05-01

    Human cytomegalovirus (HCMV) infects 50% of adults in the United States. HCMV can become a cause for concern in individuals who have a compromised immune system, which may occur after high-intensity exercise. The purpose of this preliminary study was to characterize the lymphocyte, monocyte, and neutrophil responses to exercise in HCMV+individuals. Participants were either positive (HCMV +) or negative (HCMV-) for HCMV. Participants visited the laboratory on 3 separate occasions: HCMV screening, 100% VO2max test, and 80% VO2max run. Mixed-model factorial ANOVA procedures with repeated measures on sampling condition were performed on absolute and relative circulating lymphocytes, monocytes, and neutrophils. Significant main effects for time for both absolute and relative values were seen for all leukocyte subsets regardless of virus status. Significant differences for absolute and relative values were seen between sampling conditions for all leukocyte subsets. We report for the first time that HCMV status does not affect circulating neutrophil responses to high-intensity exercise, though exercise-induced neutrocytosis is seen during the post-exercise and 60 min post-exercise sampling conditions, regardless of HCMV status. There is no HCMV effect on circulating monocyte responses to exercise, though exercise-induced monocytosis was seen during the post-exercise sampling condition regardless of HCMV status. PMID:26837931

  4. The history of fever, leukocytic pyrogen and interleukin-1

    PubMed Central

    Dinarello, Charles A

    2015-01-01

    There has been great progress in the 30 y since the reporting in 1984 of the cDNA for interleukin1 (IL1) β in the human and IL1α in the mouse. However, the history of IL1 begins in the early 1940s with investigations into the nature of an endogenous fever-producing protein released rabbit peritoneal neutrophils. Most researchers in immunology today are unaware that the field of cytokines, particularly the field of inflammatory cytokines. Toll-like receptors and innate immunity traces back to studies on fever. Researchers in infectious diseases wanted to know about an endogenous protein that caused fever, independent of infection. The endogenous fever-producing protein was called by various names: granulocyte, endogenous or leukocytic pyrogen. It is a fascinating and sometimes controversial story for biology and medicine and for the treatment of inflammatory diseases. Few imagined that this fever-producing protein would play such a major role in nearly every cell and in most diseases. This paper reviews the true background and milestones of interleukin1 from the purification of leukocytic pyrogen to the first cDNA of IL1β and the validation of cytokine biology from ill-defined factors to its present day importance.

  5. Electromagnetic wave emitting products and "Kikoh" potentiate human leukocyte functions.

    PubMed

    Niwa, Y; Iizawa, O; Ishimoto, K; Jiang, X; Kanoh, T

    1993-09-01

    Tourmaline (electric stone, a type of granite stone), common granite stone, ceramic disks, hot spring water and human palmar energy (called "Kikoh" in Japan and China), all which emit electromagnetic radiation in the far infrared region (wavelength 4-14 microns). These materials were thus examined for effects on human leukocyte activity and on lipid peroxidation of unsaturated fatty acids. It was revealed that these materials significantly increased intracellular calcium ion concentration, phagocytosis, and generation of reactive oxygen species in neutrophils, and the blastogenetic response of lymphocytes to mitogens. Chemotactic activity by neutrophils was also enhanced by exposure to tourmaline and the palm of "Kikohshi" i.e., a person who heals professionally by the laying on of hands. Despite the increase in reactive oxygen species generated by neutrophils, lipid peroxidation from unsaturated fatty acid was markedly inhibited by these four materials. The results suggest that materials emitting electromagnetic radiation in the far infrared range, which are widely used in Japan for cosmetic, therapeutic, and preservative purposes, appear capable of potentiating leukocyte functions without promoting oxidative injury. PMID:8406976

  6. Host-pathogen interactions: leukocyte phagocytosis and associated sequelae.

    PubMed

    Voyich, Jovanka M; DeLeo, Frank R

    2002-01-01

    Polymorphonuclear leukocytes (PMNs) are a critical component of the human innate immune response and are the first line of defense against invading microorganisms. Phagocytosis of invading microbes induces production of reactive oxygen species (ROS) by PMNs, which facilitates bactericidal activity. In addition to eliminating microorganisms, phagocytosis also accelerates PMN apoptosis, a process critical for resolution of inflammation. Inasmuch as leukocyte phagocytosis and ROS production are key components of the innate immune response, we developed flow cytometric methods to evaluate these processes in human PMNs. In contrast to traditional microscopy-based analyses, the methods described herein provide objective and high throughput measures of host cell-pathogen interactions. Importantly, they can be adapted for use with a number of fluorometric probes, and bacterium and host cell of choice, and each is based upon a common phagocytosis assay system. We also describe methods to measure phagocytosis-induced PMN apoptosis with this assay system. These methods entail detecting surface-exposed phosphatidylserine (early apoptosis), and measuring PMN chromatin condensation and DNA fragmentation (late apoptosis). Taken together, these assays provide rapid and accurate assessment of critical PMN processes. PMID:12815296

  7. Leukocyte subsets and neutrophil function after short-term spaceflight

    NASA Technical Reports Server (NTRS)

    Stowe, R. P.; Sams, C. F.; Mehta, S. K.; Kaur, I.; Jones, M. L.; Feeback, D. L.; Pierson, D. L.

    1999-01-01

    Changes in leukocyte subpopulations and function after spaceflight have been observed but the mechanisms underlying these changes are not well defined. This study investigated the effects of short-term spaceflight (8-15 days) on circulating leukocyte subsets, stress hormones, immunoglobulin levels, and neutrophil function. At landing, a 1.5-fold increase in neutrophils was observed compared with preflight values; lymphocytes were slightly decreased, whereas the results were variable for monocytes. No significant changes were observed in plasma levels of immunoglobulins, cortisol, or adrenocorticotropic hormone. In contrast, urinary epinephrine, norepinephrine, and cortisol were significantly elevated at landing. Band neutrophils were observed in 9 of 16 astronauts. Neutrophil chemotactic assays showed a 10-fold decrease in the optimal dose response after landing. Neutrophil adhesion to endothelial cells was increased both before and after spaceflight. At landing, the expression of MAC-1 was significantly decreased while L-selectin was significantly increased. These functional alterations may be of clinical significance on long-duration space missions.

  8. Early and delayed indium 111 leukocyte imaging in Crohn's disease

    SciTech Connect

    Navab, F.; Boyd, C.M.; Diner, W.C.; Subramani, R.; Chan, C.

    1987-10-01

    Twenty-seven patients with Crohn's disease were studied for the presence and location of activity by both early (4 h) and delayed (18-24 h) indium 111 leukocyte imaging. The results were compared with other parameters of disease activity including Crohn's disease activity index, barium studies, and endoscopy. There was a correlation between early images and Crohn's disease activity index (r = 0.78) and between delayed images and index (r = 0.82). Based upon the corresponding Crohn's disease activity index, the sensitivity of early and delayed imaging was 81.0% and 95.2%, respectively. Specificity of early and delayed imaging was 75.0% and 87.0%, respectively. Presence of activity on the early and delayed imaging agreed with activity on barium studies and colonoscopy in approximately 80% of cases. Correlation of location of disease by leukocyte imaging and x-ray was observed in 58.9% of early scans and 55.0% of delayed scans. Correlation of the location of disease by imaging and endoscopy was observed in 71.4% of early and 75.0% of delayed studies. Because of the possibility of occurrence of false-negative results in early images, delayed imaging should always be included in evaluation of disease activity in patients with Crohn's disease who are suspected of having mild activity. Delayed imaging is not required if the early imaging study clearly shows activity.

  9. Modulation of Leukocyte Behavior by an Inflamed Extracellular Matrix

    PubMed Central

    Schor, Hagai; Vaday, Gayle G.

    2000-01-01

    Inflammation is a response of the immune system to foreign insult or physical damage. Various cellular and humoral components of the immune system are recruited from the vascular system and are translocated through endothelium, and into extracellular matrix (ECM) compartments of inflamed tissues. This translocation is orchestrated by various types of accessory signals, in the form of soluble or complexed molecules, which evoke remarkable transitions in leukocyte activities. Recruited inflammatory cells give rise to mechanisms of migration, including the secretion of enzymes and other pro-inflammatory mediators and the alteration of their adhesive contacts with the ECM. Hence, migrating cells secrete enzymes, chemokines, and cytokines which interact with the ECM, and thereby, provide the cells with intrinsic signals for coordinating their responses. Resultant products of enzymatic modifications to the ECM microenvironment, such as cytokine- and ECM-derived molecules, may be also part of a cell-signaling mechanism that provides leukocytes with information about the nature of their inflammatory activity; such a mechanism may give the immune system data that can be cognitively interpreted for consequential activities. This article reviews the findings that support this notion and describe the dynamic interactions between participants of the inflammatory processes. PMID:11097214

  10. Interactions between CD44 and Hyaluronan in Leukocyte Trafficking

    PubMed Central

    McDonald, Braedon; Kubes, Paul

    2015-01-01

    Recruitment of leukocytes from the bloodstream to inflamed tissues requires a carefully regulated cascade of binding interactions between adhesion molecules on leukocytes and endothelial cells. Adhesive interactions between CD44 and hyaluronan (HA) have been implicated in the regulation of immune cell trafficking within various tissues. In this review, the biology of CD44–HA interactions in cell trafficking is summarized, with special attention to neutrophil recruitment within the liver microcirculation. We describe the molecular mechanisms that regulate adhesion between neutrophil CD44 and endothelial HA, including recent evidence implicating serum-derived hyaluronan-associated protein as an important co-factor in the binding of HA to CD44 under flow conditions. CD44–HA-mediated neutrophil recruitment has been shown to contribute to innate immune responses to invading microbes, as well as to the pathogenesis of many inflammatory diseases, including various liver pathologies. As a result, blockade of neutrophil recruitment by targeting CD44–HA interactions has proven beneficial as an anti-inflammatory treatment strategy in a number of animal models of inflammatory diseases. PMID:25741341

  11. Indium-/sup 111/ leukocyte imaging in appendicitis

    SciTech Connect

    Navarro, D.A.; Weber, P.M.; Kang, I.Y.; dos Remedios, L.V.; Jasko, I.A.; Sawicki, J.E.

    1987-04-01

    Indium-/sup 111/-labeled leukocyte scintigraphy was applied to the diagnosis of acute appendicitis. Thirty-two patients observed in the hospital for possible appendicitis were prospectively studied. Scanning was done 2 hr after radiopharmaceutical injection. Thirteen scans were positive for acute appendicitis, and all but one were confirmed at laparotomy. In addition, two cases of colitis and two cases of peritonitis were detected. Of 15 negative studies, 11 had a benign course. Four patients with negative studies had laparotomy; two were found to have appendicitis and two had a normal appendix. Of 14 proven cases of appendicitis, 12 scans were positive for appendicitis with one false-positive scan, providing a sensitivity of 86%. Specificity was 93%: all negative cases except one had negative scans. Overall accuracy was 91% (29 of 32), comparing favorably with the accepted false-positive laparotomy rate of 25%. Use of In-/sup 111/-labeled leukocyte scintigraphy serves to reduce the false-positive laparotomy rate and to shorten the clinical observation time in patients with acute appendicitis.

  12. Leukocyte responses to immobilized patterns of CXCL8.

    PubMed

    Girrbach, Maria; Rink, Ina; Ladnorg, Tatjana; Azucena, Carlos; Heißler, Stefan; Haraszti, Tamás; Schepers, Ute; Schmitz, Katja

    2016-06-01

    The attachment of neutrophils to the endothelial surface and their migration towards the site of inflammation following chemokine gradients play an essential role in the innate immune response. Chemokines adhere to glycosaminoglycans on the endothelial surface to be detected by leukocytes and trigger their movement along surface- bound gradients in a process called haptotaxis. In assays to systematically study the response of leukocytes to surface-bound compounds both the spatial arrangement of the compound as well as the mode of immobilization need to be controlled. In this study microcontact printing was employed to create patterns of hydrophobic or functionalized thiols on gold-coated glass slides and CXCL8 was immobilized on the thiol coated areas using three different strategies. Human neutrophils adhered to the CXCL8-coated lines but not to the PEG-coated background. We could show that more cells adhered to CXCL8 adsorbed to hydrophobic octadecanethiol than on CXCL8 covalently bound to amino undecanethiol or CXCL8 specifically bound to immobilized heparin on aminothiol. Likewise general cell activity such as lamellipodia formation and random migration were most pronounced for CXCL8 adsorbed on a hydrophobic surface which may be attributed to the larger amounts of protein immobilized on this type of surface. PMID:26970827

  13. Mononuclear Leukocyte Infiltrate in Extraplacental Membranes and Preterm Delivery

    PubMed Central

    Holzman, Claudia; Senagore, Patricia K.; Wang, Jianling

    2013-01-01

    Large numbers of polymorphonuclear leukocytes in the amnion and chorion define histological chorioamnionitis (HCA), a condition linked to spontaneous preterm delivery (PTD). Less is known about placental patterns of mononuclear leukocyte (MNL) density and PTD. In this prospective study (1998–2004), women were sampled from 52 clinics in 5 Michigan communities and enrolled at 16–27 weeks’ gestation. HCA and MNL distributions in delivered placentas were evaluated microscopically in a subcohort (290 preterm, 823 term). Midpregnancy biomarkers from maternal blood (i.e., C-reactive protein (CRP), corticotropin-releasing hormone, and cytokines) were compared among term and PTD subjects grouped by presence/absence of HCA and high MNL density. A density of more than 10 MNLs per high-power field in the chorion of the membrane roll, referred to as MNL-CMR, was associated with medically indicated PTD (odds ratio = 2.2, 95% confidence interval: 1.3, 3.6) and spontaneous PTD (odds ratio = 2.5, 95% confidence interval: 1.7, 3.7). Associations persisted after removal of women with HCA-positive placentas, abruption, hypertensive disorders, or obesity. HCA-associated PTD showed higher CRP and cytokine levels. MNL-CMR-associated PTD showed higher CRP and corticotropin-releasing hormone levels. These data suggest that an MNL infiltrate in the chorion of the membrane roll marks PTD pathways that are distinct from HCA and not entirely explained by pregnancy complications. PMID:23429723

  14. Leukocyte-specific protein 1 regulates T-cell migration in rheumatoid arthritis.

    PubMed

    Hwang, Seong-Hye; Jung, Seung-Hyun; Lee, Saseong; Choi, Susanna; Yoo, Seung-Ah; Park, Ji-Hwan; Hwang, Daehee; Shim, Seung Cheol; Sabbagh, Laurent; Kim, Ki-Jo; Park, Sung Hwan; Cho, Chul-Soo; Kim, Bong-Sung; Leng, Lin; Montgomery, Ruth R; Bucala, Richard; Chung, Yeun-Jun; Kim, Wan-Uk

    2015-11-24

    Copy number variations (CNVs) have been implicated in human diseases. However, it remains unclear how they affect immune dysfunction and autoimmune diseases, including rheumatoid arthritis (RA). Here, we identified a novel leukocyte-specific protein 1 (LSP1) deletion variant for RA susceptibility located in 11p15.5. We replicated that the copy number of LSP1 gene is significantly lower in patients with RA, which correlates positively with LSP1 protein expression levels. Differentially expressed genes in Lsp1-deficient primary T cells represent cell motility and immune and cytokine responses. Functional assays demonstrated that LSP1, induced by T-cell receptor activation, negatively regulates T-cell migration by reducing ERK activation in vitro. In mice with T-cell-dependent chronic inflammation, loss of Lsp1 promotes migration of T cells into the target tissues as well as draining lymph nodes, exacerbating disease severity. Moreover, patients with RA show diminished expression of LSP1 in peripheral T cells with increased migratory capacity, suggesting that the defect in LSP1 signaling lowers the threshold for T-cell activation. To our knowledge, our work is the first to demonstrate how CNVs result in immune dysfunction and a disease phenotype. Particularly, our data highlight the importance of LSP1 CNVs and LSP1 insufficiency in the pathogenesis of RA and provide previously unidentified insights into the mechanisms underlying T-cell migration toward the inflamed synovium in RA. PMID:26554018

  15. LRP5 and plasma cholesterol levels modulate the canonical Wnt pathway in peripheral blood leukocytes.

    PubMed

    Borrell-Pages, Maria; Carolina Romero, July; Badimon, Lina

    2015-08-01

    Inflammation is triggered after invasion or injury to restore homeostasis. Although the activation of Wnt/β-catenin signaling is one of the first molecular responses to cellular damage, its role in inflammation is still unclear. It was our hypothesis that the low-density lipoprotein (LDL) receptor-related protein 5 (LRP5) and the canonical Wnt signaling pathway are modulators of inflammatory mechanisms. Wild-type (WT) and LRP5(-/-) mice were fed a hypercholesterolemic (HC) diet to trigger dislipidemia and chronic inflammation. Diets were supplemented with plant sterol esters (PSEs) to induce LDL cholesterol lowering and the reduction of inflammation. HC WT mice showed increased serum cholesterol levels that correlated with increased Lrp5 and Wnt/β-catenin gene expression while in the HC LRP5(-/-) mice Wnt/β-catenin pathway was shut down. Functionally, HC induced pro-inflammatory gene expression in LRP5(-/-) mice, suggesting an inhibitory role of the Wnt pathway in inflammation. Dietary PSE administration downregulated serum cholesterol levels in WT and LRP5(-/-) mice. Furthermore, in WT mice PSE increased anti-inflammatory genes expression and inhibited Wnt/β-catenin activation. Hepatic gene expression of Vldlr, Lrp2 and Lrp6 was increased after HC feeding in WT mice but not in LRP5(-/-) mice, suggesting a role for these receptors in the clearance of plasmatic lipoproteins. Finally, an antiatherogenic role for LRP5 was demonstrated as HC LRP5(-/-) mice developed larger aortic atherosclerotic lesions than WT mice. Our results show an anti-inflammatory, pro-survival role for LRP5 and the Wnt signaling pathway in peripheral blood leukocytes. PMID:25748163

  16. Beneficial effects of secretory leukocyte protease inhibitor after spinal cord injury

    PubMed Central

    Ghasemlou, Nader; Bouhy, Delphine; Yang, Jingxuan; López-Vales, Rubèn; Haber, Michael; Thuraisingam, Thusanth; He, Guoan; Radzioch, Danuta; Ding, Aihao

    2010-01-01

    Secretory leukocyte protease inhibitor is a serine protease inhibitor produced by various cell types, including neutrophils and activated macrophages, and has anti-inflammatory properties. It has been shown to promote wound healing in the skin and other non-neural tissues, however, its role in central nervous system injury was not known. We now report a beneficial role for secretory leukocyte protease inhibitor after spinal cord injury. After spinal cord contusion injury in mice, secretory leukocyte protease inhibitor is expressed primarily by astrocytes and neutrophils but not macrophages. We show, using transgenic mice over-expressing secretory leukocyte protease inhibitor, that this molecule has an early protective effect after spinal cord contusion injury. Furthermore, wild-type mice treated for the first week after spinal cord contusion injury with recombinant secretory leukocyte protease inhibitor exhibit sustained improvement in locomotor control and reduced secondary tissue damage. Recombinant secretory leukocyte protease inhibitor injected intraperitoneally localizes to the nucleus of circulating leukocytes, is detected in the injured spinal cord, reduces activation of nuclear factor-κB and expression of tumour necrosis factor-α. Administration of recombinant secretory leukocyte protease inhibitor might therefore be useful for the treatment of acute spinal cord injury. PMID:20047904

  17. Leukocyte Cell Surface Proteinases: Regulation of Expression, Functions, and Mechanisms of Surface Localization

    PubMed Central

    Owen, Caroline A.

    2008-01-01

    A number of proteinases are expressed on the surface of leukocytes including members of the serine, metallo-, and cysteine proteinase superfamilies. Some proteinases are anchored to the plasma membrane of leukocytes by a transmembrane domain or a glycosyl phosphatidyl inositol (GPI) anchor. Other proteinases bind with high affinity to classical receptors, or with lower affinity to integrins, proteoglycans, or other leukocyte surface molecules. Leukocyte surface levels of proteinases are regulated by: 1) cytokines, chemokines, bacterial products, and growth factors which stimulate synthesis and/or release of proteinase by cells; 2) the availability of surface binding sites for proteinases; and/or 3) internalization or shedding of surface-bound proteinases. The binding of proteinases to leukocyte surfaces serves many functions including: 1) concentrating the activity of proteinases to the immediate pericellular environment; 2) facilitating pro-enzyme activation; 3) increasing proteinase stability and retention in the extracellular space; 4) regulating leukocyte function by proteinases signaling through cell surface binding sites or other surface proteins; and 5) protecting proteinases from inhibition by extracellular proteinase inhibitors. There is strong evidence that membrane-associated proteinases on leukocytes play critical roles in wound healing, inflammation, extracellular matrix remodeling, fibrinolysis, and coagulation. This review will outline the biology of membrane-associated proteinases expressed by leukocytes and their roles in physiologic and pathologic processes. PMID:18329945

  18. A simple rapid method for the removal of leukocytes from human blood.

    PubMed

    Palmer, E; Waldman, F; Dewitt, W

    1974-01-01

    Filtration of human blood cells through lamb's wool columns removed more than 96% of all leukocytes in a series of experiments, while the retention of erythrocytes by the column averaged 6.4%. This method should prove extremely useful for obtaining pure erythrocyte preparations for use in biochemical and physiological studies, and for removing leukocytes from blood prior to transfusion. PMID:4473869

  19. Seasonal variation of peripheral blood leukocyte telomere length in Costa Rica: a population based observational study

    PubMed Central

    Rehkopf, David H; Dow, William H; Rosero-Bixby, Luis; Lin, Jue; Epel, Elissa S; Blackburn, Elizabeth H

    2014-01-01

    Objectives Peripheral blood leukocyte telomere length is increasingly being used as a biomarker of aging, but its natural variation in human populations is not well understood. Several other biomarkers show seasonal variation, as do several determinants of leukocyte telomere length. We examined whether there was monthly variation in leukocyte telomere length in Costa Rica, a country with strong seasonal differences in precipitation and infection. Methods We examined a longitudinal population based cohort of 581 Costa Rican adults age 60 and above, from which blood samples were drawn between October 2006 and July 2008. Leukocyte telomere length was assayed from these samples using the quantitative PCR method. Multivariate regression models were used to examine correlations between month of blood draw and leukocyte telomere length. Results Telomere length from peripheral blood leukocytes varied by as much as 200 base pairs depending on month of blood draw, and this difference is not likely to be due to random variation. A moderate proportion of this association is statistically accounted for by month and region specific average rainfall. We found shorter telomere length associated with greater rainfall. Conclusions There are two possible explanations of our findings. First, there could be relatively rapid month-to-month changes in leukocyte telomere length. This conclusion would have implications for understanding the natural population dynamics of telomere length. Second, there could be seasonal differences in constituent cell populations. This conclusion would suggest that future studies of leukocyte telomere length use methods to account for the potential impact of constituent cell type. PMID:24615938

  20. Imaging of vascular development in early mouse decidua and its association with leukocytes and trophoblasts.

    PubMed

    Croy, B Anne; Chen, Zhilin; Hofmann, Alexander P; Lord, Edith M; Sedlacek, Abigail L; Gerber, Scott A

    2012-11-01

    In species with endometrial decidualization and hemochorial placentation (humans, mice, and others), leukocytes localize to early implant sites and contribute to decidual angiogenesis, spiral arterial remodeling, and trophoblast invasion. Relationships between leukocytes, trophoblasts, and the decidual vasculature are not fully defined. Early C57BL/6J implant sites were analyzed by flow cytometry to define leukocyte subsets and by whole-mount immunohistochemistry to visualize relationships between leukocytes, decidual vessels, and trophoblasts. Ptprc(+) (CD45(+)) cells increased in decidua between Gestational Day (GD) 5.5 and GD 9.5. Uterine natural killer (uNK) cells that showed dynamic expression of Cd (CD) 69, an activating receptor, and Klrg1 (KLRG1), an inhibitory receptor, localized mesometrially and were the dominant CD45(+) cells between GD 5.5 and GD 7.5. At GD 8.5, immature monocytes that occurred throughout decidua exceeded uNK cells numerically and many leukocytes acquired irregular shapes, and leukocyte-leukocyte conjugates became frequent. Vessels were morphologically heterogeneous and regionally unique. Migrating trophoblasts were first observed at GD 6.5 and, at GD 9.5, breached endothelium, entered vascular lumens, and appeared to occlude some vessels, as described for human spiral arteries. No leukocyte-trophoblast conjugates were detected. Whole-mount staining gave unparalleled decidual vascular detail and cell-specific positional information. Its application across murine models of pregnancy disturbances should significantly advance our understanding of the maternal-fetal interface. PMID:22954796

  1. Radiation-induced normal tissue injury: role of adhesion molecules in leukocyte-endothelial cell interactions.

    PubMed

    Quarmby, S; Kumar, P; Kumar, S

    1999-07-30

    The late onset of necrosis and fibrosis in normal tissues can be a serious consequence of radiotherapy in cancer patients. Because radiation-induced vascular injury precedes the tissue damage, vascular injury is regarded as crucial in the pathogenesis of tissue damage. An understanding of the processes responsible is essential to develop strategies for the amelioration of radiation-induced normal tissue damage. Leukocyte infiltration is commonly observed at sites of irradiation and is likely to lead to the acceleration and/or induction of parenchymal atrophy, fibrosis and necrosis in normal tissues following radiotherapy. The molecular mechanisms mediating leukocyte infiltration of tissues during inflammation have been studied extensively. It is now well established that cell adhesion molecules (CAMs) expressed on leukocytes and endothelial cells control the trafficking of leukocytes from the blood vessel lumen in these conditions. CAMs including E (endothelial), P (platelet) and L (leukocyte)-selectins, intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), beta1 and beta2 integrins and CD31 are involved in the cascade of events resulting in rolling, arrest and transmigration of leukocytes through the inflamed endothelium. Whether a similar sequence of molecular events induces leukocyte sequestration in irradiated normal tissues is not known. This review is focussed on the role of CAMs in radiation-induced leukocyte infiltration of normal tissues and the therapeutic implications of these findings. PMID:10399956

  2. Receptor cleavage and P-selectin-dependent reduction of leukocyte adhesion in the spontaneously hypertensive rat.

    PubMed

    Chen, Angela Y; Ha, Jessica N; Delano, Frank A; Schmid-Schönbein, Geert W

    2012-07-01

    The SHR, a genetic model for hypertension and the metabolic syndrome, has attenuated leukocyte adhesion to the postcapillary endothelium by an unknown mechanism. Based on recent evidence of elevated levels of MMPs in plasma and on microvascular endothelium of the SHR with cleavage of several receptor types, we hypothesize that the reduced leukocyte-endothelial interaction is a result of enhanced proteolytic cleavage of P-selectin on the postcapillary endothelium and PSGL-1 on leukocytes. The attenuated rolling interactions of SHR leukocytes with the endothelium were restored by chronic treatment with a broad-spectrum MMP inhibitor (CGS) for 24 weeks. The SHR MMP levels, in plasma and mesentery, as well as the systolic blood pressure, decreased significantly with treatment. In the SHR mesentery, labeling of P-selectin in the postcapillary venules by immunohistochemistry demonstrated, on average, a 31% lower extracellular P-selectin density compared with the normotensive WKY. A significantly lower extracellular PSGL-1 density on the membranes of SHR neutrophils compared with the WKY also supported our hypothesis. In vivo stimulation of the mesenteric postcapillary venules with histamine demonstrated that the SHR had an attenuated response, as measured by leukocyte rolling velocity on the endothelium. The reduced P-selectin and PSGL-1 density, on SHR postcapillary endothelium and on SHR leukocytes, respectively, was restored significantly by chronic MMP inhibition. The impaired ability of SHR leukocytes to reduce rolling velocity upon inflammatory stimulation led to fewer firmly adhered leukocytes to the endothelium as a contributor to immune suppression. PMID:22566571

  3. HARVESTING OF LEUKOCYTES FROM INTESTINAL LUMEN IN MURINE GIARDIASIS AND PRELIMINARY CHARACTERIZATION OF THESE CELLS

    EPA Science Inventory

    The aims of the study were to develop a method for harvesting leukocytes from the mouse small intestinal lumen and to identify leukocytes which enter the intestinal lumen of mice infected with Giardia muris. Giardia-infected and uninfected BALB/c mice were anesthetized, and the s...

  4. Fragmentation of gelatin-bound fibronectin (Fn) by inflammatory polymorphonuclear leukocytes